Heusinger-Ribeiro Juliane, Eberlein Michael, Wahab Nadia Abdel, Goppelt-Struebe Margarete
Medical Clinic IV, University of Erlangen-Nuremberg, Erlangen, Germany.
Imperial College School of Medicine, Molecular Pathology Section, London, United Kingdom.
J Am Soc Nephrol. 2001 Sep;12(9):1853-1861. doi: 10.1681/ASN.V1291853.
The induction of connective tissue growth factor (CTGF) was investigated in a human renal fibroblast cell line that exhibited many characteristics of primary human renal fibroblasts. Induction of CTGF mRNA was observed after treatment of the cells with transforming growth factor-beta (TGF-beta) or, even more prominently, lysophosphatidic acid (LPA). LPA induced a rapid transient increase in CTGF mRNA expression, with maximal levels being observed after 1 to 2 h. This increase was accompanied by CTGF protein synthesis. Induction of CTGF was insensitive to pertussis toxin and was not dependent on the activation of p42/44 mitogen-activated protein kinases. Inhibition of the proteins of the Rho family with toxin B from Clostridium difficile abrogated basal and LPA-mediated induction of CTGF. Specific targeting of RhoA with C3 exotoxin or of the Rho kinases with the inhibitor Y-27632 similarly prevented induction of CTGF, implicating RhoA as a signaling module downstream of LPA. Inhibition of RhoA depolymerized the actin cytoskeleton, as did treatment with cytochalasin D. Preincubation of the human renal fibroblasts with cytochalasin D prevented induction of CTGF by LPA, indicating a strong contribution of an intact cytoskeleton. Interference with RhoA signaling similarly inhibited the induction of CTGF by TGF-beta. Elevation of intracellular levels of cAMP and thus activation of protein kinase A prevented induction of CTGF expression. The cytoskeletal effects of cAMP, however, were reversed by LPA. These data indicate complex interactions involving LPA-mediated activation of RhoA- and protein kinase A-dependent signaling pathways. The data thus demonstrate the regulatory functions of the small GTPase RhoA and of an intact cytoskeleton in the expression of CTGF after stimulation with LPA or TGF-beta. Analogous signal transduction pathways were previously demonstrated in rat mesangial cells, suggesting a more general role for RhoA in the regulation of CTGF expression.
在一种具有原代人肾成纤维细胞诸多特征的人肾成纤维细胞系中，研究了结缔组织生长因子（CTGF）的诱导情况。在用转化生长因子-β（TGF-β）处理细胞后，观察到CTGF mRNA的诱导，甚至更显著的是在用溶血磷脂酸（LPA）处理后。LPA诱导CTGF mRNA表达迅速短暂增加，在1至2小时后观察到最高水平。这种增加伴随着CTGF蛋白合成。CTGF的诱导对百日咳毒素不敏感，且不依赖于p42/44丝裂原活化蛋白激酶的激活。用艰难梭菌的毒素B抑制Rho家族蛋白可消除CTGF的基础诱导以及LPA介导的诱导。用C3外毒素特异性靶向RhoA或用抑制剂Y - 27632靶向Rho激酶同样可阻止CTGF的诱导，这表明RhoA是LPA下游的信号模块。抑制RhoA会使肌动蛋白细胞骨架解聚，用细胞松弛素D处理也会如此。用人肾成纤维细胞预先用细胞松弛素D孵育可阻止LPA诱导CTGF，表明完整的细胞骨架起了重要作用。干扰RhoA信号同样抑制TGF-β诱导CTGF。细胞内cAMP水平升高从而激活蛋白激酶A可阻止CTGF表达的诱导。然而，cAMP的细胞骨架效应被LPA逆转。这些数据表明涉及LPA介导的RhoA和蛋白激酶A依赖性信号通路激活的复杂相互作用。这些数据因此证明了小GTP酶RhoA和完整细胞骨架在LPA或TGF-β刺激后CTGF表达中的调节功能。先前在大鼠系膜细胞中也证明了类似的信号转导途径，表明RhoA在CTGF表达调节中具有更普遍的作用。
