Departments of Pathology and Medicine, Northwestern University, Chicago, Illinois 60611, USA.
Am J Pathol. 2011 Oct;179(4):1706-18. doi: 10.1016/j.ajpath.2011.06.035. Epub 2011 Aug 18.
The mechanisms involved in tubular hypertrophy in diabetic nephropathy are unclear. We investigated the role of exchange protein activated by cAMP 1(Epac1), which activates Rap-family G proteins in cellular hypertrophy. Epac1 is expressed in heart, renal tubules, and in the HK-2 cell line. In diabetic mice, increased Epac1 expression was observed, and under high glucose ambience (HGA), HK-2 cells also exhibited increased Epac1 expression. We isolated a 1614-bp DNA fragment upstream of the initiation codon of Epac1 gene, inclusive of glucose response elements (GREs). HK-2 or COS7 cells transfected with the Epac1 promoter revealed a dose-dependent increase in its activity under HGA. Mutations in GRE motifs resulted in decreased promoter activity. HK-2 cells exhibited a hypertrophic response and increased protein synthesis under HGA, which was reduced by Epac1-siRNA or -mutants, whereas the use of a protein kinase A inhibitor had minimal effect. Epac1 transfection led to cellular hypertrophy and increased protein synthesis, which was accentuated by HGA. HGA increased the proportion of cells in the G0/G1 cell-cycle phase, and the expression of pAkt and the cyclin-dependent kinase inhibitors p21 and p27 was increased while the activity of cyclin-dependent kinase 4 decreased. These effects were reversed following transfection of cells with Epac1-siRNA or -mutants. These data suggest that HGA increases GRE-dependent Epac1 transcription, leading to cell cycle arrest and instigation of cellular hypertrophy.
糖尿病肾病中管状肥大的机制尚不清楚。我们研究了 cAMP 激活的交换蛋白 1（Epac1）的作用，它在细胞肥大中激活 Rap 家族 G 蛋白。Epac1 在心脏、肾小管和 HK-2 细胞系中表达。在糖尿病小鼠中，观察到 Epac1 表达增加，在高糖环境（HGA）下，HK-2 细胞也表现出 Epac1 表达增加。我们分离了 Epac1 基因起始密码子上游的 1614bp DNA 片段，包含葡萄糖反应元件（GREs）。用 Epac1 启动子转染的 HK-2 或 COS7 细胞在 HGA 下显示其活性呈剂量依赖性增加。GRE 基序的突变导致启动子活性降低。HK-2 细胞在 HGA 下表现出肥大反应和蛋白质合成增加，这可被 Epac1-siRNA 或 -突变体减少，而蛋白激酶 A 抑制剂的作用最小。Epac1 转染导致细胞肥大和蛋白质合成增加，HGA 进一步加剧了这一效应。HGA 增加了 G0/G1 细胞周期阶段的细胞比例，pAkt 和细胞周期蛋白依赖性激酶抑制剂 p21 和 p27 的表达增加，而细胞周期蛋白依赖性激酶 4 的活性降低。用 Epac1-siRNA 或 -突变体转染细胞可逆转这些效应。这些数据表明，HGA 增加了 GRE 依赖性 Epac1 转录，导致细胞周期停滞和细胞肥大的发生。
