IKKβ Inhibition Attenuates Epithelial Mesenchymal Transition of Human Stem Cell-Derived Retinal Pigment Epithelium. IKKβ Inhibition Attenuates Epithelial Mesenchymal Transition of Human Stem Cell-Derived Retinal Pigment Epithelium

Epithelial-mesenchymal transition (EMT), which is well known for its role in embryonic development, malignant transformation, and tumor progression, has also been implicated in a variety of retinal diseases, including proliferative vitreoretinopathy (PVR), age-related macular degeneration (AMD), and diabetic retinopathy. EMT of the retinal pigment epithelium (RPE), although important in the pathogenesis of these retinal conditions, is not well understood at the molecular level. We and others have shown that a variety of molecules, including the co-treatment of human stem cell-derived RPE monolayer cultures with transforming growth factor beta (TGF–β) and the inflammatory cytokine tumor necrosis factor alpha (TNF–α), can induce RPE–EMT; however, small molecule inhibitors of RPE–EMT have been less well studied. Here, we demonstrate that BAY651942, a small molecule inhibitor of nuclear factor kapa-B kinase subunit beta (IKKβ) that selectively targets NF-κB signaling, can modulate TGF–β/TNF–α-induced RPE–EMT. Next, we performed RNA-seq studies on BAY651942 treated hRPE monolayers to dissect altered biological pathways and signaling events. Further, we validated the effect of IKKβ inhibition on RPE–EMT-associated factors using a second IKKβ inhibitor, BMS345541, with RPE monolayers derived from an independent stem cell line. Our data highlights the fact that pharmacological inhibition of RPE–EMT restores RPE identity and may provide a promising approach for treating retinal diseases that involve RPE dedifferentiation and EMT. Overall design: We compared the transcriptomic profiles of human stem cell-derived RPE monolayer cultures co-treated with TGF–β/TNF–α and a small molecule (BAY651942) inhibitor of nuclear factor kapa-B kinase subunit beta (IKKβ)

上皮间质转化（Epithelial-mesenchymal transition, EMT）以其在胚胎发育、恶性转化及肿瘤进展中的经典作用广为人知，其亦被证实参与多种视网膜疾病的发生发展，包括增生性玻璃体视网膜病变（proliferative vitreoretinopathy, PVR）、年龄相关性黄斑变性（age-related macular degeneration, AMD）与糖尿病视网膜病变。尽管视网膜色素上皮（retinal pigment epithelium, RPE）的EMT在上述视网膜疾病的发病机制中发挥关键作用，但其分子层面的调控机制仍未被充分阐明。本团队及其他研究团队均已证实，多种分子可诱导RPE-EMT，例如联合应用转化生长因子β（transforming growth factor beta, TGF-β）与炎性细胞因子肿瘤坏死因子α（tumor necrosis factor alpha, TNF-α）处理人干细胞来源的RPE单层培养物；然而针对RPE-EMT的小分子抑制剂的相关研究仍较为匮乏。本研究证实，BAY651942作为一种靶向核因子κB激酶亚基β（nuclear factor kapa-B kinase subunit beta, IKKβ）的小分子抑制剂，可特异性调控NF-κB信号通路，能够调节TGF-β/TNF-α诱导的RPE-EMT。随后，我们对经BAY651942处理的人视网膜色素上皮（human retinal pigment epithelium, hRPE）单层细胞进行了RNA测序（RNA-seq）分析，以解析其发生改变的生物学通路与信号事件。进一步地，我们使用第二种IKKβ抑制剂BMS345541，并采用来自独立干细胞系的RPE单层细胞，验证了IKKβ抑制对RPE-EMT相关因子的作用效果。本研究数据表明，对RPE-EMT进行药理学抑制可恢复RPE细胞的固有特性，或可为涉及RPE去分化与EMT的视网膜疾病提供极具前景的治疗策略。总体实验设计：我们比较了经TGF-β/TNF-α与核因子κB激酶亚基β（IKKβ）小分子抑制剂BAY651942联合处理的人干细胞来源RPE单层培养物的转录组特征。