However, CAPE did not affect NLRP3 or IL-1beta transcription, but instead enhanced NLRP3 binding to ubiquitin molecules, promoting NLRP3 ubiquitination, and contributing to the anti-tumor effect in the AOM and DSS mouse model.

However, CAPE did not affect NLRP3 or IL-1beta transcription, but instead enhanced NLRP3 binding to ubiquitin molecules, promoting NLRP3 ubiquitination, and contributing to the anti-tumor effect in the AOM and DSS mouse model.

As shown in XREF_FIG, LPS + ATP promoted the expression of NLRP3 and pro-IL-1beta in THP-1 cells; however, real-time PCR revealed that after treatment with CAPE for 12 h, mRNA levels of NLRP3 and IL-1beta in THP-1 cells were similar to control (XREF_FIG), indicating that CAPE does not affect the transcription of NLRP3 and IL-1beta.

As shown in XREF_FIG, LPS + ATP promoted the expression of NLRP3 and pro-IL-1beta in THP-1 cells; however, real-time PCR revealed that after treatment with CAPE for 12 h, mRNA levels of NLRP3 and IL-1beta in THP-1 cells were similar to control (XREF_FIG), indicating that CAPE does not affect the transcription of NLRP3 and IL-1beta.

Moreover, CAPE decreased the mRNA levels of NLRP3, IL-1beta, IL-6, and TNF-alpha (XREF_FIG), increased the binding of NLRP3 to ubiquitin molecules and facilitated NLRP3 ubiquitination (XREF_FIG).

We then examined whether CAPE also reduces NLRP3 mRNA levels.

Altogether, these results indicate that CAPE reduces NLRP3 protein levels and suppresses NLRP3 activation in macrophages.

Western blotting showed that CAPE significantly inhibited the increased protein levels of NLRP3, caspase-1, and IL-1beta in BMDMs and THP-1 cells after LPS and ATP stimulation (XREF_FIG).

Furthermore, CAPE significantly reduced the expression of NLRP3, cleaved caspase-1, and cleaved IL-1beta, which was restored by rotenone (XREF_FIG).

CAPE enhanced the interaction between NLRP3 and Cullin1 and decreased the interaction between NLRP3 and CSN5 in THP-1 cells in a time-dependent manner ( xref ).

CAPE Suppresses Interaction Between NLRP3 and CSN5, and Enhances the Interaction Between NLRP3 and Cullin1.

CAPE enhanced the interaction between NLRP3 and Cullin1 and decreased the interaction between NLRP3 and CSN5 in THP-1 cells in a time dependent manner (XREF_FIG).

CAPE enhanced the interaction between NLRP3 and Cullin1 and decreased the interaction between NLRP3 and CSN5 in THP-1 cells in a time-dependent manner ( xref ).

Moreover, CAPE suppressed the interaction between NLRP3 and CSN5 but enhanced that between NLRP3 and Cullin1 both in vivo and in vitro .

CAPE Suppresses Interaction Between NLRP3 and CSN5, and Enhances the Interaction Between NLRP3 and Cullin1.

CAPE enhanced the interaction between NLRP3 and Cullin1 and decreased the interaction between NLRP3 and CSN5 in THP-1 cells in a time dependent manner (XREF_FIG).

Moreover, CAPE suppressed the interaction between NLRP3 and CSN5 but enhanced that between NLRP3 and Cullin1 both in vivo and in vitro .

Moreover, CAPE suppressed the interaction between NLRP3 and CSN5 but enhanced that between NLRP3 and Cullin1 both in vivo and in vitro.

NLRP3 interacts with ASC and pro-caspase-1 to form an inflammasome.

NLRP3 interacts with ASC and pro-caspase-1 to form an inflammasome.

However, CAPE did not affect NLRP3 or IL-1β transcription, but instead enhanced NLRP3 binding to ubiquitin molecules, promoting NLRP3 ubiquitination, and contributing to the anti-tumor effect in the AOM/DSS mouse model.

Moreover, CAPE decreased the mRNA levels of NLRP3, IL-1β, IL-6, and TNF-α ( xref ), increased the binding of NLRP3 to ubiquitin molecules and facilitated NLRP3 ubiquitination ( xref ).

Moreover, CAPE enhanced the binding of NLRP3 to ubiquitin molecules, promoted NLRP3 ubiquitination ( xref ), and significantly blocked the formation of NLRP3 inflammasome, which were again reversed by rotenone ( xref ).

Altogether, these results indicate that CAPE reduces NLRP3 protein levels and suppresses NLRP3 activation in macrophages.

NLRP3 triggers innate immunity by activating caspase-1 and then cleaves immune and metabolic substrates, especially the pro inflammatory cytokine interleukin-1beta (IL-1beta), which induces inflammation and promotes tumor growth.

Activated NLRP3 promotes pro-caspase-1 proteolysis into its active form, caspase-1 (p20), and then cleaves pro-IL-1beta and pro-IL-18 into their mature forms (IL-1beta and IL-18).

Moreover, NLRP3 inhibition was found to prevent CAC.

Altogether, our findings indicate that inhibition of NLRP3 inflammasome by CAPE prevents CAC.

