上海交通大学学报（医学版）

上海交通大学学报（医学版）

• 论著（基础研究） • 上一篇    下一篇

Klotho在急性缺血再灌注性肾损伤中的变化及其与凋亡的关系

钱盈盈, 严玉澄, 沈 玥, 车 琳, 关雪晶, 戴慧莉, 顾乐怡, 吴 蓓, 倪兆慧, 钱家麒   

  1. 上海交通大学 医学院附属仁济医院肾脏科， 上海 200127
  • 出版日期:2014-06-28 发布日期:2014-06-30
  • 通讯作者: 严玉澄, 电子信箱: yucheng.yan@163.com。
  • 作者简介:钱盈盈（1988—）， 女， 硕士生; 电子信箱: qyyconcer@163.com。
  • 基金资助:

    国家自然科学基金（81170687）；上海市科委医学引导项目（134119a2300）；上海市科委重大项目（12DJ1400200）

Expression change of Klotho in acute kidney injury induced by ischemia-reperfusion and its relationship with apoptosis

QIAN Ying-ying, YAN Yu-cheng, SHEN Yue, CHE Lin, GUAN Xue-jing, DAI Hui-li, GU Le-yi, WU Bei, NI Zhao-hui, QIAN Jia-qi   

  1. Renal Division, Renji Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200127, China
  • Online:2014-06-28 Published:2014-06-30
  • Supported by:

    National Natural Science Foundation of China, 81170687; Medical Guidance Project of Science and Technology Commission of Shanghai Municipality, 134119a2300; Major Project of Science and Technology Commission of Shanghai Municipality, 12DJ1400200

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摘要：

目的 观察Klotho蛋白在缺血再灌注急性肾损伤中的表达变化，探讨其在肾小管上皮细胞凋亡中的作用。方法 10只BALB/c小鼠随机分为肾脏缺血再灌注组（I/R组, n=5）和假手术组（Sham组, n=5），采用双侧肾蒂夹闭术建立肾缺血再灌注模型，于造模后24 h留取小鼠的血清及肾组织。应用酶法分别测定血清尿素氮和肌酐，ELISA法检测血清Klotho蛋白水平，TUNEL染色法检测肾脏细胞凋亡水平，Real-Time PCR及Western blotting技术分别检测肾组织Klotho、Bax、Bcl-2和Caspase-3的基因及蛋白表达，应用Pearson直线相关法分析急性肾损伤时小鼠全身及局部的Klotho水平和肾脏凋亡的关系。结果 与Sham组相比，I/R组小鼠术后24 h肾小管上皮细胞明显变性、坏死，肾脏凋亡细胞明显增加，肾组织cleaved Caspase-3蛋白水平、Bax/Bcl-2 mRNA及蛋白表达比值均显著升高。I/R组小鼠血清Klotho蛋白水平较Sham组小鼠显著降低[（669.89±136.51） pg/mL vs. (2 107.92±549.22) pg/mL，P<0.01]，肾组织Klotho mRNA及蛋白表达分别较Sham组下调约9/10 （P<0.001）及1/5 （P<0.05）。相关性分析显示肾组织bax/bcl-2 mRNA比值与血清Klotho蛋白及肾组织klotho mRNA均呈显著负相关（r=-0.833，P<0.01；r=-0.916，P<0.001），肾组织Bax/Bcl-2蛋白比值和肾组织Klotho蛋白也呈显著负相关（r=-0.637，P<0.05）。结论 缺血再灌注急性肾损伤后小鼠全身及肾组织局部Klotho表达均显著下调，Klotho水平的下降可能与凋亡诱导的肾损害密切相关。

关键词: 缺血再灌注损伤, 急性肾损伤, Klotho, Bax/Bcl-2, 凋亡

Abstract:

Objective To observe the expression change of Klotho in acute kidney injury (AKI) induced by ischemia-reperfusion and to explore the effects on the apoptosis of renal tubular epithelial cells. Methods Ten BALB/c mice were randomly divided into the ischemia-reperfusion group (I/R group, n=5) and sham-operation group (Sham group, n=5). The AKI model was established by bilateral renal pedicles clamping. The serum and renal issues of mice were collected after the model was established for 24 h. Levels of blood urea nitrogen and serum creatinine were detected by the enzymatic assay. The level of serum Klotho was measured by the ELISA. TUNEL staining was performed to assess the renal apoptosis. Expressions of genes and proteins of Klotho, Bax, Bcl-2, and Caspase-3 of renal tissues were detected by the Real-Time PCR and Western blotting. The relationship between the general and local levels of Klotho and the renal apoptosis was analyzed for mice with AKI by the Pearson line correlation method. Results Compared to the Sham group, renal tubular cells of mice of the I/R group significantly degenerated and necrotized; apoptotic cells increased significantly; and the level of cleaved Caspase-3 protein, Bax to Bcl-2 mRNA, and the protein ratios of renal tissues were significantly increased after 24 h of operations. The levels of serum Klotho of the I/R group were significantly lower than those of the Sham group [(669.89±136.51) pg/mL vs. (2 107.92±549.22) pg/mL, P<0.01]. The mRNA and protein expressions of renal Klotho were downregulated by approximately 90% (P<0.001) and 20% (P<0.05), respectively. The correlation analysis showed that the mRNA ratio of renal bax to bcl-2 was negatively correlated to both serum Klotho protein (r=-0.833, P<0.01) and renal klotho mRNA (r=-0.916, P<0.001). The renal Bax/Bcl-2 protein ratio was also significantly and negatively correlated to the renal Klotho protein (r=-0.637, P<0.05). Conclusion The general and local expressions of Klotho of mice with AKI induced by the ischemia-reperfusion decrease significantly, which may be closely related to the kidney injury induced by the apoptosis.

Key words: ischemia-reperfusion injury, acute kidney injury, Klotho, Bax/Bcl-2, apoptosis