Qian Xiaoxiao, Tan Juan, Liu Ling, Chen Sheng, You Na, Yong Huijuan, Pan Minglin, You Qiang, Ding Dafa, Lu Yibing
Department of Endocrinology, The Second Affiliated Hospital of Nanjing Medical UniversityNanjing 210000, China.
Nanjing Medical UniversityNanjing 210029, China.
Am J Transl Res. 2018 Mar 15;10(3):989-997. eCollection 2018.
Podocyte apoptosis is a typical early feature of diabetic nephropathy (DN), with loss of nephrin integrity contributing to increased proteinuria in patients with DN. Emerging evidence shows that microRNAs (miRNAs) play vital roles in the pathogenesis of DN. Thus, we aimed to further elucidate the role of miRNAs in podocyte apoptosis in DN. We used db/db and db/m mice maintained under a continuous feeding regime for 12 weeks. Using microarray analysis, we found several miRNAs potentially related to podocyte apoptosis. In addition, we cultured a conditionally immortalized human podocyte cell line in 30 mM D-glucose and found that miR-134-5p was upregulated in both db/db mice and high-glucose (HG)-treated podocytes. Upregulation of miR-134-5p was accompanied by podocyte apoptosis and downregulation of nephrin. Inhibition of miR-134-5p produced the opposite effect. Dual-luciferase reporter assays showed that miR-134-5p directly targeted the 3'-untranslated region of the B-cell lymphoma-2 gene (BCL2), and further study confirmed an increase in bcl-2 protein level in HG-treated podocytes transfected with anti-miR-134-5p. Knockdown of BCL2 impeded the antiapoptotic effect of anti-miR-134-5p. Finally, we found that miR-134-5p might regulate apoptosis in db/db mice and podocytes by targeting BCL2. Taken together, our findings suggest that miR-134-5p promotes podocyte apoptosis under HG conditions by targeting BCL2. Our study provides a meaningful approach to interpret the mechanisms of action of miRNAs involved in DN.
足细胞凋亡是糖尿病肾病（DN）的典型早期特征，nephrin完整性的丧失导致DN患者蛋白尿增加。新出现的证据表明，微小RNA（miRNA）在DN的发病机制中起重要作用。因此，我们旨在进一步阐明miRNA在DN足细胞凋亡中的作用。我们使用在持续喂养方案下饲养12周的db/db和db/m小鼠。通过微阵列分析，我们发现了几种可能与足细胞凋亡相关的miRNA。此外，我们在30 mM D-葡萄糖中培养了一种条件永生化的人足细胞系，发现miR-134-5p在db/db小鼠和高糖（HG）处理的足细胞中均上调。miR-134-5p的上调伴随着足细胞凋亡和nephrin的下调。抑制miR-134-5p产生相反的效果。双荧光素酶报告基因测定表明，miR-134-5p直接靶向B细胞淋巴瘤-2基因（BCL2）的3'-非翻译区，进一步研究证实，在用抗miR-134-5p转染的HG处理的足细胞中，bcl-2蛋白水平增加。敲低BCL2阻碍了抗miR-134-5p的抗凋亡作用。最后，我们发现miR-134-5p可能通过靶向BCL2调节db/db小鼠和足细胞的凋亡。综上所述，我们的研究结果表明，miR-134-5p在HG条件下通过靶向BCL2促进足细胞凋亡。我们的研究为解释参与DN的miRNA的作用机制提供了一种有意义的方法。
