On 2015 Jul 23, James M Heilman commented:

You sort of missed the point. The point is that Wikipedia is incredibly frequently read. This includes by the lay public, medical students, and physicians. One can warn people until they are blue in the face but I doubt it is going to change how frequently they use Wikipedia for medical information. In my opinion time is much better spent trying to improve the site. It really is not that difficult.

On 2015 May 11, David Gortler commented:

I have to say that although this data sounds promising, I would never use wikipedia as an authoritative resource for anything patient-related, and firmly instill warnings regarding wikipedia to my students as well. I was given a moderator account at one point and took time and effort to carefully compose several specialty pharmacology articles, only to have those articles edited incorrectly, filled with obvious biases, or inexplicably "re-organized" to the point that they were essentially vandalized. I had offenders blocked or banned, only to have scores more crawl out of the bowels of the internet to do the same. I have since stopped trying to correctly edit or even view my articles for accuracy.

The last time I used wikipedia was years ago and I think it was to look up something about vintage automobiles. I also looked up Gilligan's Island on it around the same time. That's about as far as I would trust it. Anyone who relies on wikipedia for anything clinically related is placing patient lives at risk. Anyone who relies on wikipedia to look up anything or research related is wasting their time and money.

On 2015 May 10, James M Heilman commented:

Yes looking at top editors I know most of those listed. A proportion are those who write a great deal of content and mostly work on medicine. Another proportion are those who do maintenance and vandalism reversion Wikipedia wide. What we are capturing is only a proportion of their edits as we just looked at medical editors. If we were to look at Wikipedia as a whole they would far surpass all those listed here.

So I agree on a global scale the top editors are more involved in maintenance. However within a specific area the top editors are more content contributors (there are of course still maintence people aswell).

On 2015 May 08, Gwinyai Masukume commented:

Thank you for engaging with this and making the data publicly available.

Five users appear on both lists (Top ten by number of bytes changed and Top ten by number of edits) and only one user is in the same position.

Perhaps the situation is a hybrid of Jimmy Wales’ and Aaron Swartz’s thoughts, the degree differing depending on the context. Looking forward to further insights from your data.

On 2015 May 13, James M Heilman commented:

And now we have data for 2014 https://en.wikipedia.org/wiki/Wikipedia:WikiProject_Medicine/Stats/Top_English_medical_editors_2014

On 2015 May 07, James M Heilman commented:

We now have data looking at the top editors for 2013 based on bytes changed here https://en.wikipedia.org/wiki/Wikipedia:WikiProject_Medicine/Stats/Top_English_medical_editors_2013b

There is a fairly good relationship with top editors for 2013 based on number of edits here https://en.wikipedia.org/wiki/Wikipedia:WikiProject_Medicine/Stats/Top_English_medical_editors_2013

So my analysis is more in line with the thoughts of Jimmy Wales rather than Aaron Swartz.

On 2015 Mar 26, James M Heilman commented:

Often people with the highest edit count also have the largest number of bytes changed.

One would really want to look at the persistent bytes contributed by a user. One also needs to keep in mind that more text is not necessarily better text. Lots of improvements involved trimming material.

Agree that their is no perfect measure of "authorship" which can be carried out in an automated fashion. Will look at running the analysis you suggest and posting the results here.

On 2015 Mar 21, Gwinyai Masukume commented:

The authors use the number of edits made to measure participation. An edit, for example, can be adding a full stop/period “.” or it can be adding a whole paragraph. Each of these actions would be counted as one edit.

Another way to measure participation is by considering the amount of text added by a contributor. This approach of considering the amount of text added has in the past yielded different results and insights compared to considering the number of edits made. Some contributors with relatively low edit counts have been found to have added a relatively high amount of text.

I am of the view that only using edit counts is a limitation. This limitation could have been remedied by considering the amount of text added by contributors from a limited number of randomly selected articles from the authors' sampling frame.

Reference

Swartz A. Who writes Wikipedia? 2006. Available: http://www.aaronsw.com/weblog/whowriteswikipedia (accessed 21 March 2015).

On 2016 Aug 23, Ben Goldacre commented:

This trial has the wrong trial registry ID associated with it on PubMed: both in the XML on PubMed, and in the originating journal article. The ID given is NCT0157384. We believe the correct ID, which we have found by hand searching, is NCT01573845.

This comment is being posted as part of the OpenTrials.net project^[1] , an open database threading together all publicly accessible documents and data on each trial, globally. In the course of creating the database, and matching documents and data sources about trials from different locations, we have identified various anomalies in datasets such as PubMed, and in published papers. Alongside documenting the prevalence of problems, we are also attempting to correct these errors and anomalies wherever possible, by feeding back to the originators. We have corrected this data in the OpenTrials.net database; we hope that this trial’s text and metadata can also be corrected at source, in PubMed and in the accompanying paper.

Many thanks,

Jessica Fleminger, Ben Goldacre*

[1] Goldacre, B., Gray, J., 2016. OpenTrials: towards a collaborative open database of all available information on all clinical trials. Trials 17. doi:10.1186/s13063-016-1290-8 PMID: 27056367

* Dr Ben Goldacre BA MA MSc MBBS MRCPsych<br> Senior Clinical Research Fellow<br> ben.goldacre@phc.ox.ac.uk<br> www.ebmDataLab.net<br> Centre for Evidence Based Medicine<br> Department of Primary Care Health Sciences<br> University of Oxford<br> Radcliffe Observatory Quarter<br> Woodstock Road<br> Oxford OX2 6GG

On 2017 Jul 13, Randi Pechacek commented:

Bubba Brooks, co-author of this paper, wrote a blog on microBEnet giving background and commentary.

On 2015 Apr 21, David Keller commented:

Clinical recommendations should not be biased by economic considerations

These recommendations for treatment of pressure ulcers ("bed sores") ignore the evidence in favor of certain mattress types, simply because of cost considerations. For example, air-fluidized beds have moderate quality evidence of reducing pressure ulcer size, compared with other surfaces, leading to an improved overall treatment effect (see Table 3 of this paper). In trying to understand why air-fluidized beds were not included in the recommended treatments for pressure ulcers, I noted that the section on "High Value Care" states that "the use of advanced support surfaces adds unnecessary costs to the health care systems". However, the evidence demonstrating improvement due to electrical stimulation was of similar quality ("moderate") as air-fluidized beds. Electrical stimulation was included in the recommended treatments for pressure ulcers. Is electrical stimulation really less expensive than an air-fluidized bed, after accounting for the professional fees charged by the clinician providing that service? How many communities have such a clinician? The cost of air-fluidized beds, like any other commodity, is flexible and based on the volume of orders received. A recommendation by ACP for air-fluidized beds would increase the number of orders and decrease the cost of such beds, due to economies of scale. Economic considerations should not bias clinical recommendations. We are physicians, not economists. We should make recommendations purely on the basis of clinical benefits, and leave the economic analysis to experts in that field, who are more qualified to provide such analysis.

On 2015 Sep 18, Sacha Laurent commented:

I've written an opinion piece on my blog where I discuss the findings of this paper and their impact on the "evolutionary success polyploids" debate. I'll be happy to get any feedback or opinion. Yours,

On 2016 Jan 01, Jaime A. Teixeira da Silva commented:

I have self-published my views on this paper.

Teixeira da Silva, J.A. (2016) Assessing the potentially misleading nature of metrics and of those who assess and create them. Self-published 4 pp. https://www.researchgate.net/publication/288835044

On 2016 Jan 11, BSH Cancer Screening, Help-Seeking and Prevention Journal Club commented:

Relevant to our current research goals, the HBRC journal club was interested to review this paper with particular interest in adding to our understanding of the role of descriptive norms in cancer risk-reducing behaviours. The paper demonstrates a strong rationale for investigating the role of descriptive norms in established risk-reducing behaviours, and in uncertain and relatively unknown behaviours. Furthermore, it offers an additional explanation to the reasons why descriptive norm literature is usually inconsistent, or every so often insignificant for health-behaviour change.

The study employed a cross-sectional design that included an opportunistic sample aged 18 to 95, and reported the use of a theoretical framework, known as the Integrative Model of Behavioural Prediction, which strengthened the methodology of the paper. Although the authors acknowledged their decision for defining uncertainty as a limitation, they attributed the uncertainty of risk-reducing behaviours using scientific knowledge rather than public perceptions of the uncertainty of the behaviours. However, we felt that the study might have benefited from the inclusion of other elements such as beliefs about causes of cancer, scientific literacy, or specifically knowledge, and controlling for age to evaluate further associations with descriptive norms and scientific uncertainty.

The journal club also discussed the possibility of controlling for individuals’ current health behaviours as another factor that could influence descriptive norms. We also deliberated on whether the established risk-reducing behaviours are also linked to other chronic illnesses, therefore; there may be differences in motivations to take up those behaviours.

Overall, the HBRC journal club enjoyed reading this article and feel that the paper is an important example of using integrated theoretical models which could improve the way we implement theoretically grounded research.

Conflicts of interest. We report no conflict of interests and note that the comments produced by the group are collective and not the opinion of any one individual.

On 2015 Mar 03, Robert Eibl commented:

Looks like a great, new approach; fascinating.

On 2015 Mar 27, James M Heilman commented:

A great analysis of this paper exists here https://en.wikipedia.org/wiki/Wikipedia:Wikipedia_Signpost/2015-03-25/Recent_research

"A paper in Advances in Physiology Education[2] claims to assess the suitability of Wikipedia's respiratory articles for medical student learning. Forty Wikipedia articles on respiratory topics were sampled on 27 April 2014. These articles were assessed by three researchers with a modified version of the DISCERN tool. Article references were checked for accuracy and typography. Readability was assessed with the Flesch–Kincaid and Coleman–Liau tools.

The paper found a wide range of accuracy scores using the modified DISCERN tool, from 14.67 for "[Nail] clubbing" to 38.33 for "Tuberculosis". Incorrect, incomplete or inconsistent formatting of references were commonly found, although these were not quantified in the paper. Readability of the articles was typically at a college level. On the basis of these findings, the paper declares Wikipedia's respiratory articles as unsuitable for medical students.

The researcher apparently uses an arbitrary unvalidated modification of the DISCERN tool to assess the accuracy of articles. The nature of this modification is not specified; nor is it available at the journal's website as claimed in the paper.

The DISCERN tool does not assess accuracy; rather, it is designed to assess "information about treatment choices specifically for health consumers". As such, the use of this tool is inappropriate to assess the suitability for medical students.

There is no acknowledgement that Wikipedia is an encyclopedia. Several of the DISCERN tool's questions are unsuitable for an encyclopedia. DISCERN questions such as "Does it describe how each treatment works?" and "Does it describe the risks of each treatment?" would be answered on other Wikipedia pages, not on the disease article's page. The author makes an a priori assumption that the medical textbooks used for comparison are perfect sources. The author does not assess those textbooks with the DISCERN tool.

The paper states: "[t]he number of citations from peer-reviewed journals published in the last 5 yr was only 312 (19%)." However this is far superior to the number of citations in the textbooks listed. The chapter on "Neoplasms of the lung" in Harrison's Principles of Internal Medicine (18th ed.) contains no citations at all. Seven sources are listed in its "Further readings" section, of which only one is from the last five years.

The claim that the article on "clubbing ... had no references or external links" is incorrect. On 27 April 2014, Wikipedia's article on "Nail clubbing" had ten references.

Several of the articles are at a rudimentary stage, containing limited information and lacking appropriate references. However two articles, "Lung cancer" and "Diffuse panbronchiolitis", were assessed by Wikipedia's editors at the highest standard and awarded "Featured article" status. Five more articles, "Asthma", "Chronic obstructive pulmonary disease", "Pneumonia", "Pneumothorax" and "Tuberculosis", reached "Good article" standard. These articles are exceptionally detailed, accurate, and well-referenced. Azer's paper makes no mention of the high quality of these articles.

The research uses an unvalidated tool for an inappropriate purpose without applying a suitable comparator, and inevitably draws incorrect conclusions.

Wikipedia is an encyclopedia. It is not a medical textbook; nor is it intended to replace medical textbooks. Rather, it should be used as a starting point by medical students. The quality of an individual article should be quickly assessed by the reader, and information can be confirmed in the references provided. Missing information should be sought from other sources, such as textbooks. Students should be encouraged to use Wikipedia alongside medical textbooks to assist their learning."

It is by User:Axl and is under a CC BY SA license

On 2016 Jul 05, STEPHEN ROPER commented:

Despite the impressive technique and the many fine figures in this paper, the findings regarding the microvascular permeability in taste buds (Fig 4) are deeply flawed. Specifically, the authors purport to show that compounds such as fluorescein injected into the blood stream have ready access to the cells within taste buds (Fig. 4g-i). Regrettably, this figure, and specifically Fig 4h, reveals a serious misconception about taste buds. This micrograph shows fluorescein-laden lingual surface epithelial squames surrounding the taste bud pore. It does NOT show taste cells. The taste bud itself lies below this plane of focus. Because the quantification in Fig 4i is taken from Fig 4h, it thus is correspondingly in error. Compounds injected into the bloodstream indeed have ready access to the stratified lingual epithelium, including superficial layers, as shown by a previous publication (Dando et al, 2015, Am J Physiol Cell Physiol. 308:C21-32. Epub 2014 Sep 10).

In fact, contrary to the conclusions reported here, there is some form of selectively permeable barrier protecting taste bud cells from many blood-borne chemicals (ibid.). Indeed, Fig. 4g of Choi et al may be consistent with such a barrier, assuming that the relatively fluorescein-free central area in this micrograph is a taste bud. [It might merely be the somewhat acellular connective tissue core of the papilla. This would also be relatively free of fluorescein].

On 2015 Mar 18, George McNamara commented:

See also

Assessing the prevalence of mycoplasma contamination in cell culture via a survey of NCBI's RNA-seq archive. Olarerin-George AO, Hogenesch JB. Nucleic Acids Res. 2015; 43(5): 2535-42. doi: 10.1093/nar/gkv136. PMID: 25712092 Mycoplasmas are notorious contaminants of cell culture and can have profound effects on host cell biology by depriving cells of nutrients and inducing global changes in gene expression. Over the last two decades, sentinel testing has revealed wide-ranging contamination rates in mammalian culture. To obtain an unbiased assessment from hundreds of labs, we analyzed sequence data from 9395 rodent and primate samples from 884 series in the NCBI Sequence Read Archive. We found 11% of these series were contaminated (defined as ≥100 reads/million mapping to mycoplasma in one or more samples). Ninety percent of mycoplasma-mapped reads aligned to ribosomal RNA. This was unexpected given 37% of contaminated series used poly(A)-selection for mRNA enrichment. Lastly, we examined the relationship between mycoplasma contamination and host gene expression in a single cell RNA-seq dataset and found 61 host genes (P < 0.001) were significantly associated with mycoplasma-mapped read counts. In all, this study suggests mycoplasma contamination is still prevalent today and poses substantial risk to research quality.

On 2015 Sep 01, Coby Viner commented:

A version of this article is now publicly available from TSpace (University of Toronto's institutional repository). The post-print version remains available from the publisher.

On 2016 Jan 16, Lewis G Halsey commented:

Fay worries that in the real world of data collection, the power of a study is not known in advance. If true, this argument would only compound our own, which is that unless power is very high (>90%) P has surprisingly low repeatability (Halsey et al., 2015), and the power of most studies calculated after the data analysis is far lower than this (Button et al., 2013, Maxwell, 2004). Therefore, researchers would not be able to design their experiment to ensure it has a very high power, and they could not rely on good fortune instead for their experiment to turn out this way.

But anyway, an integral step in testing the null hypothesis generates an estimate of the variance of the pooled population. Using this estimated parameter, obtained as the data are analysed, the researcher can immediately gauge the study’s power. The exact parameters of the population are never known. These parameters are hypothesised and then estimated, with varying certainty, according to the sample that we have. With a limited sample, these estimates can vary substantially each time an experiment is repeated. If our samples, and the estimates they generate, suggest that power is poor, then any P value that we obtain, low or not, is untrustworthy. A small P value is of little import: a repetition of the same study would give another result (our study, figure 4). This is like looking at the world through a pinhole. When the theoretical power is 0.48, P values less than 0.05 are no more likely than P values greater than 0.05. Why get excited if P is <0.01, when the next replicate experiment could give a P of 0.6?

Fay asks if there is a single better measure than P to test the likelihood that the null hypothesis is untenable. First, this brings us back to the nub of the problem - P is only a good test of the null in the ideal circumstances that study power is very high. Second, there are long-held, big concerns about the value of null hypothesis significance testing as a method for analysing and interpreting data (Cohen, 1994).

Lewis G Halsey and Gordon B Drummond

BUTTON, K., IOANNIDIS, J., MOKRYSZ, C., NOSEK, B., FLINT, J., ROBINSON, E. & MUNAFO, M. (2013) Power failure: why small sample size undermines the reliability of neuroscience. Nature Reviews Neuroscience, 14, 365-376. COHEN, J. (1994) The Earth is round (p < 0.05). American Psychologist 49, 997-1003. HALSEY, L., CURRAN-EVERETT, D., VOWLER, S. & DRUMMOND, G. (2015) The fickle P value generates irreproducible results. Nature Methods, 12, 179-185. MAXWELL, S. (2004) The persistence of underpowered studies in psychological research: Causes, consequences, and remedies. Psychological Methods, 9, 147-163.

On 2016 Jan 13, Mike Fay commented:

Halsey, Curran-Everett, Vowler, and Drummond (2015) correctly point out that P values are variable and that estimates and confidence intervals often provide more useful information. I disagree with some of the apparent implications they make from these points. For example, they state “The P value is often used without the realization that in most cases the statistical power of a study is too low for P to assist the interpretation of the data.” The problem with this statement is that in practice we never know the statistical power of a study, so we never know if it is too low. The statistical power is a function of the sample size, the statistical model and the true value of the parameters. If the statistical model describes the data generating process and we knew the true value of the parameters, then we could know the power. The problem is that we collect data precisely because we do not know the true value of the parameters, and hence do not know the power. So we never know if the power of a study is “too low”. P can assist in interpreting the data. The P value represents the probability of observing equal or more extreme data when the null hypothesis is true. So small values of the P value imply that the null hypothesis is not likely to hold.

Here is another quote from the paper: “Put simply, the P value is usually a poor test of the null hypothesis.” This statement begs the question, is there another single statistic that is better to test the null hypothesis? If you want one statistic to test the null hypothesis, the P value is just the statistic to do that. So the P value is not “flawed” (see last paragraph of the article), but is one statistic designed to perform one function. Certainly, P values should not be the whole statistical story for any data set, but they can work for what they are designed to do.

On 2015 Mar 02, William Grant commented:

For a paper on how increasing vitamin D intake and levels might reduce the mortality rates for those with disabilities, see this open access paper: Grant WB, Wimalawansa SJ, Holick MF, Cannell JJ, Pludowski P, Lappe JM, Pittaway M, May P. Emphasizing the health benefits of vitamin D for those with neurodevelopmental disorders and intellectual disabilities. Nutrients. 2015, 7, 1538-1564. http://www.mdpi.com/2072-6643/7/3/1538

On 2016 Feb 05, Christopher Southan commented:

zileuton = http://www.guidetopharmacology.org/GRAC/LigandDisplayForward?ligandId=5297 (but racemate)

MMV007384 = https://pubchem.ncbi.nlm.nih.gov/compound/17325420

MMV666023 = https://pubchem.ncbi.nlm.nih.gov/compound/2853195

MMV665805 = https://pubchem.ncbi.nlm.nih.gov/compound/16437129

On 2015 Aug 19, Bill Cayley commented:

A good example of when less-medical is better: https://lessismoreebm.wordpress.com/2015/07/02/allergy-in-children-in-hand-versus-machine-dishwashing/

On 2015 Mar 05, Ivan Oransky commented:

Mario Saad, one of the authors of the four papers subject to this expression of concern, has sued the journal to force them to remove the expression of concern, and to prevent them from retracting the papers: http://retractionwatch.com/2015/03/05/researcher-asks-court-to-overturn-motion-denying-his-request-to-quash-expressions-of-concern/

On 2016 Jan 08, Andrea Messori commented:

Drug-drug interactions in oncology

A.Messori, HTA Unit, Regional Health Service, 50100 Firenze, Italy and SIFACT (Società Italiana di Farmacia Clinica e Terapia), 20100 Milano

In the field of oncology, drug-drug interactions (DDIs) represent a topic in which a large amount of information is available. Hence, a critical selection in this area is generally thought to be needed to maximize the usefulness of recommendations and to avoid an overload of messages. In this selection process, two criteria are most frequently advocated: (1): prioritizing the DDIs showing a marked clinical relevance (that, in the main data banks, are denoted as major interactions or contra-indications); (2): prioritizing those DDIs the frequency of which is not negligible and has been documented in the real-world setting. The first point is adequately addressed by the scores assigned to single DDIs in the most authoritative data banks (e.g. Micromedex). As regards the second point, the reference list reported below indicates which DDIs have been most frequently found in the main studies conducted on this topic in cancer patients. References 1. Carcelero E, Anglada H, Tuset M, Creus N. Interactions between oral antineoplastic agents and concomitant medication: a systematic review. Expert Opin Drug Saf. 2013 May;12(3):403-20. doi: 10.1517/14740338.2013.784268. Epub 2013 Apr 16. Review. PubMed PMID: 23586848.