NLRP3 triggers innate immunity by activating caspase-1 and then cleaves immune and metabolic substrates, especially the pro inflammatory cytokine interleukin-1beta (IL-1beta), which induces inflammation and promotes tumor growth.

Altogether, our findings demonstrate that CAPE prevents CAC by post-transcriptionally inhibiting NLRP3 inflammasome.

Caffeic Acid Phenethyl Ester Prevents Colitis Associated Cancer by Inhibiting NLRP3 Inflammasome.

In this study, we provide evidence that CAPE facilitates NLRP3 ubiquitination by inhibiting ROS in THP-1 cells and inhibits enteritis and tumor burden by inhibiting NLRP3 in an AOM and DSS mouse model.

We first investigated whether CAPE inhibits the activation of NLRP3 inflammasome induced by ATP and LPS in macrophages in vitro.

We first investigated whether CAPE inhibits the activation of NLRP3 inflammasome induced by ATP and LPS in macrophages in vitro.

Lapatinib, the small molecule tyrosine kinase inhibitor which targets HER2 and EGFR, has considerable anti-tumor activity against HER2+ BC cells, including trastuzumab resistant cells.

Lapatinib, the small molecule tyrosine kinase inhibitor which targets HER2 and EGFR, has considerable anti-tumor activity against HER2+ BC cells, including trastuzumab resistant cells.

Given the fact that Lapatinib is a dual EGFR and HER2 inhibitor, we chose the HER2 overexpressing BC cell line, HCC-1954, and the EGFR overexpressing benign control cell line, MCF-10A, for further evaluation.

Given the fact that Lapatinib is a dual EGFR and HER2 inhibitor, we chose the HER2 overexpressing BC cell line, HCC-1954, and the EGFR overexpressing benign control cell line, MCF-10A, for further evaluation.

The knockdown of NLRP3 significantly reduces the proliferation, clonogenicity, invasion and migration in both Ishikawa and HEC-1A cells, while in contrast, NLRP3 overexpression enhances the proliferation, migration and invasion in both Ishikawa and HEC-1A cells and furthermore, increases caspase-1 activation and the release of IL-1beta in endometrial cancer cells.

Inhibition of NLRP3 suppresses the proliferation, migration and invasion, and promotes apoptosis in glioma cells, while in contrast, increased expression of NLRP3 significantly enhances the proliferation, migration and invasion as well as attenuating apoptosis in glioma cells (XREF_TABLE).

Similarly, NLRP3 expression levels are also correlated with the tumor size, lymph node metastatic status and IL-1beta expression in oral squamous cell carcinoma (OSCC), and downregulating NLRP3 expression markedly attenuates the proliferation, migration, and invasion of OSCC.

The knockdown of NLRP3 significantly reduces the proliferation, clonogenicity, invasion and migration in both Ishikawa and HEC-1A cells, while in contrast, NLRP3 overexpression enhances the proliferation, migration and invasion in both Ishikawa and HEC-1A cells and furthermore, increases caspase-1 activation and the release of IL-1beta in endometrial cancer cells.

Knockdown of NLRP3 suppresses UVB induced production of IL-1beta and attenuates other inflammatory mediators, such as IL-1alpha, IL-6, TNF-alpha and PGE 2.

The knockdown of NLRP3 significantly reduces the proliferation, clonogenicity, invasion and migration in both Ishikawa and HEC-1A cells, while in contrast, NLRP3 overexpression enhances the proliferation, migration and invasion in both Ishikawa and HEC-1A cells and furthermore, increases caspase-1 activation and the release of IL-1beta in endometrial cancer cells.

Dong et al. found that NLRP3 inhibits senescence and enables replicative immortality through regulating the Wnt / beta-catenin pathway via the thioredoxin-interacting protein ( TXNIP ) / NLRP3 axis ( 74 ) .

found that NLRP3 inhibits senescence and enables replicative immortality through regulating the Wnt and beta-catenin pathway via the thioredoxin interacting protein (TXNIP)/NLRP3 axis.

The knockdown of NLRP3 significantly reduces the proliferation, clonogenicity, invasion and migration in both Ishikawa and HEC-1A cells, while in contrast, NLRP3 overexpression enhances the proliferation, migration and invasion in both Ishikawa and HEC-1A cells and furthermore, increases caspase-1 activation and the release of IL-1beta in endometrial cancer cells.

Inhibition of NLRP3 suppresses the proliferation, migration and invasion, and promotes apoptosis in glioma cells, while in contrast, increased expression of NLRP3 significantly enhances the proliferation, migration and invasion as well as attenuating apoptosis in glioma cells (XREF_TABLE).

Consistently, knockdown of NLRP3 induces cell apoptosis in MCF-7 cells and decreases cell migration; nevertheless, in other cell-types, NLRP3 inflammasome may pharmacologically repress proliferation and metastasis of hepatic cell carcinoma (HCC) (XREF_TABLE).

found that NLRP3 overexpression inhibits cell proliferation and stimulates apoptosis in leukemic cells.

Similarly, NLRP3 expression levels are also correlated with the tumor size, lymph node metastatic status and IL-1beta expression in oral squamous cell carcinoma (OSCC), and downregulating NLRP3 expression markedly attenuates the proliferation, migration, and invasion of OSCC.

