中国血液净化

中国血液净化 ›› 2017, Vol. 16 ›› Issue (06): 407-412.doi: 10.3969/j.issn.1671-4091.2017.06.012

• 基础研究 • 上一篇    下一篇

Apelin-13对高糖诱导肾小管上皮细胞-间充质细胞转分化的影响

韦建辉1,廖兰2,关玉珍2,余坤2   

  1. 1. 柳州市人民医院泌尿内科
    2. 广西科技大学生物与化学工程学院
  • 收稿日期:2016-12-13 修回日期:2017-04-01 出版日期:2017-06-12 发布日期:2017-06-14
  • 通讯作者: 韦建辉 weijianhuigx@sina.com E-mail:weijianhuigx@sina.com

Effect of Apelin-13 on the epithelial-mesenchymal transition of renal tubular epithelial cells induced by high glucose

  • Received:2016-12-13 Revised:2017-04-01 Online:2017-06-12 Published:2017-06-14

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摘要： 目的  观察Apelin-13 对高糖诱导肾小管上皮细胞-间充质细胞转分化(Epithelial-mesenchymal transition,EMT)的影响并探讨其可能机制。方法  用Apelin-13(10-8、10-7和10-6 mol/L)预处理HK-2 细胞30 min 后，用含D-葡萄糖(30 mmol/L)的培养基培养HK-2 细胞48 h。免疫荧光检测HK-2 细胞中间充质细胞的标志物α-平滑肌肌动蛋白(α-SMA)的表达，透射电镜观察HK-2 细胞的超微结构，Western-Blot 检测上皮细胞的标志物E-钙粘蛋白(E-cadherin)、α-SMA 及自噬相关蛋白表达。结果
与对照组比较，高糖组的细胞呈现长梭形，细胞间隙明显增宽，E-cadherin 蛋白表达显著性下调(t=4.751，P=0.011)，而α-SMA 蛋白表达显著上调(t=3.846，P=0.016)，自噬体数量明显增多，Beclin-1、LC3-Ⅱ蛋白表达和LC3-Ⅱ/LC3-Ⅰ的比值均显著增加(t1=5.271，t2=4.695，t3=6.753，P1=0.009，P2=0.012，P3= 0.002)，p62 表达显著降低(t=4.562，P=0.012)。与高糖组比较，Apelin-13(10-7和10-6 mol/L)+高糖与高糖组比较，明显抑制了细胞形态的变化，细胞形态大多数呈椭圆型，E-cadherin 蛋白表达显著上调(t1=3.495，t2=4.124，P1=0.019，P2=0.013)，而α-SMA 蛋白表达显著下调(t1=4.014，t2=4.283，P1=0.014，P2= 0.012)，自噬体数量明显增多，Beclin-1、LC3-Ⅱ蛋白表达和LC3-Ⅱ/LC3-Ⅰ的比值均显著增加(t1=3.487，t2=4.059，t3=5.342，t4=4.271，t5=5.360，t6=5.851，P1=0.018，P2=0.014，P3=0.009，P4=0.011，P5=0.005，P6=0.004)，p62 表达显著降低(t1=4.342，t2=3.557，P1=0.012，P2=0.019)。结论Apelin-13 抑制了高糖诱导的肾小管EMT，其机制可能与Apelin-13诱导肾小管上皮细胞自噬有关。

关键词: Apelin-13, 高糖, 上皮细胞-间充质细胞转分化, 自噬, 自噬相关蛋白

Abstract: Objective To explore the effect of Apelin-13 on epithelia-mesenchymal transition (EMT) of renal tubular epithelial cell line HK-2 cells induced by high glucose. Methods After pretreatment with Apelin-13 (10-8, 10-7 and 10-6 mol/L) for 30 min, HK-2 cells were cultured in the medium containing 30 mmol/L Dglucose
for 48 h. Expression of mesenchymal cell markers alpha smooth muscle actin (α-SMA) in the HK-2 cells were tested by immunofluorescence assay. Ultrastructural changes of HK-2 cells were observed by transmission electron microscopy. Expressions of the epithelial marker E-cadherin, α-SMA, and autophagy related proteins were measured by western blot. Results Compared with the control group, HK-2 cells in the high glucose group showed long spindle shape and widened intercellular spaces, with down-regulated E-cadherin expression (t=4.751, P=0.011), up-regulated α-SMA expression (t=3.846, P=0.016), increased number of autophagy, increased expression of Beclin-1, LC3-II and LC3-II/LC3-I ratio (t=5.271, P=0.009 for Bectlin-1; t=4.695, P=0.012 for LC3- II; t=6.753, P=0.002 for LC3-II/LC3- I ratio), and decreased expression of p62 (t=4.562, P=0.012). Compared with the high glucose group, KH-2 cells in the Apelin-13 (10-7 and 10-6 mol/L) +high glucose group showed oval shape, with up-regulated E-cadherin expression (t=3.495, P=0.019 for 10- 7mol/L Apelin-13; t=4.124, P=0.013 for 10-6 mol/L Apelin-13), down-regulated α-SMA expression (t=4.014, P= 0.014 for 10-7 mol/L Apelin-13; t=4.283, P=0.012 for 10-6 mol/L Apelin-13), increased number of autophagy, increased expression of Beclin-1, and LC3-II and LC3-II/LC3-I ratio (t=3.487, P=0.018 for Beclin-1 and 10-7mol/L Apelin-13; t=4.059, P=0.014 for Beclin-1 and 10- 6 mol/L Apelin-13; t=5.342, P=0.009 for LC3-II and10- 7 mol/L Apelin-13; t=4.271, P=0.011 for LC3-II and 10- 6 mol/L Apelin-13; t=5.360, P=0.005 for LC3-II/LC3-I ratio and 10-7 mol/L Apelin-13; t=5.851, P=0.004 for LC3-II/LC3-I ratio and 10-6 mol/L Apelin-13), and decreased expression of p62 (t=4.342, P=0.012 for 10-7 mol/L Apelin-13; t=3.557, P=0.019 for 10-6 mol/L Apelin-13). Conclusions Apelin-13 inhibits the EMT of renal tubular epithelial cell line HK-2 cells induced by high glucose. The increase of autophagy induced by Apelin-13 may relate to the inhibition of EMT in HK-2
cells.

Key words: Apelin-13, High glucose, Epithelia-mesenchymal transition, Autophagy, Autophagy related protein.