敲低通过抑制视网膜缺血后NLRP3炎性小体激活来改善视网膜神经节细胞损伤。
knockdown ameliorates retinal ganglion cell injury by inhibiting NLRP3 inflammasome activation after retinal ischemia.
作者信息
Xue Lin-Ping, Feng Hai-Song
机构信息
Hubei University of Chinese Medicine, Wuhan 430065, Hubei Province, China.
Department of Encephalopathy, Hubei Provincial Hospital of Traditional Chinese Medicine, Wuhan 430070, Hubei Province, China.
出版信息
Int J Ophthalmol. 2025 Jan 18;18(1):39-50. doi: 10.18240/ijo.2025.01.05. eCollection 2025.
AIM
To explore the neuroprotective effects of high mobility group box 2 () knockdown on retinal ganglion cells (RGCs) in the retinal ischemia-reperfusion injury (RIRI).
METHODS
Oxygen-glucose deprivation (OGD)-injured RGCs from postnatal three-day C57BL/6 mice pups and high intraocular pressure (IOP)-induced RIRI mice were used as cellular and animal models of RIRI. The expression of HMGB2 in the retina of RIRI mice and OGD-injured RGCs was detected through reverse transcription-polymerase chain reaction (RT-qPCR) and Western blotting. The effects of silencing on the morphological changes, RGCs survival, and cell apoptosis in mouse retinal tissues were observed through H&E staining, immunofluorescence staining with RNA-binding protein with multiple splicing (RBPMS) antibody, and TUNEL staining, respectively. RGC viability and apoptosis were examined by CCK-8 and flow cytometry assays. The levels of proteins associated with NOD-like receptor thermal protein domain associated protein 3 (NLRP3)-mediated pyroptosis [NLRP3, Caspase-1, GSDMD-N, interleukin (IL)-1β, IL-18] and were measured by Western blotting.
RESULTS
HMGB2 protein and NLRP3 were upregulated in the retina of RIRI mice and OGD-injured RGCs (<0.001). The retina was edematous, accompanied by disorganized cell arrangement and decreased thickness of all layers, and obvious vacuoles in ganglion cell layer. silencing alleviated the reduction in total retinal thickness and the severity of retinal tissue damage as well as suppressed RGC loss and retinal cell apoptosis in RIRI mice. OGD-induced RGC apoptosis was ameliorated after downregulation of HMGB2 . Intravitreal injection of the AAV-sh-HMGB2 and si-HMGB2 resulted in significantly decrease of NLRP3, Caspase-1, GSDMD-N, IL-1β, and IL-18 protein levels in the retinal tissues of RIRI mice and OGD-injured RGCs, respectively (all <0.001).
CONCLUSION
knockdown protects against RGC apoptosis and pyroptosis after RIRI through suppressing NLRP3 inflammasome activation.
目的
探讨敲低高迁移率族蛋白盒2（HMGB2）对视网膜缺血再灌注损伤（RIRI）中视网膜神经节细胞（RGCs）的神经保护作用。
方法
将出生3天的C57BL/6小鼠幼崽中经氧糖剥夺（OGD）损伤的RGCs和高眼压（IOP）诱导的RIRI小鼠作为RIRI的细胞和动物模型。通过逆转录聚合酶链反应（RT-qPCR）和蛋白质印迹法检测RIRI小鼠视网膜和OGD损伤的RGCs中HMGB2的表达。分别通过苏木精-伊红（H&E）染色、用多种剪接的RNA结合蛋白（RBPMS）抗体进行免疫荧光染色以及TUNEL染色观察敲低HMGB2对小鼠视网膜组织形态变化、RGCs存活和细胞凋亡的影响。通过CCK-8法和流式细胞术检测RGCs活力和凋亡情况。通过蛋白质印迹法检测与NOD样受体热蛋白结构域相关蛋白3（NLRP3）介导的细胞焦亡相关的蛋白水平[NLRP3、半胱天冬酶-1（Caspase-1）、Gasdermin D-N端（GSDMD-N）、白细胞介素（IL）-1β、IL-18]和HMGB2。
结果
RIRI小鼠视网膜和OGD损伤的RGCs中HMGB2蛋白和NLRP3上调（P<0.001）。视网膜水肿，伴有细胞排列紊乱和各层厚度降低，神经节细胞层有明显空泡。敲低HMGB2可减轻RIRI小鼠视网膜总厚度的降低和视网膜组织损伤的严重程度，并抑制RGCs丢失和视网膜细胞凋亡。下调HMGB2后，OGD诱导的RGCs凋亡得到改善。玻璃体内注射腺相关病毒-sh-HMGB2（AAV-sh-HMGB2）和小干扰RNA-HMGB2（si-HMGB2）分别导致RIRI小鼠视网膜组织和OGD损伤的RGCs中NLRP3、Caspase-1、GSDMD-N、IL-1β和IL-18蛋白水平显著降低（均P<0.001）。
结论
敲低HMGB2通过抑制NLRP3炎性小体激活来保护RIRI后RGCs的凋亡和细胞焦亡。
Zotero 插件