Wu Guang-Jun, Zhao Hong-Biao, Zhang Xiao-Wei
Department of Traditional Chinese Medicine, Linyi Central Hospital, Linyi City, Shandong Province, China.
Department of Peripheral Vascular, Linyi Central Hospital, Linyi City, Shandong Province, China.
Histol Histopathol. 2021 Nov;36(11):1155-1167. doi: 10.14670/HH-18-358. Epub 2021 Jul 16.
Herein, we aimed to determine whether DAPK1 and its post-transcriptional regulator miR-361 were implicated in high glucose (HG)-induced podocyte injury and renal damage in db/db mice.
Podocytes were incubated with normal glucose (NG; 5 mM) or HG (30 mM). Podocyte apoptosis was evaluated using TUNEL staining. Lentiviral-delivered specific short hairpin RNA (shRNA) was designed to silence DAPK1 expression in podocytes. miR-361 agomir was administrated by tail intravenous injection in db/db diabetic mice to investigate the renoprotection of miR-361 in vivo.
Exposure of podocytes to HG led to a significant increase in DAPK1 mRNA and protein levels and a decrease in miR-361 expression levels. Knockdown of DAPK1 attenuated HG-triggered growth inhibition, apoptosis, DNA damage and cell membrane damage in podocytes. Mechanically, DAPK1 was a direct target of miR-361. Transfection with miR-361 mimics into podocytes resulted in a significant decrease in the DAPK1 protein expression level. In addition, HG-induced the up-regulation of the DAPK1 protein expression level in podocytes was restrained by miR-361 mimics transfection. Intriguingly, overexpression of DAPK1 in HG-stimulated podocytes muted miR-361-mediated cytoprotection, including anti-apoptosis, resistance to DNA and membrane damage. In vivo, overexpression of miR-361 protected against hyperglycemia-induced podocyte loss, tubular atrophy and interstitial fibrosis in the kidney of db/db mice. Moreover, overexpression of miR-361 inhibited the protein expression of DAPK1 in the kidney of db/db mice.
Our research presented a novel mechanism of HG-induced podocyte damage or renal lesion, supporting the miR-361/DAPK1 signaling pathway that could be used as a potential therapeutic target for the treatment of DN.
在此，我们旨在确定死亡相关蛋白激酶1（DAPK1）及其转录后调节因子微小RNA-361（miR-361）是否与高糖（HG）诱导的db/db小鼠足细胞损伤和肾损伤有关。
将足细胞分别用正常葡萄糖（NG；5 mM）或HG（30 mM）孵育。采用末端脱氧核苷酸转移酶介导的缺口末端标记法（TUNEL）染色评估足细胞凋亡。设计慢病毒传递的特异性短发夹RNA（shRNA）以沉默足细胞中DAPK1的表达。通过尾静脉注射将miR-361激动剂施用于db/db糖尿病小鼠，以研究miR-361在体内的肾脏保护作用。
足细胞暴露于HG导致DAPK1 mRNA和蛋白水平显著升高，miR-361表达水平降低。敲低DAPK1可减轻HG触发的足细胞生长抑制、凋亡、DNA损伤和细胞膜损伤。机制上，DAPK1是miR-361的直接靶点。将miR-361模拟物转染到足细胞中导致DAPK1蛋白表达水平显著降低。此外，miR-361模拟物转染可抑制HG诱导的足细胞中DAPK1蛋白表达水平上调。有趣的是，在HG刺激的足细胞中过表达DAPK1可消除miR-361介导的细胞保护作用，包括抗凋亡、抵抗DNA和膜损伤。在体内，miR-361过表达可防止db/db小鼠肾脏中高血糖诱导的足细胞丢失、肾小管萎缩和间质纤维化。此外，miR-361过表达可抑制db/db小鼠肾脏中DAPK1的蛋白表达。
我们的研究提出了HG诱导足细胞损伤或肾脏病变的新机制，支持miR-361/DAPK1信号通路可作为治疗糖尿病肾病（DN）的潜在治疗靶点。
