Zhang Chun, Liu Xiuxing, Gu Chenyang, Su Yuhan, Lv Jianjie, Liu Yidan, Gao Yuehan, Chen Hui, Xu Nanwei, Xiao Jing, Xu Zhuping, Su Wenru
Department of Ophthalmology, West China Hospital, Sichuan University, Chengdu, Sichuan 610041, China.
State Key Laboratory of Ophthalmology, Zhongshan Ophthalmic Center, Sun Yat-sen University, Guangdong Provincial Key Laboratory of Ophthalmology and Visual Science, Guangzhou 510060, China.
J Adv Res. 2025 Jun;72:633-652. doi: 10.1016/j.jare.2024.07.029. Epub 2024 Aug 6.
Autoimmune uveitis (AU) is a prevalent ocular autoimmune disease leading to significant visual impairment. However, underlying pathogenesis of AU required to develop more efficient therapy remain unclear.
We isolated peripheral blood mononuclear cells (PBMCs) from AU patients and performed single-cell RNA sequencing (scRNA-seq). Besides, experimental autoimmune uveitis (EAU) model was established and treated with histone deacetylase inhibitor (HDACi) Belinostat or vehicle. We extracted immune cells from Blank, EAU, and HDACi-treated EAU mice and used scRNA-seq, flow cytometry, siRNA, specific inhibitors, and adoptive transfer experiments to explore the role of HDACs and its downstream potential molecular mechanisms in the immune response of EAU and AU.
We found highly expressed histone deacetylases (HDACs) family in AU patients and identified it as a key factor related to CD4 effector T cell differentiation in the pathogenesis of AU. Our further studies showed that targeted inhibition of HDACs effectively alleviated EAU, restored its Th17/Treg balance, and reduced inflammatory gene expression, especially in CD4 T cells. Post-HDACs inhibition, Treg proportions increased with enhanced immunomodulatory effects. Importantly, HDACs exhibited a positive promoting role on Th17 cells. Based on scRNA-seq screening and application of knock-down siRNAs and specific inhibitors in vitro and vivo, we identified CDK6 as a key downstream molecule regulated by HDAC1/3/6 through acetyl-histone H3/p53/p21 axis, which is involved in Th17 pathogenicity and EAU development. Additionally, HDACs-regulated CDK6 formed a positive loop with ID2, inducing PIM1 upregulation, promoting Th17 cell differentiation and pathogenicity, and correlates with AU progression.
Based on the screening of clinical samples and downstream molecular functional validation experiments, we revealed a driving role for HDACs and the HDACs-regulated CDK6/ID2 axis in Th17 cell differentiation and pathogenicity in AU, proposing a promising therapeutic strategy.
自身免疫性葡萄膜炎（AU）是一种常见的眼部自身免疫性疾病，可导致严重的视力损害。然而，要开发更有效的治疗方法，AU的潜在发病机制仍不清楚。
我们从AU患者中分离出外周血单核细胞（PBMC）并进行单细胞RNA测序（scRNA-seq）。此外，建立实验性自身免疫性葡萄膜炎（EAU）模型，并用组蛋白去乙酰化酶抑制剂（HDACi）贝利司他或赋形剂进行治疗。我们从空白、EAU和HDACi治疗的EAU小鼠中提取免疫细胞，并使用scRNA-seq、流式细胞术、siRNA、特异性抑制剂和过继转移实验来探索HDACs及其下游潜在分子机制在EAU和AU免疫反应中的作用。
我们发现AU患者中组蛋白去乙酰化酶（HDACs）家族高度表达，并将其确定为AU发病机制中与CD4效应T细胞分化相关的关键因素。我们的进一步研究表明，靶向抑制HDACs可有效缓解EAU，恢复其Th17/Treg平衡，并降低炎症基因表达，尤其是在CD4 T细胞中。抑制HDACs后，Treg比例增加，免疫调节作用增强。重要的是，HDACs对Th17细胞具有正向促进作用。基于scRNA-seq筛选以及在体外和体内应用敲低siRNAs和特异性抑制剂，我们确定CDK6是HDAC1/3/6通过乙酰化组蛋白H3/p53/p21轴调控的关键下游分子，其参与Th17致病性和EAU发展。此外，HDACs调控的CDK6与ID2形成正反馈环，诱导PIM1上调，促进Th17细胞分化和致病性，并与AU进展相关。
基于临床样本筛选和下游分子功能验证实验，我们揭示了HDACs以及HDACs调控的CDK6/ID2轴在AU中Th17细胞分化和致病性中的驱动作用，提出了一种有前景的治疗策略。
