非诺贝特对人BMEC与小鼠脑微血管组织SOD3表达的影响及机制

摘要

Objective To investigate the regulating effects of fenofibrate,the agonists of peroxisome proliferator-acti-vated receptor α(PPARα),on the superoxide dismutase 3 (SOD3)expression in human brain microvascular endothelial cells (BMECs)and mouse brain microvascular tissues as well as the possible mechanism.Methods The cultured BMECs were randomly divided into two groups,which were seperately treated with 10 μg/ml of fenofibrate and the same volume of DMSO for 12 h.Fifteen mice were divided into two groups,which were treated with 30 mg/kg fenofibrate and 10 mL/kg of 10% CMC by gavage for 7 days.The tRNA was extracted and the brain microvessel was isolated.The mRNA ex-pression levels of PPARαand SOD3 were detected by RT-PCR.We constructed a luciferase reporter plasmid pGL4 mSOD3 containing SOD3 promoter.The luciferase reporter plasmid pGL4 mu mSOD3 mutated from PPRE sequence were obtained by using the mutagenesis kit.The BMECs were transfected with pGL4 mSOD3 and pGL4 mu mSOD3 for 24 h.The cells were treated with fenofibrate at a final concentration of 10 μg/mL for 12 h in order to measure the luciferase activity.Re-sults The mRNA expression of PPARαand SOD3 was increased in the BMECs and brain microvessel after the treatment of fenofibrate (all P <0.05).The luciferase activity of pGL4 mSOD3 in BMECs treated with fenofibrate was enhanced after transfection of pGL4 mSOD3 for 24 h (P <0.05).The luciferase activity of BMECs treated with fenofibrate did not signifi-cantly change after transfection of pGL4 mu mSOD3 for 24 h (P >0.05).Conclusion The SOD3 expression in the human BMECs and mouse brain microvascular tissues is regulated by fenofibrate through activating PPARα.%目的 探讨过氧化物酶体增殖物激活受体 α亚型(PPARα)激动剂非诺贝特对人脑微血管(BMEC)与小鼠脑微血管组织细胞外铜/锌超氧化物歧化酶(SOD3)表达的调节作用及其机制.方法 将培养好的BMEC随机分为两部分,分别加入终浓度为10μg/mL非诺贝特、同等体积的DMSO处理12 h;将15只小鼠随机分为两部分,分别给予30 mg/kg非诺贝特、10%的羧甲基纤维素混悬液10 mL/kg连续灌胃7 d,处死小鼠取脑组织,提取脑微血管;用RT-PCR技术检测BMEC与脑微血管组织内过氧化物酶体增殖物激活受体 α亚型(PPARα)、SOD3的mRNA表达.构建含SOD3启动子荧光素酶报告质粒pGL4 mSOD3,用突变试剂盒定点获得过氧化物酶体增殖物反应元件(PPRE)序列突变的荧光素酶报告质粒pGL4 mu mSOD3,将pGL4 mSOD3、pGL4 mu mSOD3转染BMEC 24 h,取部分细胞用终浓度10μg/mL的非诺贝特处理12 h,检测细胞荧光素酶活性.结果 经非诺贝特处理后,BMEC及小鼠脑微血管组织中PPARα、SOD3 mRNA表达均升高(P均<0.05).转染pGL4 mSOD324 h,经非诺贝特处理后BMEC内pGL4 mSOD3的荧光素酶活性增强(P<0.05);转染pGL4 mu mSOD324 h,经非诺贝特处理的BMEC荧光素酶活性变化不明显(P>0.05).结论 非诺贝特通过激活PPARα从而调节人BMEC与脑微血管组织中SOD3的表达.