大鼠青光眼应激模型中视网膜神经节细胞PTEN基因表观遗传修饰及表达的影响  

The epigenetic modification and expression of PTEN gene in retinal ganglion cells of glaucoma stress rats

作　　者：杨静[1] 史贻玉[1] 沈沛阳[1] 陈海波[1] 朱明月[2] 李伟[2] YANGJing;SHI Yi -yu;SHEN Pei-yang;CHEN Hai-bo;ZHU Ming-yue;LI Wei(Hainan Ophthalmological Hospital, Zhongshan Ophthalmic Center, Sun Yat - Sen University, Haikou 570311, Hainan, China)

机构地区：[1]中山大学中山眼科中心海南眼科医院(海南省眼科医院)海南省眼科学重点实验室(一病区),海南海口570311 [2]海南医学院基础医学院,海南海口570102

出　　处：《广东医学》2019年第8期1053-1057,共5页Guangdong Medical Journal

基　　金：海南省自然科学基金资助项目(编号:20158358)

摘　　要：目的探讨大鼠青光眼应激模型中视网膜神经节细胞PTEN基因表观遗传修饰及表达的影响。方法 (1)大鼠青光眼模型及处理。选择清洁级雌性Wistar大鼠30只,采用完全随机设计方法分为正常对照组10只、模型组10只、模型+药物干预组(甲钴胺处理)10只。除正常对照组外,其余大鼠均建立大鼠青光眼应激模型,建模完成后模型组不采取任何措施处理,正常对照组不参与建模,模型+药物干预组建模成功后采用甲钴胺进行处理,两组均干预11周。3组大鼠处死后,摘取眼球,完成视神经节细胞(RGCs)分离,用DAPI和PITC染色,荧光镜观察凋亡小体和caspase-3和PTEN细胞定位;采用免疫组织化学观察PTEN的表达。采用Western blot分析RGCs的PTEN和AKT(Ser473)表达量。(2)大鼠青光眼模型RGCs的PTEN基因表观遗传修饰的改变。造模后的1、3、5、7、9、11周处死大鼠,摘取眼球,完成RGCs分离,并将组织分为两份,一份用于测定PTEN的表达;另外一份于检测RGCs的PTEN基因启动子组蛋白(H3K9或H4K8)的乙酰化状态。结果模型+药物干预组处理后1、3、5、7、9、11周RGCs凋亡率低于模型组(P<0.05);模型+药物干预组与正常对照组PTEN阳性率差异无统计学意义(P>0.05),低于模型组(P<0.05);Western blot结果表明:模型+药物干预组PTEN表达量低于模型组和正常对照组(P<0.05);模型+药物干预组AKT(Ser473)表达量高于正常对照组(P<0.05),低于模型组(P<0.05);模型组+药物干预组PTEN基因启动子组蛋白阳性率与正常对照组比较差异无统计学意义(P>0.05),均高于模型组(P<0.05)。结论大鼠青光眼应激模型中视网膜神经节细胞PTEN基因表观遗传修饰及表达能产生明显的影响,能通过改变AKT(Ser473)表达量抑制疾病发展。Objective The epigenetic modification and expression of PTEN gene in retinal ganglion cells(RGCs) of glaucoma stress rats. Methods Thirty clean female Wistar rats were selected and divided into normal control group(n=10), model group(n=10), and intervention group(Methylcobalamin, n=10). Glaucoma model was established in model group and intervention group. Cobalamine was treated in intervention group for 11 weeks. The rats were sacrificed on the 16th day. The RGCs were isolated and collected by DAPI and PITC. The location of apoptotic bodies, caspase-3 and PTEN cells were observed by fluoroscopy. The expression of PTEN was observed by immunohistochemistry. The expression of PTEN and AKT(Ser473) in RGCs was analyzed by Western blot. At the 1st, 3rd, 5th, 7th, 9th and 11th week after model establishment, the rats were sacrificed and the eyeballs were taken for the isolation of RGCs. The PTEN expression and the acetylation of the PTEN gene promoter histone(H3 K9 or H4 K8) were analyzed in RGCs. Results The apoptosis rates of RGCs at the 1st, 3rd, 5th, 7th, 9th and 11th week after treatment in the intervention group were significantly lower than those in the model group(P<0.05). There was no significant difference in the positive rate of PTEN between the control group and intervention group(P>0.05), which was significantly lower than that of the model group(P<0.05). Western blot results showed that the expression level of PTEN was significantly lower in the intervention group than those in the model group and the normal control group(P<0.05). The expression level of AKT(Ser473) was significantly higher in the intervention group than that in the normal control group(P<0.05), but lower than that in the model group(P<0.05). There was no significant difference in PTEN gene promoter histone protein positive rate between the intervention group and the normal control group(P>0.05), which were significantly higher than that of the model group(P<0.05). Conclusion The glaucoma stress has a significant effect on the epigenetic m

关 键 词：大鼠青光眼应激模型 视网膜神经节细胞 PTEN基因表观 遗传修饰 

分 类 号：R775.2[医药卫生—眼科] Q311[医药卫生—临床医学]