NLRP3 inflammasome inactivation, driven by miR-223-3p, increases proliferation, promotes invasion and inhibits apoptosis in breast cancer cells.

The attenuation of the NLRP3 downstream pyroptosis pathway promotes apoptosis.

Consistently, knockdown of NLRP3 induces cell apoptosis in MCF-7 cells and decreases cell migration; nevertheless, in other cell-types, NLRP3 inflammasome may pharmacologically repress proliferation and metastasis of hepatic cell carcinoma (HCC) (XREF_TABLE).

Similarly, activation of NLRP3 inflammasome in mesothelial cells of lung cancer leads to an inflammatory response that fuels cancer initiation and progression and then activates the NF-kappaB-signaling pathway in lung cancer, consequently increasing proliferation and inhibiting apoptosis.

NLRP3 inflammasomes mediate both suppressions of apoptosis and progression of the cell cycle by leptin dependent ROS production in breast cancer, which is mediated via estrogen receptor alpha (ERalpha)/reduced nicotinamide adenine dinucleotide phosphate (NADPH) oxidase signaling.

proposed that significant cell death was observed only when P2X7R and NLRP3 inflammasome were both inhibited by ATP and MCC950, a specific inhibitor of NLRP3 inflammasome, and further research into safety manipulation of NLRP3 inflammasome without enhancing significant dose dependent side effects is required.

In addition, inactivation of NLRP3 inflammasome has also been found to reduce IL-1beta expression and halt development of melanoma.

Inactivation of NLRP3 inflammasome driven by miR-233-3p has been found to decrease the expression of NLRP3 inflammasome associated proteins, ASC, IL-1beta, and IL-18 in breast cancers and suppress tumor growth.

found that NLRP3 in renal tubular cells re-localizes from the cytosol to the mitochondria during hypoxia and binds to MAVS, which attenuates mtROS production and depolarization of the mitochondrial membrane potential under hypoxia.

Despite the major downstream event of NLRP3 inflammation formation of caspase-1 mediated pyroptosis, NLRP3 seems to mediate the dual-function of apoptosis and survival.

Huang et al. reported that beta-catenin promotes NLRP3 inflammasome activation, and silencing of beta-catenin impairs NLRP3 activation.

TXNIP knockdown or targeting by miR-20b resulted in a pro tumorigenic phenotype with increased cell proliferation, inhibited cell senescence reduced cell cycle modulators (p16 and p21), and decreased NLRP3 inflammasome associated proteins (NLRP3 and cleaved caspase-1).

Inhibition of NLRP3 suppresses the proliferation , migration and invasion , and promotes apoptosis in glioma cells , while in contrast , increased expression of NLRP3 significantly enhances the proliferation , migration and invasion as well as attenuating apoptosis in glioma cells ( 56 ) ( Table 2 ) .

The role of NLRP3 in promoting invasion has been demonstrated with human endometrial cancer cell lines such as Ishikawa and HEC-1A cells , where knockdown of NLRP3 significantly reduces proliferation , clonogenicity , invasion and migration .

The knockdown of NLRP3 significantly reduces the proliferation , clonogenicity , invasion and migration in both Ishikawa and HEC-1A cells , while in contrast , NLRP3 overexpression enhances the proliferation , migration and invasion in both Ishikawa and HEC-1A cells and furthermore , increases caspase-1 activation and the release of IL-1beta in endometrial cancer cells .

In contrast, overexpression of NLRP3 enhances the activities of proliferation, migration and invasion as well as increasing caspase-1 activation and IL-1beta secretion in human endometrial cancer cells.

Moreover, the silencing of NLRP3 significantly decreases the migration and invasion in OSCC cells and reduces EMT related protein expression.

The knockdown of NLRP3 significantly reduces the proliferation, clonogenicity, invasion and migration in both Ishikawa and HEC-1A cells, while in contrast, NLRP3 overexpression enhances the proliferation, migration and invasion in both Ishikawa and HEC-1A cells and furthermore, increases caspase-1 activation and the release of IL-1beta in endometrial cancer cells.

NLRP3 inflammasome inactivation, driven by miR-223-3p, increases proliferation, promotes invasion and inhibits apoptosis in breast cancer cells.

The role of NLRP3 in promoting invasion has been demonstrated with human endometrial cancer cell lines such as Ishikawa and HEC-1A cells, where knockdown of NLRP3 significantly reduces proliferation, clonogenicity, invasion and migration.

Increased activation of the NLRP3 inflammasome promotes migration and invasion activities in gastric cancer cells.

Liu et al. concluded that the upregulation of NLRP3 expression promotes the progression of endometrial cancer ; therefore , NLPR3 inflammasome might be a new therapeutic target for endometrial cancer ( 55 ) .

Collectively , these results indicate that upregulated NLRP3 expression promotes the progression of endometrial cancer ( 55 ) .

NLRP3 in the primary lesion of cancer cells drives the production of pro-IL-1beta, DC maturation, and the secretion of IL-1beta to support the evolution of tumor specific CD8 + T cells.

In the primary lesion of cancer cells, NLRP3 drives the production of pro-IL-1beta, DC maturation, and the secretion of IL-1beta to support the differentiation of tumor specific CD8 + T cells.