1. Chan A, Tan SH, Wong CM, Yap KY, Ko Y. Clinically significant drug-drug interactions between oral anticancer agents and nonanticancer agents: a Delphi-survey of oncology pharmacists. Clin Ther. 2009;31 Pt 2:2379-86. doi: 10.1016/j.clinthera.2009.11.008. PubMed PMID: 20110047.

2. Depont F, Vargas F, Dutronc H, Giauque E, Ragnaud JM, Galpérine T, Abouelfath A, Valentino R, Dupon M, Hébert G, Moore N. Drug-drug interactions with systemic antifungals in clinical practice. Pharmacoepidemiol Drug Saf. 2007 Nov;16(11):1227-33. PubMed PMID: 17879355.

3. Girre V, Arkoub H, Puts MT, Vantelon C, Blanchard F, Droz JP, Mignot L. Potential drug interactions in elderly cancer patients. Crit Rev Oncol Hematol.2011 Jun;78(3):220-6. doi: 10.1016/j.critrevonc.2010.05.004. Epub 2010 Jul 1. Review. PubMed PMID: 20594867.

4. Kotlinska-Lemieszek A, Klepstad P, Haugen DF. Clinically significant drug-drug interactions involving opioid analgesics used for pain treatment in patients with cancer: a systematic review. Drug Des Devel Ther. 2015 Sep 16;9:5255-67. doi: 10.2147/DDDT.S86983. eCollection 2015. Review. PubMed PMID: 26396499; PubMed Central PMCID: PMC4577251.

5. Kruse V, Somers A, Van Bortel L, De Both A, Van Belle S, Rottey S. Sunitinib for metastatic renal cell cancer patients: observational study highlighting the risk of important drug-drug interactions. J Clin Pharm Ther. 2014 Jun;39(3):259-65. doi: 10.1111/jcpt.12134. Epub 2014 Jan 13. PubMed PMID:24417304.

6. Miranda V, Fede A, Nobuo M, Ayres V, Giglio A, Miranda M, Riechelmann RP. Adverse drug reactions and drug interactions as causes of hospital admission in oncology. J Pain Symptom Manage. 2011 Sep;42(3):342-53. doi: 10.1016/j.jpainsymman.2010.11.014. Epub 2011 Mar 31. PubMed PMID: 21454043.

7. Riechelmann RP, Del Giglio A. Drug interactions in oncology: how common are they? Ann Oncol. 2009 Dec;20(12):1907-12. doi: 10.1093/annonc/mdp369. Epub 2009 Aug 27. Review. PubMed PMID: 19713244.

8. Stoll P, Kopittke L. Potential drug-drug interactions in hospitalized patients undergoing systemic chemotherapy: a prospective cohort study. Int J Clin Pharm. 2015 Jun;37(3):475-84. doi: 10.1007/s11096-015-0083-6. Epub 2015 Feb 25. PubMed PMID: 25711852.

9. Teo YL, Ho HK, Chan A. Metabolism-related pharmacokinetic drug-drug interactions with tyrosine kinase inhibitors: current understanding, challenges and recommendations. Br J Clin Pharmacol. 2015 Feb;79(2):241-53. doi:10.1111/bcp.12496. Review. PubMed PMID: 25125025; PubMed Central PMCID: PMC4309630.

10. Turner JP, Shakib S, Singhal N, Hogan-Doran J, Prowse R, Johns S, Bell JS. Prevalence and factors associated with polypharmacy in older people with cancer. Support Care Cancer. 2014 Jul;22(7):1727-34. doi: 10.1007/s00520-014-2171-x. Epub 2014 Mar 2. Erratum in: Support Care Cancer. 2014 Jul;22(7):1735. PubMed PMID:24584682.

11. van Leeuwen RW, Brundel DH, Neef C, van Gelder T, Mathijssen RH, Burger DM, Jansman FG. Prevalence of potential drug-drug interactions in cancer patients treated with oral anticancer drugs. Br J Cancer. 2013 Mar 19;108(5):1071-8. doi: 10.1038/bjc.2013.48. Epub 2013 Feb 14. PubMed PMID: 23412102; PubMed CentralPMCID: PMC3619066.

12. van Leeuwen RW, van Gelder T, Mathijssen RH, Jansman FG. Drug-druG interactions with tyrosine-kinase inhibitors: a clinical perspective. Lancet Oncol. 2014 Jul;15(8):e315-26. doi: 10.1016/S1470-2045(13)70579-5. Review. PubMed PMID: 24988935.

13. Voll ML, Yap KD, Terpstra WE, Crul M. Potential drug-drug interactions between anti-cancer agents and community pharmacy dispensed drugs. Pharm World Sci. 2010 Oct;32(5):575-80. doi: 10.1007/s11096-010-9410-0. Epub 2010 Jul 20.PubMed PMID: 20645002.

On 2015 Mar 30, John Cannell commented:

In Denmark, like all Scandinavian countries, the use of cod liver oil is common. Cod liver oil will raise 25(OH)D levels but also increase vitamin A intake. Increased vitamin A intake has been associated with increased mortality. This would explain the J shaped relationship found by the authors.

Bjelakovic G, Nikolova D, Gluud LL, Simonetti RG, Gluud C. supplements for prevention of mortality in healthy participants and patients with various diseases. Cochrane Database Syst Rev. 2008 Apr 16;(2):CD007176.

On 2015 Feb 25, Sundeep Salvi commented:

The sexual dysfunction seen only in Type I diabetic females could be due to use of Insulin rather than other causes as mentioned. Among Type 2 diabetic females, how many were receiving insulin? Was there a difference in sexual dysfunction between Type 2 diabetic females using insulin and those not using insulin?

On 2017 May 19, Zvi Herzig commented:

The methodology of this study would lead to unrealistic overheating producing unrealistically high levels of toxicants. The authors note:

The E-cigarette puffs were actuated by a pump and programmed to cycle every 15 seconds until the cartridge was empty. Each actuation was 6 seconds...

However, puffing an e-cigarette until the cartridge is empty would lead to overheating when liquid levels fall low, which users avoid, but entails high levels of toxicant production Farsalinos KE, 2013 Farsalinos KE, 2015.

Also, users have been shown to take four-second puffs with 20-30 s intervals Farsalinos KE, 2013. Increasing puff duration and decreasing the time of the cycle between puffs would also increase overheating.

On 2017 Sep 09, Louise B Andrew MD JD commented:

Reason(s) for withdrawal?

On 2015 Apr 21, David Keller commented:

The observed association between sauna bathing and improved cardiovascular outcomes may be due to self-selection bias

Persons at higher risk for adverse cardiac events may experience unpleasant symptoms due to the tachycardia induced by sitting in a hot sauna, such as mild dyspnea, orthostasis, or chest discomfort, at higher frequency or severity than persons in good cardiovascular health, These adverse symptoms might cause them to avoid saunas, thereby biasing the group of sauna-takers to include persons at lower risk of adverse cardiac events than the general public.

The authors suggest that, based on this study, "sauna bathing is a recommendable health habit". I disagree, and suggest that physicians should await the results of a randomized trial of sauna bathing before we recommend it for health enhancement. Only randomized trials can provide the quality of evidence required for a physician to recommend a potentially dangerous intervention for health enhancement. Observational studies such as this one are supposed to be for "hypothesis generation" only.

On 2015 Mar 04, Raphael Stricker commented:

In this one-sided opinion piece about Lyme disease, Eugene Shapiro once again finds "no evidence that viable B. burgdorferi persist in humans after conventional treatment with antimicrobials". Shapiro is a well known member of the shrinking "Lyme Denialist" cabal that views Lyme disease as a trivial illness that is "hard to catch and easy to cure", apparently ignoring the latest CDC figures showing more than 300,000 new cases per year in the USA. The fact that Lyme disease has become a major epidemic that is six times more common than HIV/AIDS in this country fails to impress Shapiro, who adheres to the dogma that persistent infection with B. burgdorferi, the Lyme spirochete, does not exist following short-course antibiotic therapy despite extensive evidence to the contrary (Stricker & Johnson, Infect Drug Resist 4: 1-9, 2011; Cameron et al, Expert Rev. Anti Infect. Ther. 12:1103-1135, 2014).

Significant controversy over Lyme disease exists for three main reasons: (1) lack of accurate and/or universally accepted testing for the disease, (2) disagreement about symptoms associated with persistent infection in chronic Lyme disease, and (3) misinterpretation and misrepresentation of underpowered Lyme antibiotic treatment trials. While many studies describe the constellation of musculoskeletal, neurocognitive and/or cardiac symptoms associated with chronic Lyme disease, Shapiro views these as "medically unexplained symptoms" not necessarily related to persistent B. burgdorferi infection. Without a universally accepted "gold standard" test, the controversy over persistent infection and optimal therapy continues to smoulder while thousands of patients continue to suffer due to the dogma espoused by Shapiro (Johnson et al. PeerJ 2:e322, 2014).

A major problem faced by Shapiro is that he is trying to prove a negative. Thus if there is any evidence that persistent infection with B. burgdorferi does exist following short-course antibiotic therapy, his opinion is obviously wrong. To address this problem, Shapiro narrows his evidence to two recent Lyme disease articles, ignoring numerous studies in animals and humans that support persistent infection or leave the issue unsettled (Cairns & Godwin, Int J Epidemiol 34: 1340-1345, 2005; Berndtson, Int J Gen Med 6: 291-306, 2013; Stricker & Johnson, PLoS Pathog 10: e1003796, 2014). Shapiro dismisses this contrary evidence as "speculative", but his narrow selection of two "convincing" studies is insufficient to support his biased conclusion.

The first study examined a group of 17 patients with recurrent erythema migrans (EM) rashes who were promptly treated for their initial episode of Lyme disease and then developed one or more EM rashes at a later date. Culture of the rashes revealed different strains of B. burgdorferi in the subsequent episodes, and Shapiro points to this as evidence for new infection rather than relapse in these patients. However as pointed out in a letter addressing the article, this is a poor model for chronic Lyme disease due to persistent infection because all patients were promptly treated for their initial illness, lived in endemic areas and most likely were reinfected with a different strain of the spirochete from a subsequent tickbite. This is a very different situation from a patient who may have been infected and never treated for months to years and develops the constellation of musculoskeletal, neurocognitive and/or cardiac symptoms that are characteristic of persistent infection with the Lyme spirochete (Donta, N Engl J Med 368:1063-1064, 2013). Thus the model for persistent infection in this study is flawed.

The second study was a xenodiagnosis safety study of 36 patients (26 Lyme patients in different stages of disease and 10 controls) who allowed ticks to feed on them, and the ticks were then examined for B. burgdorferi transmission. Shapiro states that "no viable B. burgdorferi were cultured from ticks fed on any of these patients". This conclusion is flawed for two reasons: First, 30-50% of ticks were lost during the study, rendering the transmission results uninterpretable. Second, one patient with post-treatment Lyme disease syndrome (PTLDS) was found to have a positive culture from one tick, as stated in the Results: "One nymph was found to be positive by PCR of the nymph lysate culture, but direct PCR of the nymph lysate and microscopic evaluation of the culture were negative....The original positive OspA PCR of the tick culture was confirmed by PCRs for other B. burgdorferi genes.... The DNA extracted from this culture sample was then tested by IA/PCR/ESI-MS, which was positive for 7 of the 8 assay primer pairs" (emphasis added). Thus this patient had culture-confirmed evidence of persistent infection with the Lyme spirochete in PTLDS. Shapiro generously states that this finding is "provocative" when in fact it provides definitive evidence that he is wrong.

In summary, this one-sided opinion piece will only add to the confusion and misinformation surrounding Lyme disease. With better testing and novel treatments, a solution to this tickborne disease will someday be found. Shapiro's muddled article fails to contribute to this solution.

Disclosure: RBS is a member of the International Lyme and Associated Diseases Society (ILADS) and a director of LymeDisease.org. He has no financial or other conflicts to declare.

On 2016 Sep 23, Israel Hanukoglu commented:

The link to the Proteopedia page with animations and examples: http://www.proteopedia.org/w/Rossmann_fold

On 2017 Jul 10, Randi Pechacek commented:

Rachel Adams, first author of this paper, wrote a blog post on microBEnet to provide some background.

On 2016 Mar 08, Kenneth Witwer commented:

Because of continuing interest in this topic, an extended version of these comments has been deposited on the "Negative Results" track on ResearchGate.

On 2015 Mar 04, Avinash Bidra commented:

My response to these comments can be found at: http://www.ncbi.nlm.nih.gov/pubmed/25702969

On 2016 Mar 18, ZHONGMING ZHAO commented:

My lab recently moved to the University of Texas Health Science Center at Houston. The software is now available at https://bioinfo.uth.edu/VirusFinder/.

On 2015 Mar 19, Donna Beales commented:

My understanding is that there was a change to the formulation of the suspension medium used in some trials with T. suis ova. Such alterations could conceivably have unanticipated effects on living organism therapies. Quoted as follows: All prior and very successful studies, this final formulation was of a pH value between 2.7 – 3 and had no preservative, while for the TRUST and some other newly pilot studies on other indications a formulation of a pH value of 5 plus potassium sorbate as a preservative was used... Nevertheless, the decision of changing the formulation for the drug approval studies is understandable if one takes into account what the common practice is in normal drug development according to the pharmacopeia, but a living organism does not necessarily fit into these regulations designed for single molecules and chemical compounds."

Too, 12 weeks may not be sufficient time for clinical efficacy with a living organism; even pharmaceutical immunosuppressants may require a period of several months for full effectiveness. "Another critical issue in the TRUST studies was that the endpoint was set after just 12 weeks of treatment, which is known to be the time frame where the agent just begins to unfold its mode of action effectively." Source: http://tanawisa.com/news/

These factors are relatively unknown, and should be considered in any interpretation of study conclusion.

On 2015 Apr 22, Hans-Peter Müller commented:

Response to Müller HP. Direct and CBCT dentogingival tissue thickness measurements. Oral Surg Oral Med Oral Pathol Oral Radiol 2015; 119: 482-481. http://www.ncbi.nlm.nih.gov/pubmed/25687197

On 2016 Aug 30, Ben Goldacre commented:

This trial has the wrong trial registry ID associated with it on PubMed. The ID given is NCT014182482377 - the correct ID, which we found within the text of the article itself, is NCT00962377.

This comment is being posted as part of the OpenTrials.net project^[1] , an open database threading together all publicly accessible documents and data on each trial, globally. In the course of creating the database, and matching documents and data sources about trials from different locations, we have identified various anomalies in datasets such as PubMed, and in published papers. Alongside documenting the prevalence of problems, we are also attempting to correct these errors and anomalies wherever possible, by feeding back to the originators. We have corrected this data in the OpenTrials.net database; we hope that this trial’s text and metadata can also be corrected in PubMed.

Many thanks,

Jessica Fleminger, Ben Goldacre*

[1] Goldacre, B., Gray, J., 2016. OpenTrials: towards a collaborative open database of all available information on all clinical trials. Trials 17. doi:10.1186/s13063-016-1290-8 PMID: 27056367

* Dr Ben Goldacre BA MA MSc MBBS MRCPsych<br> Senior Clinical Research Fellow<br> ben.goldacre@phc.ox.ac.uk<br> www.ebmDataLab.net<br> Centre for Evidence Based Medicine<br> Department of Primary Care Health Sciences<br> University of Oxford<br> Radcliffe Observatory Quarter<br> Woodstock Road<br> Oxford OX2 6GG

On 2015 Feb 23, William Grant commented:

Concerns over Vitamin D and clinical practice at a crossroads recommendations

In their viewpoint piece, Vitamin D and clinical practice at a crossroads, Manson and Bassuk state among other things that the Institute of Medicine (IOM) set the recommended dietary allowance for vitamin D at 600 IU/d for those living in the upper latitudes of North America aged to 70 years and 800 IU/d for those older in order to reach a 25-hydroxyvitamin D [25(OH)D] concentration of 20 ng/mL (50 nmol/L) [1]. However, it is not clear how the Dietary Reference Intakes for Calcium and Vitamin D committee arrived at that number. For example, on p. 3-20 of the vitamin D and calcium IOM report [2], Figure 3-4 from Cashman et al. [3] is given as Figure 3-4, although without the 95% confidence intervals as in the original paper. The results were based on a 22-week placebo, randomized controlled supplementation study involving men and women aged 20-40 years. Inspection of Figure 2 in Ref. 3 indicates that it would take 1155 IU/d vitamin D3 for 97.5% of the 20-40 year old population sampled to reach 50 nmol/L. In a subsequent paper based on a systematic review and meta-analysis of vitamin D intake and 25(OH)D concentrations, the same group determined that it would take 930 IU/d vitamin D3 for people living in northern Europe to reach 50 nmol/L [4]. Further complicating matters is the fact that not all of the contributions from diet are accounted for in most studies. It is becoming apparent that some food such as meat has vitamin D in the form of 25(OH)D. Thus, vegans in the UK have 25(OH)D concentrations 20 nmol/L lower than omnivores [5], so require higher vitamin D intake from non-dietary sources or solar UVB exposure.

Their comment that "while awaiting the results of the large trials now in progress, physicians would be well advised to follow current USPSTF and IOM recommendations and avoid overscreening and overprescribing supplemental vitamin D" is also not well grounded. The IOM restricted its assessment of the benefits of vitamin D to vitamin D randomized controlled trials [RCTs] with substantial benefits by 2010. A number of trials since then demonstrated health benefits, e.g., for biomarkers of inflammation, where it was found that RCTs with baseline 25(OH)D concentrations below 48 nmol/L had a 50% chance of findings benefits from vitamin D supplementation compared to 25% with baseline 25(OH)D concentrations above 50 nmol/L [6]. In addition, ecological, observational, clinical, and laboratory studies have found many health benefits of solar UVB exposure and/or vitamin D. Since the IOM report was published (29 November, 2010), 13,535 publications with vitamin D in the title or abstract have been published at PubMed.gov as of 23 February, 2015, compared with 27,775 published before that date. Many of these publications strengthen the case for vitamin D supplementation and UVB exposure.

In terms of confounding factors related to observational studies, one not mentioned in Ref. 1 is the possibility that solar UV exposure may have health benefits in addition to vitamin D production. As a result, 25(OH)D concentrations may be an index of UVB exposure. Beneficial effects of UV exposure in addition to vitamin D production have been reported for intestinal cancer [7], multiple sclerosis [8], and blood pressure [9].

As for concern about adverse effects of higher 25(OH)D concentrations based on observational studies, it should be noted that most such studies do not obtain any information from participants about vitamin D supplementation prior to having 25(OH)D concentrations measured. Thus, those with adverse health outcomes and high 25(OH)D concentrations may have started taking vitamin D supplements shortly before blood draw. For example, studies of frailty vs. 25(OH)D concentration found a U-shaded relation for elderly women [10] but a linear inverse relation for elderly men [11]. Elderly women in the U.S. are much more likely to be advised to take vitamin D supplements than men, and starting to take vitamin D late in life cannot erase the adverse effects of years of low 25(OH)D concentrations. In addition, meta-analyses of observational studies of health outcomes with respect to 25(OH)D concentrations do not show U-shaped relations for cardiovascular disease [12] or all-cause mortality rates [13].

