简介:目的观察1,25(OH)2D3对高糖诱导牛视网膜血管内皮细胞(BRECs)中血管内皮生长因子(VEGF)的表达水平变化及对细胞凋亡水平的影响。方法将分离培养的BRECs分为三组,分别为正常糖组、高糖组和高糖处理组。正常对照组细胞培养液含5mmol/L葡萄糖,高糖组细胞培养液含30mmol/L葡萄糖,高糖处理组细胞培养液含30mmol/L葡萄糖和50nM,1,25(OH)2D3。培养48h后收集细胞蛋白。蛋白免疫印迹法检测细胞VEGF及细胞凋亡相关蛋白Bax和Bcl-2表达水平;PI/Hoechst双染色法检测细胞凋亡。结果相比于正常糖组,高糖组中VEFG水平和Bax/Bcl-2比值显著增加;而在高糖处理组中表达水平远远低于高糖组,差异均有统计学意义(P〈0.05)。高糖的细胞凋亡水平高于正常糖组,而经过1,25(OH)2D3处理后,其细胞凋亡水平则有所下降,差异均有统计学意义(P〈0.05)。结论1,25(OH)2D3可以抑制高糖诱导BRECs中VEGF表达增加及细胞凋亡。
简介:目的:构建表达小鼠CD4+T细胞钙支架蛋白AHNAK1的短发夹RNA(shorthairpinRNA,shRNA)慢病毒载体,并研究其对小鼠甲状腺相关性眼病(thyroid-associatedophthalmopathy,TAO)的抑制效应。方法:设计并筛选对AHNAK1具有良好干扰效力的shRNA序列,慢病毒载体包装干扰序列,感染小鼠CD4+T细胞,检测AHNAK1静默对T细胞功能的抑制作用,采用实验动物模型观察AHNAK1体内抑制甲状腺相关性眼病的效果。结果:成功筛选出具有良好干扰效力的shRNA,并包装入慢病毒。病毒滴度为1.0伊106TU/mL,转染慢病毒的CD4+T细胞展现出失能倾向,抑制炎症免疫反应;在动物模型中抑制T细胞中AHNAK1表达可以有效控制甲状腺眼病的发生发展,显著降低治疗组T细胞中IL-2、IL-1茁和IFN-酌的表达。结论:成功构建了表达小鼠AHNAK1shRNA的慢病毒,具有抑制T细胞分泌IL-2、IL-1茁和IFN-酌的表达效应,能够有效抑制甲状腺眼病的发生发展。
简介:AIM:ToinvestigatetheinterferingeffectofY-27632,aROCK-Iselectiveinhibitor,onthesignaltransductionpathwayoftransforminggrowthfactor-β1(TGF-β1)inocularTenoncapsulefibroblasts(OTFS)invitro.METHODS:AfterOTFSfrompassages4to6invitrowereinducedbyTGF-β1andthentreatedbyY-27632,thechangesoftheOTFScellcycleswereanalyzedviaflowcytometry,andtheproteinsexpressionoftheα-smoothmuscularactin(α-SMA),connectivetissuegrowthfactor(CTGF),collagenIwerecalculatedbyWesternblot.AfterOTFStreatedbythedifferentconcentrationsofY-27632,theexpressionlevelsoftheα-SMA,CTGFandcollagenImRNAwereassayedbyRT-PCR.RESULTS:Y-27632hadnomarkedlyeffectontheOTFScellcycles.AftertreatedbyTGF-β1,OTFSinG1periodsignificantlyincreased.ThecellcyclesdistributionbybothTGF-β1andY-27632hadnoremarkabledifferencefromthatincontrolgroup.Y-27632significantlyinhibitedtheproteinsexpressionsofbothα-SMAandCTGF,whiletosomeextentinhibitedthatofcollagenI.TGF-β1significantlypromotedtheproteinsexpressionsofα-SMA,CTGFandcollagenI.AfterOTFStreatedbybothTGF-β1andY-27632,ofα-SMA,theproteinexpressionwassimilarwiththatincontrolgroup(P=0.066>0.05),buttheproteinexpressionofCTGForcollagenI,respectively,wassignificantlydifferentfromthatincontrolgroup(P=0.000<0.01).Thedifferencesofexpressionsoftheα-SMA,CTGFandcollagenImRNAin30,150,750μmol/LY-27632groupwerestatisticallysignificant,comparedwiththoseincontrolgroup,respectively(α-SMA,P=0.002,0.000,0.000;CTGF,P=0.014,0.002,0.001;collagenI,P=0.003,0.002,0.000).CONCLUSION:BlockingtheRho/ROCKsignalingpathwaybyusingofY-27632couldinhibitthecellularproliferationandtheexpressionofbothCTGFandα-SMAwhateverOTFSinducedbyTGF-β1ornot.Y-27632suppressedtheexpressionofcollagenImRNAwithoutinduction.