NLRP3 enhances IL-1beta , subsequently activating NF-kappaB , and initiates JNK signaling to cause proliferation and invasion in gastric cancer ( 21 ) .

In contrast, overexpression of NLRP3 enhances the activities of proliferation, migration and invasion as well as increasing caspase-1 activation and IL-1beta secretion in human endometrial cancer cells.

NLRP3 enhances IL-1beta, subsequently activating NF-kappaB, and initiates JNK signaling to cause proliferation and invasion in gastric cancer.

NLRP3 agonist induces Wnt and beta-catenin activation, whereas inactivation of Wnt and beta-catenin results in the inhibition of NLRP3, IL-1beta.

NLRP3 inflammasome activation induced IL-1beta and IL-18 in lung cancer may work through mechanisms other than the caspase-1 pathway, indicating that NLRP3 inflammasome can mediate the release of IL-1beta and IL-18 through caspase-1-dependent or -independent pathways.

Epistasis analysis revealed that NLRP3 variants together with polymorphisms in inflammasome related genes modulate both the frequency of inflammasome activation and the process of IL-1beta and IL-18 maturation thatinfluence HPV infection outcome and cervical cancer progression (XREF_TABLE).

NLRP3 in the primary lesion of cancer cells drives the production of pro-IL-1beta, DC maturation, and the secretion of IL-1beta to support the evolution of tumor specific CD8 + T cells.

In the primary lesion of cancer cells, NLRP3 drives the production of pro-IL-1beta, DC maturation, and the secretion of IL-1beta to support the differentiation of tumor specific CD8 + T cells.

NLRP3 inflammasome activation induced IL-1beta and IL-18 in lung cancer may work through mechanisms other than the caspase-1 pathway, indicating that NLRP3 inflammasome can mediate the release of IL-1beta and IL-18 through caspase-1-dependent or -independent pathways.

Epistasis analysis revealed that NLRP3 variants together with polymorphisms in inflammasome related genes modulate both the frequency of inflammasome activation and the process of IL-1beta and IL-18 maturation thatinfluence HPV infection outcome and cervical cancer progression (XREF_TABLE).

In contrast, overexpression of NLRP3 enhances the activities of proliferation, migration and invasion as well as increasing caspase-1 activation and IL-1beta secretion in human endometrial cancer cells.

Consistently , knockdown of NLRP3 induces cell apoptosis in MCF-7 cells and decreases cell migration ( 54 ) ; nevertheless , in other cell-types , NLRP3 inflammasome may pharmacologically repress proliferation and metastasis of hepatic cell carcinoma ( HCC ) ( 21 ) ( Table 4 ) .

Consistently, knockdown of NLRP3 induces cell apoptosis in MCF-7 cells and decreases cell migration; nevertheless, in other cell-types, NLRP3 inflammasome may pharmacologically repress proliferation and metastasis of hepatic cell carcinoma (HCC) (XREF_TABLE).

The role of NLRP3 in promoting invasion has been demonstrated with human endometrial cancer cell lines such as Ishikawa and HEC-1A cells, where knockdown of NLRP3 significantly reduces proliferation, clonogenicity, invasion and migration.

In contrast, overexpression of NLRP3 enhances the activities of proliferation, migration and invasion as well as increasing caspase-1 activation and IL-1beta secretion in human endometrial cancer cells.

The knockdown of NLRP3 significantly reduces the proliferation, clonogenicity, invasion and migration in both Ishikawa and HEC-1A cells, while in contrast, NLRP3 overexpression enhances the proliferation, migration and invasion in both Ishikawa and HEC-1A cells and furthermore, increases caspase-1 activation and the release of IL-1beta in endometrial cancer cells.

NLRP3 inflammasome inactivation, driven by miR-223-3p, increases proliferation, promotes invasion and inhibits apoptosis in breast cancer cells.

Similarly, activation of NLRP3 inflammasome in mesothelial cells of lung cancer leads to an inflammatory response that fuels cancer initiation and progression and then activates the NF-kappaB-signaling pathway in lung cancer, consequently increasing proliferation and inhibiting apoptosis.

We suggest the opposite results in NLRP3 mediated cell proliferation due to different IL-1beta levels (XREF_TABLE).

Despite the major downstream event of NLRP3 inflammation formation of caspase-1 mediated pyroptosis, NLRP3 seems to mediate the dual-function of apoptosis and survival.

Inhibition of NLRP3 suppresses the proliferation, migration and invasion, and promotes apoptosis in glioma cells, while in contrast, increased expression of NLRP3 significantly enhances the proliferation, migration and invasion as well as attenuating apoptosis in glioma cells (XREF_TABLE).

found that NLRP3 overexpression inhibits cell proliferation and stimulates apoptosis in leukemic cells.

NLRP3 enhances IL-1beta , subsequently activating NF-kappaB , and initiates JNK signaling to cause proliferation and invasion in gastric cancer ( 21 ) .

NLRP3 in the primary lesion of cancer cells drives the production of pro-IL-1beta, DC maturation, and the secretion of IL-1beta to support the evolution of tumor specific CD8 + T cells.

In the primary lesion of cancer cells, NLRP3 drives the production of pro-IL-1beta, DC maturation, and the secretion of IL-1beta to support the differentiation of tumor specific CD8 + T cells.