As to their comment regarding how interest in vitamin D could jeopardize ongoing vitamin D RCTs, that should not be the case if trial participants are screened by measuring 25(OH)D concentration prior to acceptance and including only those with 25(OH)D concentrations below 50 nmol/L then dropping any who are subsequently prescribed supplements in excess of the IOM recommendations. Over 99% of the population is not enrolled in vitamin D RCTs and should not be held hostage to ongoing or planned trials since there appear to be many health benefits and very few risks of vitamin D supplementation below 4000 IU/d [14] even by the IOM's admission [2].

References 1. Manson JE, Bassuk SS. Vitamin D research and clinical practice: at a crossroads. JAMA. 2015 Feb 19. doi: 10.1001/jama.2015.1353. [Epub ahead of print] 2. Ross AC, Taylor CL, Yaktine AL, Del Valle HB, eds. ; Committee to Review Dietary Reference Intakes for Vitamin D and Calcium; Dietary Reference Intakes for Calcium and Vitamin D. Institute of Medicine. ISBN: 0-309-16395-1, 482 pages, (2010) Available from National Academies Press at: http://www.nap.edu/catalog/13050.html 3. Cashman KD, Hill TR, Lucey AJ, et al. Estimation of the dietary requirement for vitamin D in healthy adults. Am J Clin Nutr. 2008;88(6):1535-42. 4. Cashman KD, Fitzgerald AP, Kiely M, Seamans KM. A systematic review and meta-regression analysis of the vitamin D intake-serum 25-hydroxyvitamin D relationship to inform European recommendations. Br J Nutr. 2011;106(11):1638-48. 5. Crowe FL, Steur M, Allen NE, et al. Plasma concentrations of 25-hydroxyvitamin D in meat eaters, fish eaters, vegetarians and vegans: results from the EPIC-Oxford study. Public Health Nutr. 2011;14(2):340-6. 6. Cannell JJ, Grant WB, Holick MF. Vitamin D and inflammation. Dermato-Endocrinology. 2014;6(1):e983401-1-10.<br> 7. Rebel H, der Spek CD, Salvatori D, et al.UV exposure inhibits intestinal tumour growth and progression to malignancy in intestine-specific Apc mutant mice kept on low vitamin D diet. Int J Cancer. 2015;136(2):271-7. 8. Zivadinov R, Treu CN, Weinstock-Guttman B, et al. Interdependence and contributions of sun exposure and vitamin D to MRI measures in multiple sclerosis. J Neurol Neurosurg Psychiatry. 2013;84(10):1075-81. 9. Opländer C, Volkmar CM, Paunel-Görgülü A, et al. Whole body UVA irradiation lowers systemic blood pressure by release of nitric oxide from intracutaneous photolabile nitric oxide derivates. Circ Res. 2009;105(10):1031-40. 10. Ensrud KE, Ewing SK, Fredman L, et al. Circulating 25-hydroxyvitamin D levels and frailty status in older women. J Clin Endocrinol Metab. 2010;95(12):5266-73. 11. Ensrud KE, Blackwell TL, Cauley JA, et al. Circulating 25-hydroxyvitamin D levels and frailty in older men: the osteoporotic fractures in men study. J Am Geriatr Soc. 2011;59(1):101-6. 12. Wang L, Song Y, Manson JE, et al. Circulating 25-hydroxy-vitamin D and risk of cardiovascular disease: A meta-analysis of prospective studies. Circ Cardiovasc Qual Outcomes. 2012;5(6):819-29. 13. Garland CF, Kim JJ, Mohr SB, et al. Meta-analysis of all-cause mortality according to serum 25-hydroxyvitamin D. Am J Pub Health. 2014;104(8):e43-50. 14. Vieth R. Implications for 25-hydroxyvitamin D testing of public health policies about the benefits and risks of vitamin D fortification and supplementation. Scand J Clin Lab Invest Suppl. 2012;243:144-53.

Disclosure I receive funding from Bio-Tech Pharmacal, Inc. (Fayetteville, AR), MediSun Technology (Highland Park, IL), and the Vitamin D Council (San Luis Obispo, CA).

On 2015 Feb 25, Ivan Oransky commented:

Following a great deal of criticism of this article, the journal issued a statement of concern and then reclassified the paper as "opinion." http://retractionwatch.com/2015/02/24/frontiers-lets-hiv-denier-article-stand-reclassifies-it-as-opinion/

On 2015 Oct 22, Alan Fairlamb commented:

This is an excellent and comprehensive review of the prospects for elimination of Gambiense HAT. A number of uncertainties are identified and important research questions highlighted. However, one posible threat not considered is the risk of drug resistance emerging to the limited armamentarium of drugs available to treat patients and thereby reduce transmission. Drug susceptibility testing of clinical isolates would be useful in defining optimal treatment regimens.

Antimicrob Agents Chemother. 2010 Jul;54(7):2893-900. doi: 10.1128/AAC.00332-10. Epub 2010 May 3.

On 2015 Apr 21, Helmi BEN SAAD commented:

A letter to the Editor, highlighting some methodological problems in the above article is now published: http://www.libyanjournalofmedicine.net/index.php/ljm/article/view/27760

H Ben Saad. Methodological problems in the article comparing lung function profiles and aerobic capacity of adult cigarette and hookah smokers after 12 weeks intermittent training.Libyan J Med 2015, 10: 27760 - http://dx.doi.org/10.3402/ljm.v10.27760

On 2016 Jul 14, Andrea Fuso commented:

The analysis was initially made on these two genes since those are the genes we were studying in relation to muscle differentiation (myogenin) and Alzheimer's Disease (PSEN1). In these two models we gained in the years sufficient knowledge to finely modulate methylation and to foresee that different treatments correspond to differential methylation. Moreover, we are interested in "rapid" methylation changes and it is our hypothesis that genes without CpG islands can be modulated by dynamic changes in DNA methylation that are more rapid than in CpG islands (in wich methylation-dependent silencing/activation is more stable) and possible also in non replicating tissues (brain). We are now checking promoters with and without CpG islands. Of course, designing MIPs in CpG islands could be very difficult or even impossible. That's a limitation.

On 2016 Jul 14, Long-Cheng Li commented:

Dear Andrea,

I am sorry for not having not responded earlier and thank you for your reply.

If the possibility of incomplete conversion can be excluded as you have explained above, I think the findings in your work is significant in that they makes researchers better aware of the non-CpG methylatioon issue. I do have another comment though which is why you chose two genes (myogenin and PSEN1) without CpG islands in their promoters instead of genes with CpG islands where differential methylation often occurs.

On 2016 Jul 13, Andrea Fuso commented:

None

On 2016 Jul 13, Andrea Fuso commented:

Dear Long-Cheng, thank you for your critic comments. Discussing this issue is very important and I'm sincerely happy you took some time to read our manuscript and share your thoughts. When we published the paper, we were hopeful it could raise some discussion useful to a better understanding of the non-CpG issue.

First of all, I would like to emphasize that our paper was not a criticism of MethPrimer. As a matter of fact (as disclosed in the paper) different primer design softwares, not just “MethPrimer”, allow to design bisulfite primers based on the assumption that non-CpG cytosines are transformed. MethPrimer is simply the most used and known and we defined “methprimers” the oligo designed using these different softwares. Our criticism were, on the contrary, addressed toward the concept that stands at the base of all these primer design softwares: i.e. that non-CpG cytosines are (almost) always demethylated. The idea of releasing MethPrimer was great and in line with the widely accepted concept of that era on non-CpG methylation. We should now critically consider that concept and make some effort to better understand extension and significance of non-CpG methylation and, in case, revise our tools for the study of DNA methylation. The adaptation of MethPrimer to the picking of primers in regions with few cytosines goes in this way and it is highly appreciable.

As for your comments about incomplete conversion: your considerations are correct but you are not correct when you affirm that “no efforts were made to verify the completeness of bisulfite conversion of the template DNA”. If you look at the methods and at the previous papers cited in the methods (previously published description were not repeated and just cited) you can see that we took many efforts to demonstrate that our analysis was not affected by artifacts. Our studies on myogenin methylation (some years ago) started with classical bisulphite conversion (no kits) but in the years we started to place side by side classical and kit-assisted bisulphite conversion, obtaining comparable results. The present study was made by using the Qiagen Epitect Kit but specific (high and low methylated samples) controls were modified using the ZymoResearch kit and the in-lab prepared bisulphite. These controls were real samples, not just PCR products. According to your idea that our data could be due to incomplete conversion, you should explain why incomplete conversion occurs only (or mainly) for high methylated DNA but not in low methylated DNA, since the difference between methprimers and MIPs is less when analyzing low methylated samples. In our analyses, in which samples with different (expected) methylation levels are modified in the same run, we can indeed observe both methylated and unmethylated sequences, according to the expected methylation. I think that this observation, more than any “control” witness the quality of the bisulphite conversion we make. If you suggest, or suspect, that bisulphite conversion kit (like the Qiagen and the Zymoresearch) that are today used in almost all the methylation studies, and that are preferred to the in-lab prepared bisulphite just to guarantee reliability and avoid incomplete conversion, are responsible for incomplete conversion….well, the problem is much bigger. I will be happy if you and others want to add more to this interesting discussion.

In conclusion, I completely agree with you that much more care should be put in the control of the bisulphite analysis…and as a reviewer of papers dealing with DNA methylation, I can see that few researchers take care of performing rigorous controls. But, I also invite you and the other colleagues to consider that new and unexpected results, such as relevant non-CpG methylation, deserve to be considered with unbiased attitude and methods.

On 2016 Jul 04, Long-Cheng Li commented:

As the author of MethPrimer, I thank the authors of this paper for looking into the bias issue of our program. However the conclusion reached that "MethPrimers can result in underestimation of the high methylation levels eventually biasing the detection of differences" could be misleading based on the following reasons:

1) MethPrimer was developed in an era when there was little knowledge about the existence of non-CpG methylation and under the assumption that all non-CpG cytosines are not methylated. One of the important principles used by MethPrimer in picking primers is to select primers that preferentially amplify bisulfite converted DNA, given the inherent issue of incomplete conversion of unmethylated cytosines during bisulfite treatment. This is implemented by forcing picking primers in regions which must contain a minimum number of non-CpG cytosines.

2) Using “methylation-insensitive primers” (MIPs) in which non-CpG Cs are degenerated, the authors detected higher levels of methylation compared to primers designed by MethPrimer in the promoter regions of two mouse genes (myogenin and PSEN1). However, no efforts were made to verify the completeness of bisulfite conversion of the template DNA. Although control samples with complete methylated and unmethylated DNA were included in the analysis, the use of PCR products as a control for unmethylated DNA is problematic since incomplete conversion is less an issue for PCR products compared to genomic DNA. In other words, complete conversion of the control DNA does not necessarily mean that genomic DNA was also completely converted. If that were true, the non-CpG methylation detected could be merely an artifact.

I do caution that researchers should keep in mind potential non-CpG methylation. Before more evidence is available about the prevalence of non-CpG methylation and its function, researchers should care more about potential incompleteness of bisulfite conversion and subsequent overestimation of methylation levels in the DNA.

On 2015 Apr 23, Janet Kern commented:

Because of differences noted between the study conclusion and the results, on February 24, 2015, an email was sent to Dr. Hewitson requesting the dataset. However, to date, the authors have ignored repeated requests for the dataset.

On 2015 Feb 21, Dan Laks commented:

None

On 2015 Dec 12, Donald Forsdyke commented:

LESS BP MEDICATION NEEDED IN HOT WEATHER This paper recommends “More aggressive blood pressure lowering treatment in the cold months … in high risk individuals” (1), which implies less aggressive blood pressure (BP) lowering treatment in hot months. Indeed, a well-documented “J-curve” observation is that, below a certain value, BP lowering is harmful – e.g. acute kidney injury (2). Quite rightly, the cartoon in the accompanying editorial has question marks at both the high and low ends of the temperature scale (3). However, the paper’s focus is on more treatment in winter, not on less treatment in summer, and it only scores the deaths attributed to cardiovascular disease (CVD), a number that declines in hot weather (1). Cases where mortality can be attributed to other causes are excluded.

Regarding CVD it is reported that “The excess risk was similar between people treated with blood pressure lowering agents and those without” (1). But given the widely different mechanisms of action of different BP lowering agents, surely this statement needs backing with more information on agents and their dosages? Large summertime systolic BP declines that mandate treatment adjustment are not uncommon (4). Differential responses in hot weather can depend on type and dosage of antihypertensive medication. Indeed, extrapolation from the supplementary plot (Fig 1) of Yang et al. (2015), the BP would be normal at 40°C and medication could be dispensed with (1). Taking medication at that temperature could be lethal (5). Indeed, a Canadian case study with an angiotensin receptor blocker found that medication could be dispensed with when summer temperatures reached 33°C (6).

It is obvious from Fig. 3 that the statement that “the seasonal variation in blood pressure … was abolished by the use of home central heating” (1), is incorrect. In Harbin province, where winter temperatures are similar to those in Canada, there is less BP increase in cold weather than in other provinces, and this is attributed to central heating. But the seasonal BP increase is clearly not abolished. Despite central heating, in the Canadian case study the BP decline for a 10°C increase in temperature was of the order of 20 mm Hg (6) – a far higher value than the 6 mm Hg reported here (1).

1 Yang L, Li L, Lewington S. Guo Y, Sherliker P, Bian Z, Collins R, et al (2015) Outdoor temperature, blood pressure, and cardiovascular disease mortality among 23000 individuals with diagnosed cardiovascular diseases from China. Eur Heart J 36:1178-1185. Yang L, 2015

2 Tomlinson LA, Abel GA, Chaudhry AN, Tomson CR, Wilkinson IB, Roland MO et al (2013) ACE inhibitor and angiotensin-II receptor antagonist prescribing and hospital admissions with acute kidney injury: a longitudinal ecological study. PLoS One 8:e78465.Tomlinson LA, 2013

3 Bruno RM, Taddei S (2015) ‘Tis bitter cold and I am sick at heart’: establishing the relationship between outdoor temperature, blood pressure, and cardiovascular mortality. Eur Heart J 36:1152-1154.Bruno RM, 2015

4 Stergiou GS, Myrsilidi A, Kollias A, Destounis A, Roussias L, Kalogeropoulos P (2015) Seasonal variation in meteorological parameters and office, ambulatory and home blood pressure: predicting factors and clinical implications. Hypertension Research 38(12):869-875. Stergiou GS, 2015

5 Editorial (2015) Health professionals: be prepared for heatwaves. Lancet 386:219. Anonymous, 2015

6 Forsdyke DR (2015) Summertime dosage-dependent hypersensitivity to an angiotensin II receptor blocker. BMC Res Notes 8:227. Forsdyke DR, 2015

On 2015 Feb 18, Hilda Bastian commented:

This study states as its objective "to determine the efficacy and safety of varenicline" for quitting smoking via smoking reduction. The authors point out that one limitation of the study is its generalizability to a broad population, given its stringent and extensive exclusion criteria. However, it does not stress that both this, and the size of the study, very much preclude this single study "determining" safety of varenicline. The findings in relation to serious adverse events need to be considered in the light of the lower risk for serious adverse events in this study population.

The paper does not refer readers to the safety warnings and concerns about varenicline issued by both the US FDA and European Medicines Agency (EMA), in relation both to psychiatric (FDA boxed warning) and cardiovascular events (FDA, 2012)(see also EMA). (Readers may also be interested in Singh S, 2011 on the issue of cardiovascular events.)

UPDATE: On 9 March 2015, the FDA reviewed safety data on varenicline, retaining the boxed safety warning, and including a warning on interaction with alcohol. However, in March a large meta-analysis found that varenicline increased insomnia and bad dreams, but not depression, suicide, or suicidal ideation (Thomas KH, 2015).

In terms of effectiveness, the authors rightly raise the issue of a lack of direct comparisons between varenicline and others options for smoking reduction. Readers might be interested in Asfar T, 2011, which finds that nicotine replacement therapy (NRT) achieved smoking reduction rates that were not dramatically dissimilar. Cahill K, 2010 found some (inconclusive) evidence that NRT and varenicline result in similar quit rates. Nor are pharmacological means the only successful options for reducing smoking without the risk of serious adverse events.

Note also that this study was funded by Pfizer, manufacturer of the varenicline product marketed as Chantix in the US (Champix in Europe).

On 2015 Feb 21, William Grant commented:

Outdoor workers have better health outcomes due to UVB exposure

The paper by Bauer and colleagues recommended that outdoor workers reduce their UV radiation exposure in order to reduce their risk of squamous cell carcinoma (SCC) and basal cell carcinoma (BCC) [1]. Overlooked in this paper was any discussion of the health benefits of UV exposure through outdoor work.

Solar UVB exposure reduces risk of many types of cancer as shown in geographical ecological studies from Australia, China, France, Japan, Spain, and the United States [2]. Deaths due to internal cancers are much higher than those due to non-melanoma skin cancer [3]. Solar UVB exposure is the primary source of vitamin D for most people, and there are many health outcomes linked inversely to 25-hydroxyvitamin D concentrations [4].

Perhaps most relevant for Germany is a study of cancer incidence rates with respect to occupation in Nordic countries [5]. Using a data base of nearly three million cancer incidence cases for 54 occupation categories [6], an index of UVB exposure was developed, namely, lip cancer incidence rate less lung cancer incidence rate for males. Fourteen types of cancer were significantly inversely correlated with this index for males, but only three for females. Since females wear lipstick, an independent index could not be developed for them. Interestingly, this index was inversely correlated with incidence of non-melanoma skin cancer as well as melanoma for males.

Furthermore, a study in Denmark found "Mortality was reduced among patients with BCC (10-year MRR = 0.91 (95% CI: 0.89-0.92) and did not vary by age, comorbidity, or socioeconomic status. Mortality among patients with SCC was increased and varied by age, selected chronic diseases, but not socioeconomic status." [7]. Since smoking is a risk factor for SCC [8], the finding of an increased risk of death associated with SCC is not surprising since it is also an important risk factor for many types of cancer [2] as well as cardiovascular disease and other diseases.

Since skin pigmentation has adapted to where people have lived for many generations to both reduce the adverse effects of solar UVB as well as to produce sufficient vitamin D for optimal health [9,10], it is not clear that undue protection against solar UVB exposure is warranted. However, erythema is best avoided.

References 1. Bauer A, Beissert S, Knuschke P. [Prevention of occupational solar UV radiation-induced epithelial skin cancer.] Hautarzt. 2015 Feb 17. [Epub ahead of print] 2. Moukayed M, Grant WB. Molecular link between vitamin D and cancer prevention. Nutrients. 2013;5(10):3993-4023. 3. Grant WB. In defense of the sun: An estimate of changes in mortality rates in the United States if mean serum 25-hydroxyvitamin D levels were raised to 45 ng/mL by solar ultraviolet-B irradiance. Dermatoendocrinol. 2009;1(4):207-14. 4. Pludowski P, Holick MF, Pilz S, et al. Vitamin D effects on musculoskeletal health, immunity, autoimmunity, cardiovascular disease, cancer, fertility, pregnancy, dementia and mortality- a review of recent evidence. Autoimmun Rev. 2013;12(10):976-89. 5. Grant WB. Role of solar UV irradiance and smoking in cancer as inferred from cancer incidence rates by occupation in Nordic countries. Dermatoendocrinol. 2012;4(2):203-11. 6. Pukkala E, Martinsen JI, Lynge E, et al. Occupation and cancer—follow-up of 15 million people in five Nordic countries. Acta Oncol 2009;48:646-790; 7. Jensen AØ, Lamberg AL, Jacobsen JB, et al. Non-melanoma skin cancer and ten-year all-cause mortality: a population-based cohort study. Acta Derm Venereol. 2010;90(4):362-7. 8. Leonardi-Bee J, Ellison T, Bath-Hextall F. Smoking and the risk of nonmelanoma skin cancer: systematic review and meta-analysis. Arch Dermatol. 2012;148(8):939-46. 9. Yuen AW, Jablonski NG. Vitamin D: in the evolution of human skin colour. Med Hypotheses. 2010;74(1):39-44. 10. Jablonski NG, Chaplin G. Colloquium paper: human skin pigmentation as an adaptation to UV radiation. Proc Natl Acad Sci U S A. 2010;107 Suppl 2:8962-8.