简介:目的:观察角膜缘干细胞培养液对血管内皮细胞增殖的影响。方法:分别培养角膜缘干细胞及球结膜上皮细胞,并通过免疫组化检测AE-5蛋白鉴别角膜缘干细胞。搜集两组培养细胞的上清液加入培养的人脐静脉血管内皮细胞中,并设立对照组。培养24h后通过MTT法检测细胞增殖情况并进行统计学分析。结果:角膜缘干细胞AE-5染色呈阴性,而球结膜上皮细胞为阳性。加入角膜缘干细球结膜上皮细胞和对照组培养液的血管内皮细胞MTT值分别为2.097±0.079,1.981±0.034和1.990±0.044。三组间有显著性差异(F=9.169,P=0.000)。加入角膜缘干细胞培养上清液的血管内皮细胞增殖活性明显高于其他两组(P=0.005和P=0.007)。结论:角膜缘干细胞培养液能够促进血管内皮细胞的增殖,这可能是角膜缘干细胞的特征。本研究从功能学角度为角膜缘干细胞理论提供更多的依据。
简介:AIM:TocomprehensivelyevaluatethepotentialassociationofCOL1A1polymorphismswithhighmyopiabyasystematicreviewandMeta-analysis.METHODS:AllassociationstudiesonCOL1A1andhighmyopiareporteduptoJune10,2014inPubMed,Embase,WebofScience,andtheChineseBiomedicalDatabasewereretrieved.Oddsratios(ORs)and95%confidenceintervals(95%CIs)wereanalyzedforsinglenucleotidepolymorphisms(SNPs)usingfixed-andrandom-effectsmodelsaccordingtobetween-studyheterogeneity.PublicationbiasanalyseswereconductedbyEgger’stest.RESULTS:Atotaloffourstudiesfromreportedpaperswereincludedinthisanalysis.TheMeta-analysesforCOL1A1rs2075555,composedof2304highmyopiapatientsand2272controls,failedtodetectanysignificantassociationwithhighmyopia.Atotalof971casesand649controlsweretestedforCOL1A1rs2269336.TheassociationofCOL1A1rs2269336withhighmyopiawasobservedinrecessivemodel(CCvsCG+GG,P=0.03)andinheterozygousmodel(CGvsGG,P=0.04),butnotinothermodels.CONCLUSION:ThisMeta-analysisshowsthatCOL1A1rs2269336(CCvsCG+GG)affectsindividualsusceptibilitytohighmyopia,whereasthereisnoassociationdetectedbetweenSNPsrs2075555andhighmyopia.Giventhelimitedsamplesize,furtherinvestigationsincludingmoreethnicgroupsarerequiredtovalidatetheassociation.
