【作者】 巩涛； 李勇； 范立桥； 焦志凯； 张志栋； 刘羽；

【Author】 Gong Tao Li Yong Fan Li-Qiao et al. The Third Department of Surgery, The Forth Hospital of Hebei Medical University, Shijiazhuang 050011, China

【机构】 河北医科大学第四医院暨河北省肿瘤医院胸外科；

【摘要】 目的:通过观察二十碳五烯酸(cis-5,8,11,14,17- eicosapentaenoic aicd,EPA)和二十二碳六烯酸(cis-4,7, 10,13,16,19-docosahexaenoic acid,DHA)对人胃癌BGC- 823细胞PKC βⅡ的影响,探讨ω-3脂肪酸的抗胃癌机制。方法:将人胃癌BGC-823细胞分对照组、EPA药物组和DHA 药物组,EPA组和DHA组按15 μ g／ml、30 μ g／ml、45μ g／ml加药,肿瘤细胞培养48小时后,用流式细胞仪测定胃癌细胞PKC β II蛋白含量(鼠抗人PKC βⅡ抗体,1:50),以荧光指数(Flurorescene Index,FI)表示蛋白相对含量:FI= {样品蛋白表达的平均荧光强度／正常对照样品平均荧光强度} ×100％。SP-9000免疫细胞化学检测胃癌细胞PKC βⅡ蛋白分布(鼠抗人PKC βⅡ单克隆抗体1:100),DAB显色。 Western blot检测胃癌细胞PKC βⅡ蛋白情况(鼠抗人PKC β II单克隆,1:500),DAB显色,测定蛋白印迹带的吸光度 (A)值。应用SPSS13统计软件的单因素方差分析。结果:用流式细胞仪测定人胃癌BGC-823细胞PKC β II蛋白表达在对照组中为4．71±0．92,30μg／ml和45 μg／ml的EPA 组中PKC β II蛋白分别为3．37±0．77和2．08±0．52,DHA 组中PKC β II蛋白分别为3．19±0．80和1．94±0．46。人胃癌BGC-823细胞PKC β II蛋白在药物组和对照组中的比较有显著差异,30μg／ml的EPA组或DHA组与对照组中的 PKC βⅡ蛋白比较有明显差异(p<0．05),45μg／ml的EPA 组或DHA组与对照组比较p<0．001),且45 μg／ml比30 μ g／ml下降更明显(p<0．05)。免疫细胞化学观察到对照组肿瘤细胞膜、浆中强染色,用EPA或DHA药物后PKC βⅡ在细胞膜、浆中染色减弱。Western blot检测对照组人胃癌 BGC-823细胞的PKC βⅡ蛋白印迹A值为2491．60±100． 97,30 μg／ml和45 μg／ml的EPA组PKC βII蛋白印迹 A值分别为903．28±73．64和744．00±60．08,30 μg／ml 和45 μg／ml的DHAPKC βⅡ蛋白印迹A值分别为914．31 ±80．87和755．43±64．34,药物组与对照组比较,PKC βⅡ蛋白印迹A值明显下降(p<0．001)。结论:EPA或DHA 可降低人胃癌BGC-823细胞PKC β II蛋白表达,当人胃癌 BGC-823细胞内PKC含量下降时,可能通过使其下游信号如NF-κ B含量也下降,从而使胃癌细胞生长受抑制,发生凋亡。更多还原

【Abstract】 Objective: To discuss the effect and mechanism of EPA and DHA on PKC β II of gastric cells. Methods: Human gastric BGC-823 cells divided into three groups:control group,EPA group and DHA group. Incubation of human gastric BGC-823 cells with different concentration (15 μ g/ml, 30 μ g/ml and 45 μ g/ml) of EPA and DHA for 48h, then PKC β II were detected by flow cytometry,immuno cytochemistry and Western blot. Results: PKC β II protein expression in groups of EPA 30 μ g/ml and 45 μ g/ml were 3.37 ± 0.77 and 2.08 ± 0.52 by flow cytometry, respectively. PKC β II protein expression in groups of DHA 30 μ g/ml and 45 μ g/ml were 3.19 ± 0.80 and 1.94 ± 0.46, respectively. Compared the above with control group (4.71 ± 0.92 ), PKC β II protein expression was decreased ( p<0.05 and p<0.001, respectively ). Especially compared 45 μ g/ml with 30 μ g/ml both EPA or DHA was further increased ( p<0.05, respectively ).Strong staining of PKC β II were located in cellular membrane and cytoplasm in control group by immuno cytochemistry. After treatment with EPA or DHA, staining of PKC β II were decreased. Absorbency of PKC β II in groups of EPA 30 μ g/ml and 45 μ g/ml were 903.28 ± 73.64 and 744.00 ± 60.08 by Western blot, respectively. Absorbency of PKC II in groups of DHA 30 μ g/ml and 45 μ g/ml were 914.31 ± 80.87, 755.43 ± 6434, respectively. Compared the above with control group ( 2491.60 ± 100.97 ), PKC β II was decreased ( p<0.001, respectively ). Especially compared 45 μ g/ml with 30 μ g/ml both EPA or DHA was further increased ( p<0.001, respectively ). Conclusion: EPA and DHA can induce PKC β and inhibit growth of the cultured cell.更多还原