Disclosure I receive funding from Bio-Tech Pharmacal, Inc. (Fayetteville, AR), MediSun Technology (Highland Park, IL), and the Vitamin D Council (San Luis Obispo, CA).

On 2015 Feb 24, Amanda Capes-Davis commented:

Are the authors able to comment further on the cell lines used in this study, to confirm that results are not related to the cell culture models used?

The Materials and Methods section states that cell lines (MCF-7, MCF-10A and BT549) were obtained from the ATCC and authenticated before purchase. I am very pleased to see information included on source and previous testing. However, it is not clear how long ago testing was performed. In some cases, cell lines are used by many different laboratory members and may be passed from laboratory to laboratory. That gives time for cross-contamination or other cell culture problems to arise. Mycoplasma contamination, or cross-contamination by another cell line, might result in behavioral changes at the time that morphine is introduced.

The results here have significant clinical implications, suggesting that pain control in breast cancer may contribute to chemoresistance. So it is essential to ensure that cell culture models are tested to eliminate any possibility of artefacts.

I particularly raise this issue because, as the authors note in their Discussion, previous studies have suggested that morphine decreases cell proliferation in many cancer cell types.

The International Cell Line Authentication Committee (ICLAC) has released a checklist that sets out cell line reporting requirements for manuscripts and grant applications. The checklist can be found at http://iclac.org/resources/cell-line-checklist/.

On 2015 Sep 16, Geriatric Medicine Journal Club commented:

Sleep disturbances are common in the elderly and adversely affect quality of life. Pharmacotherapy is often limited by lack of efficacy and/or adverse events. This article was critically appraised at the May 2015 Geriatric Medicine Journal Club (follow #GeriMedJC on Twitter). The full discussion can be found at: http://gerimedjc.blogspot.com/2015/05/may-2015-gerimedjc.html?spref=tw While mindfulness meditation seems to improve sleep quality in older adults, the challenge is how to operationalize this type of intervention.

On 2017 Mar 05, Atanas G. Atanasov commented:

Very good overview on the health-modulating potential of mushrooms. I have highlighted this review on: http://healthandscienceportal.blogspot.com/2017/03/mushrooms-for-improvement-of-health-and.html

On 2015 Feb 15, Jinlei Nie commented:

In RESULTS: "not significant", but and "P<0.05"???

On 2015 Jun 18, Elsamma Chacko commented:

Exercise before breakfast increases hepatic glucose production and glucose levels can go up.

On 2016 Apr 18, Tom Kindlon commented:

James C Coyne PhD has blogged here https://jcoynester.wordpress.com/2016/03/20/why-the-cochrane-collaboration-needs-to-clean-up-conflicts-of-interest/ about my comment:

"Selective reporting (outcome bias)" and White et al. (2011) I don't believe that White et al. (2011) (the PACE Trial) (3) should be classed as having a low risk of bias under "Selective reporting (outcome bias)" (Figure 2, page 15). According to the Cochrane Collaboration's tool for assessing risk of bias (21), the category of low risk of bias is for: "The study protocol is available and all of the study’s pre-specified (primary and secondary) outcomes that are of interest in the review have been reported in the pre-specified way". This is not the case in the PACE Trial. The three primary efficacy outcomes can be seen in the published protocol (22). None have been reported in the pre-specified way. The Cochrane Collaboration's tool for assessing risk of bias states that a “high risk” of bias applies if any one of several criteria are met, including that “not all of the study’s pre-specified primary outcomes have been reported” or “one or more primary outcomes is reported using measurements, analysis methods or subsets of the data (e.g. subscales) that were not pre-specified”. In the PACE Trial, the third primary outcome measure (the number of "overall improvers") was never published. Also, the other two primary outcome measures were reported using analysis methods that were not pre-specified (including switching from the bimodal to the Likert scoring method for The Chalder Fatigue Scale, one of the primary outcomes in your review). These facts mean that the “high risk of bias” category should apply.

and the response I received from one of the authors .

On 2015 Sep 14, Tom Kindlon commented:

None

On 2016 Jun 07, Tom Kindlon commented:

None

On 2015 Sep 14, Tom Kindlon commented:

(contd.)

11 National Institute for Health and Clinical Excellence. Chronic fatigue syndrome/myalgic encephalomyelitis (or encephalopathy): diagnosis and management of CFS/ME in adults and children, 2007. http://www.nice.org.uk/guidance/CG53 Accessed September 6, 2015. London: National Institute for Health and Clinical Excellence.

12 White PD, Goldsmith KA, Johnson AL, Potts L, Walwyn R, DeCesare JC, et al. Comparison of adaptive pacing therapy, cognitive behaviour therapy, graded exercise therapy, and specialist medical care for chronic fatigue syndrome (PACE): a randomised trial. The Lancet 2011;377:823-36.

13 Kindlon T. PACE Trial - 97% of the participants who didn't have a psychiatric disorder satisfied the definition of M.E. used. https://listserv.nodak.edu/cgi-bin/wa.exe?A2=ind1106A&L=CO-CURE&P=R2764 Accessed: September 6, 2015

14 Ellen Goudsmit on PubMed Commons: http://www.ncbi.nlm.nih.gov/myncbi/ellen m.goudsmit.1/comments/

15 Green CR, Cowan P, Elk R, O'Neil KM, Rasmussen AL. National Institutes of Health Pathways to Prevention Workshop: advancing the research on myalgic encephalomyelitis/chronic fatigue syndrome. Ann Intern Med. 2015; 162:860-5.

16 Haney E, Smith MEB, McDonagh M, Pappas M, Daeges M, Wasson N, et al. Diagnostic methods for myalgic encephalomyelitis/chronic fatigue syndrome: a systematic review for a National Institutes of Health Pathways to Prevention Workshop. Ann Intern Med. 2015; 162:834-40.

17 Smith MEB, Haney E, McDonagh M, Pappas M, Daeges M, Wasson N, et al. Treatment of myalgic encephalomyelitis/chronic fatigue syndrome: a systematic review for a National Institutes of Health Pathways to Prevention Workshop. Ann Intern Med. 2015; 162:841-50.

On 2015 Sep 14, Tom Kindlon commented:

Comment 2 of 2 formally submitted on September 9, 2015 using instructions here: http://www.cochranelibrary.com/help/submitting-comments-on-cochrane-reviews.html

(This is available in one piece at: http://forums.phoenixrising.me/index.php?threads/my-two-detailed-comments-on-the-cochrane-exercise-therapy-for-cfs-review-2015.39801/ )

Second comment:

I have decided to split my comment into two as it was getting very long. This is part two.

Variation in interventions

It would have been useful to have some more information on the “exercise with pacing” intervention tested in the Wallman et al. (2004) trial and how it was distinct from some other exercise interventions tested. The authors say (1): “On days when symptoms are worse, patients should either shorten the session to a time they consider manageable or, if feeling particularly unwell, abandon the session altogether” (p. 143). I don't believe the description given in the review conveys this. In the review, this approach is described as "Exercise with pacing: exercise in which the incremental increase in exercise was personally set." But Wallman et al.’s approach allows patients to decrease as well as increase how much exercise they do on the day. This approach also contrasts with how White (an investigator in two of the trials) has described graded exercise therapy: "if [after increasing the intensity or duration of exercise] there has been an increase in symptoms, or any other adverse effects, they should stay at their current level of exercise for a further week or two, until the symptoms are back to their previous levels" (2). In the PACE Trial manual White co-wrote (3), the GET intervention was guided by the principle that “planned physical activity and not symptoms are used to determine what the participant does” (p. 21); similarly, “it is their planned physical activity, and not their symptoms, that determine what they are asked to do” (p. 20). Compliance data would help us examine which approach patients are actually using: I suspect many patients are in fact doing exercise with pacing even in trials such as the PACE Trial (i.e. when they have increased symptoms, often reducing levels of exercise and sometimes doing no exercise activities at all on that day).

Bimodal versus Likert scoring in Wearden et al. (2010)

I find it odd that the fatigue scores for the Wearden et al. (2010) trial (4) are given in the 0-33 format rather than the 0-11 scoring method. The 0-11 scoring system is what is mentioned as a primary outcome measure in the protocol and is what is reported in the main paper reporting the results (4, 5). It is even what your own report says on p. 44 is the scoring method (“Fatigue Scale, FS; 11 items; each item was scored dichotomously on a 4-point scale [0, 0, 1 or 1]”). This is important because using the scoring method for which you don't report data (0-11), there is no statistically significant difference at the primary outcome point of 70 weeks (5).

Diagnostic criteria

One problem with using these trials as an evidence base, which I don't believe was mentioned, is that all the trials used the Oxford and Fukuda diagnostic criteria (6, 7). Neither of these criteria require patients to have post-exertional malaise (or something similar). Many consider this to be a core symptom of ME/CFS and it is mandatory in most of the other major criteria (8-11). [Aside: The London criteria were assessed in the PACE Trial (12) but they seem to have been operationalised in an unusual way. Ninety seven per cent of the participants who satisfied the (broad) Oxford criteria who didn't have a psychiatric disorder satisfied the definition of M.E. used (13). Ellen Goudsmit, one of the authors of the London criteria, has rejected the way they were used in the PACE Trial (14)]. So this lack of requirement for patients to have post-exertional malaise (or a similar description) means we cannot be sure that the evidence can be generalised to such patients. An independent National Institutes of Health committee this year concluded "continuing to use the Oxford definition may impair progress and cause harm. Therefore, for progress to occur, we recommend that this definition be retired" (15). An Agency for Healthcare Research and Quality review of diagnostic methods this year reached a similar conclusion: "Consensus groups and researchers should consider retiring the Oxford case definition because it differs from the other case definitions and is the least restrictive, probably including individuals with other overlapping conditions” (16). An Agency for Healthcare Research and Quality review of ME/CFS treatments said: "The Oxford CFS case definition is the least restrictive, and its use as entry criteria could have resulted in selection of participants with other fatiguing illnesses or illnesses that resolve spontaneously with time" (17).

Exclusion of some data from analyses due to baseline differences

It seems unfortunate that some data cannot be used due to baseline differences e.g. "Four trials (669 participants) contributed data for evaluation of physical functioning at follow-up (Jason 2007; Powell 2001; Wearden 2010; White 2011). Jason 2007 observed better results among participants in the relaxation group (MD 21.48, 95% CI 5.81 to 37.15). However, results were distorted by large baseline differences in physical functioning between the exercise and relaxation groups (39/100 vs 54/100); therefore we decided not to include these results in the meta-analysis". It would be good if other methods could be investigated (e.g. using baseline levels as covariates) to analyse such data.

Thank you for taking the time to read my comments.

Tom Kindlon

Conflict of Interest statement:

I am a committee member of the Irish ME/CFS Association and do a variety of unpaid work for the Association.

References

1 Wallman KE, Morton AR, Goodman C, Grove R. Exercise prescription for individuals with chronic fatigue syndrome. Med J Aust. 2005;183:142-3.

2 White P. How exercise can help chronic fatigue syndrome. Pulse: 1998. June 20:86-87.

3 Bavinton J, Darbishire L, White PD -on behalf of the PACE trial management group. Graded Exercise Therapy for CFS/ME (Therapist Manual) http://www.pacetrial.org/docs/get-therapist-manual.pdf

4 Wearden AJ, Riste L, Dowrick C, Chew-Graham C, Bentall RP, Morriss RK, Peters S, Dunn G, Richardson G, Lovell K, Powell P. Fatigue Intervention by Nurses Evaluation--the FINE Trial. A randomised controlled trial of nurse led self-help treatment for patients in primary care with chronic fatigue syndrome: study protocol. [ISRCTN74156610]. BMC Med. 2006 Apr 7;4:9.

5 Wearden AJ, Dowrick C, Chew-Graham C, Bentall RP, Morriss RK, Peters S, Riste L, Richardson G, Lovell K, Dunn G; Fatigue Intervention by Nurses Evaluation (FINE) trial writing group and the FINE trial group. Nurse led, home based self help treatment for patients in primary care with chronic fatigue syndrome: randomised controlled trial. BMJ. 2010 Apr 23;340:c1777. doi: 10.1136/bmj.c1777.

6 Sharpe M, Archard L, Banatvala J, Borysiewicz LK, Clare AW, David A, et al. Chronic fatigue syndrome: guidelines for research. Journal of the Royal Society of Medicine 1991;84 (2):118–21.

7 Fukuda K, Straus SE, Hickie I, et al. The chronic fatigue syndrome: A comprehensive approach to its definition and study. Ann Intern Med. 1994; 121: 953‑959.

8 Carruthers BM, Jain AK, De Meirleir KL, et al. Myalgic Encephalomyelitis/chronic fatigue syndrome: Clinical working case definition, diagnostic and treatments protocols. Journal of Chronic Fatigue Syndrome. 2003; 11: 7-115.

9 Carruthers BM, van de Sande MI, De Meirleir KL, et al. Myalgic encephalomyelitis: International Consensus Criteria. J Intern Med. 2011; 270: 327-338.

10 IOM (Institute of Medicine). Beyond myalgic encephalomyelitis/chronic fatigue syndrome: Redefining an illness. Washington, DC: The National Academies; 2015.

(continues)

On 2015 Sep 14, Tom Kindlon commented:

(continues)

References:

1 Turner L, Boutron I, Hróbjartsson A, Altman DG, Moher D: The evolution of assessing bias in Cochrane systematic reviews of interventions: celebrating methodological contributions of the Cochrane Collaboration. Syst Rev 2013, 2:79.

2 Chalder T, Goldsmith KA, White PD, Sharpe M, Pickles AR. Rehabilitative therapies for chronic fatigue syndrome: a secondary mediation analysis of the PACE trial. Lancet Psychiatry. 2015;2:141-152.

3 White PD, Goldsmith KA, Johnson AL, Potts L, Walwyn R, DeCesare JC, et al. Comparison of adaptive pacing therapy, cognitive behaviour therapy, graded exercise therapy, and specialist medical care for chronic fatigue syndrome (PACE): a randomised trial. The Lancet 2011;377:823-36.

4 Kindlon T. Reporting of Harms Associated with Graded Exercise Therapy and Cognitive Behavioural Therapy in Myalgic Encephalomyelitis/Chronic Fatigue Syndrome. Bull IACFS ME. 2011;19:59-111. http://iacfsme.org/ME-CFS-Primer-Education/Bulletins/BulletinRelatedPages5/Reporting-of-Harms-Associated-with-Graded-Exercise.aspx

5 Lipkin DP, Scriven AJ, Crake T, Poole-Wilson PA (1986). Six minute walking test for assessing exercise capacity in chronic heart failure. British Medical Journal 292, 653-5.

6 Kadikar A, Maurer J, Kesten S. The six-minute walk test: a guide to assessment for lung transplantation. J Heart Lung Transplant. 1997 Mar;16(3):313-9.

7 Friedberg F, Sohl S. Cognitive-behavior therapy in chronic fatigue syndrome: is improvement related to increased physical activity? J Clin Psychol. 2009 Feb 11.

8 Friedberg F. Does graded activity increase activity? A case study of chronic fatigue syndrome. Journal of Behavior Therapy and Experimental Psychiatry, 2002, 33, 3-4, 203-215

9 Results and In-depth Analysis of the 2012 ME Association Patient Survey Examining the Acceptability, Efficacy and Safety of Cognitive Behavioural Therapy, Graded Exercise Therapy and Pacing, as Interventions used as Management Strategies for ME/CFS. Gawcott, England. http://www.meassociation.org.uk/2015/05/23959/ Accessed: September 3, 2015

10 Critical Illness - A Dreadful Experience with Scottish Provident. http://forums.moneysavingexpert.com/showthread.php?t=2356683 Accessed: September 4, 2015

11 McCrone P, Sharpe M, Chalder T, Knapp M, Johnson AL, Goldsmith KA, White PD. Adaptive pacing, cognitive behaviour therapy, graded exercise, and specialist medical care for chronic fatigue syndrome: a cost-effectiveness analysis. PLoS One. 2012;7(8):e40808.

12 Rapport d’évaluation (2002-2004) portant sur l’exécution des conventions de rééducation entre le Comité de l’assurance soins de santé (INAMI) et les Centres de référence pour le Syndrome de fatigue chronique (SFC). 2006. http://health.belgium.be/internet2Prd/groups/public/@public/@shc/documents/ie2divers/14926531_fr.pdf (Starts on page 223.) Accessed September 4, 2015 (French language edition)

13 Evaluatierapport (2002-2004) met betrekking tot de uitvoering van de revalidatieovereenkomsten tussen het Comité van de verzekering voor geneeskundige verzorging (ingesteld bij het Rijksinstituut voor Ziekte- en invaliditeitsverzekering) en de Referentiecentra voor het Chronisch vermoeidheidssyndroom (CVS). 2006. http://health.belgium.be/internet2Prd/groups/public/@public/@shc/documents/ie2divers/14926531.pdf (Starts on page 227.) Accessed September 4, 2015 (Dutch language version)

14 Stordeur S, Thiry N, Eyssen M. Chronisch Vermoeidheidssyndroom: diagnose, behandeling en zorgorganisatie. Health Services Research (HSR). Brussel: Federaal Kenniscentrum voor de Gezondheidszorg (KCE); 2008. KCE reports 88A (D/2008/10.273/58) https://kce.fgov.be/sites/default/files/page_documents/d20081027358.pdf Accessed September 4, 2015

15 Wiborg JF, Knoop H, Stulemeijer M, Prins JB, Bleijenberg G. How does cognitive behaviour therapy reduce fatigue in patients with chronic fatigue syndrome? The role of physical activity. Psychol Med. 2010; 40:1281-1287.

16 Van Kessel K, Moss-Morris R, Willoughby, Chalder T, Johnson MH, Robinson E, A randomized controlled trial of cognitive behavior therapy for multiple sclerosis fatigue, Psychosom. Med. 2008; 70:205–213.

17 Powell P. FINE Trial Patient Booklet http://www.fine-trial.net/downloads/Patient PR Manual ver9 Apr05.pdf Accessed September 7, 2015

18 Twisk FNM, Maes M. A review on Cognitive Behavorial Therapy (CBT) and Graded Exercise Therapy (GET) in Myalgic Encephalomyelitis (ME)/Chronic Fatigue Syndrome (CFS): CBT/GET is not only ineffective and not evidence-based, but also potentially harmful for many patients with ME/CFS. Neuro Endocrinol Lett. 2009;30:284-299.

19 Carruthers BM et al. Myalgic Encephalomyelitis – Adult & Paediatric: International Consensus Primer for Medical Practitioners. ISBN 978-0-9739335-3-6 http://www.investinme.org/Documents/Guidelines/Myalgic Encephalomyelitis International Consensus Primer -2012-11-26.pdf Accessed September 5, 2015

20 Twisk FN. Objective Evidence of Post-exertional “Malaise” in Myalgic Encephalomyelitis and Chronic Fatigue Syndrome. J Sports Med Doping Stud 2015. 5:159. doi: 10.4172/2161-0673.1000159

21 Higgins JPT, Green S: Cochrane Handbook for Systematic Reviews of Interventions Version 5.1.0 [updated March 2011]. Table 8.5.d. The Cochrane Collaboration; 2011. http://handbook.cochrane.org/chapter_8/table_8_5_d_criteria_for_judging_risk_of_bias_in_the_risk_of.htm Accessed: September 5, 2015

22 White PD, Sharpe MC, Chalder T, DeCesare JC, Walwyn R; on behalf of the PACE trial group. Protocol for the PACE trial: A randomised controlled trial of adaptive pacing, cognitive behaviour therapy, and graded exercise as supplements to standardised specialist medical care versus standardised specialist medical care alone for patients with the chronic fatigue syndrome/myalgic encephalomyelitis or encephalopathy. BMC Neurology 2007, 7:6 http://www.biomedcentral.com/1471-2377/7/6 Accessed: September 5, 2015

On 2015 Sep 14, Tom Kindlon commented:

(contd.)

