Baoping Chen

Resistin-Like Molecule Beta (RELM-β) Regulates Proliferation of Human Diabetic Nephropathy Mesangial Cells via Mitogen-Activated Protein Kinases (MAPK) Signaling Pathway

Background Resistin-like molecule beta (RELM-β) has been reported to be associated with diabetic nephropathy (DN). However, the role of RELM-β in DN is poorly understood. This study was conducted to delineate the underlying mechanisms of action and to investigate the role of RELM-β in the primitive development of DN via MAPK signaling pathways. Material/Methods Lentivirus-mediated vectors and RNAi technology were used to establish the model of RELM-β up-regulated and down-regulated expression in human mesangial cells (HMCs). The proliferation of HMCs was detected through CCK-8 method. The cell cycle and cell proliferation of HMCs was detected through flow cytometry. The MAPKs pathway protein activity was detected through Western blotting. Results The HMCs with up-regulated and down-regulated expression of RELM-β increased or decreased significantly at 2–3 days. The HMCs with high glucose intervention reversed the proliferation inhibition. The HMCs with exogenous glucose or RELM-β protein intervention partially reversed the cell cycle inhibition. Among the MAPKs pathway, the phosphorylation activity of p38MAPK and JNK increased or decreased and ERK1/2 did not change in the overexpression or inhibition of RELM-β. The p38 MAPK pathway inhibitor SB202190 significantly inhibited the proliferation of HMCs caused by overexpression of RELM-β. Up-regulated expression of RELM-β induced the phosphorylation of p38 MAPK, JNK in HMCs and promoted HMCs proliferation and participated in early DN through the MAPKs pathway. Conclusions The results provide evidence that RELM-β is a potential molecular target for the treatment of DN.

Protocatechuic acid ameliorates high glucose-induced extracellular matrix accumulation in diabetic nephropathy

Protocatechuic acid (PCA), a phenolic compound of anthocyanins, was reported to possess various pharmacologic properties, including anti-oxidant, anti-inflammatory, anti-apoptosis, anti-diabetic and anti-tumor activities. However, the role of PCA in diabetic nephropathy remains elusive. The present study was conducted to evaluate the effects of PCA on extracellular matrix (ECM) accumulation in high glucose (HG)-induced human mesangial cells (MCs) and explore the possible mechanism. Our results demonstrated that PCA obviously inhibited HG-induced proliferation of MCs in a dose-dependent manner. In addition, PCA effectively reduced the protein expression levels of type IV collagen, laminin and fibronectin induced by HG, as well as decreased the levels of ROS and MDA in HG-stimulated MCs. Mechanistic studies showed that PCA efficiently down-regulated the phosphorylation level of p38 MAPK in HG-stimulated MCs. Taken together, our present study demonstrated that PCA protects MCs against HG damage might via inhibition of the p38 MAPK signaling pathway. Thus, PCA might be a beneficial agent for the prevention and treatment of diabetic nephropathy.

Scoparone attenuates high glucose-induced extracellular matrix accumulation in rat mesangial cells

Abstract Scoparone, a major constituent of Artemisia capillaries, has a variety of biological properties including anticoagulant, hepatoprotective, anti‐tumor, anti‐fibrosis, anti‐inflammatory, antioxidant, and antidiabetic activities. However, the renoprotective effect of scoparone under diabetic conditions remains elusive. Thus, the present study was undertaken to examine the role of scoparone in high glucose‐induced mesangial cell proliferation and extracellular matrix (ECM) accumulation and elucidate the possible mechanism of action of scoparone. Our results demonstrated that treatment with scoparone significantly inhibited the proliferation of mesangial cells under high glucose conditions. In addition, scoparone reversed high glucose‐induced fibronectin and collagen IV expression in mesangial cells, as well as suppressed reactive oxygen species production and NOX2/4 expression in high glucose‐exposed mesangial cells. Mechanistic studies revealed that scoparone prevented the activation of ERK1/2 signaling pathway in high glucose‐exposed mesangial cells, and an ERK inhibitor (U0126) protected mesangial cells treated with high glucose. Taken together, these results demonstrated that scoparone protects mesangial cells against high glucose damage in part through the inactivation of ERK signaling pathway. These findings suggest that scoparone may represent a potential drug for the treatment of diabetic nephropathy.

