鹿茸多肽介导心肌干细胞分化对终末心肌分化基因ANP和MLC-2v表达的影响

doi:10.13481/j.1671-587x.20180208

吉林大学学报(医学版) ›› 2018, Vol. 44 ›› Issue (02): 249-253.doi: 10.13481/j.1671-587x.20180208

鹿茸多肽介导心肌干细胞分化对终末心肌分化基因ANP和MLC-2v表达的影响

黄晓巍¹, 徐岩¹, 韩冬², 林贺², 律广富¹, 李晓华², 曲晓波¹, 张冰³

1. 长春中医药大学中医药与生物工程研究开发中心药理实验室, 吉林长春 130117;
2. 长春中医药大学药学院临床药学与中药药理教研室, 吉林长春 130117;
3. 北京中医药大学中药学院临床中药系, 北京 100102

收稿日期:2017-10-12 出版日期:2018-03-28 发布日期:2018-03-30
通讯作者: 张冰,教授,博士研究生导师(Tel:010-64286335,E-mail:zhangbing6@263.net) E-mail:zhangbing6@263.net
作者简介:黄晓巍(1972-),女,吉林省长春市人,主任药师,医学博士,主要从事中药药理学方面的研究。
基金资助:
国家自然科学基金面上项目资助课题（81774147）；吉林省卫计委科研计划项目资助课题（2015Z062）；高等学校青年骨干教师国内访问学者研修计划资助课题（2017）；长春中医药大学百名中青年骨干教师培养计划资助课题（2017）

Effect of differentiation of cardiac stem cells mediated by pilose antler polypeptides on expressions of terminal myocardial differentiation genes ANP and MLC-2v

HUANG Xiaowei¹, XU Yan¹, HAN Dong², LIN He², LYU Guangfu¹, LI Xiaohua², QU Xiaobo¹, ZHANG Bing³

1. Pharmacology Laboratory, Research and Development Center of Traditional Chinese Medicine and Bioengineering, Changchun University of Traditional Chinese Medicine, Changchun 130117, China;
2. Department of Clinical Pharmacy and Traditional Chinese Medicine Pharmacology, School of Pharmacy, Changchun University of Traditional Chinese Medicine, Changchun 130117, China;
3. Department of Clinical Chinese Materia, School of Chinese Materia Medicine, Beijing University of Chinese Medicine, Beijing 100029, China

Received:2017-10-12 Online:2018-03-28 Published:2018-03-30

摘要/Abstract

摘要： 目的：探讨鹿茸多肽介导心肌干细胞（CSC）分化对终末心肌分化基因心房钠尿肽（ANP）和心肌肌凝蛋白轻链2（MLC-2v）表达的影响，阐明其修复受损心肌的作用机制。方法：选取出生2d的雄性Wistar大鼠，提取CSC，免疫荧光鉴定CSC表面标志物c-Kit，流式细胞术鉴定CSC纯度。以培养皿为实验单位，分为空白对照组（加等量缓冲液）、5-氮胞苷组（终浓度3 μmol·L^-1）、鹿茸多肽组（终浓度800 mg·L^-1）和联合组（3 μmol·L^-1 5-氮胞苷＋800 mg·L^-1鹿茸多肽），于37℃、5% CO₂培养箱中培养48 h。ELISA法检测各组细胞上清液中ANP和MLC-2v表达水平，RT-PCR法检测各组细胞中ANP和MLC-2v mRNA表达水平。结果：获得纯度>95%的CSC。ELISA法检测，与空白对照组比较，5-氮胞苷组、鹿茸多肽组和联合组细胞上清中ANP和MLC-2v表达水平明显升高（P<0.05）；联合组ANP和MLC-2v表达水平高于5-氮胞苷组和鹿茸多肽组，但差异无统计学意义（P>0.05）。RT-PCR法检测，与空白对照组比较，5-氮胞苷组、鹿茸多肽组和联合组ANP和MLC-2v mRNA表达水平明显升高（P<0.05）。结论：鹿茸多肽可以通过增强心肌分化基因ANP和MLC-2v表达介导CSC分化为心肌细胞，且在一定程度上减轻5-氮胞苷所致的细胞毒性。

关键词: 心肌干细胞, 心房钠尿肽, 鹿茸多肽, 心肌肌凝蛋白轻链2

Abstract: Objective:To explore the effect of differentiation of cardiac stem cells(CSC) mediated by pilose antler polypeptides on the expressions of terminal myocardial differentiation genes atrial natriuretic polypeptide(ANP) and myosin light chain 2v(MLC-2v),and to clarify the mechanism of repairing the damaged myocardium. Methods: The healthy male Wistar rats born 2 d were selected to extract the CSC.The surface marker c-Kit and the purity of CSC were identified by immunofluorescence and flow cytometry.The culture dish was used as the experimental unit and the extracted cells were divided into the following four groups:blank control group (the same amount of buffer was added for induction),5-azacytidine group(induced with 3 μmol·L^-1 5-azacytidine),pilose antler polypeptides group(induced with 800 mg·L^-1 pilose antler polypeptides) and combined group(induced with 800 mg·L^-1 pilose antler polypeptides and 3 μmol·L^-1 5-azacytidine);the cells were incubated for 48 h in the condition of 37℃ and 5% CO₂.The expression levels of ANP and MLC-2v in supernatant of the cells in various groups were detected by ELISA method.The expression levels of ANP and MLC-2v mRNA in the cells in various groups were detected by RT-PCR method. Results: CSC were prepared with the purity>95%.The results of ELISA showed that the expression levels of ANP and MLC-2v in 5-azacytidine, pilose antler polypeptides and combined groups were significantly increased compared with blank control group(P<0.05).The expression levels of ANP and MLC-2v in combined group were increased compared 5-azacytidine and pilose antler polypeptides groups,but there were no significant differences(P>0.05).The result of RT-PCR showed that the expression levels of ANP and MLC-2v mRNA in 5-azacytidine,pilose antler polypeptides and combined groups were significantly increased compared with blank control group(P<0.05). Conclusion: Pilose antler polypeptides can induce the differentiation of CSC into cardiac cells by promoting the expressions of ANP and MLC-2v, and they can reduce the cytotoxicity induced by 5-azacytidine in some degrees.

Key words: pilose antler polypeptides, myosin light chain 2v, cardiac stem cells, atrial natriuretic polypeptide

中图分类号:

R285.5

黄晓巍, 徐岩, 韩冬, 林贺, 律广富, 李晓华, 曲晓波, 张冰. 鹿茸多肽介导心肌干细胞分化对终末心肌分化基因ANP和MLC-2v表达的影响[J]. 吉林大学学报(医学版), 2018, 44(02): 249-253.

HUANG Xiaowei, XU Yan, HAN Dong, LIN He, LYU Guangfu, LI Xiaohua, QU Xiaobo, ZHANG Bing. Effect of differentiation of cardiac stem cells mediated by pilose antler polypeptides on expressions of terminal myocardial differentiation genes ANP and MLC-2v[J]. Journal of Jilin University Medicine Edition, 2018, 44(02): 249-253.

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鹿茸多肽介导心肌干细胞分化对终末心肌分化基因ANP和MLC-2v表达的影响

Effect of differentiation of cardiac stem cells mediated by pilose antler polypeptides on expressions of terminal myocardial differentiation genes ANP and MLC-2v

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