胃癌组织细胞中ngal基因表达及其调控机制分析-analysis of ngal gene expression and its regulatory mechanism in gastric cancer tissue cells.docxVIP

应性。5.在胃癌细胞系BGC-823和SGC-7901中，NGAL基因启动子的TPA反应元件位于基因5侧翼区-85~-79。6.在生物信息学预测基础上，通过蛋白印迹实验确定，在胃癌细胞系BGC-823中，可能由ERK细胞转导信号途径介导了NGAL基因的TPA反应性，AP1可能是其中的转录激活因子。结论：1．胃癌组织细胞中，NGAL基因高表达。2．在胃癌细胞系BGC-823和SGC-7901中，NGAL基因的表达存在着较大的差别。3．在胃癌细胞系BGC-823和SGC-7901中，NGAL基因启动子的核心元件不同。4．在胃癌细胞中，NGAL基因的启动子有TPA反应性，其TPA反应元件位于基因5侧翼区-85~-79。5．AP1可能是胃癌细胞中NGAL基因的转录激活因子，介导着TPA反应性。6．ERK的磷酸化激活可能介导了NGAL基因TPA反应性信号的转导。关键词：NGAL基因，胃癌，TPA反应元件，AP1，ERK磷酸化激活ABSTRACTRecently,manyresearchessuggestthatneutrophilneutrophilgelatinase-associatedlipocalin(NGAL)wasanimportantgenerelatedwithtumor,whichwasup-expressedincoloncancer,breastcancer,pancreaticcancer,lungcancer,skincancerandesophagealcancer.However,theexpressionlevelofNGALingastriccancerremainsunclear.Inthisstudy,wehavedetectedtheNGALexpressioningastriccancertissuesandcells,andinvestigatedtheregulationalmechanismofNGALgeneexpressioningastriccancercells.ThisresearchwillprovidemorecluesforfullycognizingtheoncobiologyfunctionsofNGAL.Materialsandmethods:ExpressionofNGALproteinwasanalyzedbyimmunohistochemistryin55paraffinsectionsofgastriccancertissueandthecorrespondingsectionsofnormalgastricglandulartissue.ExpressionofNGALinBGC-823andSGC-7901celllineswasanalyzedbywesternblottingandRT-PCR.TheluciferasereportergeneexpressionvectorscontainingNGALgene5flankingregulationregionwereconstructedbysubcloningthedifferentlengthDNAfragmentsofNGALgene5flankingregulationregionintopGL3-Basicvector.IngastriccancercelllinesBGC-823andSGC-7901,thekeyregionofNGALpromoterwasidentifiedbyDual-LuciferaseReporterAssaySystem.Bytheuseofmutationanddeletiontechniques，theTPAresponsivenessofNGALpromoterwasanalyzedingastriccelllines.Byusingwesternblottingandbioinformatics,weidentifiedthetranscriptionalactivatorandapproachedthesignaltransductionpathwaysthatmediateNGALexpressionresponsetoTPAingastriccancer.Results:Inimmunohistochemicalstudy,onlyweakpositiveornegativesignalwaspresentedinnormalgastricglandepitheliumcells.Whileingastriccancercells,strongpositivestainingcouldbeobserved,awhole-cytoplasmexp