南京大学生命科学学院,南京大学生命科学学院,南京大学生命科学学院,南京大学生命科学学院,南京大学生命科学学院,南京大学生命科学学院
国家自然科学基金资助项目(81070653, 81270907, 81370926, J1103512)
State Key Laboratory of Pharmaceutical Biotechnology,School of Life Sciences,Nanjing University,State Key Laboratory of Pharmaceutical Biotechnology,School of Life Sciences,Nanjing University,State Key Laboratory of Pharmaceutical Biotechnology,School of Life Sciences,Nanjing University,State Key Laboratory of Pharmaceutical Biotechnology,School of Life Sciences,Nanjing University,School of Life Sciences, Nanjing University,State Key Laboratory of Pharmaceutical Biotechnology,School of Life Sciences,Nanjing University
This work was supported by a grant from The National Natural Science Foundation of China (81070653,81270907, 81370926,J1103512)
本实验室前期研究发现,2型糖尿病动物模型ob/ob小鼠血清中miR-122的含量较正常C57BL/6小鼠显著升高.本文进一步研究肝脏特异性miR-122及其靶蛋白AldoA(果糖1, 6-二磷酸醛缩酶A)在ob/ob小鼠肝脏代谢中的作用.首先,经qRT-PCR技术检测发现ob/ob小鼠肝脏miR-122水平较C57BL/6小鼠显著下降,而Western blotting分析发现ob/ob小鼠肝脏其靶蛋白AldoA的表达水平显著上升.进一步以miR-122分子转染293T细胞后收集其分泌的微囊泡(microvesicles,MVs),经qRT-PCR检测确认后采用特异性荧光染料DiI-C18标记MVs,以不同剂量尾静脉注射BALB/c小鼠体内,不同时间点取肝组织做冰冻切片.在荧光显微镜下观察证实,包裹有miR-122的MVs通过循环系统进入肝脏,同时qRT-PCR定量分析发现肝组织中miR-122含量显著升高,而蛋白质印迹检测发现其靶蛋白AldoA在肝脏中表达显著下降.AldoA主要催化糖酵解途径中果糖1, 6-二磷酸和磷酸二羟丙酮及甘油醛-3-磷酸之间的转变,miR-122靶向作用AldoA可能在2型糖尿病的发生发展中发挥重要作用.
In previous study, our group have found that there is a significant increased level of miR-122 in the serum of ob/ob mice, an animal model of type 2 diabetes. Here, we further investigated the role of miR-122 targeting AldoA in the liver of ob/ob mice. First, a significant decrease of miR-122 level and a notable increase of AldoA expression was found in the liver of the ob/ob mice. Second, mature miR-122 was transfected into 293T cells and then MVs isolated from 293T cells were collected; qRT-PCR was applied to confirm that miR-122 was rich in MVs. Third, specific fluorescent dye DiI-C18-labeled MVs were injected intravenously into BALB/c mice; the frozen section of liver was observed through fluorescent microscopy. Finally, miR-122 targeting AldoA in the metabolism of ob/ob mice was confirmed by qRT-PCR and Western blotting. AldoA mainly catalysed the transformation between dihydroxyacetone phosphate, glyceraldehyde-3- phosphate and fructose 1, 6 - bisphosphate in glycolytic pathway. MiR-122 may play an important role in the pathologenesis of ob/ob mice through AldoA pathway.
方志娟,李鹏,刁文丽,蒋珽,张辰宇,项阳. MiR-122在AldoA介导的ob/ob小鼠肝脏代谢中的作用研究[J].生物化学与生物物理进展,2014,41(2):185-191
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