RXRα激动剂贝萨罗汀通过调控TGF-β1/Smad2通路抑制ApoE^−/−小鼠动脉粥样硬化的形成

RXRαagonist bexarotene inhibits atherosclerosis in ApoE^−/−mice by regulating TGF-β1/Smad2 signaling pathway

目的:观察类视黄醇X受体α(RXRα)激动剂贝萨罗汀(Bex)对载脂蛋白E缺陷(ApoE^−/−)小鼠动脉粥样硬化及转化生长因子β1(TGF-β1)诱导的血管平滑肌细胞(VSMCs)增殖的影响,并探讨Bex抑制动脉粥样硬化的机制。方法:选取10只C57BL/6小鼠为正常对照组,选取30只ApoE^−/−小鼠随机分为3组:ApoE^−/−组、ApoE^−/−+Bex5(5 mg·kg^−1·d^−1 Bex)组及ApoE^−/−+Bex10(10 mg·kg^−1·d^−1 Bex)组。Bex采用灌胃法给药,每天1次,连续8周。氧化酶法检测血清甘油三酯(TG)和总胆固醇(TC)水平;选择遮蔽法测定血清低密度脂蛋白胆固醇(LDL-C)和高密度脂蛋白胆固醇(HDL-C)水平;Western blot法测TGF-β1和p-Smad2/Smad2水平;HE染色观察胸主动脉内膜硬化斑块形成情况。组织贴块法培养VSMCs,5-溴脱氧尿嘧啶核苷(BrdU)掺入法检测VSMCs增殖活性。结果:ApoE^−/−组小鼠血清TG、TC和LDL-C水平,以及胸主动脉TGF-β1和p-Smad2水平显著高于C57BL/6组(P<0.01);Bex呈剂量依赖性增加ApoE^−/−小鼠胸主动脉组织p-Smad2蛋白水平,抑制内膜斑块形成及中膜增生,显著减小斑块面积(P<0.01)。ApoE^−/−组、ApoE^−/−+Bex5组和ApoE^−/−+Bex10组之间血清TG、TC、HDL-C和LDL-C水平,以及胸主动脉TGF-β1和Smad2水平均无显著差别(P>0.05)。体外实验显示,TGF-β1(0.1~10μg/L)促进VSMCs增殖,但Bex呈浓度依赖性抑制TGF-β1(5μg/L)诱导的VSMCs增殖。Bex(10−7 mol/L)可增强TGF-β1(5μg/L)诱导的p-Smad2水平(P<0.01),却抑制TGF-β1(5μg/L)诱导的p-Smad2细胞核转位。结论:RXRα激动剂Bex抑制ApoE^−/−小鼠动脉粥样硬化的形成,其机制可能与调控TGF-β1/p-Smad2通路从而抑制VSMCs增殖有关。

AIM:To observe the effect of retinoid X receptorα(RXRα)agonist bexarotene(Bex)on the proliferation of transforming growth factor β1(TGF-β1)-induced vascular smooth muscle cells(VSMCs)and atherosclerosis in apolipoprotein E knockout(ApoE^−/−)mice,and to explore the underlying mechanism.METHODS:Ten C57BL/6 mice were selected as normal control group,and 30 ApoE^−/−mice were randomly divided into 3 groups:ApoE^−/−group,ApoE^−/−+Bex5(5 mg·kg^−1·d^−1 Bex)group and ApoE^−/−+Bex10(10 mg·kg^−1·d^−1 Bex)group.Bex was intragastrically given once a day for 8 weeks.The levels of triglyceride(TG)and total cholesterol(TC)were determined by oxidase method,and select masking method was used to determine serum levels of low-density lipoprotein cholesterol(LDL-C)and high-density lipoprotein cholesterol(HDL-C).The protein levels of TGF-β1,p-Smad2 and Smad2 were determined by Western blot.HE staining was used to observe the intima of the thoracic aorta.The VSMCs were cultured with tissue patch method,and the proliferation of VSMCs was measured by BrdU incorporation method.RESULTS:The serum levels of TG,TC and LDL-C,and the expression of TGF-β1 and p-Smad2 in thoracic aorta in ApoE^−/−group were significantly higher than those in C57BL/6 group(P<0.01).Bex increased p-Smad2 protein level in thoracic aorta in a dose-dependent manner,inhibited the intimal plaque formation and vascular medial proliferation,and decreased the plaque area in ApoE^−/−mice(P<0.01).No significant difference in serum levels of TG,TC,HDL-C and LDL-C,and TGF-β1 and Smad2 expression in thoracic aorta among ApoE^−/−group,ApoE^−/−+Bex5 group and ApoE^−/−+Bex10 group was observed.TGF-β1(0.1~10μg/L)promoted the proliferation of VSMCs,while Bex(10−9~10−7 mol/L)inhibited TGF-β1(5μg/L)-induced proliferation of VSMCs in a concentration-dependent manner.Bex(10−7 mol/L)synergistically promoted the protein level of p-Smad2 in VSMCs induced by TGF-β1(P<0.01),but inhibited TGF-β1-induced nuclear translocation of p-Smad2.CONCLUSION:RXRαagonist Bex inhibits the formation of atherosclerosis in ApoE^−/−mice,and its mechanism may be related to the regulation of TGF-β1/Smad2 pathway.