简介:AIM:Toinvestigatethelevelsofserumsolubleintercellularadhesionmolecules-1(sICAM-1)andneutrophilicexpressionofCD18inpatientswithvariousstagesofdiabeticretinopathyandtodeterminetheirdifferentexpressionpatterninthedevelopmentofdiabeticretinopathy(DR).·METHODS:LevelsofserumsICAM-1andCD18onthesurfaceofneutrophileweremeasuredin41DRpatients,theywereclassifiedinthreesubgroupsaccordingtothestageofretinopathyasdeterminedbyfund’sophthalmoscopy;10controlsubjectswerealsostudied.sICAM-1weremeasuredbyenzyme-linkedimmunosorbentassayandCD18byflowcytometry.·RESULTS:TheneutrophilicCD18expressionandserumsICAM-1levelwereallsignificantlyelevatedinalldiabeticsubgroupscomparedtocontrolsubjects(P<0.01).ThedifferencesofCD18andsICAM-1amongthediabeticsubgroupsweresignificantinCD18butnotinsICAM-1.TheprogressionofretinopathywasassociatedwithanincreasebothinCD18andinsICAM-1levelsbysimplecorrelationanalysis(β=0.74,P<0.001;β=0.38,P<0.01,respectively).ButstepwisemultipleregressionanalysisrevealedthatonlyCD18wasindependentdeterminantofretinopathy(β=1.04,P<0.01).·CONCLUSION:OurresultsconfirmthecontributionofendothelialandneutrophilicactivationinthedevelopmentofDRasindicatedbyincreasedlevelsofCD18andsICAM-1.However,adirectimplicationofCD18andICAM-1intheprogressionofDRcanbesupportedonlyintheCD18butnotICAM-1.CD18andICAM-1mayplaydifferentroleinthedevelopmentofdiabeticretinopathy.
简介:·AIM:Toinvestigatechangesinimmunogenicityofcryopreservedlimbalstemcells.·METHODS:Cryopreservedlimbalstemcells,freshprimarylimbalstemcellsandblankcontrolswereinoculatedsubcutaneouslyinC57BL-6miceandthepercentageofCD25cellsinlimbalexplantswasdeterminedbyflowcytometryatday21postinoculation.Morphologicalstudieswereperformedbylightandelectronmicroscopyoflimbalexplantsections.·RESULTS:Thenumberofregionalandsystemiclymphocytesderivedfromcryopreservedlimbalstemcellswaslowerthanthatfromfreshprimarylimbalstemcells.·CONCLUSION:Lymphocytesderivedfromcryopreservedlimbalstemcellsshowedchangesinimmunogenicity,butthesignificanceisunknown.Thecryopreservationandthawingmethodsawaitfurtherstudy.·
简介:·AIM:Toexploretheeffectofimmunizationwithcopolymer-1(COP-1)andretinalstemcells(RSCs)transplantationoninterferon-gamma(IFN-γ)levelsinaratexperimentalglaucomamodel.·METHODS:Anexperimentalglaucomawasinducedbyargonlaserphotocoagulationoftheepiscleralveinsandlimbalplexusintherighteyeofrats.Immediatelyfollowingglaucomainduction,ratswereimmunizedwithCOP-1.RSCswereculturedandtransplantedintravitreallyintotheeyesofglaucomamodelanimals1weekpost-lasertreatment.Sixexperimentalgroupswereused:COP-1/RSC,PBS/RSC,COP-1/PBS,PBS/PBS,glaucomamodelgroup,andanormalcontrolgroup.TheconcentrationofIFN-γinaqueoushumor(AH)andserumwasmeasuredbyenzyme-linkedimmunosorbentassay(ELISA)ineachofthesixgroups.Retinalganglioncell(RGC)survivalwasassessedbyquantifyingapoptosisusingHoechststaining.·RESULTS:ConcentrationsofIFN-γinAHandserumofratsthathadundergoneglaucomainductionwerehigherthanthoseofnon-inducedcontrolrats.TheconcentrationsofIFN-γinAHandserumoftheCOP-1/RSCstreatedgroupweredeterminedtobe2371.9ng/Land710.9ng/L,respectively,whichweresignificantlylowerthanthoseintheothertreatedgroups(P<0.05).Infact,IFN-γlevelsinthedualtreatedgroupwerereducedtobackgroundlevels.TheCOP-1/RSCgrouphadlowernumberofapoptoticRGCsthantheotherthreeexperimentalgroups(P<0.05).·CONCLUSION:ThereducedlevelsofIFN-γinAHandserumoftheCOP-1/RSCgroupmayberelatedtosynergisticeffectsbetweenRSCstransplantationandCOP-1immunemodulation.ItislikelythatthelowerlevelsofIFN-γpreventedRGCsglaucomatousapoptosis.·