Moreover , NLRP3 downstream , IL-1beta , also stimulates the production of ROS that , in turn , induces DNA damage and cancer development in CRC ( 42 ) ( Table 2 ) .

NLRP3 inflammasomes mediate both suppressions of apoptosis and progression of the cell cycle by leptin dependent ROS production in breast cancer, which is mediated via estrogen receptor alpha (ERalpha)/reduced nicotinamide adenine dinucleotide phosphate (NADPH) oxidase signaling.

Moreover, NLRP3 downstream, IL-1beta, also stimulates the production of ROS that, in turn, induces DNA damage and cancer development in CRC (XREF_TABLE).

Although caspase-1 activation is the major downstream event of NLRP3 inflammasome assembly, recent studies have reported that NLRP3 inflammasome could also be activated by caspase-8.

ATP, one of the major cancer metabolites and constituents of the TME, serves as a key DAMP that activates NLRP3 inflammasome via the purinergic P2X7 receptors.

Among the known human chemokines, a co-regulated set of four (chemokine (C-C motif) ligand (CCL)-4, CCL-5, chemokine (C-X-C motif) ligand (CXCL)-9, CXCL-10) chemokines is upregulated in primary PDA carcinoma and PDA liver metastasis, which regulates CD8 + T cell infiltration, activates T cells, and promotes NLRP3 mediated T cell priming and enhances anti-tumor CD8 + T cell cytotoxic activity for an effective immune checkpoint therapy response.

Furthermore, it blocked HMGB1 mediated TLR4 dependent signalling in vitro and circulating HMGB1 in vivo [XREF_BIBR].

TLR4 - / - mice are more susceptible to SARS-CoV-1 than wild-type mice with higher viral titers [ 113 ] , which means that there was impairment in the innate immune response due to the lack of TLR4 , and hence difficulty in fighting the virus .

TLR4 - / - mice are more susceptible to SARS-CoV-1 than wild-type mice with higher viral titers [ 113 ] , which means that there was impairment in the innate immune response due to the lack of TLR4 , and hence difficulty in fighting the virus .

In addition , SARS-CoV-2 may activate TLR4 to increase PI3K / Akt signalling in infected cells , preventing apoptosis and thus increasing time for viral replication .

We also briefly review the proposed use of TLR4 antagonists as antiviral treatments, including Eritoran, Resatorvid (CLI-095 and TAK242), and glycyrrhizin, as well as another compound, nifuroxazide, that interrupts TLR4 signalling.

TLR4 uses an accessory protein called MD2 for the recognition of LPS and viral proteins; MD2 initially binds to TLR4 within the cell and is also necessary for the correct trafficking of TLR4 to the cell surface [XREF_BIBR].

TLR4 uses an accessory protein called MD2 for the recognition of LPS and viral proteins; MD2 initially binds to TLR4 within the cell and is also necessary for the correct trafficking of TLR4 to the cell surface [85].

TLR4 uses an accessory protein called MD2 for the recognition of LPS and viral proteins; MD2 initially binds to TLR4 within the cell and is also necessary for the correct trafficking of TLR4 to the cell surface [85] .

COVID-19 and Toll Like Receptor 4 (TLR4) : SARS-CoV-2 May Bind and Activate TLR4 to Increase ACE2 Expression, Facilitating Entry and Causing Hyperinflammation.

We propose a model in which the SARS-CoV-2 spike glycoprotein binds TLR4 and activates TLR4 signalling to increase cell surface expression of ACE2 facilitating entry.

Indeed, we deduce that the spike glycoprotein-TLR4 interaction is stronger than the spike glycoprotein-ACE2 interaction, which is a critical finding that must be exploited.

We can confidently extrapolate the above findings in Sections 8.1 and 8.2 from SARS-CoV-1 to SARS-CoV-2; hence, we propose that SARS-CoV-2 would activate TLR4 directly, probably via its spike protein binding to TLR4 (and/or MD2).

This can be extrapolated to SARS-CoV-2, where intracellularly, its M protein may be inducing TLR4 dependent TRAF3 independent IFN-beta production.

Hence, a possible model for the interaction of SARS-CoV-2 and TLR4 is outlined in Section 11 and the graphical abstract (XREF_FIG) in which SARS-CoV-2 may activate TLR4 in the heart and lungs to cause aberrant TLR4 signalling in favour of the proinflammatory MyD88 dependent (canonical) pathway rather than the alternative TRIF and TRAM dependent anti-inflammatory and interferon pathway.

Hence , a possible model for the interaction of SARS-CoV-2 and TLR4 is outlined in Section 11 and the graphical abstract ( Figure 1 ) in which SARS-CoV-2 may activate TLR4 in the heart and lungs to cause aberrant TLR4 signalling in favour of the proinflammatory MyD88-dependent ( canonical ) pathway rather than the alternative TRIF / TRAM-dependent anti-inflammatory and interferon pathway .

We can confidently extrapolate the above findings in Sections 8.1 and 8.2 from SARS-CoV-1 to SARS-CoV-2 ; hence , we propose that SARS-CoV-2 would activate TLR4 directly , probably via its spike protein binding to TLR4 ( and / or MD2 ) .

