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ID:34634598
大小:4.39 MB
页数:60页
时间:2019-03-08
《左卡尼汀对体外高糖培养鼠视网膜muller细胞的保扩作用及机制》由会员上传分享,免费在线阅读,更多相关内容在学术论文-天天文库。
1、左卡尼汀对体外高糖培养鼠视网膜Miiller细胞的保护作用及机制中文摘要目的:观察体外培养大鼠视网膜Mtiller细胞在高糖条件下细胞活性、活性氧(ROS)和线粒体膜电位(MMP)的变化,研究左卡尼汀(L.carnitine,LC)对其作用的影响,探讨LC对Mijller细胞的保护作用及机制。方法:取出生1。3天SD大鼠视网膜,反复胰酶消化法纯化Mtiller细胞,进行GS、GFAP单标及双标免疫荧光鉴定。用台盼蓝染色法观察不同浓度葡萄糖对Miiller细胞生长状态及存活率的影响,确立高糖浓度。将
2、Mtiller细胞随机分为正常对照组、高糖组和不同浓度LC处理组,LC处理组在高糖培养液中分别加入终浓度为50、l00、200umol/LLC。MTT法检测细胞活性,Hoechst33342染核检测细胞凋亡,流式细胞仪检测细胞内ROS和MMP的变化。结果:免疫荧光显示,体外培养的细胞大部分抗GS、GFAP抗体反应阳性,该方法培养的Mtiller细胞纯度可达90%以上。MTT结果显示,高糖培养组细胞活性显著下降(P<0.05),LC处理后细胞活性上调,以100umol/LLC作用最佳。ROS检测结果
3、显示,高糖损伤后细胞内ROS水平明显高于正常对照组(P<0.05),Lc处理后,ROS值均低于高糖培养组(P<0.05)。MMP检测结果显示,高糖损伤12h后,细胞MMP较正常对照组显著下降(P<0.05),Lc处理后,MMP均高于高糖培养组(尸4、膜电位,抑制细胞的凋亡。硕士研究生.张莹(眼科学)指导教师孔庆兰教授曹玉副教授关键词:左卡尼汀;视网膜Miiller细胞;活性氧;线粒体膜电位:凋亡ProtectiveeffectandmechanismofL—CarnitineontheratretinalMiillercellsinhigh—glucoseAbstract0bjective:ToinvestigatetheprotectiveeffectofL—carnitine(LC)oncellviability,reactiveoxyg5、enspecies(ROS),mitochondrialmembranepotentials(MMP)ofculturedratsMfillercellsinducedbyhighglucoseinvitroandthepossiblemechanism.Methods:Mtillercellsweresepratedandpirifiedfromtheretinaof1—3daySDrats.Thecellswereidentifiedbyimmunofluorescencebyusingglu6、taminesynthetase(GS),glialfibrillaryacidicprotein(GFAP).TrypanbluestainingwasusedtoobservetheeffectofdifferentconcentrationsofglucoseonthegrowthandsurvivalstatusofMfillercells.TheMfillercellswererandomlydividedintocontrolgroup,highglucosegroupandtheLC7、—treatedgroupwithdifferentconcertrations.ThecellsinLC—treatedgroupwereculturedinhighglucosemediumwith50,100,200umol/LLC.CellviabilitywasdeterminedbyMTT.ApoptosisratewasmeasuredbyHoechst3342.ROSandMMPwereevaluatedbyflowcytometer.Results:Immunofluoresce8、nceshowedthatmostcellswerepositiveforanti—GS,anti—GFAPantibodyresponses,thepurityofMfillercellswasabove90%.Cellviabilitydecreasedobviouslyinhighglucose(P<0.05),anditcanbeincreasedafterinterventionbyLC,andthebesteffectisthe100umol/L.ROSlevelinh
4、膜电位,抑制细胞的凋亡。硕士研究生.张莹(眼科学)指导教师孔庆兰教授曹玉副教授关键词:左卡尼汀;视网膜Miiller细胞;活性氧;线粒体膜电位:凋亡ProtectiveeffectandmechanismofL—CarnitineontheratretinalMiillercellsinhigh—glucoseAbstract0bjective:ToinvestigatetheprotectiveeffectofL—carnitine(LC)oncellviability,reactiveoxyg
5、enspecies(ROS),mitochondrialmembranepotentials(MMP)ofculturedratsMfillercellsinducedbyhighglucoseinvitroandthepossiblemechanism.Methods:Mtillercellsweresepratedandpirifiedfromtheretinaof1—3daySDrats.Thecellswereidentifiedbyimmunofluorescencebyusingglu
6、taminesynthetase(GS),glialfibrillaryacidicprotein(GFAP).TrypanbluestainingwasusedtoobservetheeffectofdifferentconcentrationsofglucoseonthegrowthandsurvivalstatusofMfillercells.TheMfillercellswererandomlydividedintocontrolgroup,highglucosegroupandtheLC
7、—treatedgroupwithdifferentconcertrations.ThecellsinLC—treatedgroupwereculturedinhighglucosemediumwith50,100,200umol/LLC.CellviabilitywasdeterminedbyMTT.ApoptosisratewasmeasuredbyHoechst3342.ROSandMMPwereevaluatedbyflowcytometer.Results:Immunofluoresce
8、nceshowedthatmostcellswerepositiveforanti—GS,anti—GFAPantibodyresponses,thepurityofMfillercellswasabove90%.Cellviabilitydecreasedobviouslyinhighglucose(P<0.05),anditcanbeincreasedafterinterventionbyLC,andthebesteffectisthe100umol/L.ROSlevelinh
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