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It has been established that obstructive sleep apnea (OSA) is an independent risk factor for atherosclerosis. Chronic intermittent hypoxia (CIH) activates sympathoadrenal system and upregulates β3 adrenergic receptor (β3 AR). However, the effect of selective β3 AR agonist mirabegron in CIH-induced atherosclerosis remains unknown. We generated a CIH-induced atherosclerosis model through exposing ApoE−/− mice to CIH (8 h per day, cyclic inspiratory oxygen fraction 5–21%, 60-s cycle) for 6 weeks after 4-week high-fat dieting and investigated the effects of mirabegron, a selective β3 AR agonist, on CIH-induced atherosclerosis. The coronary endarterectomy (CE) specimens from coronary artery disease patients with OSA and without OSA were collected. The expression of β3 AR was significantly elevated in CIH-induced atherosclerosis model. Furthermore, treatment with mirabegron (10mg/kg per day by oral administration for 6 weeks) ameliorated atherosclerosis in ApoE−/− mice in CIH but not in normoxia. Mechanistically, mirabegron activated β3 AR and ameliorated intraplaque oxidative stress by suppressing p22phox expression and reactive oxygen species (ROS) level. In addition, in human CE specimens, β3 AR was also upregulated associated with increased p22phox expression and ROS level both in the lumen and in the plaque of coronary artery in OSA subjects. This study first demonstrated that mirabegron impeded the progression of CIH-induced atherosclerosis, at least in part, via β3 AR–mediated oxidative stress, suggesting a promising therapeutic strategy for protecting against atherosclerosis induced by CIH.

Development of skeletal muscle is a complicated biological process regulated by various regulation factors and signal pathways. MicroRNAs (miRNAs) are novel gene regulators that control muscle cell development. microRNA-143 (miR-143) is highly expressed in skeletal muscle, and we found that miR-143 level is significantly increased during bovine skeletal muscle satellite cells (MSCs) differentiation process through microarray analysis and qRT-PCR detection. However, the function of miR-143 in bovine muscle development remained unclear. In our work, the functions of miR-143 in bovine MSCs myogenic differentiation were investigated. We discovered that IGFBP5 is directly regulated by miR-143 using a dual-luciferase reporter assay. Overexpression of miR-143 led to decreased level of IGFBP5 protein and restrained cell proliferation and differentiation, while downregulation of miR-143 resulted in increased levels of IGFBP5 protein and restrained cell proliferation but improved differentiation. IGFBP5, an important component of IGF signaling pathway, contributes greatly to bovine muscle cell development. A mechanism that miR-143 can regulate the proliferation and differentiation of bovine MSCs through changing expression of IGFBP5 was elucidated by our study.

Rationale: Vascular smooth muscle cell (VSMC) proliferation and migration are crucial events involved in the pathophysiology of vascular diseases. Sirtuin 1 (SIRT1), a class III histone deacetylase (HDAC), has been reported to have the function of antiatherosclerosis, but its role in neointima formation remains unknown. Objective: The present study was designed to investigate the role of SIRT1 in the regulation of neointima formation and to elucidate the underlying mechanisms. Methods and Results: A decrease in SIRT1 expression was observed following carotid artery ligation. smooth muscle cell (SMC)–specific human SIRT1 transgenic (Tg) mice were generated. SIRT1 overexpression substantially inhibited neointima formation after carotid artery ligation or carotid artery wire injury. In the intima of injured carotid arteries, VSMC proliferation (proliferating cell nuclear antigen (PCNA)–positive cells) was significantly reduced. SIRT1 overexpression markedly inhibited VSMC proliferation and migration and induced cell cycle arrest at G1/S transition in vitro. Accordingly, SIRT1 overexpression decreased the induction of cyclin D1 and matrix metalloproteinase-9 (MMP-9) expression by treatment with serum and TNF-α, respectively, whereas RNAi knockdown of SIRT1 resulted in the opposite effect. Decreased cyclin D1 and MMP-9 expression/activity were also observed in injured carotid arteries from SMC-SIRT1 Tg mice. Furthermore, 2 targets of SIRT1, c-Fos and c-Jun, were involved in the downregulation of cyclin D1 and MMP-9 expression. Conclusions: Our findings demonstrate the inhibitory effect of SIRT1 on the VSMC proliferation and migration that underlie neointima formation and implicate SIRT1 as a potential target for intervention in vascular diseases.

Aim: To elucidate the effect and the mechanisms of curcumin on the expression of the human homeobox gene NKX3.1 in the prostate cancer cell LNCaP. Methods: The expression change of NKX3.1 in cells incubated with varying concentrations of curcumin was observed by Western blotting and RT-PCR. A dual luciferase reporter assay was used to test the effect of curcumin on the activity of the NKX3.1 1040 bp promoter. Curcumin-treated cells disposed to a designated amount of androgen analog R1881 and the androgen receptor (AR) antagonist flutamide, then the expression of NKX3.1 or the activity of the NKX3.1 promoter were investigated by Western blotting or reporter gene assay, respectively. Finally, Western blotting and electrophoretic mobility shift assay were performed to demonstrate the effect of curcumin on the expression of AR and its binding activity to the androgen response element (ARE). Results: Curcumin downregulated the expression of NKX3.1 and the activity of the NKX3.1 1040 bp promoter in LNCaP cells. R1881 increased the expression of NKX3.1 , and the AR antagonist flutamide decreased the expression of NKX3.1 in LNCaP cells, while curcumin could inhibit androgen-AR mediated induction of NKX3.1 expression. Curcumin decreased the expression of AR and the binding activity to ARE directly. Conclusion: Curcumin could downregulate NKX3.1 expression in LNCaP cells. It could also inhibit the androgen-AR mediated induction of NKX3.1 expression by downregulating AR expression and blocking its DNA binding activity.