Hence , a possible model for the interaction of SARS-CoV-2 and TLR4 is outlined in Section 11 and the graphical abstract ( Figure 1 ) in which SARS-CoV-2 may activate TLR4 in the heart and lungs to cause aberrant TLR4 signalling in favour of the proinflammatory MyD88-dependent ( canonical ) pathway rather than the alternative TRIF / TRAM-dependent anti-inflammatory and interferon pathway .

We can confidently extrapolate the above findings in Sections 8.1 and 8.2 from SARS-CoV-1 to SARS-CoV-2 ; hence , we propose that SARS-CoV-2 would activate TLR4 directly , probably via its spike protein binding to TLR4 ( and / or MD2 ) .

In addition , SARS-CoV-2 may activate TLR4 to increase PI3K / Akt signalling in infected cells , preventing apoptosis and thus increasing time for viral replication .

Moreover, HMGB1 and other DAMPS released from necrotic and lytic cells, as well as ambient LPS levels entering the lungs or released from opportunistic gram negative bacteria, can also activate TLR4, amplifying the already severe inflammation.

Specifically, the disulphide form of HMGB1 stimulates TLR4, while extracellular HMGB1 can form complexes with DNA, RNA, other DAMP or PAMP molecules; the complexes are endocytosed via RAGE and transported to the endolysosomal system.

A review by Andersson et al. suggests that HMGB1 released as a DAMP or secreted by activated immune cells could activate both TLR4 and the receptor for advanced glycation end-products (RAGE) to generate proinflammatory cytokines [XREF_BIBR].

IL-18 is also induced by the NLRP3 inflammasome in the lungs and heart.

Inhibition of the NLRP3 inflammasome signalling pathway downstream of TLR4 attenuates LPS induced acute lung injury [XREF_BIBR, XREF_BIBR].

More relevantly, TLR4 is activated by viral PAMPs to initiate an innate immune and inflammatory response.

This raises the question , is it only ACE2 that the spike protein of SARS-CoV-2 binds to or is there also another receptor involved , such as TLR4 which is known to promote cardiac hypertrophy , myocardial inflammation , and fibrosis [ 58-60 ] ?

This raises the question , is it only ACE2 that the spike protein of SARS-CoV-2 binds to or is there also another receptor involved , such as TLR4 which is known to promote cardiac hypertrophy , myocardial inflammation , and fibrosis [ 58-60 ] ?

This raises the question , is it only ACE2 that the spike protein of SARS-CoV-2 binds to or is there also another receptor involved , such as TLR4 which is known to promote cardiac hypertrophy , myocardial inflammation , and fibrosis [ 58-60 ] ?

This raises the question , is it only ACE2 that the spike protein of SARS-CoV-2 binds to or is there also another receptor involved , such as TLR4 which is known to promote cardiac hypertrophy , myocardial inflammation , and fibrosis [ 58-60 ] ?

This would potentially serve 3 simultaneous benefits : ( a ) it would increase the compliance of the lung alveoli and prevent their collapse ; ( b ) confer antiviral actions by shielding and preventing infection of naive cells , especially if TLR4 is proven to be an entry receptor or contributes to ACE2 upregulation ; and ( c ) block TLR4 to reduce inflammation and excessive cytokine production .

This would potentially serve 3 simultaneous benefits : ( a ) it would increase the compliance of the lung alveoli and prevent their collapse ; ( b ) confer antiviral actions by shielding and preventing infection of naive cells , especially if TLR4 is proven to be an entry receptor or contributes to ACE2 upregulation ; and ( c ) block TLR4 to reduce inflammation and excessive cytokine production .

For instance , ( 1 ) evidence that TLR4 has the strongest protein-protein interaction with the spike glycoprotein of SARS-CoV-2 compared to other TLRs [ 30 ] , together with ( 2 ) evidence that SARS-COV-2 strongly induces interferon-stimulated gene ( ISG ) expression in an immunopathogenic context in the respiratory tract [ 31 ] ; ( 3 ) evidence that ISG activation results in increased expression of ACE2 [ 32 ] and ( 4 ) evidence that pulmonary surfactants in the lung prevent viral infection by blocking TLR4 [ 33 ] suggest a possible mechanism in which the virus may be binding to and activating TLR4 to increase expression of ACE2 which promotes viral entry .

For instance , ( 1 ) evidence that TLR4 has the strongest protein-protein interaction with the spike glycoprotein of SARS-CoV-2 compared to other TLRs [ 30 ] , together with ( 2 ) evidence that SARS-COV-2 strongly induces interferon-stimulated gene ( ISG ) expression in an immunopathogenic context in the respiratory tract [ 31 ] ; ( 3 ) evidence that ISG activation results in increased expression of ACE2 [ 32 ] and ( 4 ) evidence that pulmonary surfactants in the lung prevent viral infection by blocking TLR4 [ 33 ] suggest a possible mechanism in which the virus may be binding to and activating TLR4 to increase expression of ACE2 which promotes viral entry .

This raises the question, is it only ACE2 that the spike protein of SARS-CoV-2 binds to or is there also another receptor involved, such as TLR4 which is known to promote cardiac hypertrophy, myocardial inflammation, and fibrosis [XREF_BIBR - XREF_BIBR]?

More relevantly, TLR4 is activated by viral PAMPs to initiate an innate immune and inflammatory response.

