年龄相关的 EZH2 抑制通过减少 H3K27me3 介导足细胞损伤。

AGE-Induced Suppression of EZH2 Mediates Injury of Podocytes by Reducing H3K27me3.

机构信息

Department of Internal Medicine III, University Hospital Jena, Jena, Germany.

Department of Internal Medicine III, University Hospital Jena, Jena, Germany,

出版信息

Am J Nephrol. 2020;51(9):676-692. doi: 10.1159/000510140. Epub 2020 Aug 27.

DOI:10.1159/000510140

PMID:32854097

Abstract

BACKGROUND

Chronic hyperglycemia, a pivotal feature of diabetes mellitus (DM), initiates the formation of advanced glycation end products (AGEs) and the dysregulation of epigenetic mechanisms, which may cause injury to renal podocytes, a central feature of diabetic kidney disease (DKD). Previous data of our group showed that AGEs significantly reduce the expression of NIPP1 (nuclear inhibitor of protein phosphatase 1) in podocytes in vitro as well as in human and murine DKD. NIPP1 was shown by others to interact with enhancer of zeste homolog 2 (EZH2), which catalyzes the repressive methylation of H3K27me3 on histone 3. Therefore, we hypothesized that AGEs can directly induce epigenetic changes in podocytes.

METHODS

We analyzed the relevance of AGEs on EZH2 expression and activity in a murine podocyte cell line. Cells were treated with 5 mg/mL glycated BSA for 24 h. To determine the meaning of EZH2 suppression, EZH2 activity was inhibited by incubating the cells with the pharmacological methyltransferase inhibitor 3-deazaneplanocin A; EZH2 expression was repressed with siRNA. mRNA expression was analyzed with real-time PCR, and protein expression with Western blot. EZH2 expression and level of H3K27 trimethylation in podocytes of diabetic db/db mice, a mouse model for type 2 DM, were analyzed using immunofluorescence.

RESULTS

Our data demonstrated that AGEs decrease EZH2 expression in podocytes and consequently reduce H3K27me3. This suppression of EZH2 mimicked the AGE effects and caused an upregulated expression of pathological factors that contribute to podocyte injury in DKD. In addition, analyses of db/db mice showed significantly reduced H3K27me3 and EZH2 expression in podocytes. Moreover, the suppression of NIPP1 and EZH2 showed similar effects regarding podocyte injury.

CONCLUSIONS

Our studies provide a novel pathway how AGEs contribute to podocyte injury and the formation of the so-called metabolic memory in DKD.

摘要

背景

慢性高血糖是糖尿病（DM）的一个关键特征，它会引发晚期糖基化终产物（AGEs）的形成和表观遗传机制的失调，这可能导致肾脏足细胞损伤，这是糖尿病肾病（DKD）的一个主要特征。我们小组之前的数据表明，AGEs 在体外以及在人和鼠 DKD 中显著降低了足细胞中 NIPP1（蛋白磷酸酶 1 的核抑制剂）的表达。其他人表明，NIPP1 与增强子结合抑制因子 2（EZH2）相互作用，EZH2 催化组蛋白 3 上 H3K27me3 的抑制性甲基化。因此，我们假设 AGEs 可以直接诱导足细胞中的表观遗传变化。

方法

我们分析了 AGEs 对小鼠足细胞系中 EZH2 表达和活性的相关性。细胞用 5mg/ml 糖化 BSA 处理 24 小时。为了确定 EZH2 抑制的意义，用药理学甲基转移酶抑制剂 3-去氮胞苷 A 孵育细胞来抑制 EZH2 活性；用 siRNA 抑制 EZH2 表达。用实时 PCR 分析 mRNA 表达，用 Western blot 分析蛋白表达。用免疫荧光法分析 2 型糖尿病模型 db/db 小鼠（db/db 小鼠，一种 2 型糖尿病的小鼠模型）足细胞中的 EZH2 表达和 H3K27 三甲基化水平。

结果

我们的数据表明，AGEs 降低了足细胞中的 EZH2 表达，从而降低了 H3K27me3。这种 EZH2 的抑制作用模拟了 AGE 的作用，并导致导致 DKD 中足细胞损伤的病理性因素的表达上调。此外，对 db/db 小鼠的分析表明，足细胞中的 H3K27me3 和 EZH2 表达明显降低。此外，NIPP1 和 EZH2 的抑制作用对足细胞损伤表现出相似的效果。