Department of Nephrology, Zhejiang Provincial People's Hospital, Hangzhou, Zhejiang 310014, P.R. China.
Department of Emergency, Zhejiang University Hospital, Hangzhou, Zhejiang 310058, P.R. China.
Mol Med Rep. 2019 Oct;20(4):3240-3248. doi: 10.3892/mmr.2019.10585. Epub 2019 Aug 9.
Diabetic kidney disease (DKD) is diagnosed increasingly frequently and represents a serious threat to human health. Krüppel‑like factor 4 (KLF4) has aroused attention due to its potential effect on podocytes and in ameliorating proteinuria associated with glomerulopathy. The purpose of the present study was to investigate the potential role of KLF4 in DKD. It was hypothesized that KLF4 impacts diabetic nephropathy by mediating the podocyte autophagic process. A KLF4 plasmid vector was constructed, and podocytes were transfected and incubated with DKD mice serum for in vitro experiments. A db/db spontaneous DKD mouse model was also established for in vivo study. After treatment, the level of serum creatinine (Scr), blood urea nitrogen (BUN), and 24‑h urinary protein was determined. Immunofluorescence and periodic acid‑Schiff staining, western blotting, flow cytometry and a TUNEL assay were performed to observe changes in glomerular morphology and the level of apoptosis, cytoskeleton proteins, epithelial‑mesenchymal transition (EMT) biomarkers, autophagic proteins and mTOR pathway proteins in each group. KLF4 overexpression significantly reduced the level of urinary albumin, Scr, BUN and attenuated mesangial matrix expansion, as well as mesangial cell proliferation in DKD mice. KLF4 overexpression also inhibited podocyte apoptosis and downregulated vimentin and α‑smooth muscle actin, and upregulated E‑cadherin and nephrin, both in vivo and in vitro. Moreover, the microtubule associated protein 1 light chain 3α (LC3)‑II/LC3‑I ratio and LC3‑II fluorescence was significantly increased in the vector‑KLF4 group compared to the negative control vector group both in vivo and in vitro. Finally, a decrease in the level of phosphorylated (p)‑mTOR and p‑S6K protein expression was observed following KLF4 overexpression in vitro. The present findings suggested that KLF4 plays a renoprotective role in DKD, which is associated with the activation of podocyte autophagy, and may be involved in the mTOR signaling pathway.
糖尿病肾病（DKD）的诊断频率越来越高，对人类健康构成严重威胁。Krüppel 样因子 4（KLF4）因其对足细胞的潜在作用以及改善与肾小球病变相关的蛋白尿而受到关注。本研究旨在探讨 KLF4 在 DKD 中的潜在作用。假设 KLF4 通过调节足细胞自噬过程影响糖尿病肾病。构建了 KLF4 质粒载体，并转染足细胞，用 DKD 小鼠血清孵育进行体外实验。还建立了 db/db 自发性 DKD 小鼠模型进行体内研究。治疗后，测定血清肌酐（Scr）、血尿素氮（BUN）和 24 小时尿蛋白水平。通过免疫荧光和过碘酸-希夫染色、Western blot、流式细胞术和 TUNEL 检测观察各组肾小球形态和细胞凋亡水平以及细胞骨架蛋白、上皮-间充质转化（EMT）标志物、自噬蛋白和 mTOR 通路蛋白的变化。KLF4 过表达可显著降低 DKD 小鼠尿白蛋白、Scr、BUN 水平，减轻肾小球系膜基质扩张和系膜细胞增殖。KLF4 过表达还可抑制足细胞凋亡，下调体内和体外的波形蛋白和α-平滑肌肌动蛋白，上调 E-钙黏蛋白和足细胞裂孔膜蛋白。此外，载体-KLF4 组与阴性对照载体组相比，体内和体外微管相关蛋白 1 轻链 3α（LC3）-II/LC3-I 比值和 LC3-II 荧光均显著增加。最后，在体外过表达 KLF4 后，观察到磷酸化（p）-mTOR 和 p-S6K 蛋白表达水平降低。这些发现表明，KLF4 在 DKD 中具有肾保护作用，与足细胞自噬的激活有关，可能涉及 mTOR 信号通路。