miR-182 enhances acute kidney injury by promoting apoptosis involving the targeting and regulation of TCF7L2/Wnt/β-catenins pathway

Abstract Acute kidney injury (AKI) is a sudden decay in renal function leading to increasing morbidity and mortality. miR‐182 has been reported to be actively involved in kidney diseases. However, the function and molecular mechanism of miR‐182 in AKI still need to be elucidated. The levels of serum creatinine (SCr), blood urea nitrogen (BUN), and urine Kim‐1 in I/R‐induced rat AKI model were detected by a Beckman Autoanalyzer. miR‐182 and transcription factor 7‐like‐2 (TCF7L2) mRNA expression were measured by qRT‐PCR. Flow cytometry and caspase‐3 colorimetry analysis were performed to determine NRK‐52E cell apoptosis. Bioinformatics and dual‐luciferase reporter were used to identify the interaction between miR‐182 and TCF7L2. miR‐182 expression was increased in both I/R‐induced rat models and hypoxia‐treated NRK‐52E cells, and miR‐182 overexpression stimulated the apoptosis of hypoxia‐induced NRK‐52E cells. Dual‐luciferase analysis disclosed that TCF7L2 was a target of miR‐182. TCF7L2 suppressed hypoxia‐induced apoptosis in NRK‐52E cells, and the inhibitory effect of TCF7L2 on cell apoptosis could be reversed with miR‐182 restoration. Moreover, the activity of Wnt/&bgr;‐catenin signaling pathway was promoted following overexpression of TCF7L2 in NRK‐52E cells with hypoxia treatment, and this effect was greatly attenuated by the increased miR‐182 expression. Finally, in vivo experiment also validated the alleviation of miR‐182 inhibitor on I/R‐induced kidney injury and apoptosis via regulating TCF7L2/ Wnt/&bgr;‐catenin pathway. miR‐182 exacerbated AKI involving the targeting and regulation of TCF7L2/Wnt/&bgr;‐catenin signaling, unveiling a novel regulatory pathway in ischemia‐reperfusion injury and elucidating a potential biomarker for AKI treatment.

miR-133b and miR-199b knockdown attenuate TGF-β1-induced epithelial to mesenchymal transition and renal fibrosis by targeting SIRT1 in diabetic nephropathy

ABSTRACT Transforming growth factor‐&bgr;1 (TGF‐&bgr;1)‐induced epithelial to mesenchymal transition (EMT) and renal fibrosis plays critical role in the development and progression of diabetic nephropathy (DN). Our study aimed to determine the detailed roles of miR‐133b & miR‐199b on TGF‐&bgr;1‐induced EMT & renal fibrosis in DN and its underlying mechanism. The expressions of miR‐133b & miR‐199b in OLETF rats, LETO rats & TGF‐&bgr;1‐treated human proximal tubule cell line (HK‐2) were examined by qRT‐PCR. Inhibition of miR‐133b or miR‐199b was realized in cells by transfection of lentivirus containing miR‐133b inhibit or miR‐199b inhibitor. The expression levels of collagen I (COL I), fibronectin (FN), &agr;‐smooth muscle actin (&agr;‐SMA), E‐cadherin & sirtuin 1 (SIRT1) were detected by western blot and immunohistochemistry. Masson staining was conducted to estimate the degree of renal fibrosis. The interaction between SIRT1 and miR‐133b, miR‐199b was explored by luciferase reporter assay and RNA immunoprecipitation (RIP) assay. miR‐133b and miR‐199b were highly expressed in the renal cortex of diabetic OLETF rats and TGF‐&bgr;1‐treated HK‐2 cells. EMT and renal fibrosis were induced in diabetic OLETF rats and TGF‐&bgr;1‐treated HK‐2 cells. Inhibition of miR‐133b and miR‐199b attenuated EMT and renal fibrosis in diabetic OLETF rats and TGF‐&bgr;1‐treated HK‐2 cells. In addition, SIRT1 was identified as a target of miR‐133b & miR‐199b in HK‐2 cells. SIRT1 knockdown dramatically reversed the suppression on TGF‐&bgr;1‐induced EMT and renal fibrosis in HK‐2 cells mediated by anti‐miR‐133b or anti‐miR‐199. Inhibition of miR‐133b & miR‐199b attenuated TGF‐&bgr;1‐induced EMT & renal fibrosis by upregulating SIRT1 shows that using different miRNAs is a potential strategy for the future treatment of DN.

