TRPV1 activation in ECs may trigger Ca(2+)-dependent PI3K/Akt/CaMKII signalling, which leads to enhanced phosphorylation of TRPV1, increased TRPV1-eNOS complex formation, eNOS activation and, ultimately, NO production.

Our findings suggest that inhibition of SR-A-mediated oxLDL uptake and promotion of ABCA1-dependent cholesterol efflux are two crucial events in suppression of cholesterol accumulation by curcumin in the transformation of macrophage foam cells.

Erythropoietin (EPO), the key hormone for erythropoiesis, also increases nitric oxide (NO) bioavailability in endothelial cells (ECs), yet the definitive mechanisms are not fully understood. Increasing evidence has demonstrated that β common receptor (βCR) plays a crucial role in EPO-mediated non-hematopoietic effects. We investigated the role of βCR in EPO-induced endothelial NO synthase (eNOS) activation in bovine aortic ECs (BAECs) and the molecular mechanisms involved. Results of confocal microscopy and immunoprecipitation analyses revealed that βCR was colocalized and interacted with EPO receptor (EPOR) in ECs. Inhibition of βCR or EPOR by neutralizing antibodies or small interfering RNA abolished the EPO-induced NO production. Additionally, blockage of βCR abrogated the EPO-induced increase in the phosphorylation of eNOS, Akt, Src, or Janus kinase 2 (JAK2). Immunoprecipitation analysis revealed that treatment with EPO increased the interaction between βCR and eNOS, which was suppressed by inhibition of Src, JAK2, or Akt signaling with specific pharmacological inhibitors. Furthermore, EPO-induced EC proliferation, migration, and tube formation were blocked by pretreatment with βCR antibody and Src, JAK2, or PI3K/Akt inhibitors. Moreover, in vivo experiments showed that EPO increased the level of phosphorylated eNOS, Src, JAK2, and Akt, as well as βCR-eNOS association in aortas and promoted the angiogenesis in Matrigel plug, which was diminished by βCR or EPOR neutralizing antibodies. Our findings suggest that βCR may play an integrative role in the EPO signaling-mediated activation of eNOS in ECs.

Evodiamine may confer novel TRPV1-dependent atheroprotection and TRPV1-independent anti-obesity action.

Background-In addition to the hematopoietic effect of erythropoietin, increasing evidence suggests that erythropoietin also exerts protective effects for cardiovascular diseases. However, the role of erythropoietin and its underlying mechanism in macrophage foam cell formation are poorly understood. Methods and Results-Compared with wild-type specimens, erythropoietin was increased in atherosclerotic aortas of apolipoprotein E-deficient (apoE Ϫ/Ϫ ) mice, mainly in the macrophage foam cells of the lesions. Erythropoietin levels in culture medium and macrophages were significantly elevated in response to oxidized low-density lipoprotein in a dose-dependent manner. Furthermore, erythropoietin markedly attenuated lipid accumulation in oxidized low-density lipoprotein-treated macrophages, a result that was due to an increase in cholesterol efflux. Erythropoietin treatment significantly increased ATP-binding cassette transporters (ABC) A1 and ABCG1 mRNA and protein levels without affecting protein expression of scavenger receptors, including scavenger receptor-A, CD36, and scavenger receptor-BI. The upregulation of ABCA1 and ABCG1 by erythropoietin resulted from liver X receptor ␣ activation, which was confirmed by its prevention on expression of ABCA1 and ABCG1 after pharmacological or small interfering RNA inhibition of liver X receptor ␣. Moreover, the erythropoietin-mediated attenuation on lipid accumulation was abolished by such inhibition. Finally, reduced lipid accumulation and marked increase in ABCA1 and ABCG1 were demonstrated in erythropoietin-overexpressed macrophages. Conclusion-Our data suggest that erythropoietin suppresses foam cell formation via the liver X receptor ␣-dependent upregulation of ABCA1 and ABCG1. (Circulation.

The transient receptor potential vanilloid type 1 (TRPV1) is crucial in the pathogenesis of atherosclerosis; yet its role and underlying mechanism in the formation of macrophage foam cells remain unclear. Here, we show increased TRPV1 expression in the area of foamy macrophages in atherosclerotic aortas of apolipoprotein E-deficient mice. Exposure of mouse bone-marrow-derived macrophages to oxidized low-density lipoprotein (oxLDL) upregulated the expression of TRPV1. In addition, oxLDL activated TRPV1 and elicited calcium (Ca2+) influx, which were abrogated by the pharmacological TRPV1 antagonist capsazepine. Furthermore, oxLDL-induced lipid accumulation in macrophages was ameliorated by TRPV1 agonists but exacerbated by TRPV1 antagonist. Treatment with TRPV1 agonists did not affect the internalization of oxLDL but promoted cholesterol efflux by upregulating the efflux ATP-binding cassette (ABC) transporters ABCA1 and ABCG1. Moreover, the upregulation of ABC transporters was mainly through liver X receptor α- (LXRα-) dependent regulation of transcription. Moreover, the TNF-α-induced inflammatory response was alleviated by TRPV1 agonists but aggravated by the TRPV1 antagonist and LXRα siRNA in macrophages. Our data suggest that LXRα plays a pivotal role in TRPV1-activation-conferred protection against oxLDL-induced lipid accumulation and TNF-α-induced inflammation in macrophages.

We investigated whether AMP-activated protein kinase (AMPK), a multifunctional regulator of energy homeostasis, is involved in transient receptor potential vanilloid type 1 (TRPV1)-mediated activation of endothelial nitric oxide synthase (eNOS) in endothelial cells (ECs) and mice. In ECs, treatment with evodiamine, the activator of TRPV1, increased the phosphorylation of AMPK, acetyl-CoA carboxylase (ACC) and eNOS, as revealed by Western blot analysis. Inhibition of AMPK activation by compound C or dominant-negative AMPK mutant abrogated the evodiamine-induced increase in phosphorylation of AMPK and eNOS and NO bioavailability, as well as tube formation in ECs. Immunoprecipitation and two-hybrid analysis demonstrated that AMPK mediated the evodiamine-induced increase in the formation of a TRPV1-eNOS complex. Additionally, TRPV1 activation by evodiamine increased the phosphorylation of AMPK and eNOS in aortas of wild-type mice but did not activate eNOS in aortas of TRPV1-deficient mice. In mice, inhibition of AMPK activation by compound C markedly decreased evodiamine-evoked angiogenesis in matrigel plugs and in a hind-limb ischemia model. Moreover, evodiamine-induced phosphorylation of AMPK and eNOS in aortas of apolipoprotein E-deficient (ApoE -/-) mice was abrogated in TRPV1-deficient ApoE -/-mice. In conclusion, TRPV1 activation may trigger AMPK-dependent signaling, which leads to enhanced activation of AMPK and eNOS and retarded development of atherosclerosis.

These findings suggest that EGb761 confers a protection from the formation of foam cells by a novel HO-1-dependent regulation of cholesterol homeostasis in macrophages.