Compliance

The review doesn't include any information on compliance. I'm not sure that there is much published information on this but I know there was a measure based on attendance at therapy sessions (which could be conducted over the phone) given for the PACE Trial (3). Ideally, it would be interesting if you could obtain some unpublished data from activity logs, records from heart-rate monitors, and other records to help build up a picture of what exercise was actually performed and the level of compliance. Information on adherence and what exercise was actually done is important in terms of helping clinicians, and indeed patients, to interpret and use the data. I mention patients because patients' own decisions about their behaviour is likely to be affected by the medical information available to them, both within and outside of a supervised programme of graded exercise; unlike with an intervention like a drug, patients can undertake exercise without professional supervision.

"Selective reporting (outcome bias)" and White et al. (2011)

I don't believe that White et al. (2011) (the PACE Trial) (3) should be classed as having a low risk of bias under "Selective reporting (outcome bias)" (Figure 2, page 15). According to the Cochrane Collaboration's tool for assessing risk of bias (21), the category of low risk of bias is for: "The study protocol is available and all of the study’s pre-specified (primary and secondary) outcomes that are of interest in the review have been reported in the pre-specified way". This is not the case in the PACE Trial. The three primary efficacy outcomes can be seen in the published protocol (22). None have been reported in the pre-specified way. The Cochrane Collaboration's tool for assessing risk of bias states that a “high risk” of bias applies if any one of several criteria are met, including that “not all of the study’s pre-specified primary outcomes have been reported” or “one or more primary outcomes is reported using measurements, analysis methods or subsets of the data (e.g. subscales) that were not pre-specified”. In the PACE Trial, the third primary outcome measure (the number of "overall improvers") was never published. Also, the other two primary outcome measures were reported using analysis methods that were not pre-specified (including switching from the bimodal to the Likert scoring method for The Chalder Fatigue Scale, one of the primary outcomes in your review). These facts mean that the “high risk of bias” category should apply.

Thank you for taking the time to read my comments.

Tom Kindlon

Conflict of Interest statement:

I am a committee member of the Irish ME/CFS Association and do a variety of unpaid work for the Association.

(continues)

On 2015 Sep 14, Tom Kindlon commented:

Comment 1 of 2 formally submitted on September 9, 2015 using instructions here: http://www.cochranelibrary.com/help/submitting-comments-on-cochrane-reviews.html

(This is available in one piece at: http://forums.phoenixrising.me/index.php?threads/my-two-detailed-comments-on-the-cochrane-exercise-therapy-for-cfs-review-2015.39801/ )

I would first like to thank those involved for their work in preparing this document. Even for those of us who have read the individual Chronic Fatigue Syndrome (CFS) papers it is useful to have the results collated, as well as details regarding the interventions. Also it is interesting to see the results of sensitivity analyses, subgroup analyses, standardised mean differences, etc.

I would like to make a few comments. I’m splitting them into two submissions as the piece had become very long. I’ve added some loose headings to hopefully make it more readable.

Objective measures

The review assessed the studies as having a high risk of bias regarding blinding, since neither participants nor assessors were blinded. Evidence suggests that subjective outcomes are more prone to bias than objective outcomes when there is no blinding (1). It is thus unfortunate that the review concentrated almost exclusively on subjective measures, failing to include results from nearly all the objective outcome measures that have been published with trials. (The exception was health resource use for which you presented follow-up data from one trial).

I hope objective outcome data can be included in a future revision or edition of this review.

Examples of objective outcomes include: exercise testing (work capacity by oxygen consumption); fitness test/step test; the six minute walking test; employment status; and disability payments.

Adding in these results would allow a more rigorous assessment of the effectiveness and relevance of the therapies, their causal mechanisms, therapeutic compliance, and safety.

On exercise testing, for example, in the PACE Trial (the largest trial in the review) there was no improvement in fitness levels as measured by a step test (2). The fitness data contrasts sharply with the many positive results from subjective self-report measures in the trial, so one is left wondering how much the subjective measures reflect reality.

On another exercise test used in the PACE Trial, the 6 minute walk test, there was a small (mean) increase from 312 metres at baseline to 379 metres at 12 months: this was 35.3 metres more than the "passive" control group when adjustments were made. However, the final result of 379 metres remains very poor compared to the more than 600 metres one would expect from healthy people of a similar age and gender make-up (3,4). By comparison, a group with Class III heart failure walked an average of 402 metres (5). A score of less than 400 metres has been suggested as the level at which somebody should be put on a lung transplant list (6). Such information from objective measures helps to add important context to the subjective measures and restraint to the conclusions that can be drawn from them.

Objective data is also needed to check compliance with a therapy. If patients diligently exercised for 12 months one would expect much better results on fitness and exercise testing than the aforementioned results in the PACE Trial. This is important when considering adverse events and safety: such trials may not give us good information on the safety of complying with such interventions if patients haven't actually complied.

Employment and receipt of disability payments are practical objective measures of general functional capacity so data on them would help establish whether patients can actually do more overall or whether they may just be doing, for example, a little more exercise but have substituted that for doing less in other areas (7,8). Also, CFS patients are sometimes pressured by insurance companies into doing graded exercise therapy (GET) programs so it would be useful to have data collated on employment outcomes to see whether pressure can in any way be justified (9,10). In the PACE Trial, there was no significant improvement in employment measures and receipt of disability payments in the GET group (11). Outside the realm of clinical trials, the quantitative and qualitative data in a major (UK) ME Association survey also found that GET didn't lead to higher levels of employment and lower levels of receipt of disability payments on average (9). Also, extensive external audits were performed of Belgian CFS rehabilitation clinics that treated using cognitive behavioural therapy (CBT) and GET. The main reports are in French and Dutch (12,13), with an English summary available (14) that says, "Employment status decreased at the end of the therapy, from an average of 18.3% of a 38h working week, to 14.9% [...] The percentage of patients living from a sickness allowance increased slightly from 54 to 57%." This contrasts with the average improvements reported in the audit for some symptoms like fatigue.

While data on (self-reported) symptoms like fatigue (one of your two primary outcomes) is interesting, arguably more important to patients is improving their overall level of functioning (and again, objective measures are needed here). Being able to work, for example, despite experiencing a certain level of fatigue would likely be more important for many than being unable to work but having slightly lower levels of fatigue.

An example of how reductions in the reported levels of fatigue may not lead to improvements in functioning can be seen in an analysis of three graded activity-oriented CBT therapy interventions for CFS (15). The analysis showed, compared to controls, there were no improvements in overall activity levels as measured by actometers despite improvements in self-reported fatigue (15). Activity in these trials was assessed using actometers. Another study that exemplifies the problem of focusing too much on fatigue scores after behavioural interventions is a study of CBT in multiple sclerosis (MS) patients with “MS fatigue”(16). The study found that following the intervention, patients with MS reported significantly lower (i.e. better) scores on the Chalder Fatigue Scale (0-33 scoring) than those in a healthy, nonfatigued comparison group! This significant difference was maintained at 3 and 6 months’ follow-up. It is difficult to believe that patients with MS fatigue (at baseline) truly subsequently had less fatigue than healthy nonfatigued controls: a much more likely scenario is that undertaking the intervention had led to response biases.

You mention that "many patient charities are opposed to exercise therapy for chronic fatigue syndrome (CFS)". One reason for concern about the way in which exercise programmes are promoted to patients is that they are often based upon models which assume that there is no abnormal physiological response to exercise in the condition, and make unsupported claims to patients. For example, in the FINE trial (Wearden et al., 2010) patient booklet (17), it is boldly asserted that: "Activity or exercise cannot harm you" (p. 49). However, a large number of studies have found abnormal responses to exercise, and the possibility of harm being done simply cannot be excluded on the basis of current evidence (discussed in 4, 18-20)."

(continues)

On 2015 Feb 24, Laurie Thomas commented:

Clinical studies of chronic fatigue syndrome are plagued by serious problems in the inclusion/exclusion criteria. These problems stem from the fact that the syndrome consists of nonspecific symptoms that are "medically unexplained." However, there is a major difference between medically unexplained and medically inexplicable. The symptoms of chronic fatigue syndrome can result from a serious circulatory problem that is easily overlooked. In 2003, Peckerman and coworkers showed that low cardiac output, as measured by impedance cardiography, predicts the severity of symptoms in CFS patients.[1] Miwa and Fujita found a small left ventricular size leading to low cardiac output in CFS patients with orthostatic intolerance.[2] Porter and coworkers reported that a case of femoral arteriovenous fistula causing high-output cardiac failure was originally misdiagnosed as chronic fatigue syndrome.[3]

The studies of graded exercise for management of CFS are based on the presumption that CFS is the result of laziness and deconditioning and that the solution to the problem is to persuade the patient to exercise. Yet in many reported cases, the real problem was unrecognized cardiac decompensation. This state of cardiac decompensation could account for the push-crash phenomenon (serious, prolonged adverse events from overexertion) among people with CFS. Thus, a graded exercise program that might be beneficial for the large number of people who are tired and achy because of major depressive disorder could be catastrophic for the relatively small number of people whose problem is due to cardiac decompensation. Unfortunately, the existing studies of exercise for management of CFS do not shed light on this problem. The patients whose exercise intolerance is too severe to allow them to participate in the exercise program might refuse to enroll or might be dismissed as noncompliant if they try but fail to exercise. Yet as a result of the positive results of graded exercise for subjects whose real problem is major depressive disorder, patients with unrecognized cardiac decompensation are being scolded for failing to exercise.

For ethical and scientific reasons, the protocol for a clinical study of subjects with CFS should be based on the best possible model for clinical management of CFS patients. It would begin with a careful assessment of the subject's circulatory status. This assessment should include a tilt-table test, or at least a measurement of supine, sitting, and standing pulse and blood pressure. Any circulatory problem should be addressed appropriately. (Note that once the patient's condition is found to be due to a circulatory problem, the patient no longer fits the inclusion criteria of "medically unexplained" symptoms.)

As improper diet is the most prevalent cause of chronic ill-health, the cardiology assessment should be followed by a run-in period of at least a week of optimal dietary management. Subjects should be fed a low-fat (<10% of calories), purely plant-based diet that excludes the most common causes of food allergies or intolerance syndromes (i.e., wheat, rye, barley, corn, soy, strawberries, and citrus fruits). To ensure adherence, the diet should be administered in a residential setting. This kind of low-fat, plant-based diet can bring about a significant drop in blood pressure in hypertensive patients within 7 days, even if the patients stop taking blood pressure medication at baseline.[4] This correction of hypertension results from the decrease in systemic resistance. Thus, this diet could lead to a significant improvement in circulation, which would be beneficial to patients whose symptoms are due to poor circulation, even if they are not hypertensive. Note also that the elimination of poorly tolerated foods is the only reliable way to establish that the patient's problem is due to a food intolerance. Of course, once the subject's problem has been shown to be dietary in origin, the subject no longer has "medically unexplained" symptoms and thus no longer fits the inclusion criteria for a study of CFS.

Many patients with a diagnosis of CFS are inactive, but they may be inactive because they are sick, rather than being sick because they are inactive. Thus, any study of exercise and CFS should be structured to establish the direction of causality. If a study of subjects with a diagnosis of CFS involves exercise, the outcome variables must involve some measurement of the subjects' overall activity levels, not just to assess compliance with the exercise program but to assess whether the subjects are merely wasting their energy on the exercises and thus become less able to perform activities of daily living. In that situation, the exercise program could actually decrease the subject's quality of life.

[1] Peckerman A, LaManca JJ, Dahl KA, Chemitiganti R, Qureishi B, Natelson BH. Abnormal impedance cardiography predicts symptom severity in chronic fatigue syndrome. Am J Med Sci. 2003 Aug;326(2):55-60.

[2] Miwa K1, Fujita M. Small heart with low cardiac output for orthostatic intolerance in patients with chronic fatigue syndrome.Clin Cardiol. 2011 Dec;34(12):782-6. doi: 10.1002/clc.20962. Epub 2011 Nov 28.

[3] Porter J1, Al-Jarrah Q1, Richardson S. A case of femoral arteriovenous fistula causing high-output cardiac failure, originally misdiagnosed as chronic fatigue syndrome. Case Rep Vasc Med. 2014;2014:510429. doi: 10.1155/2014/510429. Epub 2014 May 20.

[4] McDougall J1, Thomas LE, McDougall C, Moloney G, Saul B, Finnell JS, Richardson K, Petersen KM.Effects of 7 days on an ad libitum low-fat vegan diet: the McDougall Program cohort. Nutr J. 2014 Oct 14;13:99. doi: 10.1186/1475-2891-13-99.

On 2015 Feb 19, Joan Crawford commented:

This review states: “Chronic fatigue syndrome (CFS) is characterised by persistent, medically unexplained fatigue, as well as symptoms such as musculoskeletal pain, sleep disturbance, headaches and impaired concentration and short-term memory.”

This is important because the above description of CFS and the addition of trials in the review only requiring chronic fatigue as an inclusionary requirement (Sharpe et al, 1991) makes generalisation of the findings problematic as many patients with major depressive disorder (MDD) would also meet the above description of CFS and Sharpe et al.'s (1991) criteria if their condition was fatiguing – a common feature - along with muscular aches and pains, sleep disturbance, cognitive difficulties and so on. The high percentage of patients included in these trials suffering from depression (Table 1. Study demographics) indicates this may be their primary condition – confounding the results. Exercise, through behavioural activation programs, has a moderately positive impact on patients with depression (Cooney et al., 2013). It is unclear whether the modest improvement seen in some of these trials can be accounted for by an improvement in low mood caused by depression. Moreover, where there is data there is a high usage of antidepressants in patients included in the reviewed trials (Table 1. Study demographics).

Of the eight exercise trials included in this review, five used broad inclusion criteria (Sharpe et al, 1991) (N=1287) – 85% of all participants. Two of these studies also used a version of the London criteria, which did not exclude patients with depression and other psychiatric conditions as originally specified by the authors making it hard to assess how these criteria were operationalised. Three further trials used the CDC Fukuda (1994) CFS criteria (N=231). While these purport to be more selective, they do not necessary include patients whose primary difficulties include post exertional weakness and debility and flu-like symptoms and so on beyond broadly defined fatigue and other general symptoms which could be attributed to CFS or MDD.

There is also an issue with lack of evidence of patients’ fidelity to exercise programs using objective measures. We do not know if patients increased their activity as suggested to them by their clinicians. Without using devises such as actimeters or pedometers to track daily activity levels we have no accurate way of assessing whether an increase in activity occurred and whether this helps. Black & McCully’s (2005) study demonstrates objectively the difficulties patients face when trying to increase activity and concluded that they were exercise intolerant, unable to sustain activity targets.

The report is bold in stating “no evidence suggests that exercise therapy may worsen outcomes“. Many patient surveys from across the world report numerous instances of harm and worsening of symptoms from taking part in exercise programs. For a summary of the difficulties and limitations of the reporting of harms, in and outside of clinical trials, and why these might be underestimated please see Kindlon (2011).

References

Cooney GM, Dwan K, Greig CA, Lawlor DA, Rimer J, Waugh FR, McMurdo M, Mead GE (2013). Exercise for depression. The Cochrane Library. http://onlinelibrary.wiley.com/doi/10.1002/14651858.CD004366.pub6/abstract

Fukuda, K., Straus, S.E., Hickie, I., Sharpe, M.C., Dobbins, J.G., & Komaroff, A. (1994). The chronic fatigue syndrome: A comprehensive approach to its definition and study. International chronic fatigue syndrome study group. Annals of Internal Medicine, 121(12), 953-959.

Kindlon T. (2011). Reporting of harms associated with graded exercise therapy and cognitive behavioural therapy in Myalgic Encephalomyelitis/chronic fatigue syndrome. Bulletin of the IACFS/ME. 19(2): 59-111.

M, Archard L, Banatvala J, Borysiewicz LK, Clare AW, David A, et al. (1991). Chronic fatigue syndrome: guidelines for research. Journal of the Royal Society of Medicine, 84(2):118–21.

On 2015 Feb 14, Ellen M Goudsmit commented:

I had contact with the main author to alert her to certain misconceptions published earlier. Sadly, I found I had wasted my time.

For example, we can not tell how many, if any, patients in the PACE trial met the London criteria. Having read that the researchers planned to select individuals with ME and had listed the criteria in the protocol, I checked that Prof. White would use the original version which had not been published. I had been the Chair of the Research Working Group at AFME when they were being tested and still had a copy. They came with a questionnaire as well as a physician to establish their reliability. Prof. White was unwilling to confirm that he would use the original so in light of the uncertainty, I requested that he did not cite me as a co-author. I did not work on the lay version published in the Westcare report which I felt was deeply flawed. I was right to be cautious. The trial manual indicates that the researchers adapted the lay version and I could tell from the results that the London criteria were not used as they exclude individuals with psychological disorders so the percentage for that variable should have been nil. It wasn’t.

A second point. The review does not pay the required attention to the lack of actigraphy, an objective measure to confirm fidelity to the protocol. This has been included in most studies conducted in the USA and the Netherlands. The results from actigraphy indicate that, except for 7 individuals, there were no significant increases in activity after GET and similar therapies. According to Friedberg who assessed the phenomenon, patients on exercise trials tend to reprioritise their activities, choosing those that result in less stress etc. In short, they learn to pace themselves (Goudsmit et al 2012). That is why they feel better and less fatigued, but it's not possible to attribute improvement to an increase in activity (or fitness).

Pacing was not defined and adaptive pacing therapy (APT) refers to a programme consisting of several components including stress management, advice on sleeping etc. There are no data for pacing alone in the PACE trial, so to conclude that GET is superior to pacing therapies is premature. There is only one pacing therapy. Pacing is not a therapy. It's a simple strategy. Research by Jason suggests that people who pace themselves feel better, irrespective of the protocol they are on.

Finally, we know that many patients have adverse reactions to activity. It's a criterion for diagnosis. To dismiss them ("no evidence that exercise therapy worsens outcomes") is hard to comprehend. Every survey in every country to date has revealed that GET does have marked adverse reactions and can result in relapse. See also Sisto et al and Black and McCully, cited in Goudsmit et al 2012.

To summarise: lack of a definition of pacing resulting in confusion, repetition of incorrect information, failure to consider the findings from objective measures suggesting patients did not adhere to the protocol and ignoring consistent reports from surveys that undermine one's conclusions. I expect more objectivity and attention to detail from the Cochrane Library.

Goudsmit, EM., Jason, LA, Nijs, J and Wallman, KE. Pacing as a strategy to improve energy management in myalgic encephalomyelitis/chronic fatigue syndrome: A consensus document. Disability and Rehabilitation, 2012, 34, 13, 1140-1147. Online 19th December. doi: 10.3109/09638288.2011.635746.

On 2015 Mar 12, Donald Forsdyke commented:

ADAPTATION DECOUPLED FROM SPECIATION. The authors begin by noting that gene flow among populations, rather than assisting speciation, has a “greater impact” when acting “as a homogenizing force, reuniting populations that might otherwise have had separate evolutionary trajectories.” This blending effect, which thwarts speciation, is prevented by some form of reproductive isolation dependent upon an internal or external barrier. The nature of the barrier varies with time. The present work reflects on the order in which barriers arise, an early barrier being superceded by a later.

The two fern types, having diverged 60 million years ago, can still transfer gametes abiotically (so there are no genic incompatibilities conferring prezygotic isolation), and the resulting zygotes can still develop (so there are no genic incompatibilities conferring hybrid inviability). Thus, they are reproductively isolated to a degree sufficient to prevent blending, solely by virtue of hybrid sterility.

Following a line of reasoning that dates back to Romanes (1886), a failure of meiotic pairing, due to the accumulation of base differences in parental DNA sequences (that would not necessarily affect genes), results in sterile hybrids. The line cannot advance, so that in evolutionary terms the parents are reproductively isolated from each other. Over 60 million years, this chromosome pairing barrier would have been elevated by addition of macroscopically observable deletions, duplications or inversions. Accompanying genic changes would have affected phenotypic functions other than those affecting fertilization and development. Thus secondary adaptations would be decoupled from the primary speciation event. This is in keeping with the recent conclusion of Hedges et al. (2015) that “if adaptation is largely decoupled from speciation, we should not expect it to be a driver of speciation” (see: http://1.usa.gov/1ESNM0W where further references may be found).

