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1、第四军医大学硕士学位论文软骨细胞内，荧光显微镜下观察转染率大于90%，满足后续实验要求。4.MTT实验结果提示与对照组相比，转染miR-15a模拟物后软骨细胞增殖速率显著降低，P＜0.05。5.流式细胞术实验结果提示与对照组相比，转染miR-15a模拟物后软骨细胞凋亡率显著增高，P＜0.05。6.Westernblot结果提示与对照组相比，转染miR-15a模拟物后软骨细胞Ⅱ型胶原表达水平显著降低，P＜0.05。结论1.采用酶两步消化法可成功获得原代正常人膝关节软骨细胞与原代人骨关节炎膝关节软骨细胞，经鉴定后细胞纯度与所具有的功能表型可用于后续细胞学实验
2、。2.miR-15a在正常人膝关节软骨细胞与人骨关节炎软骨细胞中差异表达，在正常关节软骨细胞中表达量显著低于骨关节炎软骨细胞。3.化学合成的miR-15a模拟物可高效地转染进体外培养的关节软骨细胞，可在96h内显著提高细胞内miR-15a的表达量。4.miR-15a可抑制软骨细胞增殖，促进软骨细胞凋亡，提示miR-15a在骨关节炎的发病过程中扮演了重要的角色。5.miR-15a可通过抑制软骨细胞中Ⅱ型胶原的合成，影响软骨基质的完整和生物学功能的稳定，促进骨关节炎的发病。关键词：骨关节炎；软骨细胞；microRNA；miR-15a；细胞增殖；细胞凋亡-5-
3、第四军医大学硕士学位论文ExpressionofmiR-15ainHumanKneeArticularChondrocytesandEffectsofmiR-15aonProliferationandApoptosisofArticularChondrocytesCandidateforMaster:YanShijuSupervisor:Prof.FanQingyuTutor:Prof.MaBaoanDepartmentofOrthopaedics,TangduHospital,FourthMilitaryMedicalUniversity,Xi’an7
4、10032,ChinaSponsoredPrograms:NationalNaturalScienceFoundationofChina(81072194)AbstractBackgroundOsteoarthritis,alsoknownasdegenerativearthritis,isoneofthemostprevalentconnectivetissuedisorders,whichhasatremendousinfluenceonhealthofolderpeoplearoundtheworld.Osteoarthritisisakindof
5、age-relateddegenerativejointcondition,andisbecominganincreasinglycrucialriskfactorofdisabilityoftheelderly.Withtheaccelerationofagingofthepopulationinourcountry,osteoarthritishasbecomeagrowingburdenofthewholecountry.Theetiologyofosteoarthritis,however,isstillunknownyet,involvingv
6、ariousriskfactors,suchasaging,BMI(BodyMassIndex),andjointinjury.In-depthresearchofthemechanismsparticipatingintheprogressionofosteoarthritisnowisdesperatelyneededtobetterelucidatepathogenesisofosteoarthritis.Osteoarthritis,characterizedbyprogressivelossofarticularchondrocytes,ost
7、eophyteformation,ismorelikelytooccuramongweight-bearingjoints,suchashipandknee,andultimatelyleadstothedestructionofarticularcartilagestructure,jointdeformityandlossoffunctionofjoint.MicroRNAs,afamilyofendogenousnoncodingRNAmoleculeswith17-25-6-第四军医大学硕士学位论文nucleotides,ishighlycons
8、ervedandprevalentineukaryoticcells.Micro
