C6细胞热休克蛋白抗原肽复合物的提纯及抑瘤作用的研究

Restrain tumor ability of HAC purified from C6 glioma cells

目的 提纯大鼠脑胶质瘤C6细胞热休克蛋白抗原肽复合物(HAC),免疫SD大鼠,观察HAC的抑瘤作用。方法 采用免疫亲和层析方法提纯大鼠脑胶质瘤C6细胞HAC,免疫20只大鼠为实验组,以另20只大鼠作为对照组,于免疫后1周,采用立体定向脑内接种方法,以C6细胞攻击两组大鼠,于肿瘤细胞攻击后第二周,取两组动物外周静脉血,测定外周静脉血淋巴细胞计数,并应用流式细胞仪技术测定外周血中CD3+CD4+和CD3+CD8+T淋巴细胞的比例。观察饲养过程中实验动物出现的症状、体征和第四周实验动物存活率。于第四周处死存活动物,取脑组织进行HE染色病理组织学检查,并用免疫组化方法分析脑胶质瘤浸润区T淋巴细胞分布情况。结果 实验组大鼠外周血淋巴细胞计数显著高于对照组(P<0.01),CD3+CD4+和CD3+CD8+T淋巴细胞的比例实验组均显著高于对照组(P<0.01)。实验组动物症状出现时间显著晚于对照组动物(P<0.01),实验组动物第四周末存活率显著高于对照组(P<0.05)。实验组胶质瘤局部浸润的CD3+和CD4+细胞数均显著高于对照组(P<0.01),实验组胶质瘤局部浸润的CD8+细胞数与对照组比较无显著差异(P>0.05),实验组胶质瘤局部浸润T淋巴细胞CD4+/CD8+显著高于对照组(P<0.01)。结论 C6细胞中HAC可以诱导大鼠产生对C6细胞的细胞免疫,提高大鼠存活率。

Objective Purify heat shock protein-antigen complex (HAC) , immunize SD rats with HAC, observe its supressing tumor ability. Methods Adopt immune affinity chromatography to purify HAC from C6 cells. Immunize 20 rats with HAC as experimental group, and other 20 rats as control group, one week after immunization inoculate C6 cell into the brain of the rats by stereotaky tedaique. After 2 weeks obtain peripheral vein blood, count the amount of lymphocyte, and count CD3+ CD4+andCD3+CD8+T lymphocyte with flow cytometer. Observe the symptoms that the animals present, and the survival rate of the rats at the end of 4th week. Kill all rats at the end of 4th week; obtain the brain tissue for histopathologic examination and immunohistochemistry examination. Result The amount of peripheral blood lymphocyte of the experiment group rats was significantly higher than that of the control group (P<0.01 ) , the rate of CD3+CD4+and CD3+CD8+T lymphocyte of the experiment group rats was significantly higher than that of the control group ( P<0.01 ) . The time that the experiment group rats present the symptoms was significant later than that be the control group ( P<0.01 ) , and the survival rate of the experiment group rats was significant higher than that of the control group ( P< 0.05 ) . The number of CD3+andCD4+T lymphocytes that infiltrated in the tumor tissue of the experiment group rats was significantly more than that of the control group. There was no significantly difference of CD8T lymphocytes infiltration between the two groups. The ratio of CD4+/CD8+ T lymphocytes of the experiment group rats was significantly higher than that of the control group ( P<0.01 ) . Conclusion HAC purified from C6 cells can induce the rats to develop the immune ability against C6 cells, and rise the survival rate of the rats.