沉默调节蛋白1通过Smads系统促进小梁网细胞迁移并减少转化生长因子-β诱导的细胞外基质积累。

Sirtuin1 Promotes the Migration of Cells and Reduces the Accumulation of Extracellular Matrix in Trabecular Meshwork Cells Induced by Transform Growth Factor-β via Smads System.

作者信息

Tie Jinjun, Guo Junhong, Yuan Jiemei, Wang Jiantao

机构信息

Department of Ophthalmology, The Affiliated Hospital of Guizhou Medical University, 550000 Guiyang, Guizhou, China.

Shenzhen Eye Hospital, Jinan University, Shenzhen Eye Institute, 518040 Shenzhen, Guangdong, China.

出版信息

Discov Med. 2025 Apr;37(195):685-694. doi: 10.24976/Discov.Med.202537195.59.

DOI:10.24976/Discov.Med.202537195.59

PMID:40287804

Abstract

BACKGROUND

Primary open-angle glaucoma (POAG) is one of the common types of glaucoma, an eye disease that causes irreversible blindness. Fibrosis of the trabecular meshwork (TM) caused by the accumulation of extracellular matrix (ECM) induced by transform growth factor-β (TGF-β) is closely related to high intraocular pressure (IOP). Deacetylase Sirtuin1 (Sirt1) plays an anti-oxidation and anti-fibrosis role in many diseases, including glaucoma; however, its mechanisms have not been fully revealed. In this study, we analyzed the anti-fibrotic role of Sirt1 in TM fibrosis induced by TGF-β to investigate potential mechanisms.

METHODS

Transcriptome sequencing of trabecular meshwork cells (TMCs) was performed after transfection with the adenovirus-Sirt1-green fluorescent protein (Adv-Sirt1-GFP). Then, 5 ng/mL TGF-β was used to induce overexpression of ECM in TMCs . The expression of target proteins was detected by Western blot and immunofluorescence, and cytokine expression was detected by enzyme-linked immunosorbent serologic assay (ELISA). At the same time, we detected the functional changes in cell proliferation, adhesion, and migration.

RESULTS

After treatment with TGF-β, we found that the accumulation of ECM was increased (fibronectin (FN), collagen I (COL I), laminin (LN), < 0.05), and the phosphorylation (activation) of Smad2/3 and the expression of Smad4 were increased ( < 0.001). The results of transcriptome sequencing suggested that Sirt1 inhibits the expression of ECM by regulating the functions of co-Smad and co-COL binding proteins, thus participating in the regulation of cell adhesion. Finally, we confirmed that: (1) Sirt1 reduced the accumulation of ECM in TMCs by inhibiting the phosphorylation of Smad2/3 ( < 0.05) and the expression of Smad4 ( < 0.05), and (2) Sirt1 decreased the adhesive ability of TMCs by reducing the secretion of integrins (integrin-α3 (ITGα3), < 0.01; integrin-β1 (ITGβ1), < 0.001) and cadherins (E-cadherin, < 0.01; N-cadherin, < 0.01), and promoted cell migration ( < 0.05).

CONCLUSION

Sirt1 promotes the migration of cells and reduces the accumulation of ECM in TMCs induced by TGF-β by inhibiting the activation of Smad2/3 and the expression of Smad4.

摘要

背景

原发性开角型青光眼（POAG）是青光眼的常见类型之一，是一种可导致不可逆性失明的眼部疾病。转化生长因子-β（TGF-β）诱导的细胞外基质（ECM）积累所引起的小梁网（TM）纤维化与高眼压密切相关。去乙酰化酶沉默调节蛋白1（Sirt1）在包括青光眼在内的多种疾病中发挥抗氧化和抗纤维化作用；然而，其机制尚未完全阐明。在本研究中，我们分析了Sirt1在TGF-β诱导的TM纤维化中的抗纤维化作用，以探究潜在机制。

方法

用腺病毒-Sirt1-绿色荧光蛋白（Adv-Sirt1-GFP）转染小梁网细胞（TMCs）后进行转录组测序。然后，用5 ng/mL TGF-β诱导TMCs中ECM的过表达。通过蛋白质免疫印迹法和免疫荧光检测靶蛋白的表达，通过酶联免疫吸附测定（ELISA）检测细胞因子表达。同时，我们检测了细胞增殖、黏附及迁移方面的功能变化。

结果

用TGF-β处理后，我们发现ECM的积累增加（纤连蛋白（FN）、I型胶原（COL I）、层粘连蛋白（LN），<0.05），并且Smad2/3的磷酸化（激活）及Smad4的表达增加（<0.001）。转录组测序结果表明，Sirt1通过调节共-Smad和共-COL结合蛋白的功能抑制ECM的表达，从而参与细胞黏附的调节。最后，我们证实：（1）Sirt1通过抑制Smad2/3的磷酸化（<0.05）和Smad4的表达（<0.05）减少TMCs中ECM的积累，以及（2）Sirt1通过减少整合素（整合素-α3（ITGα3），<0.01；整合素-β1（ITGβ1），<0.001）和钙黏蛋白（E-钙黏蛋白，<0.01；N-钙黏蛋白，<0.01）的分泌降低TMCs的黏附能力，并促进细胞迁移（<0.05）。