On 2017 May 09, Yang K Xiang commented:

In the their commentary, Dr. Santulli has noted the differences in glucose tolerance tests between two studies (1, 2). In our study, 5-6 week old WT and β2AR -/- were fed a high fat diet (60% fat) for 6 months; both strains develop diabetes and glucose intolerance when compared to animals of same genotypes fed with a control chow (10% fat). We did not observe differences in glucose homeostasis between WT and β2AR -/- fed with the control chow. This contrasts with the Santulli study, in which β2AR -/- strain used in their studies develop diabetes and glucose intolerance at 6-months of age when fed a chow diet. Several factors may contribute to the differences in diabetic phenotypes observed.

1. In the Stanlulli study, β2AR -/- were backcrossed to the C57Bl6/N strain. In our study, the β2AR -/- is backcrossed into the C57Bl6/J strain.
2. Our study used a defined control chow with 10% fat whose composition with the exception of fat and sucrose content matched that of the high-fat diet. The possibility therefore exists that the “chow” diet in the Santulli study, whose composition is not described in detail could contribute in part to some of the metabolic changes observed. In addition, our study does not exclude that β2AR -/- mice may have metabolic issues relative to WT after feeding with the defined control chow.

The primary focus of our work was to understand the cardiac response to obesity and long-term hyperinsulinemia. In this regard the β2AR -/- mice on a high fat diet developed hyperglycemia and hyperinsulinemia, which therefore enabled us to determine if the absence of β2ARs in the heart could modulate the cardiac maladaptation that develops in wild type animals. We reported fasted insulin concentrations to demonstrate the existence of hyperinsulinemia in response to high fat feeding. However, we did observe in data not presented in the manuscript that insulin concentrations in β2AR -/- mice after intraperitoneal administration glucose were statistically lower than those in high fat fed WT, suggesting a reduced insulin release from islets, consistent with the conclusions of the in Santulli study. The Muzzin study, mentioned in the commentary is an animal with complete absence of all three β adrenergic receptors and as such caution is advised in comparing that model to mice with selective loss of the β2AR.

A study published by Jiang and colleagues was also discussed, which reported that β2AR -/- mice display a diabetic retinopathy phenotype. Although the authors of this study did not provide background information of glucose and insulin levels, they suggest that β adrenergic signaling is essential for maintaining retinal muller cell viability. Thus the observed retinopathy might not be related to diabetes per se. Taken together, these data suggest that β2AR signaling is associated with glucose metabolism and complications that may be modulated in a tissue-specific manner in different tissues in diabetes. Ultimately, transgenic approaches with tissue-specific deletion of β2AR may offer more insight into the underlying mechanism of these tissue-specific phenotypes.

Reference

1) Inhibiting Insulin-Mediated β2-Adrenergic Receptor Activation Prevents Diabetes-Associated Cardiac Dysfunction. Circulation. 2017;135:73-88.

2) Age-related impairment in insulin release: the essential role of β2-adrenergic receptor. Diabetes. 2012;61:692-701.

On 2017 Apr 28, Gaetano Santulli commented:

In the present article, Wang, Liu, Fu and colleagues report that β2-adrenergic receptor (β2AR) plays a key role in hyperinsulinemia-induced cardiac dysfunction (1). Overall, the data are very interesting and compelling. However, we noticed that in this paper β2AR-/- mice do not exhibit glucose intolerance; in fact, they seem to have a response to intraperitoneal glucose that is even better than wild-type mice (though a statistical analysis comparing these two groups is not provided). Although surprisingly not reported by the Authors, mounting evidence indicates that the deletion of β2AR has detrimental effects on glucose metabolism (2-4). Indeed, we have demonstrated that β2AR-/- mice display impaired insulin release and significant glucose intolerance (2). Muzzin and colleagues found that the ablation of βARs mechanistically underlies impaired glucose homeostasis (3). Other groups have confirmed these results, also showing that β2AR-/- mice develop diabetic-related microvascular complications (i.e. retinopathy)(4). Nonetheless, the Authors fail to at least discuss previous relevant literature describing the alterations in glucose metabolism observed in β2AR-/- mice and do not accurately circumstantiate their findings. Furthermore, the Authors do not provide any measurement (not in vivo nor in isolated islets) of insulin levels following glucose challenge, showing just baseline serum levels. We believe that for the sake of scientific appropriateness the Readers of Circulation will appreciate a clarification, in particular regarding the fact that pertinent literature in the field has been overlooked.

A formal e-Letter has been published by Circulation.

Competing Interests: None.

References 1) Inhibiting Insulin-Mediated beta2-Adrenergic Receptor Activation Prevents Diabetes-Associated Cardiac Dysfunction. Circulation. 2017;135:73-88.

2) Age-related impairment in insulin release: the essential role of β2-adrenergic receptor. Diabetes. 2012;61:692-701.

3) The lack of beta-adrenoceptors results in enhanced insulin sensitivity in mice exhibiting increased adiposity and glucose intolerance. Diabetes. 2005;54:3490-5.

4) Beta2-adrenergic receptor knockout mice exhibit a diabetic retinopathy phenotype. PLoS One. 2013;8:e70555.