Silencing of PAQR3 suppresses extracellular matrix accumulation in high glucose-stimulated human glomerular mesangial cells via PI3K/AKT signaling pathway

ABSTRACT Progestin and AdipoQ Receptor 3 (PAQR3), a member of the PAQR family, was involved in multiple biological processes, including tumorigenesis, cholesterol homeostasis, autophagy, obesity, insulin sensitivity and energy metabolism. However, the role of PAQR3 in diabetic nephropathy is still unclear. Therefore, in this study, we investigated the effects of PAQR3 on cell proliferation and extracellular matrix (ECM) accumulation in human glomerular mesangial cells (MCs) cultured under high glucose (HG), and explored the underlying mechanism. Our results demonstrated that HG significantly up‐regulated the expression of PAQR3 in human MCs. In addition, knockdown of PAQR3 efficiently suppressed MC proliferation and ECM production in HG‐stimulated MCs. Furthermore, knockdown of PAQR3 markedly reversed HG‐induced PI3K/AKT activation in MCs. In summary, our present study demonstrated that knockdown of PAQR3 suppressed HG‐induced the proliferation and ECM accumulation in human MCs, via inhibiting the PI3K/AKT signaling pathway. Thus, PAQR3 may be a potential therapeutic target for the treatment of diabetic nephropathy.

Knockdown of NLRC5 inhibits renal fibroblast activation via modulating TGF-β1/Smad signaling pathway

&NA; NLRC5, the largest member of the Nucleotide‐binding domain and leucine‐rich repeat (NLR) protein family, is recently proven to be a critical modulator in fibrogenesis. However, the role of NLRC5 in renal fibrosis remains unknown. In the present study, we investigated the effects of NLRC5 on transforming growth factor &bgr;1 (TGF‐&bgr;1)‐stimulated rat renal fibroblasts in vitro. Our results showed that the expression of NLRC5 was also obviously upregulated in renal fibrosis tissues and TGF‐&bgr;1‐treated NRK‐49F cells. Knockdown of NLRC5 inhibited the proliferation of NRK‐49F cells induced by TGF‐&bgr;1, as well as suppressed the accumulation of extracellular matrix (ECM) in NRK‐49F cells induced by TGF‐&bgr;1. Furthermore, knockdown of NLRC5 inhibited the expression of phosphorylated Smad3 in TGF‐&bgr;1‐treated NRK‐49F cells. In conclusion, our results show that knockdown of NLRC5 inhibits renal fibroblast activation via modulating TGF‐&bgr;1/Smad signaling pathway. Therefore, NLRC5 may act as a key mediator in renal fibroblast activation and fibrogenesis.

