含金属硫蛋白蛋奶粉对小鼠肝细胞实体瘤的治疗作用

Therapeutic effects of egg-milk with metallothionein on H-22 hepatoma in BALB/c mice

目的：观察含金属硫蛋白（MT）蛋奶粉对荷瘤小鼠肝癌有无治疗作用。方法：70只BALB/c小鼠随机分为正常对照组10只、荷瘤鼠组20只、低剂量和高剂量含MT蛋奶粉肿瘤治疗组各20只。除正常对照组小鼠外，在其余小鼠体内移植H-22小鼠肝癌细胞株形成移植性肝癌模型，连续用含MT蛋奶粉给荷瘤小鼠灌胃2周后，每组断头处死10只小鼠，取其脾脏，采用MTT法检测各组小鼠淋巴细胞转化功能和IL-2活性，及流式细胞术检测脾脏CD4＋、CD8＋和CD25＋T细胞百分率以及淋巴细胞周期进程和凋亡的变化，以及3H-TDR掺入法检测CTL和NK细胞杀伤活性，并采用细胞病变抑制法检测小鼠IFN-γ活性以及L929体外杀伤法检测TNF-α活性。并于开始灌胃MT蛋奶粉后的第21-31天观察小鼠肿瘤大小。结果：在BALB/c小鼠形成肝癌第27、29及31天，含低剂量和高剂量MT蛋奶粉组小鼠肿瘤体积均明显小于肿瘤对照组（P〈0.05）；含低剂量和高剂量MT蛋奶粉肿瘤治疗组淋巴细胞刺激指数（SI）较肿瘤对照组均显著增加（P〈0.01），含高剂量MT蛋奶粉肿瘤治疗组小鼠脾脏CD4^＋细胞百分率及CD4^＋/CD8^＋比值较肿瘤对照组均显著增加（P〈0.01，P〈0.05），其CD25^＋T细胞百分率较肿瘤对照组显著降低（P〈0.01）；与肿瘤对照组比较，含高剂量MT蛋奶粉肿瘤治疗组小鼠淋巴细胞G0/G1期细胞百分率显著降低（P〈0.05），G2/M期细胞百分率显著增加（P〈0.05）；同时，含高剂量MT蛋奶粉肿瘤治疗组小鼠淋巴细胞凋亡百分率较肿瘤对照组（荷瘤鼠组）显著降低（P〈0.05）；荷瘤小鼠脾脏CTL细胞毒活性显著低于正常对照组（P〈0.05），含高剂量MT组CTL细胞毒活性显著高于荷瘤小鼠（P〈0.05）；荷瘤小鼠脾脏NK细胞毒活性明显低于正常对照组（P〈0.05），而含MT肿瘤治疗组NK细胞毒活性明显高于荷瘤组小鼠，尤其以高剂量组为显著（P〈0.01）。与肿瘤对照组比较，含高剂量MT蛋奶粉显著增加了荷瘤小鼠的IL-2和TNF-α活性（P〈0.01），含低剂量MT蛋奶粉对荷瘤小鼠的TNF-α活性亦有明显提高作用（P〈0.01），对IFN-γ活性并无显著影响。结论：含MT蛋奶粉对肿瘤的治疗性效应可能是通过增强荷瘤小鼠的免疫功能实现的。

Objective To explore the therapeutic effects of metallothionein （MT） on the H-22 liver neoplasm of BALB/c mice after administration with different concentrations of egg-milk with MT. Methods Seventy BALB/c mice were randomly divided into four groups： normal control （n=10）, tumor control （n=20）, low dose MT egg-milk group （n=20） and high dose MT egg-milk group （n= 20）. The mice excluding the normal control were injected with H-22 cells subcutaneously to build liver neoplasm models before they were administrated with egg-milk with MT for continuous two weeks. Lymphocyte proliferation ability and IL-2 activity in their spleen were measured with MTT method, the percentages of CD^4＋ , CD^8＋ and CD25＋ T cells were measured with flow cytometry, the killing activities of CTL and NK cells were detected with 3 H-TdR method, the activities of IFN-γ and TNF-α were measured with cytopathic effect inhibition （CPEI） and L929 killed in vitro methods, respectively. The tumor volume of mice was detected at the 21th to 31th day. Results As compared with those in the tumor group, the tumor volume of the mice with H-22 cell hepatoma after administration with egg-milk with low dose and high dose MT decreased significantly at the 27 th, 29 th and 31 th day （P〈0.05）. The lymphocyte stimulating indexes in the low dose and high dose MT groups were significantly higher than that in the tumor control （P〈0.01）. Compared with the tumor control, the positive percentage of CD4^＋ T cells and the ratio of CD4^＋/CD8^＋ T cells increased significantly （P〈0. 01）, and the positive percentage of CD25^＋ T cells decreased significantly （P〈0. 05）. The percentage of Go/G1 cells and the percentage of apoptotic cells in the high dose MT group were obviously lower than those in the tumor control （P〈0.05）, and the percentage of G2/M cells in the high dose MT group was obviously higher than that in the tumor control （P〈0.05）. The killing percentages of NK cells and CTL cells and the IL-2 and TNF-a activities in the high dose MT group were significantly higher than those in the tumor control （P〈0.01 or P〈0. 05）. While MT egg-milk had no effect on the IFN-γ activity. Conclusion Egg-milk with MT could build up the immune function of the mice with liver neoplasm and play a therapeutic effect on the tumor to some extent.