Department of Ophthalmology, The Second Affiliated Hospital of Fujian Medical University, Engineering Research Centre of Assistive Technology for Visual Impairment, Fujian Province University, Quanzhou, Fujian Province, China.
https://orcid.org/0000-0001-7195-3624.
Invest Ophthalmol Vis Sci. 2024 Jan 2;65(1):49. doi: 10.1167/iovs.65.1.49.
To elucidate the influence of dopamine receptor 1 (DRD1) on the proliferation of mouse corneal epithelial cells (MCECs) under inflammatory conditions.
In vitro, immortalized MCECs (iMCECs) were treated with IL-1β, with and without pcDNA3.1_DRD1. Primary MCECs (pMCECs) were exposed to IL-1β, with and without DRD1 agonist (A68930). Cell proliferation was quantified using the Cell Counting Kit-8 (CCK-8) assay and immunofluorescence staining for Ki-67 and p63. Expression levels of NOD-like receptor protein 3 (NLRP3), IL-1β, and IL-6 were assessed. To establish a corneal injury model in mice, a 2-mm superficial keratectomy was performed. Either 0.1% A68930 or PBS was topically administered three times daily to the injured eyes for up to 5 days post-injury. Immunofluorescence analysis was employed to evaluate the expression of Ki-67, p63, and CD45 in mouse corneas. Western blotting and real-time quantitative PCR were utilized for quantitative analysis of DRD1, NLRP3, IL-1β, and IL-6 in mouse corneas. Corneal epithelial regeneration was monitored through fluorescein sodium staining for a duration of up to 5 days following the injury.
Overexpression of DRD1 and A68930 promoted MCEC proliferation and suppressed the expression of NLRP3, IL-1β, and IL-6 in vitro. Topical application of the 0.1% A68930 following mechanical corneal injury in mice led to increased Ki-67 and p63 expression compared to PBS treatment. Furthermore, topical administration of the 0.1% A68930 reduced the expression of CD45, NLRP3, IL-1β, and IL-6. Analysis with fluorescein sodium indicated accelerated corneal epithelial regeneration in the 0.1% A68930 treatment group.
DRD1 treatment counteracts NLRP3-associated inflammation and facilitates epithelial repair of corneal injury.
阐明多巴胺受体 1（DRD1）在炎症条件下对小鼠角膜上皮细胞（MCEC）增殖的影响。
在体外，用白细胞介素-1β（IL-1β）处理永生化 MCEC（iMCECs），并分别转染 pcDNA3.1_DRD1 和空载体。用 IL-1β 处理原代 MCEC（pMCECs），并分别用 DRD1 激动剂（A68930）和 DMSO 预处理。用细胞计数试剂盒-8（CCK-8）检测细胞增殖，用 Ki-67 和 p63 免疫荧光染色检测细胞增殖。检测 NOD 样受体蛋白 3（NLRP3）、白细胞介素-1β（IL-1β）和白细胞介素-6（IL-6）的表达水平。通过 2-mm 浅层角膜切除术建立小鼠角膜损伤模型，伤后每日 3 次向损伤眼滴 0.1%A68930 或 PBS，持续 5 天。免疫荧光分析评价 Ki-67、p63 和 CD45 在小鼠角膜中的表达。用 Western blot 和实时定量 PCR 检测小鼠角膜中 DRD1、NLRP3、IL-1β 和 IL-6 的表达。用荧光素钠染色监测损伤后 5 天内角膜上皮的再生情况。
DRD1 过表达和 A68930 处理促进 MCEC 增殖，并抑制体外 NLRP3、IL-1β 和 IL-6 的表达。与 PBS 处理相比，小鼠机械性角膜损伤后局部应用 0.1%A68930 可增加 Ki-67 和 p63 的表达。此外，局部应用 0.1%A68930 可降低 CD45、NLRP3、IL-1β 和 IL-6 的表达。荧光素钠分析表明，0.1%A68930 处理组角膜上皮再生加快。
DRD1 治疗可拮抗 NLRP3 相关炎症，促进角膜损伤的上皮修复。
