Xiu Yang1, Ming Yang2, Yuemei Chen3, Yingying Qian1, Xiao Fei1, Chanjuan Gong2, Ming Wang1, Xiangcheng Xie1, Zhen Wang4. 1. Department of Nephrology, Affiliated Hangzhou First People's Hospital, School of Medicine, Zhejiang University Hangzhou, Zhejiang Province, China. 2. Department of Nephrology, Shanghai Changzheng Hospital Shanghai City, China. 3. Department of Nephrology, The Second Affiliated School of Medicine, Hospital of Zhejiang University Hangzhou, Zhejiang Province, China. 4. Department of Nephrology & Rheumatology, Shanghai Tenth People's Hospital, Tongji University School of Medicine Shanghai, China.
Abstract
OBJECTIVE: Diabetic nephropathy (DN) is a serious kidney-based complication of diabetes, wherein podocyte injury is deemed crucial in the development of early stage. Various miRNAs, as report goes, is involved in the pathogenesis of varieties of kidney diseases including DN. In this study, we found a target relationship between miR-30a-5p and Becn1, of which there are few studies about the role in podocyte injury. We therefore used immortalized rat podocyte cell line to explore the role and molecular mechanism of miR-30a-5p targeting Becn1 gene in high-glucose-induced glomerular podocyte injury. METHODS: The mRNA and protein expressions of miR-30a-5p and Becn1 were detected respectively by quantitative reverse transcriptase PCR and western blotting. The proliferation, apoptosis, and the levels of interleukin (IL)-6 and tumor necrosis factor (TNF)-α were detected by MTT assay, flow cytometry, and enzyme-linked immuno sorbent assay, respectively. Intracellular reactive oxygen species (ROS), superoxide dismutase (SOD) and malondialdehyde (MDA) levels were also determined. RESULTS: Compared with normal group, miR-30a-5p in model groups were down-regulated, while Becn1 expression was significantly up-regulated, with slower proliferation, higher apoptosis rate, lower SOD level, and significantly higher ROS, MDA, IL-6, and TNF-α levels (all P<0.05). Overexpression of miR-30a-5p or Becn1 knock-out could lower Becn1 expression, apoptosis rate, promote proliferation, with relatively higher SOD level and lower ROS, MDA, Il-6, and TNF-α levels of model cells (all P<0.05). CONCLUSION: Up-regulation of miR-30a-5p can suppress the expression of Becn1 to increase the growth and inhibit the apoptosis of immortalized rat podocyte cell line, therefore ameliorating podocyte injury induced by high glucose in vitro. AJTR
OBJECTIVE:Diabetic nephropathy (DN) is a serious kidney-based complication of diabetes, wherein podocyte injury is deemed crucial in the development of early stage. Various miRNAs, as report goes, is involved in the pathogenesis of varieties of kidney diseases including DN. In this study, we found a target relationship between miR-30a-5p and Becn1, of which there are few studies about the role in podocyte injury. We therefore used immortalized rat podocyte cell line to explore the role and molecular mechanism of miR-30a-5p targeting Becn1 gene in high-glucose-induced glomerular podocyte injury. METHODS: The mRNA and protein expressions of miR-30a-5p and Becn1 were detected respectively by quantitative reverse transcriptase PCR and western blotting. The proliferation, apoptosis, and the levels of interleukin (IL)-6 and tumor necrosis factor (TNF)-α were detected by MTT assay, flow cytometry, and enzyme-linked immuno sorbent assay, respectively. Intracellular reactive oxygen species (ROS), superoxide dismutase (SOD) and malondialdehyde (MDA) levels were also determined. RESULTS: Compared with normal group, miR-30a-5p in model groups were down-regulated, while Becn1 expression was significantly up-regulated, with slower proliferation, higher apoptosis rate, lower SOD level, and significantly higher ROS, MDA, IL-6, and TNF-α levels (all P<0.05). Overexpression of miR-30a-5p or Becn1 knock-out could lower Becn1 expression, apoptosis rate, promote proliferation, with relatively higher SOD level and lower ROS, MDA, Il-6, and TNF-α levels of model cells (all P<0.05). CONCLUSION: Up-regulation of miR-30a-5p can suppress the expression of Becn1 to increase the growth and inhibit the apoptosis of immortalized rat podocyte cell line, therefore ameliorating podocyte injury induced by high glucose in vitro. AJTR
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