il6和lps对hacat细胞il15表达的调控研究及其启动子区调控元件的鉴定-regulation of il6 and lps on il15 expression in ha cat cells and identification of regulatory elements in promoter region.docx

英文缩略词表缩写英文全称中文全称AKTSerine-threonineProteinKinaseB丝氨酸-苏氨酸蛋白激酶BAPSAmmoniumPersulfate过硫酸铵BCABicinchoninicAcid二喹啉甲酸Beta-actinBeta-actinBeta-肌动蛋白CKCytokine细胞因子DEPCDiethypyrocarbonate焦碳酸二乙酯DulbeccosModifiedEaglesDMEMMedium改良的Eagles培养基EBEthidiumBromide溴乙锭ECLElectrochemiluminescence化学发光ERK1/2ExogenouslyRegulatedKinases1/2胞外信号调节激酶1/2FBSFetalBovineSerum胎牛血清GlyceraldehydePhosphateGAPDHDehydrogenase磷酸甘油醛脱氢酶HaCaTThehumankeratinocytecellline人皮肤角质形成细胞株ILInterlukin白细胞介素JakJanusTyrosineKinase蛋白酪氨酸激酶LPSLipopolysaccharide脂多糖LSPLongSignalPeptide长信号肽2-(4-morpholinyl)-8-phenyl-4H-1-BLY294002MAPKenzopyran-4-oneMitogen-ActivatedProteinKinase特异性PI3K抑制剂丝裂原激活的蛋白激酶mRNAMessengerRNA信使RNA-1-NF-κBNuclearFactor-κB核因子-κBODOpticalDelnsity光密度PBSPhosphateBufferedSaline磷酸盐缓冲液PCRPolymeraseChainReaction聚合酶链式反应PD980592-(2-Amino-3-methoxyphenyl)-4H-1-Benzopyran-4-one特异性MAPK抑制剂PI3KPhosphotidylinsitol-3-Kinase磷脂酰肌醇激酶PMSFPhenylmethanesulfonylfluoride苯甲基磺酰氟PVDFPolyvinylideneFluoride聚偏二氟乙烯膜Real-TimePCRReal-TimePolymeraseChainReaction实时PCRRT-PCRReversedTranscriptivePolymeraseChainReaction逆转录PCRSDSSodiumdodecylsulfate十二烷基硫酸钠SDSSodiumDodecylSulfatePolyacrylamideGelElectrophoresis十二烷基硫酸钠-聚丙烯酰胺凝胶电泳SSPShortSignalPeptide短信号肽STATSignalTransducerandActivatorofTranscription信号转导子和转录激活子TEMEDTetramethylethylenediamine四甲基二乙胺TrisMethyltris(methylethylketoxime)Silane三羟甲基氨基甲烷盐酸盐TBSTrisbufferredsalineTris缓冲生理盐水TLRTollLikeReceptorToll样受体UVBUltravioletLight,MidrangeSunbeamSpectrum紫外线光谱WBWestern-Blot免疫印迹IL-6和LPS对HaCaT细胞IL-15表达的调控研究及其启动子区调控元件的鉴定中文摘要目的（1）探讨IL-6和LPS对HaCaT细胞IL-15表达的影响及其相关信号通路；（2）构建IL-15基因系列启动子荧光素酶报告基因载体，分析IL-15基因启动子活性从而鉴定其关键调控元件。方法（1）培养HaCaT细胞，用IL-6和LPS分别处理，Real-timePCR分析IL-6和LPS对IL-15mRNA水平表达的影响；Western-Blot分析IL-6和LPS对IL-15蛋白水平表达的影响。同时Western-Blot分析IL-6和LPS对HaCaT细胞信号通路MAPKs-ERK1/2和PI3K-AKT激活的改变，再使用其特异性抑制剂阻断IL-6和LPS激活的相关信号通路，检测IL-15的表达改变，从而探讨这些信号通路与HaCaT细胞IL-15表达之间的关系。（2）通过PCR方法从HaCaT细胞基因组中克隆七段逐步缺失的IL-15基因启动子序列；分别重组到含萤火虫荧光素酶的报告基因载体pGL3-Basic质粒上，构建IL-15基因系列启动子荧光素酶报告基因载体；用脂质体转染法将其转染至HaCaT细胞，同时并转染海肾荧光素酶报告基因载体作为内参照，给予IL-6和L