摘要
目的探讨PI3K/Akt信号通路对高糖状态下视网膜Müller细胞的影响及机制。方法本实验分两部分,动物实验雄性SD大鼠随机分成对照组、糖尿病组、胰岛素样生长因子-1(insulin-like growth factors-1,IGF-1)组、IGF-1+LY294002(PI3K/Akt信号通路抑制剂)组。后三组采用链脲佐菌素(streptozotocin,STZ)(50 mg·kg-1)诱导成为糖尿病模型。模型诱导成功后,IGF-1组给予IGF-1(80 g·L^-1)5μL玻璃体内注射给药,IGF-1+LY294002组给予5μL(IGF-1+LY294002)(80μg·L^-1)玻璃体内注射给药。细胞实验对培养的Müller细胞进行不同处理,分为对照组(空白对照)、高糖组(30 mmol·L^-1葡萄糖)、IGF-1组(30 mmol·L^-1葡萄糖+80μg·L^-1 IGF-1)、IGF-1+LY294002组30 mmol·L^-1葡萄糖+80μg·L^-1(IGF-1+LY294002)。HE染色检测大鼠视网膜内核层(inner nuclear layer,INL)细胞密度,免疫荧光染色检测大鼠视网膜神经胶质纤维酸性蛋白(glial fibrillary acid protein,GFAP)表达及Müller细胞Caspase-3表达,TUNEL染色检测细胞凋亡情况,Western blot检测p-Akt、NF-κB、Caspase-3蛋白相对表达水平。结果动物实验中,与对照组相比,糖尿病组及IGF-1+LY294002组GFAP表达均明显增加,INL细胞密度均明显降低(均为P<0.05);与糖尿病组相比,IGF-1组GFAP表达明显下降,INL细胞密度明显增加(均为P<0.05)。细胞实验中,与对照组相比,高糖组及IGF-1+LY294002组细胞凋亡率和NF-κB、Caspase-3蛋白相对表达水平均明显增加,p-Akt蛋白相对表达均明显降低(均为P<0.05);而与高糖组相比,IGF-1组细胞凋亡率、NF-κB及Caspase-3蛋白相对表达均明显下降,p-Akt蛋白相对表达明显增加(均为P<0.05)。结论激活PI3K/Akt通路可对抗高糖状态下视网膜Müller细胞凋亡,其机制可能与下调NF-κB、Caspase-3蛋白表达有关。
Objective To investigate the effect and mechanism of PI3K/Akt signaling pathway on retinal Müller cells in diabetic rats.Methods The experiment was divided into two parts.As for animal experiment,male SD rats were randomly divided into control group,diabetic group,insulin-like growth factors(IGF-1)group and IGF-1+LY294002(PI3K/Akt signaling pathway inhibitor)group.Diabetes model induced by streptozotocin(STZ)(50 mg·kg-1)in the latter three groups.After successful induction of the model,the IGF-1 group was given intravitreal injection of 5μL IGF-1(80μg·L^-1),and the IGF-1+LY294002 group was given 5μL(IGF-1+LY294002)(80μg·L^-1).As for cell experiment,the cultured Müller cells were divided into control group(blank control),high glucose group(30 mmol·L^-1 glucose),IGF-1 group(30 mmol·L^-1 glucose+80μg·L^-1 IGF-1),IGF-1+LY292 group 30 mmol·L^-1 glucose+80μg·L^-1(IGF-1+LY294002).HE staining was used to detect the density of retinal inner nuclear layer(INL)cells,immunofluorescence staining was used to detect the expression of glial fibrillary acidic protein(GFAP)in rat retina and the expression of Caspase-3 in Müller cells,TUNEL staining was used to detect apoptosis of Müller cells,and Western blot was used to detect the relative expression of p-Akt,NF-κB and Caspase-3 proteins.Results In the animal experiment,when compared with the control group,the expression level of GFAP in diabetes group and IGF-1+LY294002 group increased significantly,and the density of INL cells decreased significantly(both P<0.05).Compared with diabetes group,the expression level of GFAP in IGF-1 group decreased significantly,and the density of INL cells increased significantly(both P<0.05).In the cell experiment,when compared with the control group,the apoptotic rate and the expression levels of NF-κB and Caspase-3 in high glucose group and IGF-1+LY294002 group increased significantly,while the expression level of p-Akt decreased significantly(both P<0.05);compared with high glucose group,the apoptotic rate and the expressio
作者
刘文强
冯闯
左中夫
刘学政
LIU Wenqiang;FENG Chuang;ZUO Zhongfu;LIU Xuezheng(Department of Anatomy,School of Basic Medicine,Jinzhou Medical University,Jinzhou 121001,Liaoning Province,China;Graduate School of Liaoning University of Traditional Chinese Medicine,Shenyang 110847,Liaoning Province,China)
出处
《眼科新进展》
CAS
北大核心
2020年第11期1024-1028,共5页
Recent Advances in Ophthalmology
基金
国家自然科学基金资助(编号81571383)
中国博士后科学基金资助(编号2017M612870)。