On 2015 Jun 10, Paul Golding commented:

Baggott and Tamura imply that my experiment was flawed because: “The study design by Golding (2014) should have included a balanced supplementation of micronutrients at the level of generally recommended dietary intakes.” and advise that “Therefore, the data presented by Golding (2014) must be cautiously interpreted”.

My full response is available here.

Was the 500 mg/day vitamin C supplement excessive, and could it have abnormally extended the time taken to deplete the liver folate store?

Baggott and Tamura claim that my 500 mg/day vitamin C supplement was excessive, and that my saturated vitamin C status was abnormal. They propose that by abnormally protecting the reduced (unstable) form of folate from oxidation, which they claim would not have been preserved by recommended vitamin C intakes, the “excessive” vitamin C preserved the liver folate store. I strongly disagree with these assertions.

Carr et al (2012), Levine et al. (1996) and Lykkesfeldt and Poulsen (2010) recommend that vitamin C intake should be sufficient to ensure plasma saturation. If the plasma vitamin C concentration is insufficient to prevent oxidation of fully reduced folate, although sufficient to prevent scurvy, there would be sub-optimal vitamin C status.

There is no evidence that, once there is sufficient vitamin C present to ensure plasma saturation, and thereby protect the fully reduced folates, additional amounts will affect the time taken to develop folate deficiency.

Levine et al. (1996) recommend 200 mg vitamin C daily, very significantly higher than the 70 mg taken by Herbert (Herbert 1962), and a level that ensures plasma saturation. They plotted plasma vitamin C concentration against the daily vitamin C dose (Levine et al. 1996 Figure 1C); this produced a sigmoid curve with 200 mg the lowest dose to reach a plateau.

I used data from Levine et al. (1996 Table 1) to produce a chart to illustrate the relationship between vitamin C dose and the plasma concentration, at relevant levels. At Herbert’s dose of 70 mg/day, plasma concentration is 32 µmol/L; at 200 mg/day recommended by Levine et al. (1996), plasma concentration is 66 µmol/L; at my dose of 500 mg/day, plasma concentration is 71 µmol/L. A 500 mg dose of vitamin C will not produce a significantly higher plasma concentration than the 200 mg dose recommended by Levine et al. (1996), but a 70 mg dose will produce a very significantly lower plasma concentration. The Excel file, high-resolution PDF and Microsoft PowerPoint slide for my chart are available at the link above.

The predicted plasma vitamin C concentration of 71 µmol/L, produced by my intake of 500 mg/day, is very close to the value considered by Carr et al. (2012) to be necessary for good health.

The vitamin C supplement used by me was integrated with the iron tablets taken to prevent the iron deficiency reported by Herbert. As stated in Golding (2014), the iron tablets (Abbott Australia Ferro-Grad C) comprised 325 mg ferrous sulphate (equivalent to 105 mg elemental iron), with 562 mg sodium ascorbate (equivalent to 500 mg vitamin C). Sodium ascorbate is necessary to ensure adequate absorption of the iron (Hurrell 2002, McCurdy 1968).

Was the 1000 µg/day vitamin B12 supplement excessive, and could it have abnormally extended the time taken for me to deplete my liver folate store?

Although Baggott and Tamura raise the possibility of such a confounding effect, they have not suggested any mechanism for it, or cited any published reports of such findings.

I found no published reports of vitamin B12 used to overcome a dietary folate deficiency, or any findings of interference by vitamin B12 supplements with folate metabolism where there was no initial vitamin B12 deficiency. An initially untreated vitamin B12 deficiency can cause a secondary functional folate deficiency; the “methylfolate trap” (Tisman and Herbert 1973). Such a secondary folate deficiency might be corrected by vitamin B12 supplements, depending on the cause of the vitamin B12 deficiency, but there is no published evidence that increasing the vitamin B12 intake beyond that will have any effect on folate metabolism.

As explained in Golding (2014), the 1000 µg/day vitamin B12 supplement was necessary to prevent vitamin B12 deficiency. Extensive testing during my previous vitamin B12 investigation showed that my vitamin B12 is absorbed and transported to cells normally but is not utilised adequately within the cells, presumably because of a deficiency of one of the B12 cofactors. At least 250 µg was needed to avoid deficiency but 1000 µg is the most readily available, allows a reasonable margin for error, and is within the recommended range for the oral vitamin B12 dose (Kuzminski et al. 1998).

Did the experiment include appropriate supplementation of micronutrients and was the title adequate?

Baggott and Tamura have not provided any evidence that taking either of these supplements could have interfered with the development of folate deficiency in my experiment, and have not offered any plausible mechanism for such confounding effects. In addition, my supplementation with vitamin C and vitamin B12 was reasonable, in the doses used, to prevent deficiencies.

I therefore disagree with their advice that “Therefore, the data presented by Golding (2014) must be cautiously interpreted”, and with what they imply in their conclusion; that my experiment was flawed because the study design failed to include “a balanced supplementation of micronutrients at the level of generally recommended dietary intakes.”

I also disagree with their assertion that “The title … should have contained words clearly indicating that the subject had a high initial folate status and was saturated with ascorbate.” My replete folate status was clearly stated in Objective, Experiment Design and The Subject. According to Levine et al. (1996), Carr et al. (2012) and Lykkesfeldt and Poulsen (2010), plasma saturation is the normal optimal vitamin C status.

References

Baggott JE, Tamura T (2014) The second study of experimental human folate deficiency. SpringerPlus 3:719

Carr AC, Pullar JM, Moran S, Vissers MC (2012) Bioavailability of vitamin C from kiwifruit in non-smoking males: determination of 'healthy' and 'optimal' intakes. J Nutr Sci 1:e14

Golding PH (2014) Severe experimental folate deficiency in a human subject – a longitudinal study of biochemical and haematological responses as megaloblastic anaemia develops. SpringerPlus 3:442

Herbert V (1962) Experimental nutritional folate deficiency in man. Trans Assoc Am Physicians 75:307–320

Herbert V (1987) Recommended dietary intakes (RDI) of folate in humans. Am J Clin Nutr 45(4):661–670

Hurrell RF (2002) Fortification: overcoming technical and practical barriers. J Nutr 132(4 Suppl):806S–812S

Kuzminski AM, Del Giacco EJ, Allen RH, Stabler SP, Lindenbaum J (1998) Effective treatment of cobalamin deficiency with oral cobalamin. Blood 92(4):1191-1198

Levine M, Conry-Cantilena C, Wang Y, Welch RW, Washko PW, Dhariwal KR, Park JB, Lazarev A, Graumlich JF, King J, Cantilena LR (1996) Vitamin C pharmacokinetics in healthy volunteers: evidence for a recommended dietary allowance. Proc Natl Acad Sci 93(8):3704–3709

Lykkesfeldt J, Poulsen HE (2010) Is vitamin C supplementation beneficial? Lessons learned from randomised controlled trials. Br J Nutr 103(9):1251-1259

McCurdy PR, Dern RJ (1968) Some therapeutic implications of ferrous sulfate-ascorbic acid mixtures. Am J Clin Nutr 21(4):284–288

Tisman G, Herbert V (1973) B12 dependence of cell uptake of serum folate: an explanation for high serum folate and cell folate depletion in B 12 deficiency. Blood 41(3):465–469

On 2015 Mar 10, Ben A Inglis commented:

The parameters for the magnetization preparation (MP) RF pulse are not given in the paper, the supplemental material nor in the authors' previous work (Franklin et al 2013, Franklin et al 2014) on blood flow effects in gray matter measured with MP-RAGE. Was a non-selective or slab-selective inversion pulse used? The latter is expected to have more flow weighting than the former. Also, what was the inversion time (TI)? The value of TI will also influence flow weighting. In this report as well as the two prior reports, the TR quoted is considerably shorter than the TR typically employed when using a TI of 700-900 ms at 3 T. Was the magnetization preparation actually enabled? Finally, given that the paper contends there is an effect of blood flow in assessing gray matter with MP-RAGE, discussion of the effects of the MP scheme on flow weighting is essential. As given the paper fails to explain the mechanism of the blood flow weighting and whether there might be ways to minimize effects, e.g. by using a non-selective inversion and particular inversion times.

On 2016 Jan 12, Tamás Ferenci commented:

I have replied to this paper criticizing certain aspects of it (Ferenci T, 2017), available for download here, to which the original authors also responded (Diamond DM, 2017), available for download here.

On 2015 Nov 01, John Tucker commented:

The authors correctly point out that the use of relative risk information in isolation can be misleading, but this is true of their preferred measure of absolute risk reduction as well. The examples chosen by the authors to illustrate their point are 2 to 5 year trials of drugs that are taken prophylactically for much longer periods. Further, two were stopped early for ethical reasons after crossing the threshold for showing clear clinical benefit, which placed an upper limit on the level of benefit that could be demonstrated. The survival curves from longer term trials such as 4S and WOSCOPS show a progressively larger absolute benefit with longer duration of treatment.

Presented in isolation, both relative and absolute risk reduction can be misleading.

On 2015 Feb 17, Ryan Radecki commented:

Post-publication commentary: "Inappropriately Promoted tPA "Drip and Ship" Safety"

“More community hospitals are giving a powerful clot-busting medication to stroke victims, improving their chances of survival and recovery, new research shows.”

This statement comes from coverage of the American Heart Association release regarding this synopsis of the Get-With-the-Guideline Registry. Part of this statement is true – more community hospitals are using tPA for acute ischemic stroke. In this review of 44,667 patients treated with tPA over the past decade, 23.5% received tPA outside of a specialized stroke or academic center.

The second half of this press statement is false.

Patients treated by the “drip and ship” method, as community administration of tPA is described, did not have an improved chance of survival. Patients treated at community hospitals were younger, had less-severe strokes, and had fewer prior strokes – yet their in-hospital mortality was 10.9%, compared with 9.7%....

http://www.emlitofnote.com/2015/02/inappropriately-promoted-tpa-drip-and.html

On 2015 Feb 17, Ryan Radecki commented:

Post-publication commentary: "Christmas Comes Early for Endovascular Therapy in Stroke"

Quite literally, in fact, considering the timing of the publication of MR-CLEAN – and, now, the triple fall-out from those results.

Due to the positive findings presented by the MR-CLEAN investigators in December 2014, three other ongoing major endovascular trials used this opportunity to check their results early. ESCAPE, EXTEND-IA, and SWIFT-PRIME – all products of the Covidien Solitaire FR clinical trial machine – ceased recruitment and looked to see if they’d met statistical measures of efficacy. Obviously, if you’re reading about all three of these trials here today – presented at the International Stroke Conference yesterday – they all found the results they hoped. ESCAPE and EXTEND-IA were published simultaneously in the NEJM, while SWIFT-PRIME is still in the manuscript drafting stages.

The more robust of the two trials is ESCAPE, whose original target enrollment was 500 patients based on a primary outcome of “ordinal shift analysis” on the modified Rankin scale.....

http://www.emlitofnote.com/2015/02/christmas-comes-early-for-endovascular.html

On 2015 Feb 17, Ryan Radecki commented:

Post-publication commentary: "Christmas Comes Early for Endovascular Therapy in Stroke"

Quite literally, in fact, considering the timing of the publication of MR-CLEAN – and, now, the triple fall-out from those results.

Due to the positive findings presented by the MR-CLEAN investigators in December 2014, three other ongoing major endovascular trials used this opportunity to check their results early. ESCAPE, EXTEND-IA, and SWIFT-PRIME – all products of the Covidien Solitaire FR clinical trial machine – ceased recruitment and looked to see if they’d met statistical measures of efficacy. Obviously, if you’re reading about all three of these trials here today – presented at the International Stroke Conference yesterday – they all found the results they hoped. ESCAPE and EXTEND-IA were published simultaneously in the NEJM, while SWIFT-PRIME is still in the manuscript drafting stages.

EXTEND-IA was a much smaller trial, targeting only 100 patients, with coprimary outcomes of reperfusion and NIHSS improvement at 24 hours....

http://www.emlitofnote.com/2015/02/christmas-comes-early-for-endovascular.html

On 2015 Apr 22, Melissa Raven commented:

Inappropriate analysis of mortality in mental disorders

There are serious problems with this analysis of mortality in mental disorders. Walker et al. have very inappropriately generalized mortality estimates derived from their analysis of clinical samples (primarily tertiary treatment samples), which include high proportions of patients (often inpatients) with psychotic disorders, to cases of common mental disorders (primarily anxiety disorders and mood disorders) identified in population surveys (World Mental Health (WMH) surveys). The authors have thereby grossly exaggerated the population-level mortality due to mental disorders. Unfortunately this has been accepted uncritically by the reviewers, the editors, and many readers.

I have posted a more detailed comment on JAMA Psychiatry http://archpsyc.jamanetwork.com/article.aspx?articleid=2110027

On 2015 Mar 10, Harri Hemila commented:

Vitamin C may help against exercise-induced bronchoconstriction

Boulet LP, 2015 reviewed the treatment options for exercise-induced bronchoconstriction (EIB) and stated that there is insufficient data to recommend vitamins or antioxidants as treatment. This statement is misleading.

Three randomized trials examined the effect of vitamin C on exercise-induced FEV1 decline in asthmatics, see Schachter EN, 1982, Cohen HA, 1997, and Tecklenburg SL, 2007. Each trial found that vitamin C halved the FEV1 decline, with the pooled estimate indicating a reduction of postexercise FEV1 decline by 48% (95%CI: 33% to 64%), see Hemilä H, 2013 and Hemila H, 2014. The studies were carried out in three different decades and on two different continents, and the mean ages of the participants were 14 yr in Cohen HA, 1997 and 25-26 yr in Schachter EN, 1982 and Tecklenburg SL, 2007. It is not evident how far the estimate of effect can be generalized, but similar findings from such dissimilar studies indicate that vitamin C may be effective for a wider population who suffer from EIB.

Boulet LP, 2015 refers to the Cochrane review by Wilkinson M, 2014 as a justification for the statement that there are insufficient data on vitamins or antioxidants as treatment of EIB. However, Wilkinson M, 2014 does not include any of the above mentioned three RCTs in their analyses, since their analysis is restricted to studies that used both vitamin C and vitamin E together. Those studies are not relevant to the question whether vitamin C alone has effects on EIB.

Given the safety and low cost of vitamin C and the consistency in the findings of the three RCTs on vitamin C and EIB, it would seem reasonable to encourage physically active people to test whether vitamin C is beneficial on an individual basis, if they suffer from EIB.

On 2015 Apr 02, Marcus Munafò commented:

Carter and colleagues [1] report that 17% of the excess mortality among smokers is due to causes to yet established. This is an important insight – we know that smoking kills, but do not fully understand how. Nevertheless, there are reasons for being cautious about ascribing a direct causal role to smoking for some of the outcomes reported. The supplementary appendix indicates substantial excess mortality among smokers for “all other mental diseases”, including suicide. However, homicide is related to smoking in a similar way to suicide [2], which casts doubt on the possibility that smoking may be acting via a biological pathway. Mendelian randomization [3] offers a possible solution, protecting against confounding and reverse causality and enabling stronger causal inference. Since we do not yet fully understand why smoking kills, we have established the Causal Analyses for Research on Tobacco and Alcohol (CARTA) consortium to use Mendelian randomization to elucidate the causal role of smoking on a range of health outcomes and intermediate markers (http://www.bris.ac.uk/expsych/research/brain/targ/research/collaborations/carta/), which has failed to provide evidence that smoking causes depression [4].

Marcus Munafò, Amy Taylor and George Davey Smith

1. Carter, B.D., et al., Smoking and mortality--beyond established causes. N Engl J Med, 2015. 372(7): p. 631-40.
2. Davey Smith, G., A.N. Phillips, and J.D. Neaton, Smoking as "independent" risk factor for suicide: illustration of an artifact from observational epidemiology? Lancet, 1992. 340(8821): p. 709-12.
3. Davey Smith, G. and S. Ebrahim, 'Mendelian randomization': can genetic epidemiology contribute to understanding environmental determinants of disease? Int J Epidemiol, 2003. 32(1): p. 1-22.
4. Taylor, A.E., et al., Investigating the possible causal association of smoking with depression and anxiety using Mendelian randomisation meta-analysis: the CARTA consortium. BMJ Open, 2014. 4(10): p. e006141.

On 2015 May 05, Arthur Yin Fan commented:

The original trial by Hinman's has many methodology flaws, please read the comments under the original trial report.

On 2015 Feb 11, Robert Eibl commented:

Better understanding of the immune system appears to be the key element for cancer cure, but not all cancer research agencies share this priority.

On 2015 Oct 24, Geriatric Medicine Journal Club commented:

There is already strong evidence for comprehensive geriatric assessment in the inpatient setting. This orthogeriatrics trial also incorporates a cost-effectiveness evaluation. This article was critically appraised at the October 2015 Geriatric Medicine Journal Club (follow #GeriMedJC on Twitter). The full discussion can be found at: http://gerimedjc.blogspot.com/2015/10/october-2015-gerimedjc.html?spref=tw Despite more resourcing, when patients are cared for on a comprehensive geriatric care ward, outcomes of mobility of function are better. An important point raised during the journal club discussion is that by creating specialized wards, we may be distracting from a larger effort to make an entire hospital senior-friendly.

On 2015 Jul 13, Graham Coop commented:

After our paper appeared we were made aware of a great potential example of paternal control of female meiotic transmission (Herrera et al. 1996). In many species dispensable supernumerary B-chromosomes are preferentially transmitted to offspring, with this preferential segregation (drive) often occurring through female meiosis (Jones 1991, Burt and Trivers 2006). Herrera et al. studied a widespread B chromosome polymorphism in the grasshopper, Eyprepocnemis plorans. In crosses between 1B females and males from the same focal population in which B-chromosomes were present, B-chromosomes were transmitted following Mendelian ratios. However, when the same females were crossed to males from a population where the B chromosome was not present, they transmitted their B-chromosome to much more than half of their progeny. There was no obvious reduction in fertility, suggesting that this was not due to lethality and potentially due to meiotic drive. Herrera et al suggested that the male control of female meiosis is exerted during the first meiotic division, perhaps due to an effect of substances in the male ejaculate. These results are consistent with our hypothesis that sperm-based suppressors of drive may arise and spread in response to the spread of female meiotic drive elements (such as B-chromosomes), such that female meiotic drive can re-emerge when eggs are exposed to specific sperm from a population where drive suppression had not evolved. We thank Juan Pedro M. Camacho (Universidad de Granada) for kindly bringing this example to our attention and for feedback on this note.

Yaniv Brandvain and Graham Coop

Cited References:

Jones, R.N. 1991. B-Chromosome Drive. American Naturalist 137: 430-442.

Burt, A. and R. Trivers, 2006. Genes in conflict. Belknap Press, Cambridge.

Herrera, J. A., M. D. López-León, J. Cabrero, M. W. Shaw and J. P. M. Camacho. 1996. Evidence for B chromosome drive suppression in the grasshopper Eyprepocnemis plorans. Heredity 76: 633–639.

On 2015 Jun 16, Andrew Brown commented:

Read the commentary by Nwudu VC, 2015 for important considerations regarding the quality and completeness of this systematic review.

On 2015 Feb 13, Heping Zhang commented:

Thanks for your comments. Our work regarding the comparative genomic analysis of genus Bifidobacterium began in 2013, which was finished in early 2014. As we found, the manuscript from your side had not been published elsewhere when we were submitting the manuscript or revising the manuscript according to the reviewers’ suggestions from PLOS ONE. Anyway, it’s very glad that consistent conclusions were reached by two teams independently, which proved the reliability of both researches. In future, Bifidobacterium genomics is likely to be one of our interests. We sincerely wish to cooperate with the team of Prof. Ventura, and all of the colleagues worldwide with enthusiasm on promoting advancement of this certain area.

