高浓度葡萄糖条件下肾小球系膜细胞内源性烟酰胺磷酸核糖转移酶对波形蛋白表达的调控作用

doi:10.13481/j.1671-587x.20170503

吉林大学学报(医学版) ›› 2017, Vol. 43 ›› Issue (05): 867-873.doi: 10.13481/j.1671-587x.20170503

高浓度葡萄糖条件下肾小球系膜细胞内源性烟酰胺磷酸核糖转移酶对波形蛋白表达的调控作用

翟晓雅¹, 冯乔², 韦日明³, 陈叶¹, 胡婷婷³, 冯乐平¹

1. 桂林医学院公共卫生学院营养与食品卫生学教研室, 广西桂林 541004;
2. 桂林医学院附属医院优生遗传科, 广西桂林 541004;
3. 桂林医学院生物技术学院生物技术教研室, 广西桂林 541004

收稿日期:2017-04-02 出版日期:2017-09-28 发布日期:2017-09-29
通讯作者: 冯乐平,教授,硕士研究生导师(Tel:0773-5835125,E-mail:lpfeng1226@163.com) E-mail:lpfeng1226@163.com
作者简介:翟晓雅(1994-),女,山西省原平市人,医学硕士,主要从事糖尿病发病机制的细胞生物学方面的研究。
基金资助:
国家自然科学基金资助课题（81460164，31060161）；广西壮族自治区科技厅自然科学基金资助课题（2015GXNSF）

Regulation effect of endogenous nicotinamide phosphoribosyltransferase on Vimentin expression of glomerular cells in high concentration of glucose

ZHAI Xiaoya¹, FENG Qiao², WEI Riming³, CHEN Ye¹, HU Tingting³, FENG Leping¹

1. Department of Nutrition and Food Hygiene, School of Public Health, Guilin Medical University, Guilin 541004, China;
2. Department of Prepotency, Affiliated Hospital, Guilin Medical University, Guilin 541004, China;
3. Department of Biotechnology, School of Biotechnology, Guilin Medical University, Guilin 541004, China

Received:2017-04-02 Online:2017-09-28 Published:2017-09-29

摘要/Abstract

摘要： 目的：探讨高浓度葡萄糖条件下肾小球系膜细胞过度表达的内源性烟酰胺磷酸核糖转移酶（Nampt）对波形蛋白（Vimentin）表达的调控作用，阐明糖尿病并发肾脏炎症纤维化的形成机制。方法：取患严重自发性糖尿病的C57/BL6小鼠肾脏，以野生型C57/L86小鼠作为对照组，行病理切片和组织荧光染色，应用免疫共聚焦方法检测肾小球系膜细胞中内源性Nampt和Vimentin表达及定位；肾小球膜HBZY-1细胞随机分为4组，即0.56 mmol·L^-1低浓度葡萄糖（LG）对照组、200 mmol·L^-1高浓度葡萄糖（HG）处理组、HG+FK866组和HG+NMN组；高浓度葡萄糖干预5 d后，分别施加FK866（10 μmol·L^-1）和NMN（1 mmol·L^-1）作用24 h后，通过免疫荧光和免疫印迹法检测细胞内源性Nampt、Vimentin、核因子κB p65（NF-κBp65）和依赖于烟酰胺腺嘌呤二核苷酸（NAD+）的组蛋白去乙酰化酶1（Sirt1）表达水平；应用RT-PCR和免疫印迹等方法检测细胞Nampt和Vimentin表达水平。结果：与野生型C57/BL6小鼠组比较，严重糖尿病组小鼠肾小球明显萎缩，肾小球细胞内Vimentin表达水平随内源性Nampt表达水平升高而增加（P < 0.01）；与0.56 mmol·L^-1低浓度葡萄糖对照组比较，高浓度葡萄糖冲击细胞Nampt、NF-κBp65和Vimentin表达水平明显升高（P < 0.01），Sirt1表达水平明显降低（P < 0.01）。HG+FK866和HG+NMN组肾小球系膜细胞中Nampt、Vimentin和NF-κBp65表达水平均较对照组明显降低（P < 0.01）。结论：高浓度葡萄糖冲击条件下肾小球系膜细胞过度表达的内源性Nampt能够通过NF-κBp65和Sirt1信号通路促进Vimentin表达。

关键词: 高浓度葡萄糖, 核因子κBp65, 去乙酰化酶1, 波形蛋白, 烟酰胺磷酸核糖转移酶

Abstract: Objective: To investigate the regulation effect of endogenous nicotinamide phosphoribosyl transferase (Nampt) on the Vimentin expression of glomerular cells in high concentration glucose,and to clarify the mechanism of formation of diabetic kidney inflammation fibrosis. Methods: The C57/BL6 diabetic mice were selected and the kidney tissues were collected, and the wild C57/L86 mice were used as control group;the pathological section and tissue fluorescence staining were performed.The expression and location of endogenous Nampt and Vimentin in the glomerular cells were detected by immuno-focused technology. The HBZY-1 cells were randomly divided into 4 groups:low concentration of glucose (LG, 0.56 mmol·L^-1) control group,high concentration glucose (HG, 200 mmol·L^-1) group,HG + FK866 group and HG+nicotinamide mononucleotide (NMN) group.In HG group, the cells were treated with FK866 (10 μmol·L^-1) and NMN (1 mmol·L^-1) for 24 h after cultured with HG for 5 d. The expression levels of Nampt, Vimentin, nuclear factor-kappa B p65 (NF-κBp65) and sirtuin type 1 (Sirt1) were detected by immunofluorescence and Western blotting methods. The expression levels of Nampt and Vimentin were detected by RT-PCR and Western blotting methods. Results: The shape and size of glomerulus had obvious atrophy of the mice in severe diabetic group compared with normal C57/BL6 mice group. The expression level of Vimentin in glomerular cells was increased with the increasing of endogenous Nampt (P<0.01). When the HBZY-1 cells were cultured in HG condition,the exprssion levels of Nampt, Vimentin and NF-kB p65 were obviously increased while the Sirt1 expression levels was significantly decreased compared with control group (P<0.01).The expression levels of Nampt, Vimentin and NF-κB p65 in glomerular cells in HG+FK866 HG+NMN groups were singnificartyly decreased compared with control group (P<0.01). Conclusion: The endogenous Nampt over-expression in glomerular cells can enhance the expression of Vimentin under high concentration of glucose stress through NF-κBp65 and Sirt1 signal pathway.

Key words: high concentration of glucose, nicotinamide phosphoribosyltransferase, sirtuin type 1, vimentin, nuclear factor-kappa Bp65

中图分类号:

R363.21

翟晓雅, 冯乔, 韦日明, 陈叶, 胡婷婷, 冯乐平. 高浓度葡萄糖条件下肾小球系膜细胞内源性烟酰胺磷酸核糖转移酶对波形蛋白表达的调控作用[J]. 吉林大学学报(医学版), 2017, 43(05): 867-873.

ZHAI Xiaoya, FENG Qiao, WEI Riming, CHEN Ye, HU Tingting, FENG Leping. Regulation effect of endogenous nicotinamide phosphoribosyltransferase on Vimentin expression of glomerular cells in high concentration of glucose[J]. Journal of Jilin University Medicine Edition, 2017, 43(05): 867-873.

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高浓度葡萄糖条件下肾小球系膜细胞内源性烟酰胺磷酸核糖转移酶对波形蛋白表达的调控作用

Regulation effect of endogenous nicotinamide phosphoribosyltransferase on Vimentin expression of glomerular cells in high concentration of glucose

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