·实验研究·Nestin在高眼压大鼠视网膜神经胶质细胞中的表达薛黎萍丁鹏吴开力江春光胡竹林肖丽波胡世兴TheexpressionofnestininretinalglialcellsinrathypertentioneyeXueLiping,DingPeng,WuKaili,JiangChunguang,HuZhulin,XiaoLibo,HuShixing.DepartmentofOphthalmology,SecondPeople’sHospitalofYunnanProvince,Kunming650021,ChinaAbstractBackgroundElevatedintraocularpressureleadstothelossofretinalganglioncellsandvigorousreactionofretinalglialcells.Theexpressionofnestininretinalglialcellssecondarytohypertentionanditssignificanceareunclear.ObjectiveThisstudyaimtoinvestigatetheexpressionofnestininretinalglialcells(RGCs)inocularhypertentionrats.MethodsTheocularhypertentionmodelswereestablishedbycauterizingthelimbus2drainingveinsintherighteyesof42SDrats,andaconjunctivalincisioninthelefteyesoftheratsservedastheshamgroup.Theintraocularpressure(IOP)wasmeasuredwiththeTono2PenXLtonometer.ThenumberofRGCsintheratswithocularhypertentionwascounted.TheexpressionofthenestinproteininRGCswassemi2quantitativelyanalyzedusingWesternblot,andtheinductionoftheexpressionofnestininMüllercellswasdetectedbyimmunochemistry.Doubleimmunofluorescencewascarriedouttoevaluatetheexpressionofnestin/GSornestin/GFAPinthefootplateofMüllercellsundertheconfocallaserscaningmicroscope.ResultsSignificantdifferenceswerefoundintheIOPbetweenthemodelgroupandtheshamgroupatvarioustimepoints(P<0105).In1weekto3weeksafteroperation,thenumberofRGCssignificantlydeclinedinthemodelgroupcomparedwiththeshamgroup(P<0105).Immunochemistryshowedthatfrom2hoursthrough1weekafteroperation,theexpressionofnestinwasgraduallyenhancedinthemodelgroupincomparisonwiththeshamgroup.Westernblotrevealedthattheexpressionofthenestinproteinreflectedasimilartendencytothatofimmunofluorescence.ThepresentresultsdeterminedthatMüllerglialcellsreactedvigorouslytoincreasedintrocularpressureasmanifestedbytheinducedexpressionofnestin.ImmunoelectronmicroscopyalsoconfirmedtheinducedexpressionofnestininMüllerglialcellsinocularhypertention.ConclusionThepresentresultsuggeststhattheinducedexpressionofnestininMüllerglialcellsisaresponsetoincreasedIOP.TheinducedexpressionofnestininMüllerglialcellsespeciallyattheirend2feetsuggestsapotentialneuroprotectivemechanisminneuronaldegeneration.Nestinmaybeausefulbiomarkerforretinalinjurystudy.Keywordsintraocularpressure;Müllercell;nestin;retinalganglioncells摘要目的探讨第六类中间丝蛋白nestin在高眼压大鼠视网膜神经胶质细胞中的表达情况及意义。方法42只SD大鼠采用烧灼右眼涡静脉的方法制作高眼压模型为模型组,剪开左眼结膜作为假手术组。测量并记录眼压,计数术后不同时间点视网膜神经节细胞(RGCs)数。Westernblot半定量分析术后视网膜内nestin蛋白含量。免疫电镜检测nestin在高眼压大鼠视网膜Müller细胞上的诱导表达。行nestin/谷氨酰胺合成酶(GS)或nestin/胶质纤维酸性蛋白(GFAP)免疫荧光双标,共焦显微镜观察nestin/GFAP在Müller细胞上的表达情况。结果术后各时间点模型组与假手术组眼压的差异有统计学意义(P<0105)。术后1～3周模型组RGCs数量显著减少,与假手术组比较差异有统计学意义(P<0105)。免疫荧光结果显示术后2h～3周,模型组nestin在Müller细胞上表达的荧光强度逐渐增强,与假手术组比较差异有统计学意义。Westernblot半定量分析结果与免疫荧光染色结果一致。免疫电镜结果进一步证实眼压升高后Müller细胞上出现nestin的诱导表达。结论Nestin在Müller细胞上的诱导表达是Müller细胞对眼压升高产生的一种反应,nestin的表达量与病程进展相一致。眼压升高时,nestin在Müller细胞上诱导表达,尤其在足板处的强烈表达,可能对RGCs...