On 2016 May 10, Morten Oksvold commented:

Please pay attention to the following report by ORI (Office of Research Integrity) before reading this article:

https://ori.hhs.gov/content/case-summary-walker-kenneth

On 2016 Dec 17, John Tucker commented:

The problem with the conclusions of this article, is of course, that the most impassioned arguments made from first principles and in vitro results carry little weight when used to argue against the results of multiple phase 3 trials involving tens of thousands of patients.

Statins reduce MI and cardiovascular death. The data is clear.

On 2015 Aug 17, Guangchuang Yu commented:

the original idea of using bubble chart to compare annotations was published in OMCIS, 2012, http://www.ncbi.nlm.nih.gov/pubmed/22455463 by clusterProfiler package.

On 2016 May 31, Marcelo G Bonini commented:

Thank you Rick!

On 2016 May 26, FREDERICK DOMANN commented:

References for the MnSOD over-expressing MCF-7 cells used in this study can be found here:

1. http://www.ncbi.nlm.nih.gov/pubmed/11929863
2. http://www.ncbi.nlm.nih.gov/pubmed/16170370

On 2016 May 31, David C. Norris commented:

Following Hardin's model¹ , Roman² advances an argument characteristic of that "intolerance of reason" which stands "not often on the side of liberty"³ : A crisis looms, having no technical solution; my analysis of human nature proves coercion indispensable.

Hardin's crisis was the 1960s' looming Malthusian apocalypse. "The necessity of abandoning the commons in breeding" demanded "mutual coercion, mutually agreed upon"¹ -- words that could hardly sound more chilling except against the background of Hardin's eerie silence on all details of said "coercion". Roman for his part volunteers specifics, but only to conflate as "coercion" such divergent measures as cost-sharing (a decidedly non-coercive remedy for the moral hazard which Roman misreads as a 'tragedy of the commons') and outright healthcare rationing: "giv[ing] local communities responsibility for governance of common-pool resources."²

Ironically, Hardin's crisis now yields to technical innovation⁴ plus the spread of liberal values and liberty itself.⁵ Can these same forces not solve also an artificial crisis in which -- unlike Nature's common-pool oceans and atmosphere -- our Treasury has become a healthcare commons purely by an accident of our politics?

[1] Hardin G, 1968 Full text

[2] Roman BR, 2015

[3] Hayek, F. A. The Road to Serfdom: Text and Documents--The Definitive Edition. Edited by Bruce Caldwell. 1 edition. Chicago: University Of Chicago Press, 2007. See p. 200 in Ch. 13, titled 'The Totalitarians in Our Midst'. The fuller quotation is as follows:

The influence of these scientist-politicians was of late years not often on the side of liberty: the "intolerance of reason" so frequently conspicuous in the scientific specialist, the impatience with the ways of the ordinary man so characteristic of the expert, and the contempt for anything which was not consciously organized by superior minds according to a scientific blueprint were phenomena familiar in German public life for generations before they became of significance in England.

[4] Whitty CJ, 2013

[5] Bongaarts J, 2011

On 2017 Jan 11, Thomas E Sussan commented:

Extrapolation of the levels of exposure in this model to those of human users is difficult due to a number of factors, including differences in respiratory mechanics, altered metabolic rates of nicotine and other constituents, and the fact that mice are passively breathing rather than actively puffing. Development of an appropriate animal model that is both representative of a human condition and also amenable to investigation can be challenging. As a result, animal models often push the limits of a typical human exposure in order to obviate the need for a protracted disease course. Additionally, due to interspecies differences, toxicity studies generally employ a 10-fold uncertainty factor when extrapolating from laboratory animals to humans, and further uncertainty factors are also typically used to account for vulnerable/susceptible populations. Findings in animal toxicity studies are generally applicable to humans, and animal studies have been used for decades to provide the basis for human studies.

Mice are frequently more resistant than humans to toxicant exposures, as is the case for cigarette smoke. The most widely accepted mouse model of cigarette smoke-induced lung disease utilizes daily exposure to high levels of cigarette smoke for 4-7 hours per day for 6 mo, resulting in a phenotype that is morphologically similar to that of a chronic smoker with mild COPD. Additionally, unlike humans, mice rarely progress beyond mild stages of disease. Due to the relative short history of e-cigarettes, we don’t have the ability to compare the pulmonary responses in mice to those of chronic e-cig users. Thus, while animal models have been indispensible for providing the basis of human studies, they all have limitations, and our model is no different. Based on duration of exposure, we believe that our published model is reasonable, although we do not dispute that the level of exposure is elevated compared to that of a typical e-cig user.

In our exposure model, mice breathed freely in a chamber containing a constant mix of 20% e-cig vapor/80% air for 1.5h twice per day. In 3 hours, a human would take approximately 2160-3600 breaths (12-20 breaths/minute), and thus a 20% e-cig vapor would be equivalent to 432-720 breaths. However, it is notable that a typical human breath is substantially larger than a typical e-cig puff. Thus, breaths and puffs are not equivalent, and our exposure is higher than that of a typical human e-cig user.

Drs. Mukhin and Rose used a different approach to estimate exposure in our model based on our measured cotinine levels and previously published values of metabolic rates for nicotine and cotinine in mice and humans, a published equation to calculate steady state cotinine, published steady state cotinine levels in mice after nicotine infusion, and a published volume of e-cig liquid consumption per puff. Based on these values, they determined that our model was equivalent to 3600-4600 puffs by a human user (although our e-cigs contained 1.8% nicotine, not 1.6%, which reduces these estimates slightly to 3333-4111). This puff estimate is approximately 10x higher than the average number of daily puffs taken by a typical e-cig user. These published estimates for nicotine and cotinine metabolism can vary considerably due to genetic variation within and between species, age, sex, diet, etc, making it difficult to accurately compare exposures based on nicotine. Regardless, I am not disputing the validity of their estimate nor questioning their intentions. In fact, our exposure estimate (based on number of breaths) is consistent with their estimate (based on nicotine metabolism). The reported serum cotinine concentrations demonstrated that these levels were consistent with those of human e-cig users, but it is correct that this is actually indicative of an elevated exposure due to differences in nicotine metabolism between mice and humans.

Drs. Mukhin and Rose also stated that due to lack of clarity in the methods, it is possible that the puff estimate could be as high as 13000 puffs per day. To clarify this area of confusion that was not clear in our methods, blood was collected from 5-10 mice almost immediately after conclusion of exposure, but the process of collecting blood from each mouse required a certain amount of time. Thus, we wrote in the methods that blood collection was made within 60 min of exposure, but we strived to collect it as soon as possible.

Regarding the claim by PHE that we simply made the mice sick and thus reduced their recovery, the mice displayed no overt signs of sickness (ie vomiting, diarrhea, dizziness, lethargy, etc). They exhibited only mild effects in the absence of infection. However, in response to infection they demonstrated immunosuppression, which was evident even when the airway macrophages were infected ex vivo. This demonstrates a clear impairment at the cellular level.

The relevance of this study’s findings is unclear. The phenotypes demonstrated in this model certainly warrant further exploration of potential immunosuppressive effects in human e-cig users, especially considering the well-published impaired immune responses observed in both humans and mice exposed to cigarette smoke, which are similar to the findings in the current study. Our current study should not serve as the definitive body evidence, but should instead guide future studies. There is some more recent evidence to suggest that human e-cig users exhibit gene expression signatures indicative of immunosuppression (PMID: 27288488), although it’s noteworthy that this commenter has also criticized this human study and several other e-cig studies. Another group has shown in a retrospective study that switching from cigarettes to e-cigs may reduce the number of COPD exacerbations (PMID: 27986085), suggesting that the immune response is improved as a result of switching. Additional studies are still greatly needed to determine the relevance of our current animal study to e-cig users.

The point by Pruen is interesting. We previously did some preliminary testing to compare the disposable NJOY e-cigs versus the rechargeable NJOY e-cigs, and we noted considerable differences between the two with respect to the average life of a cartridge. We expected them to be similar, but were surprised to see that the rechargeable e-cigs lasted much longer than the disposables. Our animal exposure used the rechargeable e-cigs. Our exposure included a real-time light scattering monitor connected to the inlet of the exposure chamber to determine when the output of each e-cig began to wane. E-cig cartridges were replaced whenever the monitor detected a drop in vapor density. No one was puffing the e-cigs to ensure that the vapor remained pleasant to the taste, but we can definitively state that there was no dry puffing and thus overheating. We also puffed each e-cig for 2 seconds, once per minute to further prevent overheating of the coil and dry wicking.

Our study compared e-cig exposure to room air, but did not directly compare e-cig vapor to cigarette smoke. The questions of whether e-cigs are safe and whether they are safer than cigarettes are both valid questions, but our current study only addressed the first question. While our study demonstrated an effect on the immune response that was similar in nature to that previously seen with cigarette smoke, we did not directly compare the relative effects between cigarettes and e-cigs. No individual study can address all questions, and as stated above, further studies are needed to determine the effect of e-cigs on never-smokers, former-smokers, and dual users.

Dr Farsalino’s comment is simply a restatement of a paragraph in the Discussion of our publication, which stated the following: “Cigarette smoke contains 1014 free radicals per puff…We determined that E-cig vapor contains 7x1011 free radicals per puff…this concentration is several orders of magnitude lower than in cigarette smoke.”

On 2016 Dec 20, Zvi Herzig commented:

Important methodological issues have been raised in response to this study.

• Mukhin and Rose calculate, based on cotinine levels, that aerosol exposure levels in this study are the mouse equivalent of 11,000-13,000 puffs per day! link
• Public Health England's 2015 report on e-cigarettes notes that suddenly exposing nicotine-naive subjects to quantities of nicotine tolerated only by heavy smokers would be expected to result in stress, sickness and vomiting, which explains the reduced recovery from infection in exposed mice. link
• Mayer notes that it has been known for decades that nicotine has anti-inflammatory effects associated with immune suppression in mice and rats, but this is apparently not reflected in humans. Thus, this study's findings of reduced recovery after infection are not surprising, but lack relevance. link
• Pruen explains that this study's design protected poorly from inadvertent overheating, likely resulting in unrealistic toxicant exposures from pyrolysis. link
• Without cigarette smoke exposed controls it's quite difficult to estimate magnitude of effect in relation to real-world human consumption.

Additionally, Farsalinos notes:

"Concerning free radicals, the authors found 7x1011 spins/puff compared to 1014 spins/puff for smoking (other reports have measured up to 1017 spins/puff for smoking). That is about 150 times lower compared to tobacco cigarettes." link

On 2015 Feb 05, William Grant commented:

Increasing vitamin D levels can reduce unplanned hospital readmissions

The paper by Merkow and colleagues points out that readmissions after surgery are largely due to new postdischarge complications related to the procedure [1]. The information about the characteristics of the patients and the factors associated with unplanned readmission provide important clues as to the causes of the readmissions. This comment examines the possible role of low 25-hydroxyvitamin D [25(OH)D] concentration as an important underlying cause of readmission.

One aspect is that blacks have higher unplanned readmissions than whites (Table 4). Blacks have much lower mean 25(OH)D concentrations than whites [2] due to the facts that most vitamin D is made from solar UVB exposure and blacks have darker skin.

The next clue is the types of disease with significantly higher unplanned readmission rates (Table 4). Risk of congestive heart failure death is significantly inversely correlated with 25(OH) concentrations [3]. Treating those with heart failure increases heart function [4]. Prevalence of chronic pulmonary obstructive disease is inversely correlated with 25(OH)D concentration [5]. Those with COPD and severe vitamin D deficiency are more likely to be hospitalized [6].

Incidence of diabetes is inversely correlated with 25(OH)D concentration [7]. Incidence and mortality rates for many types of cancer are inversely correlated with solar UVB doses and 25(OH)D concentrations [8]. Would infection: an observational study in Massachusetts found that surgical site infections for those undergoing Roux-en-Y gastric bypass surgery were linearly inversely correlated with 25(OH)D concentrations below about 30 ng/mL (75 nmol/L) [9]. Vitamin D has a number of antimicrobial actions such as inducing production of cathelicidin and defensins [10]. It has been proposed that increasing 25(OH)D concentrations would reduce the risk of hospital acquired infections [11]. Renal failure: A study in Boston found that preadmission 25(OH)D concentrations below 20 ng/mL is predictive of acute kidney injury [12]. Peripheral vascular disease: 25(OH)D concentrations are significantly inversely correlated with peripheral vascular disease [3]. Sepsis: 25(OH)D concentrations are significantly inversely correlated with risk of sepsis [13, 14].

The general finding in many observational studies is that 25(OH)D concentrations below 30 ng/mL have increased risk of many chronic and infectious diseases [15, 16].

Looking at Table 2, the two most frequent reasons for unplanned readmissions directly related to 25(OH)D concentrations are surgical site infection and sepsis. Together they account for about 22% of readmissions. This portion of the readmissions can be reduced significantly if those undergoing surgery in hospitals have higher 25(OH)D concentrations.

To reduce this portion of readmissions, it would be worthwhile to measure 25(OH)D concentrations of those planning surgery or admitted to the hospital. Giving loading doses of vitamin D3 should increase 25(OH)D concentrations rapidly. A loading dose of 250,000 IU vitamin D3 increased 25(OH)D concentrations by 58 nmol/L (23 ng/mL) after 5 days in a study with healthy adults in one study [17]. A study in Austria found that giving 550,000 IU of vitamin D3 shortly after admission to the intensive care unit followed by 90,000 IU/mo thereafter found a significant reduction in mortality rates between 28 days and 6 months after admission for those with 25(OH)D concentrations below 12 ng/mL [18]. There were no significant adverse effects associated with this dose of vitamin D in this patient population although some of the patients had elevated serum calcium levels. 550,000 IU is higher than would typically be given to raise 25(OH)D concentrations rapidly. Prescription grade 50,000 IU capsules of vitamin D3 are now available.

References 1. Merkow RP, Ju MH, Chung JW, et al. Underlying Reasons Associated With Hospital Readmission Following Surgery in the United States. JAMA. 2015;313(5):483-495 2. Ginde AA, Liu MC, Camargo CA Jr. Demographic differences and trends of vitamin D insufficiency in the US population, 1988-2004. Arch Intern Med. 2009;169(6):626-32. 3. Anderson JL, May HT, Horne BD, et al. Relation of vitamin D deficiency to cardiovascular risk factors, disease status, and incident events in a general healthcare population. Am J Cardiol. 2010;106(7):963-8. 4. Dalbeni A, Scaturro G, Degan M, et al. Effects of six months of vitamin D supplementation in patients with heart failure: a randomized double-blind controlled trial. Nutr Metab Cardiovasc Dis. 2014;24(8):861-8. 5. Skaaby T, Husemoen LL, Thuesen BH, et al. Vitamin D status and chronic obstructive pulmonary disease: a prospective general population study. PLoS One. 2014;9(3):e90654. 6. Malinovschi A, Masoero M, Bellocchia M, et al. Severe vitamin D deficiency is associated with frequent exacerbations and hospitalization in COPD patients. Respir Res. 2014;15(1):131. 7. Song Y, Wang L, Pittas AG, et al. Blood 25-hydroxy vitamin D levels and incident type 2 diabetes: a meta-analysis of prospective studies. Diabetes Care. 2013;36(5):1422-8. 8. Moukayed M, Grant WB. Molecular link between vitamin D and cancer prevention. Nutrients. 2013;5(10):3993-4023. 9. Quraishi SA, Bittner EA, Blum L, et al. Association between preoperative 25-hydroxyvitamin D level and hospital-acquired infections following Roux-en-Y gastric bypass surgery. JAMA Surg. 2014;149(2):112-8. 10. Youssef DA, Miller CWT, El-Abbassi AD, et al. Antimicrobial implications of vitamin D. Dermatoendocrinol, 2011;3(4):220 – 9. 11. Youssef DA, Ranasinghe T, Grant WB, Peiris AN. The potential of vitamin D to reduce the risk of hospital-acquired infections, Dermatoendocrinol. 2012;4(2):167-75. 12. Braun AB, Litonjua AA, Moromizato T, et al. Association of low serum 25-hydroxyvitamin D levels and acute kidney injury in the critically ill. Crit Care Med. 2012;40(12):3170-9. 13. Moromizato T, Litonjua AA, Braun AB, et al. Association of low serum 25-hydroxyvitamin D levels and sepsis in the critically ill. Crit Care Med. 2014;42(1):97-107. 14. Rech MA, Hunsaker T, Rodriguez J. Deficiency in 25-hydroxyvitamin D and 30-day mortality in patients with severe sepsis and septic shock. Am J Crit Care. 2014;23(5):e72-9. 15. Hossein-Nezhad A, Holick MF. Vitamin D for health: A global perspective. Mayo Clin Proc. 2013;88(7):720-55. 16. Pludowski P, Holick MF, Pilz S, et al. Vitamin D effects on musculoskeletal health, immunity, autoimmunity, cardiovascular disease, cancer, fertility, pregnancy, dementia and mortality- a review of recent evidence. Autoimmun Rev. 2013;12(10):976-89. 17. Kearns MD, Binongo JN, Watson D, et al. The effect of a single, large bolus of vitamin D in healthy adults over the winter and following year: a randomized, double-blind, placebo-controlled trial. Eur J Clin Nutr. 2015;69(2):193-7.<br> 18. Amrein K, Schnedl C, Holl A, et al. Effect of high-dose vitamin D3 on hospital length of stay in critically ill patients with vitamin D deficiency: The VITdAL-ICU Randomized Clinical Trial. JAMA. 2014;312(15):1520-30.

Disclosure I receive funding from Bio-Tech Pharmacal, Inc. (Fayetteville, AR), MediSun Technology (Highland Park, IL), and the Vitamin D Council (San Luis Obispo, CA).

On 2015 May 12, University of Lausanne Genomics, Ecology and Evolution Journal Club commented:

The article nicely highlights the need for a reference genome panel consisting of African variants and understanding African population history. This is a summary of the discussion at our journal club. By sequencing a large number of single sperm DNA molecules, the authors showed that meiosis is an important source not only of genetic diversity of gametes with different alleles combinations as products of chromosomal crossovers (COs), but it is also the source of sequence variation, knowing that recombination may cause local mutagenic effect at crossover sites with recurrent double strand breaks (DSBs). This study contributes to the understanding of the sequence evolution at recombination hotspots. Observed de novo mutations changed strong (S) CG into weak (W) TA base pairs and they all occurred mainly at CpG sites. As it is shown that GC base pairs are preferentially transferred during crossover, the authors suggested that GC-biased gene conversion (gBGC) is the dominant force shaping the nucleotide composition at hotspots and potentially in other recombination products, which might explain the high GC content associated with recombination. It is possible that gBGC is an adaptation to reduce the opposing mutational load of recombination, knowing that mutation favors weak over strong nucleotides. Still, small sample size gives little power for detecting potential differences between COs and NCOs.

On 2015 Feb 04, Robert Eibl commented:

fascinating!

On 2015 May 20, NephJC - Nephrology Journal Club commented:

This study was discussed on May 12th and 13th in the open online nephrology journal club, #NephJC, on twitter.

Introductory comments are available at the NephJC website. The discussion was quite detailed, with more than 65 participants, including nephrologists, gastroenterologists and also input from the senior author, Professor Paulo Angeli.

A transcript and a curated (i.e. Storified) version of the tweetchat are available at the NephJC website.

The highlights of the tweetchat were:

• The authors, with research funding from the University of Padova, should be commended for conducting this elegant and well-thought out trial, which was stopped early after the interim analysis reported a superiority of the intervention (terlipressin) compared to the standard therapy (midodrine+ octreotide).

• There was some discussion around a few factors/issues: the fact that acute kidney injury in hepatorenal syndrome is a heterogenous entity and whether using biomarkers in conjunction to guide patient selection would have been helpful; concern about the early stopping and whether power/sample size should have taken interim analysis into account; the possibility that the continuous terlipressin infusion, used in this study, perhaps is more effective than bolus dosing used previously (and which will be tested in an ongoing trial by the same group of authors).

• There was a transatlantic divide in the approach to such patients. Terlipressin is not available in North America, and the evidence from this trial did not convince most participants in the American chat; on the other hand, terlipressin is available in most of Europe and UK, and the participants in that chat felt this data supported their standard of care.

Interested individuals can track and join in the conversation by following @NephJC or #NephJC, or visit the webpage at NephJC.com.

