胆汁泡相与微胶粒相的蛋白质表达谱对比观察

目的 采用比较蛋白质组学技术研究胆固醇结石患者胆囊汁汁的胆汁泡相与微胶粒相的蛋白质表达谱的差异,为探索胆石成因提供实验依据.方法 密度梯度超速离心法制备胆汁泡相和微胶粒相,提取蛋白,构建相应的蛋白质表达谱,ImageMaster图像分析软件获得差异蛋白点信息,质谱鉴定差异表达蛋白.结果 胆汁泡相与微胶粒相总蛋门浓度分别为(1.5358 4±0.0682)mg/ml、(7.1222 4±0.2022)mg/ml(P<0.01).两组组内蛋白质平均匹配点数分别为120 4±24和198 4±37,组间72 4±16个蛋白质点相互匹配,匹配率为45.30%.胆汁泡相与微胶粒相成功鉴定出差异蛋白质点8个,泡相中6个点表达上调,2个点表达下调.视黄醇结合蛋白(RBP)和白蛋白(HSA)丰度差异为免疫印迹法证实.结论 初步建立了胆同醇结石患者胆囊胆汁的胆汁泡相和微胶粒相蛋白表达谱,分析、鉴定了差异蛋白点,为筛选胆固醇结石形成关键调控子提供依据.

Differential analysis of protein profiles of vesicular phase and micellar phase of gallbladder bile from cholesterol gallstone patients

Objective To identify the proteins which play key roles during the formation of cholesterol gallstone.difie.rential analysis was carried out that the proteome of vesicular phase and micellar phase of gallbladder bile from cholesterol gallstone patients.Methods Vesicular and micellar phases were isolated by the density gradient ultracentrifugation method.Total proteins from the two phases were extracted.and the protein expressional profiles were established by two-dimensional electrophoresis respectively.The differentially expressed protein spots analyzed by ImageMaster two-dimensional mass spectrometry(MALDI-TOF-MS).Results The concentrations of proteins from vesicular phase and micellar phase of gallbladder bile from cholesterol gallstone patients were(1.5358 4±0.0682)me,/ml and (7.1222 4±0.2022)mg/ml(P<0.01)respectively.The average matched protein spots were 120±24 and 198±37 in the two groups respectively.There were 72±16 matched spots in the two representative gels maps and the matched rate was 45.30%.Eiisat differentially expressed protein spots were identified from the two cholesterol-carrier phases.Among them,6 were up-regulated with 2 down-regulated in vesicular phase comnared with micellar phase.The abundance difierentiation of RBP and HSA Was Confirmed by immunoblotting.Conclusions The difierential protein profiles of vesicular phase and micellar phase of gallbladder bile from cholesterol gallstone patients were established and 8 difierentiaI protein spots were identified successfully.The data may be a basis for further screening the key regulators of formation of cholesterol gallstone.