To study the role of sirtuin 1 (SIRT1) in Fas ligand (FasL) expression regulation during vascular lesion formation and to elucidate the potential mechanisms.SIRT1 and FasL protein levels were detected by Western blotting in either mouse arteries extract or the whole rat aortic vascular smooth muscle cell (VSMC) lysate. Smooth muscle cell (SMC)-specific human SIRT1 transgenic (Tg) C57BL/6 mice and their littermate wild-type (WT) controls underwent complete carotid artery ligation (ligation groups) or the ligation-excluded operation (sham groups). The carotid arteries were collected 1 day after operation. Reverse transcription-polymerase chain reaction was performed to detect the mRNA levels of SIRT1 and FasL. Luciferase reporter assays were performed to detect the effect of WT-SIRT1, a dominant-negative form of SIRT1 (SIRT1H363Y), and GATA-6 on the promoter activity of FasL. Flow cytometry assay was applied to measure the hypodiploid DNA content of VSMC so as to monitor cellular apoptosis.SIRT1 was expressed in both rat aortic VSMCs and mouse arteries. Forced SIRT1 expression increased FasL expression both in injured mouse carotid arteries 1 day after ligation (P0.001) and VSMCs treated with serum (P0.05 at the transcriptional level, P0.001 at the protein level). No notable apoptosis was observed. Furthermore, transcription factor GATA-6 increased the promoter activity of FasL (P0.001). The induction of FasL promoter activity by GATA-6 was enhanced by WT-SIRT1 (P0.001), while SIRT1H363Y significantly relieved the enhancing effect of WT-SIRT1 on GATA-6 (P0.001).Overexpression of SIRT1 up-regulates FasL expression in both flow-restricted mouse carotid arteries and serum-stimulated VSMCs. The transcription factor GATA-6 participates in the transcriptional regulation of FasL expression by SIRT1.

Angiotensin II (Ang II) stimulates vascular smooth muscle cell (VSMC) hypertrophy as a critical event in the development of vascular diseases such as atherosclerosis. Sirtuin (SIRT) 1, a nicotinamide adenine dinucleotide dependent deacetylase, has been demonstrated to exert protective effects in atherosclerosis by promoting endothelium-dependent vascular relaxation and reducing macrophage foam cell formation, but its role in VSMC hypertrophy remains unknown. In this study, we tried to investigate the effect of SIRT1 on Ang II-induced VSMC hypertrophy. Results showed that adenoviral-mediated over-expression of SIRT1 significantly inhibited Ang II-induced VSMC hypertrophy, while knockdown of SIRT1 by RNAi resulted in an increased [(3)H]-leucine incorporation of VSMC. Accordingly, nicotinamide adenine dinucleotide phosphate oxidase 1 (Nox1) expression induced by Ang II was inhibited by SIRT1 in VSMCs. SIRT1 activator resveratrol decreased, whereas endogenous SIRT1 inhibitor nicotinamide increased Nox1 expression in A7r5 VSMCs. Furthermore, transcription factor GATA-6 was involved in the down-regulation of Nox1 expression by SIRT1. These results provide new insight into SIRT1's anti-atherogenic properties by suppressing Ang II-induced VSMC hypertrophy.

The proinflammatory cytokine TNF-alpha plays an important role in stimulating inflammatory responses of vascular smooth muscle cells (VSMCs). The anti-inflammatory function of Sirtuin 1 (SIRT1), a NAD-dependent class III histone/protein deacetylase, has been well documented, but how SIRT1 is regulated under inflammatory conditions is largely unknown. In the present research, we showed that levels of SIRT1 mRNA and protein expression increased in TNF-alpha-treated VSMCs. Overexpression of the p65/RelA subunit of NF-kappaB, a TNF-alpha-activated inflammatory transcription factor, in A7r5 cells, upregulated SIRT1 mRNA and protein expression as well as SIRT1 promoter activity, while knockdown of endogenous p65/RelA expression by RNAi not only led to a decrease in SIRT1's basal protein expression and promoter activity, but almost abolished the TNF-alpha-induced elevation of SIRT1 protein expression and SIRT1 promoter activity. Furthermore, using promoter deletion analysis and chromatin immunoprecipitation assays, we found that p65/RelA bound to the SIRT1 promoter at a consensus NF-kappaB binding site. Our study indicates that p65/RelA mediates the TNF-alpha-induced elevated expression of SIRT1 in VSMCs, shedding new light on the regulation of SIRT1 under inflammatory conditions.