Mechanism of the inhibitory effect of atorvastatin on leptin expression induced by angiotensin II in cultured human coronary artery smooth muscle cells

Abstract

Leptin contributes to the pathogenesis of atherosclerosis. Ang II (angiotensin II), a proatherogenic cytokine, increases leptin synthesis in cultured adipocytes. Statin suppresses leptin expression in adipocytes and human coronary artery endothelial cells. However, the effect of Ang II and statin on leptin expression in VSMCs (vascular smooth muscle cells), the major cell types in atheroma, is poorly understood. Thus the aim of the present study was to investigate the molecular mechanism of atorvastatin for reducing leptin expression after Ang II stimulation in VSMCs. VSMCs from human coronary artery were cultured. Ang II stimulation increased leptin protein and mRNA and phospho-JNK (c-Jun N-terminal kinase) expression. Exogenous addition of Dp44mT (2,2'-dipyridyl-N,N-dimethylsemicarbazone) and mevalonate increased leptin protein expression similarly to Ang II. Atorvastatin, SP600125, JNK siRNA (small interfering RNA) and NAC (N-acetylcysteine) completely attenuated the leptin and phospho-JNK protein expression induced by Ang II. Ang II significantly increased ROS (reactive oxygen species) formation in human VSMCs. Addition of atorvastatin and NAC significantly attenuated the formation of ROS induced by Ang II. Addition of atorvastatin and SP600125 inhibited the phosphorylation of Rac1 induced by Ang II. The gel shift and promoter activity assay showed that Ang II increased AP-1 (activator protein-1)-binding activity and leptin promoter activity, while SP600125, NAC and atorvastatin inhibited the AP-1-binding activity and leptin promoter activity induced by Ang II. Ang II significantly increased the migration and proliferation of cultured VSMCs, while addition of atorvastatin, SP600125, NAC and leptin siRNA before Ang II stimulation significantly inhibited the migration and proliferation of VSMCs induced by Ang II. Ang II significantly increased secretion of leptin from human VSMCs, and addition of SP600125, atorvastatin and NAC before Ang II stimulation almost completely inhibited the leptin secretion induced by Ang II. In conclusion, Ang II induces leptin expression in human VSMCs, and atorvastatin could inhibit the leptin expression induced by Ang II. The inhibitory effect of atorvastatin on Ang II-induced leptin expression was mediated by Rac, ROS and JNK pathways.

Effects of angiotensin II on leptin and downstream leptin signaling in the carotid body during acute intermittent hypoxia

“…Furthermore, blocking OB-Rs in these car-529 diomyocytes prevented the hypertrophic effects ANG II 530 normally has on these cells suggesting that leptin may 531 mediate effects normally attributed to direct ANG II action 532 (Rajapurohitam et al, 2012). Finally, in human vascular 533 smooth muscle cells, it has been previously found that 534 increased ANG II results in increased leptin protein and 535 mRNA expression (Shyu et al, 2012). Thus, it is not 536 unreasonable to suggest that leptin may be involved in 537 mediating some of the changes in carotid body chemosen-538 sitivity normally attributed to ANG II (Eyzaguirre and 539 Zapata, 1968;Bock, 1980;Gomez-Nino et al, 1990).…”

B-type natriuretic peptide inhibits angiotensin II-induced proliferation and migration of pulmonary arterial smooth muscle cells

“…In order to measure the production of reactive oxygen species (ROS), an assay using 2′,7′‐dichlorodihydrofluorescein diacetate (DCFH‐DA, Molecular Probes) was performed as previously described . After serum starvation for 48 hr, PASMCs (1 × 10 5 cells) were stimulated by Ang II (10 nM) for 48 hr with or without pretreatment of BNP (1–1,000 nM, 30 min).…”

“…In order to measure the production of reactive oxygen species (ROS), an assay using 2 0 ,7 0 -dichlorodihydrofluorescein diacetate (DCFH-DA, Molecular Probes) was performed as previously described. 20,21 After serum starvation for 48 hr, PASMCs (1 Â 10 5 cells) were stimulated by Ang II (10 nM) for 48 hr with or without pretreatment of BNP (1-1,000 nM, 30 min). One group of PASMCs were pretreated with Rp-cGMPS (50 mM, 30 min) before treatment of BNP (1,000 nM, 30 min), followed by stimulation with Ang II (10 nM, 48 hr).…”

Gene expression signature of human HepG2 cell line

“…IL-10 has been found to be over-expressed in liver cancer tissues with AXIN1 and PIK3CA mutations (Boyault et al, 2007), therefore, its down-regulation in HepG2 cells suggests that it is indicative of the inflammatory process specifically related to HCC because it is found in the absence of viral and genetic complications. As regards adipokines, AGT (angiotensin) is produced constitutively and released into the circulation mainly by the liver; it increases the synthesis of LEP (leptin) that plays a key role in regulating energy intake and energy expenditure (Shyu et al, 2012) and, on the other hand, down-regulates NAMPT (Nicotinamide phosphoribosyltransferase) that activates insulin receptor and has insulin-mimetic effects, by lowering the blood glucose and improving the insulin sensitivity (Benigni et al, 2009). Our data are, as expected, consistent with their close inter-correlation, however, their concomitant down-regulation has also been found in liver tissues with HCC associated to other complications such as HBV, HCV, hematochromatosis, and Wilson's disease (Budhu et al, 2006;Chan et al, 2006), thus, these adipokines cannot be used as liver cancer marker.…”