Department of Ophthalmology, Shanghai General Hospital (Shanghai First People's Hospital), Shanghai Jiao Tong University, School of Medicine, Shanghai, China.
National Clinical Research Center for Eye Diseases; Shanghai Key Laboratory of Ocular Fundus Diseases; Shanghai Engineering Center for Visual Science and Photomedicine; Shanghai Engineering Center for Precise Diagnosis and Treatment of Eye Diseases, Shanghai, China.
Exp Mol Med. 2022 May;54(5):673-684. doi: 10.1038/s12276-022-00778-0. Epub 2022 May 27.
Subretinal fibrosis remains a major obstacle to the management of neovascular age-related macular degeneration. Choroidal pericytes were found to be a significant source of subretinal fibrosis, but the underlying mechanisms of pericyte-myofibroblast transition (PMT) remain largely unknown. The goal of this study was to explore the role and potential mechanisms by which PMT contributes to subretinal fibrosis. Choroidal neovascularization (CNV) was induced by laser photocoagulation in transgenic mice with the collagen1α1-green fluorescent protein (Col1α1-GFP) reporter, and recombinant adeno-associated virus 2 (rAAV2)-mediated TGF-β2 (rAAV2-TGF-β2) was administered intravitreally to further induce PMT. Primary mouse choroidal GFP-positive pericytes were treated with TGF-β2 in combination with siRNAs targeting Smad2/3, the Akt inhibitor MK2206 or the mTOR inhibitor rapamycin to examine cell proliferation, migration, and differentiation into myofibroblasts. The involvement of the Akt/mTOR pathway in PMT in subretinal fibrosis was further investigated in vivo. Intraocular TGF-β2 overexpression induced GFP-positive pericyte infiltration and PMT in subretinal fibrosis, which was mimicked in vitro. Knockdown of Smad2/3 or inhibition of Akt/mTOR decreased cell proliferation, PMT and migration in primary mouse pericytes. Combined inhibition of Smad2/3 and mTOR showed synergistic effects on attenuating α-smooth muscle actin (α-SMA) expression and cell proliferation. In mice with laser-induced CNV, the administration of the Akt/mTOR inhibitors suppressed pericyte proliferation and alleviated the severity of subretinal fibrosis. Our results showed that PMT plays a pivotal role in subretinal fibrosis, which was induced by TGF-β2 through the Smad2/3 and Akt/mTOR pathways. Thus, inhibiting PMT may be a novel strategy for the treatment of subretinal fibrosis.
视网膜下纤维化仍然是新生血管性年龄相关性黄斑变性治疗的主要障碍。脉络膜周细胞被发现是视网膜下纤维化的重要来源，但周细胞-肌成纤维细胞转化（PMT）的潜在机制仍知之甚少。本研究旨在探讨 PMT 促进视网膜下纤维化的作用和潜在机制。通过激光光凝在 Col1α1-GFP 报告转基因小鼠中诱导脉络膜新生血管（CNV），并通过重组腺相关病毒 2（rAAV2）介导的 TGF-β2（rAAV2-TGF-β2）玻璃体腔给药进一步诱导 PMT。用 TGF-β2 联合靶向 Smad2/3 的 siRNA、Akt 抑制剂 MK2206 或 mTOR 抑制剂雷帕霉素处理原代小鼠脉络膜 GFP 阳性周细胞，以检测细胞增殖、迁移和分化为肌成纤维细胞。进一步在体内研究了 Akt/mTOR 通路在视网膜下纤维化 PMT 中的作用。眼内 TGF-β2 过表达诱导 GFP 阳性周细胞浸润和视网膜下纤维化，在体外也得到了模拟。Smad2/3 敲低或 Akt/mTOR 抑制降低了原代小鼠周细胞的增殖、PMT 和迁移。Smad2/3 和 mTOR 的联合抑制对降低α-平滑肌肌动蛋白（α-SMA）表达和细胞增殖显示出协同作用。在激光诱导的 CNV 小鼠中，Akt/mTOR 抑制剂的给药抑制了周细胞增殖，并减轻了视网膜下纤维化的严重程度。我们的结果表明，PMT 通过 TGF-β2 诱导的 Smad2/3 和 Akt/mTOR 通路在视网膜下纤维化中起关键作用。因此，抑制 PMT 可能是治疗视网膜下纤维化的一种新策略。
