Pioglitazone modulates the proliferation and apoptosis of vascular smooth muscle cells via peroxisome proliferators-activated receptor-gamma.

BACKGROUND: PPARγ is a member of the nuclear hormone receptor superfamily. It has been considered as a mediator regulating metabolism, anti-inflammation, and pro-proliferation in the Vascular Smooth Muscle Cells (VSMCs). Thiazolidinediones (TZDs), synthetic ligands of PPARγ, have anti-proliferative and pro-apoptotic effects on VSMCs, which prevent the formation and progression of atherosclerosis and restenosis following percutaneous coronary intervention (PCI). However, the underlying mechanism remains elusive. This present study therefore aimed to investigate the signaling pathway by which pioglitazone, one of TZDs, inhibits proliferation and induces apoptosis of VSMCs. METHODS: The effects of pioglitazone on VSMC proliferation and apoptosis were studied. Cell proliferation was determined using BrdU incorporation assay. Cell apoptosis was monitored with Hoechst and Annexin V staining. The expression of caspases and cyclins was determined using real-time PCR and Western blot. RESULTS: Pioglitazone treatment and PPARγ overexpression inhibited proliferation and induced apoptosis of VSMCs, whereas blocking by antagonist or silencing by siRNA of PPARγ significantly attenuated pioglitazone's effect. Furthermore, pioglitazone treatment or PPARγ overexpression increased caspase 3 and caspase 9 expression, and decreased the expression of cyclin B1 and cyclin D1 in VSMCs. CONCLUSIONS: Pioglitazone inhibits VSMCs proliferation and promotes apoptosis of VSMCs through a PPARγ signaling pathway. Up-regulation of caspase 3 and down-regulation of cyclins mediates pioglitazone's anti-proliferative and pro-apoptotic effects. Our results imply that pioglitazone prevents the VSMCs proliferation via modulation of caspase and cyclin signaling pathways in a PPARγ-dependent manner.

MicroRNA-34a regulates high glucose-induced apoptosis in H9c2 cardiomyocytes.

Hyperglycemia is an important initiator of cardiovascular disease, contributing to the development of cardiomyocyte death and diabetic complications. The purpose of the present study was to investigate whether high glucose state could induce apoptosis of rat cardiomyocyte cell line H9c2 through microRNA-mediated Bcl-2 signaling pathway. The expression of miR-34a and Bcl-2 mRNA was detected by using real-time PCR. Western blotting was used to examine the changes in apoptosis-associated protein Bcl-2. Apoptosis of H9c2 cells was tested by using flow cytometry. The results showed that the expression of miR-34a was significantly elevated and that of Bcl-2 was strongly reduced, and apoptosis of cardiomyocytes was apparently increased in the high-glucose-treated H9c2 cells as compared with normal-glucose-treated controls. In addition, we identified Bcl-2 gene was the target of miR-34a. miR-34a mimics reduced the expression of Bcl-2 and increased glucose-induced apoptosis, but miR-34a inhibitor acted as the opposite mediator. Our data demonstrate that miR-34a contributes to high glucose-induced decreases in Bcl-2 expression and subsequent cardiomyocyte apoptosis.

C-reactive protein induces interleukin-6 and thrombospondin-1 protein and mRNA expression through activation of nuclear factor-ĸB in HK-2 cells.

BACKGROUND: Although C-reactive protein (CRP) is significantly increased in patients with diabetic nephropathy, whether CRP exerts direct proinflammatory effects on human renal tubular epithelial cells (HK-2 cells) is still unclear. METHODS: HK-2 cells were incubated with purified CRP at clinically relevant concentrations (0, 5, 10, 20 and 40 µg/ml). The protein and transcript levels of thrombospondin-1 (TSP-1) and interleukin-6 (IL-6) were determined by ELISA and RT-PCR. Phosphorylation of p38MAPK was investigated through Western blot analysis in HK-2 cells induced by CRP. The activation of nuclear factor-kappa B (NF-κB) was studied via EMSA. A specific p38MAPK inhibitor (SB203580) and an NF-κB inhibitor (PDTC; pyrrolidine dithiocarbamate) were used to analyze the signal transduction in CRP induction. To explore the direct or indirect role of CRP in HK-2 cells, IL-6 or TSP-1 antibodies were used. The expression of IL-6, TSP-1 and transforming growth factor-ß(1 )(TGF-ß(1)) were determined through Western blot analysis in HK-2 cells. RESULTS: In HK-2 cells, purified CRP significantly induced protein release and mRNA expression of IL-6 and TSP-1 in a dose- and time-dependent manner. TGF-ß(1) protein was overexpressed in HK-2 cells induced by CRP, which cannot be inhibited by IL-6 or TSP-1 antibodies. CRP triggered phosphorylation of p38MAPK and activation of NF-κB-mediated signal transduction. SB203580 (5 µM) and PDTC (50 µM) efficiently suppressed those effects of CRP in HK-2 cells. CONCLUSIONS: CRP induces IL-6 and TSP-1 protein release and mRNA expression from HK-2 cells via activation of the p38MAPK and NF-κB signaling pathways and TGF-ß(1) was highly expressed in HK-2 cells, suggesting that CRP plays an important role in the propagation and prolongation of inflammation in renal fibrosis.

PPARgamma gene C161T substitution alters lipid profile in Chinese patients with coronary artery disease and type 2 diabetes mellitus.

BACKGROUND: Peroxisome proliferator-activated receptor gamma (PPARgamma) is a ligand-activated transcription factor, which regulates gene expression of the key proteins involved in lipid metabolism, vascular inflammation, and proliferation. PPARgamma may contribute to attenuating atherogenesis and postangioplasty restenosis. PPARgamma C161-->T substitution is associated with a reduced risk of coronary artery disease (CAD). Whether or not the gene substitution alters the risk of CAD in type 2 diabetes mellitus (T2DM) patients remains unclear. METHODS: A total of 556 unrelated subjects from a Chinese Han population, including 89 healthy subjects, 78 CAD patients, 86 T2DM patients, and 303 CAD combined with T2DM patients, were recruited to enroll in this study. PPARgammaC161-->T gene polymorphism was determined by polymerase chain reaction and restriction fragment length polymorphisms. Plasma levels of lipoproteins, apolipoproteins, glucose, and insulin were measured by ELISA or radioimmunoassay (RIA). The coronary artery lesions were evaluated by coronary angiography. RESULTS: The frequency of the 161T allele in CAD, T2DM, and CAD combined with T2DM patients was similar to that observed in the healthy control group. However, in CAD combined with T2DM patients, the group with angiographically documented moderate stenoses had a higher frequency of the 161T allele in comparison to the group with severe stenoses (P < 0.05). Moreover, in CAD with T2DM patients, the triglyceride levels and apoB in CC homozygote carriers were significantly higher than those in "T" allele carriers. CONCLUSIONS: PPARgammaC161-->T genotypes weren't significantly associated with the risk of CAD, but were markedly correlated with severity of disease vessels in patients with CAD and T2DM. Furthermore, PPARgammaC161-->T substitution was associated with an altered adipose, but not glucose metabolism. These results indicate that the PPARgamma C161-->T polymorphism may reduce the risk of severe atherogenesis by modulation of adipose metabolism, especially triglycerides and apoB, in Chinese patients with CAD and T2DM.