PDF(Pubmed)
Abstract：
Objective: The level of tumor necrosis factor-α (TNF-α) is upregulated during the development of pulmonary vascular remodeling and pulmonary hypertension. A hallmark of pulmonary arterial (PA) remodeling is the excessive proliferation of PA smooth muscle cells (PASMCs). The purpose of this study is to investigate whether TNF-α induces PASMC proliferation and explore the potential mechanisms. Methods: PASMCs were isolated from 8-week-old male Sprague-Dawley rats and treated with 0, 20, or 200 ng/mL TNF-α for 24 or 48 h. After treatment, cell number, superoxide production, histone acetylation, DNA methylation, and histone methylation were assessed. Results: TNF-α treatment increased NADPH oxidase activity, superoxide production, and cell numbers compared to untreated controls. TNF-α-induced PASMC proliferation was rescued by a superoxide dismutase mimetic tempol. TNF-α treatment did not affect histone acetylation at either dose but did significantly decrease DNA methylation. DNA methyltransferase 1 activity was unchanged by TNF-α treatment. Further investigation using QRT-RT-PCR revealed that GADD45-α, a potential mediator of DNA demethylation, was increased after TNF-α treatment. RNAi inhibition of GADD45-α alone increased DNA methylation. TNF-α impaired the epigenetic mechanism leading to DNA hypomethylation, which can be abolished by a superoxide scavenger tempol. TNF-α treatment also decreased H3-K4 methylation. TNF-α-induced PASMC proliferation may involve the H3-K4 demethylase enzyme, lysine-specific demethylase 1 (LSD1). Conclusions: TNF-α-induced PASMC proliferation may be partly associated with excessive superoxide formation and histone and DNA methylation.
摘要：
目的:肿瘤坏死因子-α(TNF-α)水平在肺血管重构和肺动脉高压的发生发展过程中上调。肺动脉(PA)重塑的标志是PA平滑肌细胞(PASMC)的过度增殖。本研究的目的是研究TNF-α是否诱导PASMC增殖并探讨其可能的机制。方法：从8周龄雄性Sprague-Dawley大鼠中分离PASMC，并用0、20或200ng/mLTNF-α治疗24或48h。治疗后，单元格编号,超氧化物生产,组蛋白乙酰化，DNA甲基化，和组蛋白甲基化进行评估。结果:TNF-α处理增加了NADPH氧化酶的活性,超氧化物生产,和与未处理对照相比的细胞数量。超氧化物歧化酶模拟tempol挽救了TNF-α诱导的PASMC增殖。TNF-α治疗在任一剂量下都不影响组蛋白乙酰化，但确实显着降低了DNA甲基化。通过TNF-α处理,DNA甲基转移酶1活性不变。使用QRT-RT-PCR的进一步调查显示GADD45-α，DNA去甲基化的潜在介质,TNF-α治疗后增加。单独对GADD45-α的RNAi抑制增加了DNA甲基化。TNF-α损害了导致DNA低甲基化的表观遗传机制，它可以被超氧化物清除剂tempol废除。TNF-α处理也降低了H3-K4甲基化。TNF-α诱导的PASMC增殖可能涉及H3-K4去甲基酶,赖氨酸特异性去甲基酶1（LSD1）。结论:TNF-α诱导的PASMC增殖可能与超氧化物过度形成、组蛋白和DNA甲基化有关。
