MicroRNA-140-5p ameliorates the high glucose-induced apoptosis and inflammation through suppressing TLR4/NF-κB signaling pathway in human renal tubular epithelial cells.

Hyperglycemia-induced renal tubular cell injury is thought to play a critical role in the pathogenesis of diabetic nephropathy (DN). However, the role of miRNAs in renal tubular cell injury remains to be fully elucidated. The aim of the present study was to investigate the role and mechanisms of miRNAs protecting against high glucose (HG)-induced apoptosis and inflammation in renal tubular cells. First, we analyzed microRNA (miRNA) expression profiles in kidney tissues from DN patients using miRNA microarray. It was observed that miRNA-140-5p (miR-140-5p) was significantly down-regulated in kidney tissues from patients with DN. An inverse correlation between miR-140-5p expression levels with serum proteinuria was observed in DN patients, suggesting miR-140-5p may be involved in the progression of DN. HG-induced injury in HK-2 cells was used to explore the potential role of miR-140-5p in DN. We found that miR-140-5p overexpression improved HG-induced cell injury, as evidenced by the enhancement of cell viability, and inhibition of the activity of caspase-3 and reactive oxygen species (ROS) generation. It was also observed that up-regulation of miR-140-5p suppressed HG induced the expressions of pro-inflammatory cytokines, such as tumor necrosis factor-α (TNF-α), interleukin (IL)-1β and IL-6 in HK-2 cells. In addition, TLR4, one of the upstream molecules of NF-κB signaling pathway, was found to be a direct target of miR-140-5p in the HK-2. Moreover, the HG-induced activation of NF-κB signaling pathway was inhibited by miR-140-5p overexpression. These results indicated that miR-140-5p protected HK-2 cells against HG-induced injury through blocking the TLR4/NF-κB pathway, and miR-140-5p may be considered as a potential prognostic biomarker and therapeutic target in the treatment of DN.

Su J ，Ren J ，Chen H ，Liu B ... -  《-》

MicroRNA-25 inhibits high glucose-induced apoptosis in renal tubular epithelial cells via PTEN/AKT pathway.

Diabetic nephropathy (DN) has become the major cause of end-stage renal disease (ESRD). It has been demonstrated that apoptosis of renal tubular epithelial cells induced by hyperglycemia contributes to the pathogenesis of DN. Recent researches have corroborated the critical roles of microRNAs (miRNAs) in the apoptosis of various types of cells including renal tubular epithelial cells. However, the eff ; ;ect of miRNAs on the hyperglycemia-induced apoptosis of renal tubular epithelial cells remains unclear. The aim of this study is to explore the eff ; ;ect of miRNAs on the hyperglycemia-induced apoptosis of renal tubular epithelial cells and its molecular mechanism. Using a miRNA microarray, miRNAs putatively associated with DN were examined in renal biopsy tissue samples from DN patients and healthy controls. Validation analysis of miR-25 level in serum samples and renal biopsy tissue samples was performed using quantitative reverse transcription PCR (qRT-PCR). Then, gain- and loss- of function experiments were performed to determine the protective roles of miR-25 in high glucose-induced damage to renal tubular epithelial cells. Furthermore, the target gene of miR-25 and the downstream signaling pathway were also investigated. Microarray analysis and qRT-PCR revealed that miR-25 was significantly downregulated in renal biopsy tissue and serum samples from DN patients. We also observed that an inverse relationship between serum miR-25 level and proteinuria in DN patients. Meanwhile, miR-25 was decreased in human kidney (HK-2) cells subjected to HG treatment in a time dependent manner. Its overexpression reduced production of reactive oxygen species (ROS), suppressed cell apoptosis in HG-induced cell damage model, which was coupled with the decreased expression of cleaved caspase-3 and activity of caspase-3. Subsequent analyses demonstrated that phosphatase and tensin homolog deleted on chromosome ten (PTEN) was a direct and functional target of miR-25, which was validated by the dual luciferase reporter assay. Most importantly, the overexpression of PTEN effectively reversed the protective effects of miR-25 mimics on renal tubular epithelial cell injury. We also found that the anti-apoptotic effects of miR-25 are dependent on the activation of PTEN/Akt pathway. In addition, we observed that PTEN was upregulated in renal biopsy tissue samples from patients with DN, and an inverse relationship was found between PTEN and miR-25 expression, suggesting that miR-25 may exert its function through regulation of PTEN in DN. Taken together, our study proved that overexpression of miR-25 could ameliorate HG-induced oxidative stress and apoptosis in renal tubular epithelial cells through activation of PTEN/AKT pathway, suggesting that overexpression of miR-25 might provide a potential therapeutic approach for DN.

Li H ，Zhu X ，Zhang J ，Shi 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 ... -  《-》

lncRNA MALAT1 Promotes Diabetic Nephropathy Progression via miR-15b-5p/TLR4 Signaling Axis.

The long noncoding RNA metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) are closely associated with the pathogenesis of diabetic nephropathy (DN). But a complete mechanism for MALAT1 in DN has yet to be identified. This study investigated the effect of MALAT1 on DN through the regulation of miR-15b-5p/TLR4 signaling. Renal tissues were collected from DN patients. Human renal tubular epithelial cells (HK-2) were used as a model of DN induced by high glucose (HG). We then measured the viability, apoptosis, and inflammatory cytokine levels of HK-2 cells using the corresponding assays. Following transfections of si-MALAT1, si-MALAT1+miR-15b-5p inhibitor, or si-MALAT1+vector TLR4 into HG-stimulated HK-2 cells, cell viability, apoptosis, and inflammatory cytokines were again measured. Furthermore, dual-luciferase reporter assay validated the interactions of MALAT1/miR-15b-5p and miR-15b-5p/TLR4. In addition, the interaction between MALAT1 and miR-15b-5p was investigated by RNA immunoprecipitation (RIP). A significant upregulation of MALAT1 was observed in DN kidney tissues, as well as in HG-stimulated HK-2 cells. MALAT1 knockdown attenuates the inhibition of cell viability, apoptosis, and inflammatory response induced by HG in HK-2 cells. Moreover, a miR-15b-5p inhibitor or TLR4 overexpression reversed the above effects induced by MALAT1 knockdown. These results indicate that reduced MALAT1 ameliorates HG-stimulated HK-2 cell damage through an inhibition of the miR-15b-5p/TLR4 axis. MALAT1 may serve as a biomarker and potential therapeutic target for DN.

Yang Z ，Song D ，Wang Y ，Tang L ... -  《-》