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1、重庆医科大学博士研究生学位论文成细胞在自然、连续、动态的成长过程中存在差异表达蛋白。鉴定的21个候选蛋白功能各异，它们通过参与细胞的基础能量代谢；影响蛋白质的合成、折叠、降解过程；调控细胞的生长、增殖、分化；参与细胞骨架蛋白的构建；参与细胞的凋亡、免疫调控。使细胞增殖能力降低，加速细胞的凋亡；损伤细胞之间的连接，分泌各种细胞因子，通过各种通路，加速胶原的降解，导致氧化和抗氧化的失衡，形成了错综复杂的网络体系，在自然老化和（或）光老化的过程发挥一定作用。我们探寻到其中差异表达的蛋白或特异表达的蛋白有可能成为皮肤老化的标志性
2、蛋白，初步探讨了角质形成细胞在皮肤老化过程中的作用及分子机制，为揭示衰老的本质奠定了基础。但要确定某种蛋白是皮肤老化的标志性蛋白，尚需进一步验证。关键词角质形成细胞，皮肤老化，蛋白质组7万方数据重庆医科大学博士研究生学位论文ANALYSISANDIDENTIFICATIONOFSKINKERATINOCYTESPROTEOMICINAGINGPROCESSABSTRACTOBJECTIVETheyarestilllackofunderstandinginskinagingprocessandskinagingobject
3、ivemarkerinclinicalpractice.Thisstudyistoexplorethedifferentiallyexpressedproteinoraspecificproteinexpressionthatcanbeamarkerofskinagingfromanexposedandnon-exposedskinofyouth,middle-agedandelderly.Exploretheroleandmolecularmechanismofkeratinocytesintheskinagingpr
4、ocess.Tolayafoundationforafurtherstudyinacomprehensiveexplanationofthemechanismofskinagingandtorevealthenatureofskinaging.METHODByusingaproteomicstechnologyplatform,utilizingDisposeⅡtoisolateepidermallayerfromskin,optimizeandimprovesampleprocessingmethods,addacoc
5、ktail,RNase,DNaselysisliquidtoextractproteinsfromepidermis,acquired2-DEimagesofepidermaltissuekeratinocytebyusinganisoelectricandverticalfocusingelectrophoresiswithsilverstaining,tosetupatwo-dimensionalgelelectrophoresisthatsuitableforstudyepidermaltissue.Through
6、outGS800scannertoscanstainedgeldensity,imageanalysiswithPDQuest7.4,intensitycorrection,backgroundreduction,spotdetection,matching,IDcorrection,inorderto8万方数据重庆医科大学博士研究生学位论文determinedifferencesinproteinspots.Theselectedproteinwillfurtheranalyzeanddatabasequeryusin
7、gMALDI-TOFmassspectrometry,andfinallyidentificationofdifferencesinthecandidateprotein.RESULTObtainedtwo-dimensionalgelelectrophoresismapswereclear;completeseparationoftheproteinspots;mapsthatwereobtainedby3repeatedtimesineachgroupswereverysimilar;carriedoutthreei
8、nter-gelproteinspotmatching,thedeviationresultoffirstmeasurementbetweenproteingelbyIEFwas(1.798±1.232)mmwithsecondmeasurementbySDS-PAGEdeviationwas(1.063±1.031