Inhibition of the NLRP3 inflammasome signalling pathway downstream of TLR4 attenuates LPS induced acute lung injury [XREF_BIBR, XREF_BIBR].

As mentioned previously , TLR4 is activated by its typical ligand , LPS .

As mentioned previously , TLR4 is activated by its typical ligand , LPS .

These proteins cause TLR4 activation , to induce an inflammatory response during acute viral infection .

These proteins cause TLR4 activation , to induce an inflammatory response during acute viral infection .

DAMP Mediated TLR4 Activation.

Even if the virus infects cardiomyocytes via ACE2 only , the subsequent immune-mediated myocardial injury and inflammation is likely mediated via TLR4 due to the DAMPs released from the lysed cardiomyocytes .

Even if the virus infects cardiomyocytes via ACE2 only , the subsequent immune-mediated myocardial injury and inflammation is likely mediated via TLR4 due to the DAMPs released from the lysed cardiomyocytes .

TLR4 activation by LPS on cardiomyocytes leads to subsequent reduction in myocardial contractility [XREF_BIBR, XREF_BIBR], and the predominant view in the literature is that TLR4 activation on cardiac structural fibroblasts and cardiac macrophages leads to a profibrotic and proinflammatory response, respectively [XREF_BIBR, XREF_BIBR].

TLR4 activation by LPS on cardiomyocytes leads to subsequent reduction in myocardial contractility [ 77 , 81 ] , and the predominant view in the literature is that TLR4 activation on cardiac structural fibroblasts and cardiac macrophages leads to a profibrotic and proinflammatory response , respectively [ 78 , 82 ] .

Furthermore, while LPS increased TLR4 content in the lungs and heart by at least twofold, nifuroxazide was shown to significantly reduce TLR4 content by 45.7% in the lungs and 31.2% in the heart in the curative regimen and more so in the prophylactic regimen, compared to the LPS control [XREF_BIBR].

TLR4 activation by LPS on cardiomyocytes leads to subsequent reduction in myocardial contractility [ 77 , 81 ] , and the predominant view in the literature is that TLR4 activation on cardiac structural fibroblasts and cardiac macrophages leads to a profibrotic and proinflammatory response , respectively [ 78 , 82 ] .

TLR4 can be activated by LPS ( classical PAMP ) , DAMPs , or viral PAMPs .

Additionally, it inhibited RSVF-, DENV-NS1-, and EBOV glycoprotein mediated TLR4 activation.

Mechanistically, LCZ696 prevents LPS induced activation of the TLR4 and Myd88 pathway and nuclear translocation of nuclear factor kappa-B (NF-kappaB) p65 factor.

Mechanistically, LCZ696 prevents LPS induced activation of the TLR4 and Myd88 pathway and nuclear translocation of nuclear factor kappa-B (NF-kappaB) p65 factor.

The priming step triggers nuclear factor-kappaB (NF-kappaB)-dependent upregulation of NLRP3 and pro-IL-1beta expression and lowers the activation threshold of NLRP3 by additional post-translational modification (PTMs).

NLRP3 activation is negatively regulated by tyrosine phosphorylation and protein tyrosine phosphatase non receptor 22 (PTPN22) dephosphorylates NLRP3 upon its activation.

This suggests that post-translational modifications of NLRP3 (such as phosphorylation and ubiquitination) and subsequent autophagic removal of inflammasome components upon their activation serves as a negative-feedback loop to prevent excessive inflammatory response.

Autophagic removal of NLRP3 inflammasome activators, such as intracellular DAMPs, NLRP3 inflammasome components, and cytokines can reduce inflammasome activation and inflammatory response.

Furthermore, silencing of NLRP3 or ASC in microglia suppressed caspase-1 activation and enhanced autophagy after PrP-106-126 stimulation.

As mentioned above, p62 expression negatively regulates NLRP3 activation by mediating mitophagic elimination of damaged mitochondria upon NLRP3 activation.

The pharmacological inhibition of autophagy and loss of p62 greatly enhanced the NLRP3 inflammasome activation.

This is supported by increasing number of studies demonstrating that impaired mitophagy enhances NLRP3 activation , whereas induction of mitophagy reduces NLRP3 activation ( 87-90 ) .

NLRP3 activation is negatively regulated by tyrosine phosphorylation and protein tyrosine phosphatase non receptor 22 (PTPN22) dephosphorylates NLRP3 upon its activation.

It was suggested that pyrin interacts with NLRP3 inflammasome component in inhibitory manner and FMF-associated mutations prevent these inhibitory interactions ( xref , xref ).

As mentioned above, pyrin interacts with NLRP3 inflammasome components and leads to their autophagic interaction, thereby acting as anti-inflammatory factor ( xref ).

The first hypothesis involves interaction between thioredoxin-interacting protein (TXNIP) and NLRP3 after an increase in ROS caused by NLRP3 activators, such as MSU.

Upon detecting specific stimuli sensor protein NLRP3 interacts with ASC via homotypic PYD-PYD domain interaction ( xref ) and nucleates ASC into prion-like filaments, thereby forming a single ASC “speck” within activated cell.

The NEK7-NLRP3 interaction was shown to be dependent on potassium efflux ( xref ).

