摘要
目的 探讨乳脂肪球表皮生长因子8(MFG-E8)在青光眼大鼠视神经保护中的作用与机制.方法 构建青光眼大鼠模型,将MFG-E8或D89E注射至大鼠玻璃体腔内.检测大鼠的眼压变化.通过HE染色、TUNEL染色、免疫荧光染色分别检测大鼠视网膜组织病理损伤、节细胞凋亡和小胶质细胞激活水平;通过Western blotting检测视网膜组织中cleaved caspase-3、caspase-3、cleaved caspase-9、caspase-9和BAX蛋白表达;通过ELISA和实时定量PCR检测视网膜组织中IL-10、TGF-β和NGF的表达水平.结果 与对照组相比,青光眼大鼠的眼压显著增加,视网膜神经节细胞复合体(GCC)层变薄,凋亡节细胞增加,cleaved-caspase 3/caspase-3、cleaved caspase-9/caspase-9和BAX水平升高,IBA1阳性细胞数增加,且IL-10、TGF-β、NGF水平增高;经MFG-E8治疗后,青光眼大鼠视网膜GCC层厚度增加,cleaved caspase-3/caspase-3、cleaved caspase-9/caspase-9、BAX水平降低,IBA1阳性细胞数和IL-10、TGF-β、NGF水平均增加;而MFG-E8失活异构体D89E处理后,大鼠青光眼进展进一步加重.结论 MFG-E8能够介导小胶质细胞激活,抑制神经节细胞凋亡,减缓大鼠青光眼的进展.
Abstract
Objective To investigate the role and mechanism of milk fat globule-EGF factor 8(MFG-E8)in neuroprotection in glaucoma rats.Methods A glaucoma rat model was constructed,and MFG-E8 or D89E was injected into the vitreous cavity of the rats.The intraoc-ular pressure of the rats was measured.HE staining,TUNEL staining and immunofluorescence staining were used to detect the patholog-ical injury,apoptosis and microglia activation of the retina.The protein expressions of cleaved caspase-3,caspase-3,cleaved caspase-9,caspase-9 and Bax were detected by Western blotting,and the expression levels of IL-10,TGF-βand NGFwere detected by ELISA and RT-qPCR.Results Compared with the control group,the intraocular pressure of glaucoma rats significantly increased,the retinal GCC layer became thinner,the apoptotic cells increased,the levels of cleaved caspase-3/caspase-3,cleaved caspase-9/caspase-9 and Bax increased,and the number of IBA1 positive cells increased,after treatment with MFG-E8,retinal GCC layer thickness increased,and the levels of cleaved caspase-3/caspase-3,cleaved caspase-9/caspase-9,Bax decreased,the number of IBA1 positive cells and the levels of IL-10,TGF-β,NGFincreased,but the progression of glaucoma was aggravated after the treatment of MFG-E8 inactivated isomer D89E.Conclusion MFG-E8 can mediate microglia activation,inhibit apoptosis of ganglion cells,and slow down the progression of glaucoma in rats.
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