β-Amyrin ameliorates diabetic nephropathy in mice and regulates the miR-181b-5p/HMGB2 axis in high glucose-stimulated HK-2 cells.

Diabetic nephropathy (DN) is a diabetic complication that can cause renal failure. β-amyrin has been identified to possess anti-diabetic property. This study was designed to evaluate the potential role of β-amyrin in DN and its underlying mechanism. Streptozotocin-induced diabetic mice were used as the in vivo model, and high glucose (HG)-stimulated human proximal tubular HK-2 cells were utilized as the in vitro model. Renal histological changes in mice were assessed by hematoxylin-eosin and periodic acid-Schiff staining. HK-2 cell viability and apoptosis were detected by Cell Counting Kit-8 assay and flow cytometry analysis, respectively. β-amyrin was found to ameliorate kidney injury in DN mice and suppressed inflammatory response as well as apoptosis of HG-stimulated HK-2 cells. miR-181-5p expression in murine renal tissues and HK-2 cells was detected by in situ hybridization (ISH) and fluorescence in situ hybridization (FISH). MiR-181b-5p, a previously identified target for diabetic kidney disease, was downregulated in renal tissues and HG stimulated HK-2 cells, and β-amyrin induced the upregulation of miR-181b-5p. Binding relationship between miR-181b-5p and high mobility group box 2 (HMGB2) was confirmed by luciferase reporter assay. MiR-181b-5p bound to 3' untranslated region of HMGB2 to suppress its expression. As shown by immunohistochemical staining and immunofluorescence staining, HMGB2 was upregulated in the in vivo and in vitro models of DN, and β-amyrin induced the downregulation of HMGB2. Moreover, HMGB2 overexpression neutralized the suppressive effects of miR-181b-5p elevation on the inflammatory response and apoptosis of HG-treated HK-2 cells. Overall, β-amyrin ameliorates DN in mice and suppresses inflammatory response and apoptosis of HG-stimulated HK-2 cells via the miR-181b-5p/HMGB2 axis.

Xu W ，Zhang H ，Zhang Q ，Xu J ... -  《-》

Downregulation of Salusin-β protects renal tubular epithelial cells against high glucose-induced inflammation, oxidative stress, apoptosis and lipid accumulation via suppressing miR-155-5p.

Chen H ，Jin G 《-》

MiRNA-133a-3p Attenuates Renal Tubular Epithelial Cell Injury via Targeting MALM1 and Suppressing the Notch Signaling Pathway in Diabetic Nephropathy.

Diabetic nephropathy (DN) is a serious microvascular complication of diabetes characterized by structural and functional changes of kidneys. Human renal tubular epithelial (HK-2) cells are important for kidney recovery post injury and usually used for establishment of DN cell models. The study explored the role of microRNA (miR)-133a-3p in DN cell model and animal model. A cell model for DN was established via high glucose (HG) stimulation to HK-2 cells. Cell viability and apoptotic rate were measured by cell counting kit 8 and flow cytometry. Polymerase chain reaction was performed to quantify levels of miR-133a-3p and targets. Luciferase reporter assay was conducted to verify the binding of miR-133a-3p and MAML1. After establishment of a mouse model of DN, levels of renal function indicators were measured by biochemical analysis. Hematoxylin-eosin and periodic acid-schiff staining of kidney samples were performed to analyze histological changes. Western blotting was conducted to quantify levels of apoptotic markers, MAML1, and factors related to Notch signaling. Results showed that HG induced HK-2 cell apoptosis and the reduction of cell viability. MiR-133a-3p was lowly expressed in HG-stimulated HK-2 cells. Overexpressed miR-133a-3p improved HK-2 cell injury by increasing cell viability and hampering apoptosis under HG condition. In addition, miR-133a-3p directly targets MAML1 3'-untranslated region. MAML1 overexpression countervailed the repressive impact of miR-133a-3p on cell apoptosis in the context of HG. Moreover, miR-133a-3p inhibited the activity of Notch pathway by downregulating MAML1. MiR-133a-3p inhibits DN progression in mice, as evidenced by reduced fasting blood glucose level, improved levels of renal function parameters, and alleviation of kidney atrophy. In conclusion, miR-133a-3p improves HG-induced HK-2 cell injury and inhibits DN progression by targeting MAML1 and inactivating Notch signaling.

