5-Aza-2′-deoxycytidine induces human Tenon's capsule fibroblasts differentiation and fibrosis by up-regulating TGF-β type I receptor
5-Aza-2′-deoxycytidine induces human Tenon's capsule fibroblasts differentiation and fibrosis by up-regulating TGF-β type I receptor
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DOI：
10.1016/j.exer.2017.09.004
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发表时间：
2017-12-01
影响因子：
3.4
通讯作者：
Ye, Wen
中科院分区：
文献类型：
--
作者：
Fu, Shuhao;Sun, Li;Ye, Wen
青光眼滤过手术失败率高的主要原因是术后瘢痕形成导致滤过功能丧失。我们研究了DNA甲基转移酶抑制剂5-氮杂-2 '-脱氧胞苷（5-Aza-dc）对人Tenon囊成纤维细胞（HTFs）分化和纤维化的影响及其作用机制，特别是与转化生长因子（TGF）-β 1信号传导的关系。TGF-β 1用于诱导培养的HTF的分化。5-Aza-dc抑制DNA甲基转移酶（DNMT）活性6小时后，治疗过程对应于TGF-β 1诱导的DNMT活性降低，而不影响细胞活力（通过细胞计数试剂盒-8测定）。5-Aza-dc还降低了24至48小时的DNMT 1和DNMT 3a蛋白表达。通过划痕试验评估的HTFs迁移在5-Aza-dc处理后24小时显著增加，时间过程与TGF-β 1相似。5-Aza-dc处理显著增加了α-平滑肌肌动蛋白（α-SMA）、胶原蛋白-1A1（Col 1A 1）、纤连蛋白（FN）和TGF-β I型受体（TGE β RI）的mRNA和蛋白水平。此外，5-Aza-dc对DNMT活性抑制、细胞迁移和纤维化的作用均被TGF β RI抑制剂SB-431542逆转。同时，DNMT 1的敲低上调了TGF β RI的表达，并在HTFs中具有相同的纤维化诱导作用，这也被SB-431542抑制。因此，结果表明DNA低甲基化通过上调TGEBRI诱导HTFs分化和纤维化。DNA甲基化状态在结膜下伤口愈合中起重要作用。(C)2017爱思唯尔有限公司版权所有
The principle reason of high failure rate of glaucoma filtration surgery is the loss of filtration function caused by postoperative scar formation. We investigated the effects of 5-aza-2'-deoxycytidine (5-Aza-dc), a DNA methyltransferases inhibitor, on human Tenon's capsule fibroblasts (HTFs) differentiation and fibrosis and its mechanism of action, especially in relation to transforming growth factor (TGF)-beta 1 signaling. TGF-beta 1 was used to induce differentiation of cultured HTFs. 5-Aza-dc suppressed DNA methyltransferases (DNMTs) activity 6 h after treatment with a course corresponding to that of TGF-beta 1-induced reduction of DNMT activity without affecting cell viability as measured by Cell Counting Kit-8 assay. 5-Aza-dc also reduced DNMT1 and DNMT3a protein expression from 24 to 48 h. HTFs migration evaluated by scratch-wound assay were significantly increased 24 h after 5-Aza-dc treatment, a time course similar to that of TGF-beta 1. Treatment with 5-Aza-dc significantly increased the mRNA and protein levels of alpha-smooth muscle actin (alpha-SMA), collagen-1A1 (Col1A1), fibronectin (FN) and TGF-beta type I receptor (TGE beta RI). Furthermore, the effects of 5-Aza-dc on DNMT activity suppression, cell migration, and fibrosis were all reversed by a TGF beta RI inhibitor- SB-431542. Meanwhile, knockdown of DNMT1 upregulated TGF beta RI expression and had the same fibrosis-inducing effect in HTFs, which was also inhibited by SB-431542. Thus, the results indicate that DNA hypomethylation induces HTFs differentiation and fibrosis through up-regulation of TGEBRI. DNA methylation status plays an important role in subconjunctival wound healing. (C) 2017 Elsevier Ltd. All rights reserved.