Rationale: Excessive Ang II (angiotensin II) levels lead to a profibrotic and hypertrophic milieu that produces deleterious remodeling and dysfunction in hypertension-associated heart failure. Agents that disrupt Ang II–induced cardiac dysfunction may have clinical utility in the treatment of hypertension-associated heart failure. Objective: We have examined the potential effect of celastrol—a bioactive compound derived from the Celastraceae family—on Ang II–induced cardiac dysfunction. Methods and Results: In rat primary cardiomyocytes and H9C2 (rat cardiomyocyte-like H9C2) cells, celastrol attenuates Ang II–induced cellular hypertrophy and fibrotic responses. Proteome microarrays, surface plasmon resonance, competitive binding assays, and molecular simulation were used to identify the molecular target of celastrol. Our data showed that celastrol directly binds to and inhibits STAT (signal transducer and activator of transcription)-3 phosphorylation and nuclear translocation. Functional tests demonstrated that the protection of celastrol is afforded through targeting STAT3. Overexpression of STAT3 dampens the effect of celastrol by partially rescuing STAT3 activity. Finally, we investigated the in vivo effect of celastrol treatment in mice challenged with Ang II and in the transverse aortic constriction model. We show that celastrol administration protected heart function in Ang II–challenged and transverse aortic constriction–challenged mice by inhibiting cardiac fibrosis and hypertrophy. Conclusions: Our studies show that celastrol inhibits Ang II–induced cardiac dysfunction by inhibiting STAT3 activity.

Evidence indicates that Ang II (angiotensin II) activates STAT3 (signal transducer and activator of transcription 3) in cardiomyocytes. However, the mechanisms underlying STAT3 activation and downstream responses are not fully known. In this study, we show that Ang II caused biphasic STAT3 activation in cardiomyocytes. A rapid and early activation was mediated by direct association between TLR4 (toll-like receptor-4) and STAT3. This early activation increased IL-6 (interleukin-6) production, which in turn, induced the second STAT3 activation through the IL-6/gp130 (glycoprotein 130)/JAK2 (Janus-family tyrosine kinases 2) pathway, resulting in unregulated expression of genes for cardiac remodeling. Moreover, STAT3 inhibition or TLR4 knockout in mice protected against Ang II–induced hypertrophy, fibrosis, and cardiac functional deficits. Thus, Ang II–induced STAT3 activation in cardiomyocytes was biphasic, providing a sequential induction of IL-6 and myocardial remodeling genes, respectively. This work supports a novel mechanism on STAT3 activation in Ang II–induced cardiac dysfunction and remodeling.

Background: Atherosclerosis is a chronic inflammatory disease. Although Toll-like receptor 4 (TLR4) has been involved in inflammatory atherosclerosis, the exact mechanisms by which oxidized-low-density lipoproteins (ox-LDL) activates TLR4 and elicits inflammatory genesis are not fully known. Myeloid differentiation factor 2 (MD2) is an extracellular molecule indispensable for lipopolysaccharide recognition of TLR4. Method: Apoe À/À Md2 À/À mice and pharmacological inhibitor of MD2 were used in this study. We also reconstituted Apoe À/À mice with either Apoe À/À or Apoe À/À Md2 À/À marrow-derived cells. Mechanistic studies were performed in primary macrophages, HEK-293T cells, and cell-free system. Finding: MD2 levels are elevated in atherosclerotic lesion macrophages, and MD2 deficiency or pharmacological inhibition in mice reduces the inflammation and stunts the development of atherosclerotic lesions in Apoe À/À mice fed with high-fat diet. Transfer of marrow-derived cells from Apoe-Md2 double knockout mice to Apoe knockout mice confirmed the critical role of bone marrow-derived MD2 in inflammatory factor induction and atherosclerosis development. Mechanistically, we show that MD2 does not alter ox-LDL uptake by macrophages but is required for TLR4 activation and inflammation via directly binding to ox-LDL, which triggers MD2/TLR4 complex formation and TLR4-MyD88-NFkB pro-inflammatory cascade. Interpretation: We provide a mechanistic basis of ox-LDL-induced macrophage inflammation, illustrate the role of macrophage-derived MD2 in atherosclerosis, and support the therapeutic potential of MD2 targeting in atherosclerosis-driven cardiovascular diseases.