简介:目的:探讨胎牛血清对小鼠角膜上皮细胞生长和分化的影响。方法:分别在无血清低钙培养基(KSFM)和含100mL/L胎牛血清的KSFM中培养小鼠角膜上皮细胞。比较两种培养基中角膜上皮细胞的群体倍增(PD)。通过RT-PCR和Westernblotting方法检测p63、角蛋白19以及involucrin的表达。结果:在KSFM中,小鼠角膜上皮细胞可稳定传代超过25代,但在含胎牛血清的培养基中,细胞仅能存活3代。在KSFM中,培养细胞呈典型铺路石样外观,RT-PCR和Westernblotting结果显示细胞表达p63、角蛋白19以及involucrin。当培养基中加入胎牛血清后,细胞形态明显增大,呈扁平状;involucrin表达显著增强。结论:胎牛血清可抑制小鼠角膜上皮细胞增生、诱导其分化。
简介:AIM:Todeterminetheproliferativepotentialandthemaintenanceofstemcellactivityinstoredhumanlimbaltissues,andcorrelatethiswiththepreservationtime,cellviabilityandtheexpressionofstemcellmarkers.METHODS:Thirtylimbalrimsweresplitinto4partsandstoredincornealpreservationmediumat4℃for0,1,4,or7days.ThelimbalstemcellandmitoticmarkersP63,CK19,proliferatingcellnuclearantigen(PCNA),andKi67weredeterminedbyimmunohistochemicalstaining.Theproliferativepotentialoflimbalepithelialcellswasassessedbycellviability,theabilityofgeneratingstratifiedepithelium,andcolonyformingassay.RESULTS:Thestoredtissuesmaintainedlimbalstratifiedstructureto7daysandexhibitedcomparableexpressionlevelofstemcellandmitoticmarkers.Theproportionofviablecellsdecreasedwiththeprolongedpreservationtime,whilecolonyformingefficiencydecreasedfromthe1stdayanddisappearedatthe4thday.Wheninoculatedonamnioticmembrane,thecellspreservedfor1dayformedastratifiedepithelium,whilethecellsfrom4days’preservationformedadiscontinuouslayer.CONCLUSION:Thecolonyformingefficiencyoflimbalepithelialstem/progenitorcellsdecreasedrapidlywiththeincreasingpreservationtime,whiletheexpressionlevelofmarkersandcapacityofformingepithelialmonolayeronamnioticmembranedecreasedgradually.Thelimbalepithelialstemcellslosttheirfunctionearlierthanthelostexpressionlevelofstemcellmarkers.Thismayhelpustobetterchoosetheappropriatepreservationgraftsforfuturelimbalstemcelltransplantation.
简介:目的研究高眼压及其持续时间对大鼠视网膜caspase-3表达的影响。方法通过前房灌注平衡盐液建立大鼠急性高眼压模型:高眼压持续时间均为4h,依据眼内压的不同将sD大鼠60只随机分为正常对照组、40mmHg(1mmHg=0.133kPa)组、60mmHg组、80mmHg组、100mmHg组,每组12只。将眼内压为80mmHg的48只sD大鼠随机分为正常对照组、2h组、4h组、8h组,每组12只。正常对照组不给予前房灌注,其他各组分别给予不同的高眼压或高眼压持续不同时间。各组灌注结束后快速摘除眼球,分别采用Western印迹法和免疫组织化学法检测视网膜caspase-3的表达。结果与对照组相比,40mmHg组大鼠视网膜caspase-3的表达无增加(P〉0.05);60mmHg组、80mmHg组和100mmHg组大鼠视网膜caspase-3的表达均强于正常对照组(q值分别为4.87、5.28和6.71;P〈0.01),但各组之间差异无统计学意义(P〉0.05)。与对照组相比,2h组大鼠视网膜caspase-3的表达差异无统计学意义(P〉0.05),4h组和8h组大鼠视网膜caspase-3的表达均增加(q值分别为2.81和3.67;P〈0.01)。表达caspase-3的视网膜细胞主要位于神经节细胞层、外丛状层和内丛状层。结论采用前房灌注平衡盐液制作急性高眼压模型研究高眼压大鼠视网膜caspase-3的表达时,眼内压为80mmHg、高眼压持续4h即可。
简介:AIM:Tocharacterizetemporalpatternofresolutionandrecurrenceofnaivechoroidalneovascularization(CNV)secondarytowetage-relatedmaculardegeneration(AMD)treatedwithintravitrealbevacizumabonasneededregimen,andtoanalyzebaselineriskfactorsforCNVresolutionorrecurrence.METHODS:Ninety-oneeyesof80patientswithnewlydiagnosedwetAMDwereretrospectivelystudied.Alleyesweretreatedwitharoundofthreemonthlyintravitrealbevacizumabinjections,followedbyoneadditional’bonus’injectionafterresolutionofCNVactivity.Duringfollow-up,eyesweremonitoredwithfluoresceinangiography,opticalcoherencetomography,andbest-correctedvisualacuity(BCVA).IncaseofrecurrencesofCNVactivity,eyeswereretreatedwithotherroundsofbevacizumabinjectionsfollowingthesametreatmentprotocol.RESULTS:Overamedianfollow-upof532d,themedianresolutiontimeofCNVactivityinthefirst,second,andthirdtreatmentroundwas98d,126d,and111d,respectively.Themedianrecurrencetimeforthethreeroundswas154d,126d,and151d,respectively.Nosignificantdifferenceinresolutiontime(P=0.09)orinrecurrencetime(P=0.11)wasdetectedamongtreatmentrounds.Age(P=0.0082)andlensstatus(P=0.035)werefoundtobeassociatedwithCNVresolution;forevery1-yearincreaseinagetherewas4%greaterchanceofCNVresolution;Phakiceyesdemonstrateda33%betterchancetoexperienceCNVresolutionthanpseudophakiceyes.ForCNVrecurrence,lensstatus(P=0.0009)andgender(P=0.0446)werefoundtobepredictive;pseudophakiceyeshada3.69-foldgreaterrisktoexperiencerecurrenceofCNVactivitycomparedtophakiceyes;maleshada2.19-foldgreaterrisktoexperiencerecurrenceofCNVactivitythanfemales.NosignificantBCVAchangesamongthreetreatmentroundswerenoted(P=0.56).CONCLUSION:ResolutiontimeandrecurrencetimeofCNVactivitywerenotsignificantlydifferentamongtreatmentrounds,suggestingabsenceoftachyphylaxistobevacizumab.Acautiousdecisionshouldbemadeupondisco