On 2015 Mar 22, Gholson J Lyon commented:

In regards to TAF1 reported in this paper, we also published a finding involving TAF1 in this paper:

http://www.ncbi.nlm.nih.gov/pubmed/?term=seqhbase

And we also posted the TAF1 discovery on BioRxiv: http://biorxiv.org/content/early/2015/01/21/014050

We are looking for other families with TAF1 mutations and any phenotypes overlapping with our family, so please contact me if you find any. My contact info can be found on my website: http://lyonlab.labsites.cshl.edu/

On 2015 Feb 03, David Mage commented:

From the perspective of a chemical engineer, a most critical parameter for calculating the extent of rebreathing of exhalations with high CO2 and low O2 from the presence of a facial obstruction is the exact millimeters of clearance, if any, between the infant's nostrils and the obstruction. Because the usual natural practice of the shocked discoverer of an apparently moribund infant is to immediately pick the infant up to revive it, that critical measurement is lost forever.

On 2016 Feb 27, Jomar Rabajante commented:

Corrigendum: The authors regret the following minor errors

In Fig.5a, the caption should state n=5 instead of n>=5. This is correct in the accepted manuscript but was changed in the published manuscript. We were not able to correct this in the galley proof. Moreover, in the captions for Fig. 5a and Fig 5b, X5 should be X{i not equal to 5} and X4 should be X{i not equal to 4}, respectively. The readers could immediately correct these errors by looking at Fig. 5a and 5b. We also rearranged the first sentences in Fig. 5 for clarity. In summary, the caption for Fig. 5 should be read

“Fig. 5. Examples of oscillating pathways. See Supplementary Figs. 13-15 and 17 for the stochastic simulations and for the parameter values used. (a-b) There are no deep valleys only continuous zigzag canals. (a) n=5; X{i not equal to 5} are initially silenced. (b) n=4; X{i not equal to 4} are initially silenced. (c) Damped oscillations towards multipotency or pluripotency. The rates of decline of the timescale factors are all equal to 0.001. (d) Damped oscillations resulting in partial differentiation and reversal of dominant GRF. The rates of decline of the timescale factors are not all equal. The initial dominant regulatory factor is GRF 5 but eventually becomes inferior as oscillations dampen.”

In Supplementary Figure 20, we were not able to change the legend in the graph when we were rearranging the figures. The legend should state that the red broken line pertains to the entropy corresponding to Fig. 5c (instead of 3d), and the blue broken line pertains to the entropy corresponding to Fig. 5d (instead of 3e). The caption for Supplementary Figure 20 is correct.

These two errors do not affect or change the information in the figures, the results and the discussion in the manuscript.

On 2015 Feb 06, Ryan Radecki commented:

Post-publication commentary: "Which Review of Tamiflu Data Do You Believe?"

Ever since its introduction, there have been skeptics regarding the utility of oseltamivir and other neuraminidase inhibitors for the treatment of influenza. Roche has profited tremendously off strategic stockpiling by many governments as a response to pandemic influenza – yet, nearly all the data comes from Roche-conducted trials, and the data has been persistently cloaked from independent review. This past year, after much strife and public shaming, the Cochrane Collaboration received some access to clinical trial reports to conduct an independent review. This review found, on average, adults receiving early treatment with oseltamivir benefited by reduction in symptom duration from 7 days to 6.3 days. No benefit was found for reduction in respiratory infectious complications or hospitalization, the truly critical need during influenza outbreaks.

However, a second group also conducted an independent review – the “Multiparty Group for Advice on Science”. Their results, based on an individual-patient meta-analysis, are published in the Lancet and offer similar – yet wildly different – conclusions....

http://www.emlitofnote.com/2015/02/which-review-of-tamiflu-data-do-you.html

On 2015 Apr 16, Paul Brookes commented:

Figure 5 of this paper contains images that appear more similar to each other than one would expect by coincidence.

http://imgur.com/tXdWQwE

A corresponding post. has been made at PubPeer... https://pubpeer.com/publications/BBC278257761C0C12E5D38844873DF

The journal has been notified.

On 2015 Feb 18, David Keller commented:

Should Parkinson patients with low LDL cholesterol increase dietary cholesterol intake?

Patients with Parkinson Disease (PD) tend to have lower LDL and total cholesterol levels than matched subjects without PD (and concomitantly lower rates of cardiovascular atherosclerotic diseases). The association of elevated cholesterol levels with reduced risk for PD incidence suggests that PD patients with abnormally low LDL and total cholesterol levels may benefit by elevating these lipid levels into the normal range, perhaps by consuming more dietary saturated fat or cholesterol itself. Of course, increasing LDL levels would also tend to increase cardiovascular risk; if increasing LDL by diet is found to slow down PD progression, then each PD patient will have to decide how much cardiovascular risk increase they are willing to accept in exchange for the concomitant neurological benefits. A randomized trial of atherogenic diet in PD patients would require careful attention to fully-informed consent, and should not raise the cardiovascular risk profile of any subject higher than the age-adjusted lower limit of normal for that patient.

On 2015 Feb 18, David Keller commented:

Evidence that high cholesterol, not statin use, is protective against Parkinson disease.

Prior studies have demonstrated decreased incidence of Parkinson disease (PD) with statin use, and with elevated LDL and total cholesterol. Subjects who are taking a statin are generally doing so because of a history of elevated LDL cholesterol. The reduced incidence of PD in these patients is difficult to interpret; is it due to their use of statins, or to their history of underlying elevated cholesterol levels? This uncertainty is due to "indication bias", so named because one factor of interest (elevated cholesterol) is an indication for the other factor of interest (a statin drug) to be prescribed. Prior studies which attributed a decreased risk for PD to statin use instead of the underlying elevated cholesterol may have suffered from indication bias.

Huang and colleagues addressed this problem by adjusting statin use for prior cholesterol levels, and adjusting cholesterol levels for current statin use. They found that high LDL and total cholesterol are associated with lower risk of incident Parkinson Disease (PD), and that statin use is actually a risk factor for PD. These findings imply that prior studies which reported a lower risk of PD with statin use may have suffered from indication bias, due to higher statin use among subjects with a past history of high cholesterol, and to lower current cholesterol levels among patients currently taking a statin. Huang's findings suggest that statin drugs are not candidates to slow or stop the progression of PD, based on the 56 PD cases analyzed.

Would it be possible to apply the statistical adjustments employed in this study to re-analyze the data used in prior studies, to reduce the effects of indication bias and determine whether prior reports of a protective effect of statin use were actually measuring the effects of the history of high cholesterol in these subjects?

On 2015 Feb 23, William Grant commented:

Vitamin D disparities may explain some of the findings regarding risk of primary cesarean delivery

The paper by Min and colleagues found many interesting results regarding correlations with primary cesarean delivery [1]. However, one additional important factor was overlooked: 25-hydroxyvitamin D [25(OH)D] concentration. This comment briefly summarizes how 25(OH)D concentrations are related to the findings of Ref. 1.

First, 25(OH)D concentrations have been found inversely correlated with primary cesarean delivery [2]. Second, black Americans have much lower 25(OH)D concentrations than white Americans due to dark skin pigmentation and the fact that most vitamin D comes from solar UVB exposure. In the period 2001-4, 25(OH)D concentrations for women aged 20-39 years were 14 ng/mL for blacks and 28 ng/mL for whites [3]. Vitamin D disparities has been proposed to explain black-white disparities in adverse birth outcomes [4]. Low 25(OH)D concentrations are also associated with adverse pregnancy outcomes such as preterm birth [5,6], and obesity [7]. Parathyroid hormone concentration has been found directly correlated with gestational diabetes [8], but parathyroid hormone concentrations are inversely correlated with 25(OH)D concentrations although also modified by age [9].

A vitamin D randomized controlled trial conducted with black, Hispanic, and white pregnant women in South Carolina found that it took 4000 IU/d vitamin D3 to raise 25(OH)D concentrations to above 40 ng/mL and normalize 1,25-dihydroxyvitamin D concentrations [10]. 1,25-dihydroxyvitamin D controls expression of many genes, so this function is very important during fetal development. No adverse effects were reported such as hypercalcemia.

References 1. Min CJ, Ehrenthal DB, Strobino DM. Investigating racial differences in risk factors for primary cesarean delivery. Am J Obstet Gynecol. 2015 Jan 28. pii: S0002-9378(15)00085-X. 2. Merewood A, Mehta SD, Chen TC, Bauchner H, Holick MF. Association between vitamin D deficiency and primary cesarean section. J Clin Endocrinol Metab. 2009;94(3):940-5. 3. Ginde AA, Liu MC, Camargo CA Jr. Demographic differences and trends of vitamin D insufficiency in the US population, 1988-2004. Arch Intern Med. 2009;169(6):626-32. 4. Bodnar LM. Simhan HN. Vitamin D may be a link to black-white disparities in adverse birth outcomes. Obstet Gynecol Surv. 2010; 65(4): 273–284. 5. Bodnar LM, Klebanoff MA, Gernand AD, et al. Maternal vitamin D status and spontaneous preterm birth by placental histology in the US Collaborative Perinatal Project. Am J Epidemiol. 2014;179(2):168-76. 6. Wagner CL, Baggerly C, McDonnell SL, et al. Post-hoc comparison of vitamin D status at three timepoints during pregnancy demonstrates lower risk of preterm birth with higher vitamin D closer to delivery. J Steroid Biochem Mol Biol. 2014 Nov 13. pii: S0960-0760(14)00268-4. doi: 10.1016/j.jsbmb.2014.11.013. [Epub ahead of print] 7. Drincic AT, Armas LA, Van Diest EE, Heaney RP. Volumetric dilution, rather than sequestration best explains the low vitamin D status of obesity. Obesity (Silver Spring). 2012;20(7):1444-8. 8. Kramer CK, Swaminathan B, Hanley AJ, et al. Vitamin D and parathyroid hormone status in pregnancy: effect on insulin sensitivity, β-cell function, and gestational diabetes mellitus. J Clin Endocrinol Metab. 2014;99(12):4506-13. 9. Valcour A, Blocki F, Hawkins DM, Rao SD. Effects of age and serum 25-OH-vitamin D on serum parathyroid hormone levels. J Clin Endocrinol Metab. 2012;97(11):3989-95. 10. Hollis BW, Johnson D, Hulsey TC, et al. Vitamin D supplementation during pregnancy: double-blind, randomized clinical trial of safety and effectiveness. J Bone Miner Res. 2011;26(10):2341-57.

Disclosure I receive funding from Bio-Tech Pharmacal, Inc. (Fayetteville, AR), MediSun Technology (Highland Park, IL), and the Vitamin D Council (San Luis Obispo, CA).

On 2015 Feb 12, Brian Clark commented:

We (Brian C. Clark and Jack Blazyk) published an opinion article in the “In My View” section of the August 2014 issue of The Journal of The American Osteopathic Association (JAOA) titled “Research in the Osteopathic Medical Profession: Roadmap to Recovery.” Four of our clinical colleagues in the Department of Osteopathic Manipulative Medicine (OMM) at our home institution, the Ohio University Heritage College of Osteopathic Medicine, responded to our article with the letter-to-the-editor indexed above. As stated in the “Editor’s Note” at the end of their letter-to-the-editor, "The JAOA declined to publish the response submitted by Drs. Clark and Blazyk.”

Since the JAOA denied us the opportunity to reply to the letter in print, we are posting our response here on PubMed Commons (see follow-up post below). When the journal editors attempted to alter our reply, we stated that "we are not willing to make any further changes to our response. Since our article was clearly defined as an opinion piece, we feel that we are entitled to state our opinion, particularly with regard to our response to Walkowski et al.'s letter-to-the-editor. Is it the JAOA's intent to edit or censor their letter? If not, we believe that we should be afforded the opportunity to reply according to our beliefs and principles. If the decision is not to publish our response, we request a brief statement that the authors and the journal were unable to agree on an acceptable response.”

On 2015 Feb 12, Brian Clark commented:

We welcome the correspondence from Walkowski and colleagues submitted in response to our article titled “Research in the Osteopathic Medical Profession: Roadmap to Recovery.” We address their questions and criticisms as follows:

Use of NIH Funding Data to Compare Research by Institutional Type

While it might be useful to compare different disciplines according to the number of institutional research FTE, these data are not easily obtainable nor publicly available. It seems highly unlikely, though, that any normalization procedure would reveal serious research strength within osteopathic medical schools in this regard, as our NIH funding levels are negligible. Is not this criticism by the correspondents an admission of the unfortunate reality that our osteopathic medical schools lack sufficient numbers of faculty trained in and committed to research?

We recognize that there are many impactful research findings arising from non-NIH funded studies and did not mean to imply, that for research to be valued, it must be funded by NIH. We simply chose this metric because the NIH peer-review process is rigorous and respected, and NIH funding is the typical standard by which medical schools as well as other health professional schools are compared for stature in research and scholarly activity.

OMM & EBM

Due to word limitations we were unable to provide more examples of unsubstantiated applications of osteopathic manipulation. We agree that there is clearly growing evidence for the efficacy of OMM to treat certain conditions, particularly low back pain, and that more work needs to be done. An example of the tension between reliance on historical tradition vs. evidence-based practice, however, is the intense training that many osteopathic medical schools offer in the use of diagnostic palpatory tests. For more than three decades, scientists and clinicians have investigated the reliability of many diagnostic tests, with systematic reviews of the literature finding that motion and landmark location diagnostic tests have poor inter-rater reliability [1-3]. Even the most rigorous reliability studies fail to demonstrate acceptable reliability rates for most diagnostic palpatory tests [4]. In light of the poor inter-rater reliability of palpatory diagnosis, validity cannot possibly be addressed. This raises the question of why so much time and effort is invested in palpatory diagnosis training and in promoting its value in osteopathic clinical decision-making. We simply provide this as one example to support our argument.

Why hold OMM to a different standard than other fields of medicine?

We did not intend to single out OMM as the only area that lacks a sufficient evidence base to warrant acceptance as a medical standard of care. The following example was included in our original manuscript, but was excised by the JAOA editors. The AOA recognizes a specialty college in ‘Prolotherapy Regenerative Medicine’ despite the lack of evidence of its efficacy for the treatment of musculoskeletal pain (e.g., to our knowledge, all MedLine-indexed systematic reviews or meta-analysis reports suggest that prolotherapy is either ineffective or inconclusive, at best) [5-13]. Since the osteopathic profession is ideally positioned and has an inherent responsibility to take the lead in musculoskeletal research, examining the utility of interventions such as prolotherapy by an objective, systematic approach is certainly reasonable. Whether or not physicians should be using prolotherapy to treat their patients may be debatable, but is it appropriate for the AOA itself to recognize Prolotherapy Regenerative Medicine as a specialty college? We strongly encourage the AOA, as well as osteopathic medical schools, to examine the balance between creating a unique identity versus the importance of accepting and assimilating scientific and clinical evidence that must ultimately determine best treatments and advances in healthcare.

Should OPP (osteopathic principles and practices) research be encouraged and supported?

While we concur that OPP research should be encouraged and supported, we questioned whether the AOA Council on Research should focus all of its attention and resources on OPP-related research at the expense all other areas of research critical to advancing human health, particularly related to the delivery of primary care.

Other thoughts:

• We did not intend for this article to be perceived as divisive or territorial (as stated by Walkowski and colleagues). In fact, one of our peer reviewers in the JAOA review process noted that “…every critical statement is substantiated with data and cannot possibly be taken as mean-spirited or as an attack on the profession. On the contrary, the tone of this paper is one of genuine concern for the DO profession, attempting to help in a positive way to shape the future research agenda.” To be explicitly clear, our goal in publishing this paper was not only to point out the relative lack of scholarly activity and research focus in osteopathic medical schools, but to offer specific steps to improve this situation.

• The AOA Commission on Osteopathic College Accreditation (COCA) must begin to enforce its research accreditation standards. Many of the existing osteopathic medical schools have little or no infrastructure to support research. Moreover, there are an additional 13 proposed osteopathic medical schools on the drawing board (as of 17-June-2014, 13 entities that have requested ‘Applicant status’, which is the first step in seeking accreditation). Will COCA continue to approve new schools, as well as renew accreditation to existing schools, that do not significantly contribute to developing new knowledge and improving healthcare through scientific inquiry, as mandated by COCA Standard Seven? The time is long overdue for the leadership of the osteopathic profession to act by putting teeth into its accreditation standards.

• Lastly, we agree wholeheartedly with Walkowski and colleagues that a team-based approach that synthesizes the respective expertise of clinicians and scientists is critical to impactful health-related research; however, we strongly disagree with their suggestion that: “We are first osteopathic, and it is that quality that serves to primarily employ and support our basic science faculty. To that end, it is the job of each researcher in our colleges to support the philosophy and practice of osteopathic medicine by actively designing and engaging in studies that support or refute the current science around that practice.” The notion that non-physician researchers at osteopathic institutions should confine their research to the philosophy and practice of osteopathic medicine is antithetical to free and unfettered scientific inquiry and discovery. As we stated in our article, “this approach marginalizes our profession by ceding new developments in the vast panoply of modern health care not only to MDs but also to all other research-driven health care professionals.” We argue that the job of folks like us (i.e., PhD researchers) is not to support any particular philosophy, but rather to constructively criticize and question the rational basis underlying any area of medical practice that might advance medical care. This is what medically-oriented PhD scientists, like ourselves, are trained to do.

Are osteopathic medical schools, the AOA, and COCA ready and willing to take the first steps toward improving our stature in research and scholarly activity in order to elevate our standing among our healthcare colleagues and the general public? Now is the time for action.

Brian C. Clark, PhD and Jack Blazyk, PhD Ohio University

On 2015 Feb 12, Brian Clark commented:

REFERENCES

1.Seffinger et al. Spine, 2004. 29(19): p. E413-25.

2.Stochkendahl et al. J Manip Physiol Thera, 2006. 29(6): p. 475-85, 485 e1-10.

3.van der Wurff et al. Manual Therapy, 2000. 5(1): p. 30-6.

4.Degenhardt et al. J Am Osteo Assoc, 2010. 110(10): p. 579-86.

5.Chou et al. Spine, 2009. 34(10): p. 1078-93.

6.Dagenais et al. Spine J, 2005. 5(3): p. 310-28.

7.Dagenais et al. Spine J, 2008. 8(1): p. 203-12.

8.Krogh et al. Am J Sports Med, 2012.

9.Rabago et al. Clin J Sport Med, 2005. 15(5): p. 376-80.

10.Rabago et al. Br J Sports Med, 2009. 43(7): p. 471-81.

11.Staal et al. Cochrane Rev, 2008(3): p. CD001824.

12.Staal et al. Spine, 2009. 34(1): p. 49-59.

13.Yelland et al. Spine, 2004. 29(19): p. 2126-33.

On 2015 Feb 12, Brian Clark commented:

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On 2015 Feb 12, Brian Clark commented:

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On 2013 Jun 13, Karl Broman commented:

A hugely influential paper, worth spending the time to work out the unmentioned details. It covers basically everything except the permutation test (see Churchill GA, 1994)

On 2015 Apr 27, David Keller commented:

How much conflict of interest is acceptable for an author arguing for a controversial treatment?

The authors of the review paper “Testosterone and Cardiovascular Risk” [1] argue for the safety and therapeutic benefits of treating aging men with supplemental testosterone (which they call "T"), claiming health-enhancement beyond the simple correction of any T deficiency which may be present. Unfortunately, there is a lack of data from randomized, controlled trials to prove the safety or efficacy of widespread T supplementation for aging men. The authors use the existing observational data to argue that the benefits of routine testosterone treatments will far outweigh the faint signals of cardiovascular harm which have been detected. Their implied message is that aging men should not be denied the benefits of testosterone supplementation while we await the definitive large, randomized study confirming the observed associations of “T” with positive health outcomes.

Their arguments are reminiscent of those favoring routine post-menopausal estrogen replacement, prior to the Women's Health Initiative's disappointing results. Is it possible that the decline of sex hormone levels with age in both men and women is a natural process that should not be tampered with? Apparently not, according to the many pharmaceutical companies which market testosterone products, listed in the “Competing Interests” section of this paper.