Artesunate ameliorates high glucose-induced rat glomerular mesangial cell injury by suppressing the TLR4/NF-κB/NLRP3 inflammasome pathway

Inflammatory response is important for the development and progression of diabetic nephropathy (DN). Artesunate (ART), an antimalarial drug, possesses anti-inflammatory effect and exhibits protective effect on chronic kidney diseases. However, the effect of ART on DN is unknown. The aim of the present study was to evaluate the effect and the molecular mechanism of ART on DN in an in vitro model. The rat mesangial cell line, HBZY-1, was induced by high glucose (HG; 30u202fmM d-glucose) in the presence or absence of ART (15 and 30u202fμg/ml) and incubated for 24u202fh. We found that HG induced the proliferation of HBZY-1u202fcells, while treatment with ART inhibited the cell proliferation. Treatment with ART inhibited HG-induced inflammatory cytokines production and expression of extracellular matrix (ECM). Besides, HG induced reactive oxygen species (ROS) and malondialdehyde (MDA) levels, and inhibited the superoxide dismutase (SOD) activity of HBZY-1u202fcells, and the effects were attenuated by ART treatment. ART decreased HG-induced the expression levels of toll-like receptor 4 (TLR4), myeloid differentiation primary response gene 88 (MyD88), nuclear factor κB (NF-κB) p-p65, and nod-like receptor protein 3 (NLRP3). Inhibition of the TLR4/NF-κB pathway suppressed NLRP3 inflammasome in HBZY-1u202fcells. In conclusion, ART exhibited protective effect on HG-induced HBZY-1u202fcells by inhibiting the inflammatory response, oxidative stress and ECM accumulation. The TLR4/NF-κB/NLRP3 inflammasome pathway was involved in the protective effect of ART. The results suggested that ART might be a potential therapy agent for the DN treatment.

CTRP6 inhibits cell proliferation and ECM expression in rat mesangial cells cultured under TGF-β1

C1q/tumor necrosis factor-related protein 6 (CTRP6), a member of CTRPs family, was involved in fibrosis. However, the biological function of CTRP6 in renal fibrosis remains elusive. This study aimed to examine the role of CTRP6 in renal fibrosis and explore the possible mechanism. Our results demonstrated that the expression of CTRP6 was significantly downregulated in renal fibrotic tissues and TGF-β1-treated NRK-49F cells. In addition, overexpression of CTRP6 inhibited the proliferation, migration, and ECM expression in TGF-β1-treated NRK-49F cells. Furthermore, overexpression of CTRP6 attenuated TGF-β-induced phosphorylation of ERK1/2 in NRK-49F cells. The ERK1/2 inhibitor U0126 enhanced the inhibitory effects of CTRP6 overexpression on cell proliferation, migration and ECM expression in TGF-β1-stimulated NRK-49F cells. In conclusion, we have demonstrated that CTRP6 suppressed ECM expression in renal fibroblasts induced by TGF-β1 through the ERK signaling pathway. Therefore, CTRP6 may be a potential therapeutic target for the treatment of renal fibrosis.

Study of the Correlation between the Level of CRP and Chemerin of Serum and the Occurrence and Development of DN

Abstract The aim of the study was to find the correlation between CRP and chemerin in development of DN. We choose 90 type-2 diabetic patients between February 2010 and February 2013, who were then divided into DN group and healthy control group. The results of BP showed that there is no difference in SBP and DBP of patients in the three groups. HDL-C of patients in diabetic group and DN group is lower compared with control. CRP in diabetic group and DN group is higher than that of patients in control group. Comparing the patients in DN group with that in diabeteic group, CRP was significantly higher. Chemerin level in the diabetic group and DN group is higher than control group. When comparing the patients in DN group with those in diabeteic group, serum level of chemerin was significantly higher. Serum level of chemerin is negatively correlated with HDL-C and positively correlated with FPG, HbA1c, LDL-C, BUN and Scr. Serum CRP is negatively correlated with HDL-C and positively correlated with FPG, HbA1c, LDL-C, BUN and Scr. Serum level of chemerin is positively correlated with CRP (r=0.701, P<0.05). CRP and chemerin of the DN patients rose significantly, and may participate in the occurrence and development of DN.