The NLRP3 inflammasome activation triggers the interaction of NLRP3 with NEK7, leading to the inflammasome assembly, the ASC speck formation and caspase 1 activation.

Many of NLRP3 activators, such as nigericin, ATP, pore forming toxins, and particulate stimuli, are known to induce potassium efflux and decrease intracellular potassium levels, which is required for direct binding of NEK7 to NLRP3 inflammasome.

NLRP3 lacking phosphorylation site did not interact with p62 and was not sequestered into phagophore ( xref ).

It was reported that only phosphorylated NLRP3 interacted with p62 in ASC-dependent manner and was sequestered into phagophore.

The molecular mechanism for MSU-induced NLRP3 inflammasome activation is not yet fully elucidated.

The NLPR3 inflammasome assembly was disrupted due to reduced NLRP3 protein levels , which resulted in decreased caspase-1 activation and IL-1beta production upon the NLRP3 inflammasome activation after Ka treatment ( 101 ) .

Furthermore, silencing of NLRP3 or ASC in microglia suppressed caspase-1 activation and enhanced autophagy after PrP-106-126 stimulation.

Downregulation of voltage dependent anion channels (VDAC), which are required for ROS production, or ROS scavengers impaired and reversed NLRP3 mediated caspase-1 activation and IL-1beta release in response to NLRP3 activators nigericin, MSU, alum, and silica.

Furthermore, a few other studies have shown that NLRP3 deficient mice have increased autophagy levels at baseline and under stress conditions, such as hypoxia or hyperoxia in different tissues and epithelial cells.

Furthermore, silencing of the NLRP3 downregulated autophagy and LC3-I conversion to LC3-II induced by MSU in osteoblasts.

Silencing core molecules of the NLRP3 inflammasome complex in human macrophages reduced the autophagy response after infection with Pseudomonas aeruginosa, however it enhanced the macrophage mediated killing of internalized P. aeruginosa.

NLRP3 was shown to modulate autophagy.

NLRP3 was shown to modulate autophagy .

Autophagy Targets NLRP3 Inflammasome Components.

Downregulation of voltage dependent anion channels (VDAC), which are required for ROS production, or ROS scavengers impaired and reversed NLRP3 mediated caspase-1 activation and IL-1beta release in response to NLRP3 activators nigericin, MSU, alum, and silica.

The first hypothesis involves interaction between thioredoxin interacting protein (TXNIP) and NLRP3 after an increase in ROS caused by NLRP3 activators, such as MSU.

Mitochondrial dysfunction and release of mitochondrial ROS (mtROS) and mitochondrial DNA (mtDNA) are another important triggers for NLRP3 inflammasome activation and some of NLRP3 activators induce increased mtROS and cytosolic ROS generation.

Downregulation of voltage dependent anion channels (VDAC), which are required for ROS production, or ROS scavengers impaired and reversed NLRP3 mediated caspase-1 activation and IL-1beta release in response to NLRP3 activators nigericin, MSU, alum, and silica.

Even though the exact mechanism of NLRP3 inflammasome activation mediated by generation of mitochondrial ROS is not yet fully elucidated, several hypotheses have been suggested.

In the cytosol bacterial LPS activated caspase-4 and caspase-11 and induced pyroptosis and consequently activated NLRP3 inflammasome.

Chemotherapy has been the current standard adjuvant treatment for early-stage non-small-cell lung cancer (NSCLC) patients, while recent studies showed benefits of epidermal growth factor receptor-tyrosine kinase inhibitor (EGFR-TKI).

Chemotherapy has been the current standard adjuvant treatment for early-stage non-small-cell lung cancer (NSCLC) patients, while recent studies showed benefits of epidermal growth factor receptor-tyrosine kinase inhibitor (EGFR-TKI).

Growth Hormone Modulation of Hepatic Epidermal Growth Factor Receptor Signaling.

Growth Hormone Modulation of Hepatic Epidermal Growth Factor Receptor Signaling.

For instance, ubiquitination of NLRP3 by FBXL12, TRIM1, ARIH2 or the dopamine induced E3 ligase MARCH7 promotes the proteasomal degradation of NLRP3 in resting macrophages, whereas deubiquitylation of NLRP3 LRR domain on K63 by BRCC3 triggers ASC oligomerization and inflammasome activation (XREF_FIG).

Indeed, the LBD of VDR is able to physically interact with the NACHT-LRR domain of NLRP3 thus inhibiting the association of NLRP3 with BRCC3 and preventing NLRP3 deubiquitination (XREF_FIG).

For instance, ubiquitination of NLRP3 by FBXL12, TRIM1, ARIH2 or the dopamine-induced E3 ligase MARCH7 promotes the proteasomal degradation of NLRP3 in resting macrophages ( xref ), whereas deubiquitylation of NLRP3 LRR domain on K63 by BRCC3 triggers ASC oligomerization and inflammasome activation ( xref , xref ) ( xref ).

For instance, ubiquitination of NLRP3 by FBXL12, TRIM1, ARIH2 or the dopamine induced E3 ligase MARCH7 promotes the proteasomal degradation of NLRP3 in resting macrophages, whereas deubiquitylation of NLRP3 LRR domain on K63 by BRCC3 triggers ASC oligomerization and inflammasome activation (XREF_FIG).