Hydrogen peroxide increases extracellular matrix mRNA through TGF-β in human mesangial cells
Hydrogen peroxide increases extracellular matrix mRNA through TGF-β in human mesangial cells
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DOI：
10.1046/j.1523-1755.2001.00469.x
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发表时间：
2001-01-01
影响因子：
19.6
通讯作者：
Rodríguez-Puyol, D
中科院分区：
文献类型：
--
作者：
Iglesias-de la Cruz, MC;Ruiz-Torres, P;Rodríguez-Puyol, D
背景资料。在病理状态下，包括许多肾脏疾病在内的许多疾病中，都会产生过量的ROS。转化生长因子-β可能介导肾纤维化，ROS可能通过转化生长因子-β途径发挥促纤维化作用。用葡萄糖氧化酶(GO)孵育培养的人肾小球系膜细胞(HMCs)，检测转化生长因子-β1和细胞外基质(ECM)成分的表达。加入中和抗转化生长因子-β抗体，以检验过氧化氢通过激活转化生长因子-β途径刺激细胞外基质表达的假设。Northern印迹分析显示，转化生长因子-β1和细胞外基质蛋白(I型、III型和IV型胶原，以及纤维连接蛋白)的稳态水平显著增加约两倍。虽然GO处理后对mRNA的稳定性没有显著影响，但其他应用启动子-报告实验、竞争-定量逆转录-聚合酶链式反应、水貂肺上皮细胞增殖实验和转化生长因子-β1酶联免疫吸附试验的研究都表明，GO(>；1.5倍)分别显著刺激了转化生长因子-β1的启动子活性、mRNA水平、生物活性和蛋白产生。过氧化氢酶预处理可抑制GO诱导的转化生长因子-β1基因的表达。当与全选择性中和抗转化生长因子-β抗体孵育时，可防止GO刺激的ECM分子的表达。GO诱导HMCs产生过氧化氢可诱导转化生长因子-β1的合成，从而增加ECM基因的表达。这些细胞反应可能是以氧化应激为特征的肾脏疾病的发生和发展的基础。
Background. Reactive oxygen species (ROS) are excessively produced in pathologic states, including many renal diseases. Transforming growth factor-beta (TGF-beta) may mediate renal fibrotic injury, and ROS may act through the TGF-beta pathway to exert a profibrotic effect.Methods. The expression of TGF-beta1 and extracellular matrix (ECM) components were assessed in cultured human mesangial cells (HMCs) incubated with glucose oxidase (GO), an enzyme that continuously generates hydrogen peroxide from glucose. A neutralizing anti-TGF-beta antibody was added to test the hypothesis that hydrogen peroxide acts through activation of the TGF-beta pathway to stimulate ECM expression.Results. Northern blot analysis revealed significantly increased steady-state levels of TGF-beta1 and ECM proteins (collagen types I, III, and IV, and fibronectin) by approximately twofold. While no significant effect on mRNA stability after treatment with GO was observed, other studies employing promoter-reporter assays, competitive-quantitative reverse transcription-polymerase chain reaction, mink lung epithelial cell proliferation assay, and TGF-beta1 enzyme-linked immunosorbent assay all demonstrated significant stimulation by GO (>1.5-fold) of TGF-beta1 promoter activity, mRNA level, bioactivity, and protein production, respectively. Catalase pretreatment prevented the GO-induced stimulation of TGF-beta1 mRNA. When incubations were performed with a panselective neutralizing anti-TGF-beta antibody, the GO-stimulated expression of ECM molecules was prevented.Conclusions. GO-induced hydrogen peroxide production induces TGF-beta1 synthesis and thereby increases ECM gene expression in cultured HMCs. These cellular responses may underlie the development and progression of renal diseases characterized by oxidative stress.