TGR5 suppresses high glucose-induced upregulation of fibronectin and transforming growth factor-β1 in rat glomerular mesangial cells by inhibiting RhoA/ROCK signaling
TGR5 suppresses high glucose-induced upregulation of fibronectin and transforming growth factor-β1 in rat glomerular mesangial cells by inhibiting RhoA/ROCK signaling
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TGR5 通过抑制 RhoA/ROCK 信号传导来抑制高葡萄糖诱导的大鼠肾小球系膜细胞中纤连蛋白和转化生长因子-β1 的上调
DOI：
10.1007/s12020-016-1032-4
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发表时间：
2016-12-01
期刊：
影响因子：
3.7
通讯作者：
Huang, Heqing
中科院分区：
文献类型：
--
作者：
Xiong, Fengxiao;Li, Xuejuan;Huang, Heqing
RhoA/ROCK可通过激活核因子-κ B（NF-κ B）在糖尿病背景下引起肾脏炎症和纤维化。已知TGR 5在维持代谢稳态和抗炎中的作用，其与NF-κ B抑制密切相关。鉴于TGR 5在肾脏中高度富集，我们的目的是研究TGR 5对高糖（HG）处理的大鼠肾小球系膜细胞（GMC）中纤维连接蛋白（FN）和转化生长因子-β 1（TGF-β 1）的调节作用。这两种因子均与肾脏炎症密切相关，并由NF-κ B B介导。此外，我们的研究确定这种调节是否通过抑制RhoA/ROCK和随后的NF-κ B抑制来实现。采用聚合酶链反应检测TGR 5 mRNA水平。Western blot检测TGF-5、FN、TGF-β 1、p65、I κ B α、磷酸化MYPT 1（Thr 853）和MYPT 1的蛋白表达。GlucoS-transferase-pull down和免疫荧光分别检测RhoA的活化、TGR 5和p65的分布。采用电泳迁移率变动法测定NF-κ B的DNA结合活性。在GMCs中，TGR 5激活或过表达显著抑制HG或转染组成型活性RhoA诱导的FN和TGF-β 1蛋白表达、NF-κ B和RhoA/ROCK激活。TGF-β 1、FN蛋白表达增强，RhoA/ROCK激活增强。然而，当使用选择性蛋白激酶A（PKA）抑制剂时，TGR 5不能抑制RhoA/ROCK活化。这项研究表明，在HG处理的GMC中，TGR 5显著抑制NF-κ B介导的FN和TGF-β 1的上调，这是糖尿病肾病的标志。这些功能与通过PKA抑制RhoA/ROCK密切相关。
RhoA/ROCK can cause renal inflammation and fibrosis in the context of diabetes by activating nuclear factor-kappa B (NF-kappa B). TGR5 is known for its role in maintaining metabolic homeostasis and anti-inflammation, which is closely related to NF-kappa B inhibition. Given that TGR5 is highly enriched in kidney, we aim to investigate the regulatory role of TGR5 on fibronectin (FN) and transforming growth factor-beta 1 (TGF-beta 1) in high glucose (HG)-treated rat glomerular mesangial cells (GMCs). Both the factors are closely related to renal inflammations and mediated by NF-kappa B. Moreover, our study determines whether such regulation is achieved by the inhibition of RhoA/ROCK and the subsequent NF-kappa B suppression. Polymerase chain reaction was taken to test the mRNA level of TGR5. Western blot was used to measure the protein expressions of TGR5, FN, TGF-beta 1, p65, I kappa B alpha, phospho-MYPT1 (Thr853), and MYPT1. Glutathione S-transferase-pull down and immunofluorescence were conducted to test the activation of RhoA, the distribution of TGR5, and p65, respectively. Electrophoretic mobility shift assay was adopted to measure the DNA binding activity of NF-kappa B. In GMCs, TGR5 activation or overexpression significantly suppressed FN and TGF-beta 1 protein expressions, NF-kappa B, and RhoA/ROCK activation induced by HG or transfection of constitutively active RhoA. By contrast, TGR5 RNA interference caused enhancement of FN, TGF-beta 1 protein expressions, increase of RhoA/ROCK activation. However, TGR5 cannot suppress RhoA/ROCK activation when a selective Protein kinase A (PKA) inhibitor was used. This study suggests that in HG-treated GMCs, TGR5 significantly suppresses the NF-kappa B-mediated upregulation of FN and TGF-beta 1, which are hallmarks of diabetic nephropathy. These functions are closely related to the suppression of RhoA/ROCK via PKA.