Li Jinjian, Yang Kun, Pan Xinghui, Peng Hui, Hou Chenting, Xiao Jie, Wang Qing
Ophthalmology, Affiliated Hospital of Qingdao University, Qingdao, 266500, People's Republic of China.
Medical Research Center, Affiliated Hospital of Qingdao University, Qingdao, 266500, People's Republic of China.
J Inflamm Res. 2022 Jun 2;15:3269-3283. doi: 10.2147/JIR.S361541. eCollection 2022.
The biological role and mechanism of long noncoding RNA (lncRNA) myocardial infarction-associated transcript (MIAT) in dry eye remain to be illustrated. Pyroptosis is a noticeable form of inflammatory activation, which is characteristic of gasdermin D (GSDMD)-driven cell death. The present study was designed to explore the role of MIAT in pyroptosis and apoptosis induced by hyperosmolarity stress (HS) in human corneal epithelial cells (HCECs).
HCECs were cultured in 70-120 mM hyperosmotic medium for 24 h to create a dry eye model in vitro. The level of the pyroptosis marker GSDMD was measured, and the cell inflammatory response was evaluated by detecting IL-1β and IL-18 levels. Exogenous caspase-1 inhibitor Ac-YVAD-CHO was used. The pyroptosis in HCECs was examined by caspase-1 activity, immunofluorescent staining, and Western blotting. Flow cytometry was performed to test the apoptosis rate of HCECs. Cell migration and proliferation were detected. The expression of the lncRNA MIAT in HCECs was detected by quantitative real-time PCR. MIAT was knocked down by small interfering RNA (siRNA) transfection. The effects of caspase-1 inhibition on pyroptosis, apoptosis, migration, and proliferation were observed.
HS promoted pyroptosis in HCECs by elevating caspase-1, GSDMD, and the active cleavage of GSDMD (N-terminal domain, N-GSDMD), and increased the release of IL-1β, IL-18, LDH and the rate of apoptosis, with reduced cell migration. These changes were prevented by the inhibition of caspase-1. The expression of MIAT was significantly increased in HCECs exposed to a hyperosmotic medium. Silencing MIAT increased the expression of GSDMD, caspase-1, and inflammatory chemokines IL-1β and IL-18, and promoted apoptosis while inhibiting migration and proliferation in HCECs.
The lncRNA MIAT is involved in HS-induced pyroptosis and apoptosis and the inflammatory response of HCECs and provides a new understanding of the pathogenesis of dry eye.
长链非编码RNA（lncRNA）心肌梗死相关转录本（MIAT）在干眼发病机制中的生物学作用和机制仍有待阐明。细胞焦亡是一种显著的炎症激活形式，其特征是由gasdermin D（GSDMD）驱动的细胞死亡。本研究旨在探讨MIAT在高渗应激（HS）诱导的人角膜上皮细胞（HCEC）焦亡和凋亡中的作用。
将HCEC在70 - 120 mM的高渗培养基中培养24小时，以建立体外干眼模型。检测细胞焦亡标志物GSDMD的水平，并通过检测IL - 1β和IL - 18水平评估细胞炎症反应。使用外源性半胱天冬酶 - 1抑制剂Ac - YVAD - CHO。通过半胱天冬酶 - 1活性、免疫荧光染色和蛋白质免疫印迹法检测HCEC中的细胞焦亡。采用流式细胞术检测HCEC的凋亡率。检测细胞迁移和增殖情况。通过定量实时PCR检测HCEC中lncRNA MIAT的表达。通过小干扰RNA（siRNA）转染敲低MIAT。观察半胱天冬酶 - 1抑制对细胞焦亡、凋亡、迁移和增殖的影响。
HS通过升高半胱天冬酶 - 1、GSDMD以及GSDMD的活性裂解产物（N端结构域，N - GSDMD）促进HCEC的细胞焦亡，并增加IL - 1β、IL - 18、乳酸脱氢酶（LDH）的释放以及凋亡率，同时降低细胞迁移能力。这些变化可通过抑制半胱天冬酶 - 1来预防。在暴露于高渗培养基的HCEC中，MIAT的表达显著增加。沉默MIAT可增加GSDMD、半胱天冬酶 - 1以及炎症趋化因子IL - 1β和IL - 18的表达，并促进HCEC的凋亡，同时抑制其迁移和增殖。
lncRNA MIAT参与HS诱导的HCEC焦亡、凋亡及炎症反应，为干眼发病机制提供了新的认识。