Li Y ，Tan P ，Liu Q ，Liu M ，Wang Y ，Kong W ，Sun H ，Shao X ... -  《-》

Circ_WBSCR17 aggravates inflammatory responses and fibrosis by targeting miR-185-5p/SOX6 regulatory axis in high glucose-induced human kidney tubular cells.

Diabetic nephropathy (DN), a severe microvascular complication of diabetes, has complex pathogenesis. Circular RNAs (circRNAs) exert broad biological functions on human diseases. This study intended to explore the role and mechanism of circ_WBSCR17 in DN. DN mice models were constructed using streptozotocin injection, and DN cell models were assembled using high glucose (HG) treatment in human kidney 2 cells (HK-2). The expression of circ_WBSCR17, miR-185-5p and SRY-Box Transcription Factor 6 (SOX6) was detected by quantitative real-time polymerase chain reaction (qRT-PCR). The protein levels of SOX6 and fibrosis markers were examined by western blot. The release of inflammatory cytokines, cell proliferation and apoptosis, were assessed by enzyme-linked immunosorbent assay (ELISA), cell counting kit-8 (CCK-8) assay and flow cytometry assay, respectively. The predicted interaction between miR-185-5p and circ_WBSCR17 or SOX6 was verified by dual-luciferase reporter assay and RNA immunoprecipitation (RIP) assay. Circ_WBSCR17 was highly expressed in DN mice models and HG-induced HK-2 cells. Circ_WBSCR17 knockdown or SOX6 knockdown promoted cell proliferation and blocked cell apoptosis, inflammatory responses and fibrosis, while circ_WBSCR17 overexpression or SOX6 overexpression conveyed the opposite effects. MiR-185-5p was a target of circ_WBSCR17 and directly bound to SOX6. MiR-185-5p could reverse the role of circ_WBSCR17 or SOX6. Moreover, the expression of SOX6 was modulated by circ_WBSCR17 through intermediating miR-185-5p. Circ_WBSCR17 triggered the dysfunction of HG-induced HK-2 cells, including inflammatory responses and fibrosis, which was accomplished via the miR-185-5p/SOX6 regulatory axis.

Li G ，Qin Y ，Qin S ，Zhou X ，Zhao W ，Zhang D ... -  《-》

Long non-cording RNA XIST promoted cell proliferation and suppressed apoptosis by miR-423-5p/HMGA2 axis in diabetic nephropathy.

This research studied the effect of long non-coding RNA X-inactive-specific transcript (XIST) on DN. The effect of high glucose (HG) on the expression of XIST and miR-423-5p was detected by quantitative real-time PCR (qRT-PCR) in human kidney (HK) cells (human glomerular mesangial cells (HMCs) and human kidney-2 (HK-2) cells). The effect of XIST depletion and miR-423-5p inhibition or overexpression on high mobility group protein A2 (HMGA2) protein level was examined by western blot in HG-induced HK cells. The impacts of XIST depletion on viability and apoptosis were assessed by 3-(4, 5-dimethyl-2-thiazolyl)-2, 5-diphenyl-2-H-tetrazolium bromide (MTT) and flow cytometry assays in HG-induced HK cells. We found the expression of XIST and HMGA2 protein was significantly upregulated in DN tissues and cells. Moreover, HG treatment induced the upregulation of XIST and HMGA2 protein level in HK cells. Besides, both XIST depletion and HMGA2 depletion decreased cell proliferation but increased apoptosis in HG-treated HK cells. Furthermore, HMGA2 upregulation or miR-423-5p inhibition partly eliminated the effects of XIST depletion on cell proliferation, apoptosis of HG-treated HK cells. Interestingly, HMGA2 upregulation partly reversed miR-423-5p overexpression-mediated suppression on viability and promotion on apoptosis in HG-treated HK cells. Mechanistically, XIST sponged miR-423-5p to regulate HMGA2 expression in DN cells. Taken together, XIST depletion suppressed proliferation and promoted apoptosis via miR-423-5p/HMGA2 axis in HG-treated HK cells, which may provide a potential therapeutic target for DN.

Chen H ，Guo Y ，Cheng X 《-》