Department of Ophthalmology, The First Hospital of Shijiazhuang City, Shijiazhuang 050000, Hebei Province, China.
Department of Ophthalmology, The First Hospital of Shijiazhuang City, Shijiazhuang 050000, Hebei Province, China.
Gene. 2020 Dec 30;763:145030. doi: 10.1016/j.gene.2020.145030. Epub 2020 Aug 2.
To investigate the impact and the mechanism of Gadd45α mediating p38MAPK pathway on the retinal ganglion cells (RGCs) injury in chronic ocular hypertension (COH) rats.
COH model in rats were established and intraocular pressure (IOP) was tested. Retrograde labeling was used for counting RGCs and TUNEL staining was performed for RGCs apoptosis. Western Blotting was conducted to examine the expression of Gadd45α and p38MAPK pathway. Besides, RGC-5 cells cultured in vitro were treated with HO. Cell viability was detected by CCK-8, ROS level tested by DCFH-DA assay, and cell apoptosis examined by flow cytometry.
COH rats had increased expression of Gadd45α and p-p38/p38 protein 1-4 weeks after surgery. Rats in COH group enhanced obviously in IOP, RGC apoptosis rate and the protein expression of Gadd45α, p-p38/p38, Bax/Bcl-2 and cleaved caspase-3, but declined appreciably in RGC counting. However, the above indicators of COH rats were effectively improved by Gadd45α shRNA treatment. Additionally, RGC-5 cells in HO group reduced in cell viability and went up in ROS level and apoptosis rate. The HO-induced RGC-5 cells treated with Gadd45α shRNA were improved apparently in those indicators, and cells treated with pcDNA Gadd45α showed an opposite trend. Moreover, p38 MAPK inhibitor SB203580 can effectively reverse the damage of pcDNA Gadd45α from HO-induced RGC-5 cells.
Silencing Gadd45α can reduce the RGC damage in COH rats by inhibiting p38MAPK pathway and such a protective role may be associated with the suppression of RGC apoptosis and the mitigation of oxidative stress.
探讨生长停滞 DNA 损伤可诱导蛋白α（Gadd45α）通过 p38MAPK 通路对慢性高眼压（COH）大鼠视网膜神经节细胞（RGCs）损伤的影响及其机制。
建立 COH 大鼠模型并检测眼内压（IOP）。采用逆行标记法计数 RGCs，TUNEL 染色检测 RGCs 凋亡。Western blot 检测 Gadd45α 和 p38MAPK 通路的表达。此外，体外培养 RGC-5 细胞，用 H2O2（HO）处理。用 CCK-8 检测细胞活力，用 DCFH-DA 法检测 ROS 水平，用流式细胞术检测细胞凋亡。
术后 1-4 周，COH 大鼠 Gadd45α 和 p-p38/p38 蛋白表达增加。COH 组大鼠 IOP 明显升高，RGC 凋亡率及 Gadd45α、p-p38/p38、Bax/Bcl-2 和 cleaved caspase-3 蛋白表达增加，RGC 计数明显减少。然而，Gadd45α shRNA 处理可有效改善 COH 大鼠的上述指标。此外，HO 组 RGC-5 细胞活力降低，ROS 水平和凋亡率升高。HO 诱导的 RGC-5 细胞经 Gadd45α shRNA 处理后，上述指标明显改善，而 pcDNA Gadd45α 处理的细胞则呈相反趋势。此外，p38MAPK 抑制剂 SB203580 可有效逆转 pcDNA Gadd45α 对 HO 诱导的 RGC-5 细胞的损伤。
沉默 Gadd45α 可通过抑制 p38MAPK 通路减轻 COH 大鼠的 RGC 损伤，这种保护作用可能与抑制 RGC 凋亡和减轻氧化应激有关。
