COG1410对视网膜缺血-再灌注损伤小鼠视网膜神经节细胞的保护作用及其机制  

Protective effect of COG1410 on the survival of retinal ganglion cells in mice with retinal ischemia-reperfusion injury and its mechanism

作　　者：赵茹 罗晋媛 贺涛[1] 邢怡桥[1] Zhao Ru;Luo Jinyuan;He Tao;Xing Yiqiao(Eye Center,Renmin Hospital of Wuhan University,Wuhan 430060,China)

机构地区：[1]武汉大学人民医院眼科中心,武汉430060

出　　处：《中华实验眼科杂志》2023年第11期1065-1075,共11页Chinese Journal Of Experimental Ophthalmology

基　　金：湖北省自然科学基金青年项目(2021CFB109);武汉大学自主科研项目(青年教师资助项目)(2042021kf0131)。

摘　　要：目的探索载脂蛋白E模拟肽COG1410对小鼠视网膜缺血-再灌注(IR)损伤后M1/M2小胶质细胞极化及视网膜神经节细胞(RGCs)存活的影响及其可能的机制。方法将18只8周龄C57BL/6J雄性小鼠按照随机数字表法分为正常对照组6只、IR 3 d组6只、IR 7 d组3只和IR 14 d组3只,其中IR组小鼠使用生理盐水进行前房灌注,将眼压提高至100 mmHg(1 mmHg=0.133 kPa)并维持1 h,以建立视网膜IR损伤模型。取正常对照组和IR 3 d组各3只,采用视网膜冰冻切片免疫荧光染色法观察视网膜小胶质细胞的分布情况。取正常对照组、IR 3 d组、IR 7 d组和IR 14 d组各3只,通过视网膜铺片免疫荧光染色法观察视网膜M1型、M2型小胶质细胞随IR损伤后时间的变化。另取91只C57BL/6J小鼠按照随机数字表法随机分为正常对照组19只、IR组24只、生理盐水组24只、COG1410组24只,其中正常对照组小鼠维持正常眼压,其余3个组均建立IR损伤模型,且COG1410组和生理盐水组在造模后分别尾静脉注射1 mg/kg COG1410和等体积生理盐水。采用视网膜铺片免疫荧光染色法观察各组小胶质细胞表型及RGCs存活率;采用实时荧光定量PCR法检测各组视网膜肿瘤坏死因子α(TNF-α)、白细胞介素-1β(IL-1β)mRNA的相对表达量;采用TUNEL法观察视网膜神经细胞的凋亡情况;采用Western blot法检测各组视网膜核因子κB(NF-κB)、B淋巴细胞瘤2(Bcl-2)、Bcl-2关联X蛋白(Bax)蛋白的表达水平。结果正常对照组与IR 3 d组视网膜小胶质细胞主要分布在神经节细胞层、内丛状层和外丛状层。正常对照组、IR 3 d组、IR 7 d组和IR 14 d组M1型、M2型小胶质细胞数目总体比较差异均有统计学意义(F=29.83、57.62;均P<0.001);与正常对照组相比,IR 3 d组M1型小胶质细胞数量增多,IR 7 d组M2型小胶质细胞数量增多,差异均有统计学意义(均P<0.05)。正常对照组、IR组、生理盐水组、COG1410组M1型小胶质细胞比例分别为(Objective To explore the effects of apolipoprotein E-mimetic peptide COG1410 on M1/M2 microglia polarization and retinal ganglion cells(RGCs)survival after ischemia-reperfusion(IR)injury in the mouse retina and its possible mechanisms.Methods Eighteen 8-week-old C57BL/6J male mice were divided into control group(6 mice),IR 3 days group(6 mice),IR 7 days group(3 mice),and IR 14 days group(3 mice)according to the randomized number table method.Mice in IR group were perfused in the anterior chamber using saline,and the intraocular pressure(IOP)was raised to 100 mmHg(1 mmHg=0.133 kPa)and maintained for 1 hour in order to establish a model of IR injury in the retina.Three mice from control group and 3 mice from IR 3 days group were taken to observe the distribution of retinal microglia by immunofluorescence staining of retinal frozen sections.Three mice were taken from normal control,IR 3 days,IR 7 days,and IR 14 days groups respectively to observe the changes of retinal M1-type and M2-type microglial cells with time after IR injury by immunofluorescence staining of retina.Another 91 C57BL/6J mice were randomly divided into normal control group(19 mice),IR group(24 mice),saline group(24 mice),and COG1410 group(24 mice)according to the random number table method.Mice in normal control group maintained a normal IOP,and the IR injury model was established in the other three groups.In addition,COG1410 group and saline group were injected with 1 mg/kg COG1410 and an equal volume of saline by tail vein injection,respectively.The microglia phenotype and survival rate of RGCs were observed by immunofluorescence staining of retinal wholemount.The relative expressions of retinal tumor necrosis factor-ɑ(TNF-ɑ)and interleukin-1β(IL-1β)mRNA were detected by real-time fluorescence quantitative PCR.The apoptosis of retinal neuronal cells was observed by the TUNEL assay.The expression levels of retinal nuclear factor-κB(NF-κB),B lymphocyte-2(Bcl-2),and Bcl-2-associated X protein(Bax)proteins were detected by Western blot.Use an

关 键 词：视网膜 再灌注损伤 COG1410 小胶质细胞极化 青光眼 载脂蛋白E 

分 类 号：R774.1[医药卫生—眼科]