二甲双胍通过AMPK/PPAR-γ通路诱导自噬抑制高糖对血管平滑肌细胞的增殖作用

张紫微; 郑甲林; 许晓宇; 王红

doi:10.12259/j.issn.2095-610X.S20231026

二甲双胍通过AMPK/PPAR-γ通路诱导自噬抑制高糖对血管平滑肌细胞的增殖作用

doi: 10.12259/j.issn.2095-610X.S20231026

张紫微¹,
郑甲林¹,
许晓宇^{1, 2},
王红^1, ,

1.
云南省阜外心血管病医院冠心病中心，云南昆明　650032
2.
昆明医科大学研究生院，云南昆明　650500

基金项目: 云南省科技厅-昆明医科大学应用基础研究联合专项基金资助项目（81170250）；云南省医学创新团队基金资助项目（202005AE160020）

详细信息

作者简介:
张紫微（1985～），男，江西九江人，医学博士，副主任医师，主要从事冠心病基础研究与介入诊疗工作

通讯作者:
王红，E-mail：wangh43@126.com

中图分类号: R54
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出版历程
- 收稿日期: 2023-08-23
- 网络出版日期: 2023-09-12
- 刊出日期: 2023-10-25

Metformin Inhibits high Glucose-induced Vascular Smooth Muscle Cells Proliferation by Promoting Autophagy via AMPK/PPAR-γ

Ziwei ZHANG¹,
Jialin ZHENG¹,
Xiaoyu XU^{1, 2},
Hong WANG^{1
, ,}

1.
Center of Coronary Heart Disease，Fuwai Yunnan Cardiovascular Hospital， Kunming Yunnan 650032，China
2.
Graduate School of Kunming Medical University， Kunming Yunnan 650500，China

摘要

摘要: 目的探讨二甲双胍（Metformin）对高糖培养的小鼠主动脉血管平滑肌细胞（vascular smooth muscle cell，VSMCs）自噬、细胞增殖的影响及其调控机制。方法用Western blot检测不同处理组微管相关蛋白轻链-3-Ⅱ（LC3-Ⅱ），Becline-1蛋白水平变化，透射电镜观察各处理组VSMCs自噬小体水平；用Western blot检测不同处理组p-AMPK/AMPK、PPAR-γ蛋白表达水平；用EDU法检测不同处理组VSMCs增殖情况。结果二甲双胍、雷帕霉素逆转高糖抑制VSMCs自噬的作用（P < 0.001）；Compound C、GW9662增加高糖抑制VSMCs自噬的作用（P < 0.05）；二甲双胍逆转高糖对AMPK/PPAR-γ通路的负调控（P < 0.01）；二甲双胍、雷帕霉素抑制高糖促进的VSMCs增殖（P < 0.001），Compound C、GW9662逆转二甲双胍抑制高糖促VSMCs增殖的作用（P < 0.05）。结论高糖促进VSMCs增殖可能是自噬调节相关的，二甲双胍可通过激活AMPK/PPAR-γ通路上调VSMCs自噬水平，抑制高糖促进的VSMCs异常增殖。
- 二甲双胍 /
- 高糖 /
- 自噬 /
- 血管平滑肌细胞 /
- 增殖
Abstract: Objective To investigate the effect and regulatory mechanism of metformin on autophagy and cell proliferation of vascular smooth muscle cells （VSMCs） cultured in high glucose in mice aortic. Methods Western blot was used to detect the changes in microtubule-associated protein light chain 3-II （LC3-II） and Beclin-1 protein levels in different treatment groups. Transmission electron microscope was used to observe the the level of autophagosomes in VSMCs of each treatment group. Western blot was used to detect the expression levels of p-AMPK/AMPK and PPAR-γ proteins in different treatment groups. The EDU method was used to detect the proliferation of VSMCs in different treatment groups. Results Metformin and rapamycin reversed the inhibitory effect of high glucose on VSMC autophagy （P < 0.001）. Compound C and GW9662 enhanced the inhibitory effect of high glucose on VSMC autophagy （P < 0.05）. Metformin reversed the negative regulation of high glucose on the AMPK/PPAR-γ pathway （P < 0.01）. Metformin and rapamycin inhibited high glucose-induced VSMC proliferation （P < 0.001）, and Compound C and GW9662 reversed the inhibitory effect of metformin on high glucose-induced VSMC proliferation （P < 0.05）. Conclusion High glucose promotes the proliferation of VSMCs, which may be regulated by autophagy. Metformin can upregulate autophagy levels in VSMCs by activating the AMPK/PPAR-γ pathway, thereby inhibiting the abnormal proliferation of VSMCs induced by high sugar.
- Metformin /
- High glucose /
- Autophagy /
- Vascular smooth muscle cell /
- Proliferation

HTML全文

图 1 二甲双胍对高糖培养下的VSMCs自噬水平的影响

A：Western blot检测对照组、HG组、HG+MET组及HG+RAP组LC3-Ⅱ/LC3-Ⅰ、Becline-1的表达情况；B：定量分析不同处理组LC3-Ⅱ/LC3-Ⅰ的表达水平；C：定量分析不同处理组Becline-1蛋白的表达水平；D：电镜检测对照组、HG组、MET+HG组自噬小体（12000×）（黑色箭头示自噬小体）。^*P < 0.05，^**P < 0.01，^***P < 0.001 。

Figure 1. Effect of metforin on autophagy in HG-cultured VSMCs

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图 2 AMPK、PPAR-γ抑制剂对VSMCs自噬的影响

A：Western blot检测对照组、HG组、HG+CC组及HG+GW组LC3-Ⅱ/LC3-Ⅰ、Becline-1的表达情况；B：定量分析不同处理组LC3-Ⅱ/LC3-Ⅰ的表达水平；C：定量分析不同处理组Becline-1的表达水平。^*P < 0.05，^**P < 0.01，^***P < 0.001。

Figure 2. Effect of AMPK or PPAR-γ inhibitor on autophagy in VSMCs

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图 3 二甲双胍对高糖培养下VSMCs p-AMPK/AMPK、PPAR-γ表达的影响

A：Western blot检测对照组、HG组、HG+MET组及HG+CC组p-AMPK/AMPK的表达；B：定量分析不同处理组p-AMPK/AMPK的表达水平；C：Western blot检测对照组、HG组、HG+MET组及HG+GW组PPAR-γ的表达；D：定量分析不同处理组PPAR-γ的表达水平。^*P < 0.05，^**P < 0.01，^***P < 0.001。

Figure 3. Effect of metformin and high glucose on p-AMPK/AMPK、PPAR-γexpression of VSMCs

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图 4 二甲双胍抑制高糖诱导的VSMCs增殖

A：EDV法检测不同处理组VSMCs增殖情况；B：EDV阳性细胞统计比较。^* P < 0.05，^** P < 0.01，^***P < 0.001 VS CTRL；^#P < 0.05，^###P < 0.001 VS HG，^&P < 0.05，^&&P < 0.01，^&&&P < 0.001 VS HG+MET。

Figure 4. Effect of metformin on high glucose-induced proliferation of VSMCs

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