Expression and function of Ets-1 during experimental acute renal failure in rats
Expression and function of Ets-1 during experimental acute renal failure in rats
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DOI：
10.1097/01.asn.0000145459.54236.d3
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发表时间：
2004-12-01
影响因子：
13.6
通讯作者：
Sasaki, S
中科院分区：
文献类型：
--
作者：
Tanaka, H;Terada, Y;Sasaki, S
转录因子Ets家族由保守的DNA结合Ets结构域定义，该结构域形成有翼的螺旋-转角-螺旋结构基序。Ets家族参与多种生物学功能，包括细胞生长、迁移和分化。本研究假设Ets-1在急性肾功能衰竭（ARF）后再生过程中重新表达，并在细胞周期蛋白D1的转录调控和肾小管细胞的细胞周期进程中起关键作用。为了阐明Ets-1在ARF中的意义，使用大鼠ARF体内模型和LLC-PK 1细胞作为体外模型。夹闭大鼠左肾动脉1 h后，于6、12 h取全肾匀浆，提取总RNA。再灌注后24、48和72 h，通过Western印迹分析和实时逆转录-PCR。Ets-1 mRNA和蛋白表达在缺血后6 ~ 24 h均显著增加。缺氧诱导因子-1 α的表达在缺血再灌注后6 h即显著增加，在缺血再灌注后48 h和72 h下降。免疫组化显示Ets-1在近端肾小管表达，并与增殖细胞核抗原（PCNA）共表达。Ets-1过表达可促进LLC-PK 1细胞周期的进程，增加cyclin D1启动子活性和蛋白表达。Ets-1启动子活性增加3和6小时之间的缺氧，缺氧也诱导LLC-PK 1细胞中的Ets-1蛋白水平的变化。缺氧诱导Ets-1的表达可被显性负性缺氧诱导因子-1 α（hypoxia-inducible factor-1 alpha）的转染所阻断。凝胶位移分析表明，在缺血再灌注条件下，Ets-1结合到细胞周期蛋白D1启动子的ets-1结合位点。过表达Ets-1对LLC-PK 1细胞caspase 3活性和细胞死亡ELISA值无明显影响。总而言之，这些数据表明Ets-1在ARE中肾小管的细胞周期进程中发挥关键作用。Ets-1途径可能调节细胞周期蛋白D1的转录并控制ARF中肾小管的再生。
The Ets family of transcription factors is defined by a conserved DNA-binding Ets domain that forms a winged helix-turn-helix structure motif. The Ets family is involved in a diverse array of biologic functions, including cellular growth, migration, and differentiation. The hypothesis in this study was that Ets-1 is re-expressed during regeneration after acute renal failure (ARF) and plays a key role in the transcriptional regulation of cyclin D1 and the cell cycle progression in renal tubular cells. For clarifying the significance of Ets-1 in ARF, a rat ARF model in vivo,and LLC-PK1 cells as an in vitro model were used. After the left rat renal artery was clamped for 1 h, the whole kidney homogenate was examined and total RNA was extracted at 6, 12. 24, 48, and 72 h after reperfusion by Western blot analysis and real-time reverse transcription-PCR. Ets-1 mRNA and protein expression were strongly increased at 6 to 24 h after the ischemia, respectively. The expression of hypoxia-inducible factor-1alpha was increased dramatically as early as 6 h after ischemia-reperfusion and decreased at 48 and 72 h after ischemia-reperfusion. In the immunohistologic examination, Ets-1 was expressed in the proximal tubules and coexpressed with proliferating cell nuclear antigen (PCNA). Furthermore, overexpression of Ets-1 promoted the cell cycle and increased the promoter activity and protein expression of cyclin D1 in LLC-PK1 cells. Ets-1 promoter activity increased between 3 and 6 h in hypoxia, and hypoxia also induced changes in the Ets-1 protein level in LLC-PK1 cells. The Ets-1 induction by hypoxia was abolished by the transfection of dominant-negative hypoxia-inducible factor-1alpha. A gel shift assay demonstrated that Ets-1 binds to the ets-1 binding site of the cyclin D1 promoter in the ischemia-reperfusion condition. Overexpression of Ets-1 did not significantly change the caspase 3 activity or the value of cell death ELISA in LLC-PK1 cells. Taken together, these data suggest that Ets-1 plays a key role in the cell-cycle progression of renal tubules in ARE The Ets-1 pathway may regulate the transcription of cyclin D1 and control the regeneration of renal tubules in ARF.