On 2017 Apr 28, Gaetano Santulli commented:

In the present article, Wang, Liu, Fu and colleagues report that β2-adrenergic receptor (β2AR) plays a key role in hyperinsulinemia-induced cardiac dysfunction (1). Overall, the data are very interesting and compelling. However, we noticed that in this paper β2AR-/- mice do not exhibit glucose intolerance; in fact, they seem to have a response to intraperitoneal glucose that is even better than wild-type mice (though a statistical analysis comparing these two groups is not provided). Although surprisingly not reported by the Authors, mounting evidence indicates that the deletion of β2AR has detrimental effects on glucose metabolism (2-4). Indeed, we have demonstrated that β2AR-/- mice display impaired insulin release and significant glucose intolerance (2). Muzzin and colleagues found that the ablation of βARs mechanistically underlies impaired glucose homeostasis (3). Other groups have confirmed these results, also showing that β2AR-/- mice develop diabetic-related microvascular complications (i.e. retinopathy)(4). Nonetheless, the Authors fail to at least discuss previous relevant literature describing the alterations in glucose metabolism observed in β2AR-/- mice and do not accurately circumstantiate their findings. Furthermore, the Authors do not provide any measurement (not in vivo nor in isolated islets) of insulin levels following glucose challenge, showing just baseline serum levels. We believe that for the sake of scientific appropriateness the Readers of Circulation will appreciate a clarification, in particular regarding the fact that pertinent literature in the field has been overlooked.

A formal e-Letter has been published by Circulation.

Competing Interests: None.

References 1) Inhibiting Insulin-Mediated beta2-Adrenergic Receptor Activation Prevents Diabetes-Associated Cardiac Dysfunction. Circulation. 2017;135:73-88.

2) Age-related impairment in insulin release: the essential role of β2-adrenergic receptor. Diabetes. 2012;61:692-701.

3) The lack of beta-adrenoceptors results in enhanced insulin sensitivity in mice exhibiting increased adiposity and glucose intolerance. Diabetes. 2005;54:3490-5.

4) Beta2-adrenergic receptor knockout mice exhibit a diabetic retinopathy phenotype. PLoS One. 2013;8:e70555.

On 2017 May 09, Yang K Xiang commented:

In the their commentary, Dr. Santulli has noted the differences in glucose tolerance tests between two studies (1, 2). In our study, 5-6 week old WT and β2AR -/- were fed a high fat diet (60% fat) for 6 months; both strains develop diabetes and glucose intolerance when compared to animals of same genotypes fed with a control chow (10% fat). We did not observe differences in glucose homeostasis between WT and β2AR -/- fed with the control chow. This contrasts with the Santulli study, in which β2AR -/- strain used in their studies develop diabetes and glucose intolerance at 6-months of age when fed a chow diet. Several factors may contribute to the differences in diabetic phenotypes observed.

1. In the Stanlulli study, β2AR -/- were backcrossed to the C57Bl6/N strain. In our study, the β2AR -/- is backcrossed into the C57Bl6/J strain.
2. Our study used a defined control chow with 10% fat whose composition with the exception of fat and sucrose content matched that of the high-fat diet. The possibility therefore exists that the “chow” diet in the Santulli study, whose composition is not described in detail could contribute in part to some of the metabolic changes observed. In addition, our study does not exclude that β2AR -/- mice may have metabolic issues relative to WT after feeding with the defined control chow.

The primary focus of our work was to understand the cardiac response to obesity and long-term hyperinsulinemia. In this regard the β2AR -/- mice on a high fat diet developed hyperglycemia and hyperinsulinemia, which therefore enabled us to determine if the absence of β2ARs in the heart could modulate the cardiac maladaptation that develops in wild type animals. We reported fasted insulin concentrations to demonstrate the existence of hyperinsulinemia in response to high fat feeding. However, we did observe in data not presented in the manuscript that insulin concentrations in β2AR -/- mice after intraperitoneal administration glucose were statistically lower than those in high fat fed WT, suggesting a reduced insulin release from islets, consistent with the conclusions of the in Santulli study. The Muzzin study, mentioned in the commentary is an animal with complete absence of all three β adrenergic receptors and as such caution is advised in comparing that model to mice with selective loss of the β2AR.

A study published by Jiang and colleagues was also discussed, which reported that β2AR -/- mice display a diabetic retinopathy phenotype. Although the authors of this study did not provide background information of glucose and insulin levels, they suggest that β adrenergic signaling is essential for maintaining retinal muller cell viability. Thus the observed retinopathy might not be related to diabetes per se. Taken together, these data suggest that β2AR signaling is associated with glucose metabolism and complications that may be modulated in a tissue-specific manner in different tissues in diabetes. Ultimately, transgenic approaches with tissue-specific deletion of β2AR may offer more insight into the underlying mechanism of these tissue-specific phenotypes.

Reference

1) Inhibiting Insulin-Mediated β2-Adrenergic Receptor Activation Prevents Diabetes-Associated Cardiac Dysfunction. Circulation. 2017;135:73-88.

2) Age-related impairment in insulin release: the essential role of β2-adrenergic receptor. Diabetes. 2012;61:692-701.