State Key Laboratory Cultivation Base, Shandong Provincial Key Laboratory of Ophthalmology, Shandong Eye Institute, Shandong Academy of Medical Sciences, Qingdao, China.
Mol Pharmacol. 2013 Mar;83(3):671-80. doi: 10.1124/mol.112.081059. Epub 2013 Jan 2.
Trichostatin A (TSA) has been shown to prevent fibrosis in vitro and in vivo. The present study aimed at investigating the role of reactive oxygen species (ROS) scavenging by TSA on transforming growth factor-β (TGF-β)-induced myofibroblast differentiation of corneal fibroblasts in vitro. Human immortalized corneal fibroblasts were treated with TGF-β in the presence of TSA, the NAD(P)H oxidase inhibitor diphenyleneiodonium (DPI), the antioxidant N-acetyl-cysteine (NAC), the NF-E2-related factor 2-antioxidant response element (Nrf2-ARE) activator sulforaphane, or small interfering RNA. Myofibroblast differentiation was assessed by α-smooth muscle actin (α-SMA) expression, F-actin bundle formation, and collagen gel contraction. ROS, H(2)O(2), intracellular glutathione (GSH) level, cellular total antioxidant capacity, and the activation of Nrf2-ARE signaling were determined with various assays. Treatment with TSA and the Nrf2-ARE activator resulted in increased inhibition of the TGF-β-induced myofibroblast differentiation as compared with treatment with DPI or NAC. Furthermore, TSA also decreased cellular ROS and H(2)O(2) accumulation induced by TGF-β, whereas it elevated intracellular GSH level and cellular total antioxidant capacity. In addition, TSA induced Nrf2 nuclear translocation and up-regulated the expression of Nrf2-ARE downstream antioxidant genes, whereas Nrf2 knockdown by RNA interference blocked the inhibition of TSA on myofibroblast differentiation. In conclusion, this study provides the first evidence implicating that TSA inhibits TGF-β-induced ROS accumulation and myofibroblast differentiation via enhanced Nrf2-ARE signaling.
曲古抑菌素 A（TSA）已被证明可在体外和体内预防纤维化。本研究旨在探讨 TSA 通过清除活性氧（ROS）在体外抑制转化生长因子-β（TGF-β）诱导的角膜成纤维细胞肌成纤维细胞分化中的作用。用人永生化角膜成纤维细胞在 TSA、NAD（P）H 氧化酶抑制剂二苯基碘（DPI）、抗氧化剂 N-乙酰半胱氨酸（NAC）、核因子 E2 相关因子 2-抗氧化反应元件（Nrf2-ARE）激活剂萝卜硫素或小干扰 RNA 的存在下用 TGF-β处理。通过α-平滑肌肌动蛋白（α-SMA）表达、F-肌动蛋白束形成和胶原凝胶收缩来评估肌成纤维细胞分化。用各种测定法测定 ROS、H2O2、细胞内谷胱甘肽（GSH）水平、细胞总抗氧化能力和 Nrf2-ARE 信号转导的激活。与 DPI 或 NAC 处理相比，用 TSA 和 Nrf2-ARE 激活剂处理导致 TGF-β诱导的肌成纤维细胞分化的抑制增加。此外，TSA 还降低了 TGF-β诱导的细胞 ROS 和 H2O2 积累，而增加了细胞内 GSH 水平和细胞总抗氧化能力。此外，TSA 诱导 Nrf2 核易位并上调 Nrf2-ARE 下游抗氧化基因的表达，而 RNA 干扰的 Nrf2 敲低阻断了 TSA 对肌成纤维细胞分化的抑制。总之，本研究首次证明 TSA 通过增强 Nrf2-ARE 信号抑制 TGF-β诱导的 ROS 积累和肌成纤维细胞分化。
