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目的探讨沉默信息调节因子1(silent mating type information regulation 2 homolog-1,SIRT1)对氧化型低密度脂蛋白(oxidized low density lipoprotein,ox-LDL)诱导的血管平滑肌细胞(vascular smooth muscle cell,VSMC)泡沫化的作用和相关的分子机制。方法用组织贴块法体外培养原代VSMC,用80μg/mL ox-LDL刺激VSMC诱导泡沫细胞形成。检测SIRT1在ox-LDL刺激不同时间(24、48、72 h)的表达变化。SIRT1的活性调控分别应用SIRT1激动剂(SRT1720,SRT,1μmol/L)和抑制剂(nicotinamide,Nic,100μmol/L)进行干预。干预24 h后采用Western blot检测VSMC过氧化物酶体增殖物激活受体γ(peroxisome proliferator-activated receptorγ,PPARγ)、酰基辅酶A:胆固醇酰基转移酶1(A-cholesterol acyltransferase 1,ACAT1)的蛋白表达,干预48 h后采用油红O染色检测细胞内脂质沉积和泡沫细胞形成情况。应用PPARγ激动剂(Rosiglitazone,RSG,50μmol/L)和抑制剂(GW9662,10μmol/L)调控PPARγ的表达,Western blot检测VSMC中ACAT1的表达。结果 1ox-LDL诱导的VSMC泡沫细胞中,SIRT1蛋白表达在48 h降低约47%(P<0.01);油红O染色显示,SRT可以抑制VSMC泡沫细胞形成,而Nic可逆转SRT的抑制作用;2ox-LDL诱导的VSMC泡沫细胞中ACAT1的表达显著增加[(2.77±0.70),P<0.01],SIRT1激动剂SRT可显著抑制ACAT1的表达[(0.90±0.27),P<0.01],而SIRT1抑制剂Nic可阻断这一作用,升高ACAT1的表达[(2.25±0.37),P<0.05];3ox-LDL诱导的VSMC泡沫细胞中PPARγ的表达减少[(0.73±0.12),P<0.05],SIRT1激动剂SRT可上调VSMC中PPARγ的表达[(0.98±0.16),P<0.05],SIRT1抑制剂Nic抑制PPARγ的表达[(0.47±0.13),P<0.01];4PPARγ激动剂RSG可抑制ACAT1的表达[(0.73±0.09),P<0.01],而PPARγ抑制剂GW9662则上调ACAT1表达[(1.68±0.09),P<0.01]。结论 SIRT1通过上调VSMC中PPARγ表达而抑制ACAT1表达,从而抑制ox-LDL诱导的VSMC泡沫样变。
Objective To investigate the effects of silent mating type information regulation 2 homolog-1 (SIRT1) on vascular smooth muscle cell (VSMC) induced by oxidized low density lipoprotein (ox-LDL) The role of foam and related molecular mechanisms. Methods Primary VSMCs were cultured in vitro by tissue patch method and stimulated with 80 μg / mL ox-LDL to induce the formation of foam cells. The change of SIRT1 expression in ox-LDL stimulated at different time (24,48,72 h) was detected. The activity of SIRT1 was controlled by SIRT1 agonist (SRT1720, SRT, 1μmol / L) and nicotinamide (Nic, 100μmol / L) respectively. Western blot was used to detect the protein expression of peroxisome proliferator-activated receptor gamma (PPARγ), acyl-CoA: ACAT1 in VSMCs 24 h after intervention Expression and intervention 48 h after the use of oil red O staining intracellular lipid deposition and foam cell formation. The expression of PPARγ was regulated by PPARγ agonist (Rosiglitazone, RSG 50μmol / L) and inhibitor (GW9662,10μmol / L). The expression of ACAT1 in VSMC was detected by Western blot. Results The expression of SIRT1 protein in VSMC foam cells induced by 1ox-LDL decreased about 47% at 48 h (P <0.01). Oil red O staining showed that SRT inhibited the formation of foam cells in VSMCs, The expression of ACAT1 was significantly increased in VSMC foam cells induced by 2ox-LDL [(2.77 ± 0.70), P <0.01]. The STI1 agonist SRT significantly inhibited the expression of ACAT1 [(0.90 ± 0.27), P <0.01] Inhibition of Nic could block this effect and increase the expression of ACAT1 [(2.25 ± 0.37), P <0.05]; PPARγ expression decreased in VSMC foam cells induced by 3ox-LDL [(0.73 ± 0.12), P <0.05 SIRT1 agonist SRT up-regulated the expression of PPARγ in VSMC [(0.98 ± 0.16), P <0.05], SIRT1 inhibitor Nic inhibited the expression of PPARγ [(0.47 ± 0.13), P <0.01]; 4PPARγ agonist RSG (0.73 ± 0.09), P <0.01], while PPARγ inhibitor GW9662 up-regulated the expression of ACAT1 [(1.68 ± 0.09), P <0.01]. Conclusion SIRT1 inhibits ACAT1 expression by up-regulating PPARγ expression in VSMC, thereby inhibiting ox-LDL-induced foamy change in VSMC.