HSV1-tk自杀基因系统对小鼠黑色素瘤的杀伤和抑制效应

Killing and Suppressing Effect of HSV1-tk Suicide Gene System on Murine Malignant Melanoma

【目的】检测自杀基因系统单纯疱疹病毒Ⅰ型胸苷激酶/丙氧鸟苷(HSV1-tk/GCV)对小鼠黑色素瘤的杀伤和抑制效应。【方法】将B16/tk(tk+)细胞和未经修饰的B16(tk-)细胞按tk+细胞占总细胞数的0%、10%、20%、40%、80%比例分别进行混合,各组细胞于96孔板中培养,加入丙氧鸟苷(GCV)至15.7μmol/L,采用四甲基偶氮唑盐(MTT)法检测不同tk+细胞比例的HSV1-tk/GCV系统在体外对肿瘤细胞的杀伤率。各组混合细胞分别接种于小鼠腋下,以GCV注射液按100 mg.kg-1.d-1进行治疗,检测瘤体体积、湿质量,测定抑瘤效应。【结果】tk+比例10%以上治疗组,HSV1-tk/GCV系统对黑色素瘤B16细胞均有明显杀伤效应,但旁杀伤作用不明显;从移植瘤体积看,20%以上tk/GCV治疗组肿瘤呈现明显生长抑制状态(P<0.05),以80%tk/GCV治疗组作用最显著(P<0.01);从移植瘤质量看,80%tk/GCV治疗组肿瘤呈现明显生长抑制状态(P<0.01),其他组虽随着tk+细胞比例的增高瘤体平均质量下降,但差异无显著性意义。【结论】HSV1-tk/GCV系统对小鼠黑色素瘤的杀伤和抑制效应均随着B16/tk细胞比例的增高而增强,已建立显示出体外杀伤效应和体内抑瘤活性的小鼠黑色素瘤自杀基因系统。

Objective To explore the killing and suppressing effect of HSV1-tk/GCV suicide gene system on murine malignant melanoma.Methods After mixing B16 /tk（tk^＋） cells and B16（tk^-） cells in the proportion of tk^＋ cells being 0%,10%,20%,40% and 80%,we cultured the melanoma tumor cells in the 96-well plate,and added ganciclovir（GCV） into the culture till the final concentration of GCV was 15.7μmol/L.The in-vitro antitumor effect of HSV1-tk suicide gene system with different proportions of tk^＋ cells was detected by methyl thiazolyl tetrazolium（MTT） assay.After subaxillary subcutaneous injection of the above-mentioned mixed cells,C57 mice were treated with GCV at 100 mg.kg-1.d-1.The tumor-suppressing effect was evaluated by measuring tumor mass size and tumor mass wet weight.Results HSV1-tk suicide gene system with the proportion of tk^＋ cells being more than 10% showed obvious in-vitro suppressing effect on malignant melanoma cells,while the bystander effects were not evident.The results of tumor mass size test showed that tumor growth was inhibited obviously in the group treated by GCV with tk^＋ cells being more than 20%（P〈0.05）,particularly in the group with tk^＋ cellsbeing 80%（P〈0.01）.The results of tumor mass weight test showed that tumor growth was inhibited obviously inthe group treated by GCV with tk^＋ cells being 80%（P〈0.01）.The average tumor weight in other groups decreased with the increase of tk^＋ cells proportion,but there was no statistical significance.Conclusion The killing and suppressing effect of HSV1-tk/GCV suicide gene system on murine malignant melanoma increases with the increase of tk^＋ cells proportion,indicating that the HSV1-tk/GCV suicide gene system with in-vitro killing effect and in-vivo suppressing effect is obtained successfully.