小檗碱对心肌细胞microRNA24的干预作用和其可能机制.pdfVIP

Research Articles 基础研究 365 小檗碱对心肌细胞microRNA24的干预作用及其可能机制 1 2 1 2 1* 刘锐锋 何湘伟 张幼怡 袁 谷 徐 明 【摘要】 目的 探讨天然小分子探针小檗碱干预microRNA24(miR24) 表达水平的作用以及其可能机制。方法体外采用质谱法分析小檗 碱与miR24 的前体rno-mir-24-1 的相互结合作用；采用实时定量PCR 法检测小檗碱对腺病毒携带的rno-mir-24-1 及其成熟体miR24 表达水 平的调控、以及对miR24 的靶基因p27 的mRNA 表达水平的影响；采用Western blot 检测小檗碱对p27 蛋白表达水平的影响。结果小檗碱 与rno-mir-24-1 相互结合，对其表达水平没有显著影响；小檗碱以剂量依赖方式明显抑制miR24 的表达；小檗碱能抑制miR24 过表达引起 的p27 下调。结论小檗碱与rno-mir-24-1 的结合并明显下调miR24 的表达，提示天然小分探针有望成为一种有效干预microRNA 的手段。 【关键词】天然小分子；小檗碱；microRNA ；miR24 Berberine Downregulates MicroRNA24 in Rat Cardiomyocyte and Its Possible Mechanism 1 2 1 2 1* LIU Ruifeng , HE Xiangwei , ZHANG You-yi , YUAN Gu , XU Ming (1. Key Laboratory of Cardiovascular Molecular Biology and Regulatory peptides, Ministry of Health, Institute of Vascular Medicine, Peking University Third Hospital, Beijing 100191, China, 2.Beijing National Laboratory for Molecular Sciences, Key Laboratory of Bioorganic Chemistry and Molecular Engineering of Ministry of Education, Department of Chemical Biology, College of Chemistry and Molecular Engineering, Peking University, Beijing 100871, China). 【Abstract】Objective To understand if natural small molecule probe Berberine could downregulate the expression of microRNA24(miR24) and to explore its possible mechanism. Methods To investigate the non-covalent interaction of rno-mir-24-1 and Berberine by ESI mass spectrometry. And the cardiomyocytes isolated from neonatal Sprague-Dawley rat were transfected with Adenovirus which carried the coding sequences of rno-mir-24-1 and GFP, at the same time the cells were treated with Berberine. Then the expression of rno-mir-24-1 and miR24 was determined by Real time PCR. The expression of P27,a target of miR24 ,was also exam