重组金黄色葡萄球菌肠毒素C3生物学活性的初步研究

目的:表达和纯化重组金黄色葡萄球菌肠毒素C3(rSEC3),并初步探讨rSEC3的生物学活性。方法:将转化有pET-32a(+)-SEC3重组质粒的大肠杆菌BL21经IPTG诱导表达、纯化后,MTT法检测0.1、0.5、1、5、10、50、100μg/ml的rSEC3促进人外周血单个核细胞(PBMC)增殖及抑制人宫颈癌细胞(Hela细胞)、人食管癌细胞(KYSE细胞)的作用。结果:重组金黄色葡萄球菌肠毒素C3在大肠杆菌BL21中成功表达。与空白对照相比,0.1～100μg/ml的rSEC3对PBMC均有显著的促增殖作用(P<0.05),1μg/ml时促增殖作用最强(t=113.851,P=0.000),和100μg/ml PHA作用相似(t=0.693,P=0.527)。以Hela细胞为靶细胞,培养48小时后,与空白对照相比,0.5、1、5μg/ml的rSEC3能显著增强PBMC对Hela细胞杀伤作用(P<0.05),抑瘤率分别为(70.44±9.27)%、(93.65±8.05)%、(103.40±6.65)%;以KYSE细胞为靶细胞,与空白对照相比,3个浓度的rSEC3也都有抑瘤作用(P<0.05),抑瘤率分别为(28.10±2.72)%、(46.62±4.17)%、(19.35±3.05)%。结论:rSEC3蛋白具有良好促人外周血单个核细胞增殖活性,并能够增强PBMC对肿瘤细胞的杀伤活性。

Expression and preliminary bioactivity analysis of the recombinant Staphylococcus aureus enterotoxin C3

Objective:To express the recombinant Staphylococcus aureus enterotoxin C3(rSEC3) and preliminary investigate its activity.Methods:E.coli BL21 containing the recombinant plasmids pET-32a(+)-SEC3 was expressed and purified.Then the human PBMCs was stimulated with different concentration of rSEC3 at 0.1,0.5,1,5,10,50,100 μg/ml and the promoting proliferation activity was detected by MTT method.Last,the tumor-cytotoxicity effects of Hela and KYSE were examined after being induced 48 h.Results:The recombinant Staphylococcus aureus enterotoxin C3 was expressed efficiently in E.coli BL21.Compared with blank control,0.1-100 μg/ml of rSEC3 could significantly promote proliferation of human PBMCs(P<0.05).1 μg/ml of rSEC3 was the most efficient dose(t=113.851,P=0.000) and showed a similar effect with 100 μg/ml PHA(t=0.693,P=0.527).After cultured 48 h,rSEC3 at 0.5,1,5 μg/ml had a strong depressant effect on Hela by activating PBMC compared with blank control(P<0.05).The inhibition rates were(70.44±9.27)%,(93.65±8.05)%,and(103.40±6.65)%.Furthermore,the rSEC3 also enhanced the killing activity to KYSE cells(P<0.05),and the inhibition rates were(28.10±2.72)%,(46.62±4.17)%,and(19.35±3.05)%,respectively.Conclusion:The recombinant protein rSEC3 has activity on human PBMCs proliferation,and it can enhance PBMCs killing effect on tumor cells.