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目的研究幽门螺杆菌(Helicobacter pylori,Hp)活菌对体外培养的胃上皮细胞形态与生长的影响.方法采用 Hp 与细胞体外共培养技术,结合细胞计数、MTT、检测及 Ki-67抗原的免疫组化分析,观察 Hp 活菌对人胃腺癌细胞系 SGC-7901生长增殖的影响,同时参照病毒的致细胞病变效应实验计算 Hp 引起50%细胞空泡变性所需的最低浓度结果①细胞毒效应实验表明,3.26×10~6 CFU/mL 的 Hp 能使50%的细胞出现空泡变性;②细胞计数结果显示,在培养的细胞中加入3.2×10~4 CFU/mL Hp 时,72 h 内 SGC-7901细胞的数量与未加 Hp 的对照组基本一样;当 Hp 浓度增加到1.6×10~5CFU/ml 时,细胞数量较对照组显著增多;但是 Hp浓度继续增加5倍后,细胞增殖受到抑制,且随 Hp 浓度进一步加大,抑制效应愈大,差异有显著性(P<0.01).③MTT 检测结果与细胞计数结果十分相似,当终浓度为1.6×10~5CFU/mL 的 Hp 与细胞共培养时,可显著促进细胞生长(P<0.01):但是 Hp 浓度5倍增加(8×10~5 CFU/mL)以后,细胞的增殖减少(P<0.05);当 Hp 浓度达2×10~7CFU/mL 和1×10~8CFU/mL 时,细胞的生长明显被抑制(P<0.01).①Ki-67抗原的免疫组化分析显示,阳性细胞的胞核呈棕黄色着色,低浓度的 Hp(3.2×10~4 CFU/mL 和1.6×10~5 CFU/mL)作用后,SGC-7901细胞内 Ki-67抗原阳性率较对照明显增高(P<0.01);相反,较高浓度 Hp(4×10~6 CFU/mL 和2×10~7CFU/mL)作用后,细胞内 Ki-67抗原阳性率较对照显著减少(P 值分别为0.05与0.01).结论较低浓度的 Hp 不仅对细胞未显示明显的毒性,反而对细胞的生长增殖有一定的促进作用;而较高浓度的 Hp 却明显抑制细胞增殖,且浓度越大,抑制作用越显著.Hp 对细胞增殖的双向效应可能与其同时具有促进和抑制细胞增殖两类因素有关.本实验结果可用以部分解释人体感染 Hp 后所表现的不同结局.
Objective To study the effect of Helicobacter pylori (Hp) viable cells on the morphology and growth of gastric epithelial cells cultured in vitro.Methods Hp was co-cultured with cells in vitro and cell count, MTT assay and Ki-67 antigen immunization Histopathological analysis of Hp viable cells on human gastric adenocarcinoma cell line SGC-7901 growth and proliferation, while referring to the virus’s cytopathic effect experiment calculated Hp 50% cell vacuolar degeneration caused by the minimum concentration results ① cytotoxic effect Experiments showed that Hp at 3.26 × 10 ~ 6 CFU / mL resulted in vacuolar degeneration in 50% of the cells. ②Cell counting results showed that within 72 h after adding 3.2 × 10 ~ 4 CFU / mL Hp to cultured cells, The number of SGC-7901 cells was almost the same as that of the control group without Hp. When the concentration of Hp was increased to 1.6 × 10 ~ 5CFU / ml, the number of cells increased significantly compared with the control group. However, when the Hp concentration was increased by 5 times, (P <0.01) .③MTT assay was very similar to the cell counting results. When the final concentration of Hp was 1.6 × 10 ~ 5CFU / mL, Co-culture can be significantly promoted (P <0.01). However, the cell proliferation was decreased (P <0.05) when the concentration of Hp was increased by 5 times (8 × 10 ~ 5 CFU / mL) The cell growth was significantly inhibited at 10 ~ 8CFU / mL (P <0.01). ①Ki-67 antigen immunohistochemical analysis showed that the nucleus of positive cells showed brownish yellow and low concentration of Hp (3.2 × 10 ~ 4 CFU / mL and 1.6 × 10 ~ 5 CFU / mL), the positive rate of Ki-67 antigen in SGC-7901 cells was significantly higher than that in control (P <0.01) / mL and 2 × 10-7CFU / mL), the positive rate of Ki-67 antigen in the cells was significantly lower than that of the control (P = 0.05 and 0.01, respectively) .Conclusion Hp not only has no obvious cytotoxicity , But on the contrary, the cell growth and proliferation have a certain role in promoting; and higher concentrations of Hp significantly inhibited cell proliferation, and the greater the concentration, the more significant inhibition.Hp on cell proliferation two-way effect may be both with the promotion and inhibition of cells Proliferation of two types of factors.The results of this experiment can be used to partially explain the human body after the infection of Hp showed different outcomes.