制备淋巴瘤多基因肿瘤疫苗的方法学探索

Studies on the Preparation of Polygene Tumor Vaccine for Lymphoma

目的探讨用于淋巴瘤的多基因肿瘤疫苗的制作方式及淋巴瘤的基因治疗方法。方法采用腺病毒转染、脂质体辅助转染、脂质体辅助腺病毒转染的方式,将带GFP的载体转染B系淋巴瘤A20细胞株,并以Hela细胞作为对照,荧光显微镜下观察转染效率。将B7-1与CD40L表达载体联合导入A20荷瘤小鼠肿瘤内,观察肿瘤生长情况,应用肿瘤病理切片和HE染色技术观察肿瘤组织形态及淋巴细胞浸润情况。结果对A20细胞而言,单纯脂质体和单纯腺病毒转染效率均非常低,脂质体辅助腺病毒转染效率有所提高。但均显著低于采用腺病毒转染Hela细胞。瘤内联合注射B7-1和CD40L表达载体可导致肿瘤生长延缓及体积缩小,肿瘤消退效应较明显。肿瘤形态学观察表明,治疗小鼠肿瘤局部有炎性细胞浸润并伴有大面积的坏死,肿瘤局限化。结论B系淋巴瘤细胞株难于通过体外基因导入制作肿瘤疫苗,脂质体辅助腺病毒转染效果较好,质粒载体瘤内注射可作为一种安全有效的肿瘤疫苗作用方式。

Objective To study the preparation polygene tumor vaccine and the genetherapy for lymphoma. Methods GFP were transected into A20 cells with adenovirus, lipofectin and adenovirus with lipofectin assistant, and Hela cell lines were used as control. After 48 ～ 72 hours, the cells were observed under the fluorescence microscope to determine the efficiency of transfection. B7-1 and CD40L expression vectors in combination were directly injected into lymphomas of A420 mice model, the histological characteristics and the cell infiltration in lymphoma were observed. Results In the A20 cells, either the cells transected by lipofectin or adenovirus alone resulted in a very low efficiency. When adenovirus transected GFP were used with lipofectin assistant, the efficiency became higher. However, it is always far lower than that in Hela＇ s. Intratumoral injection of B7-1 and CD40L plasmids which resulted in reduction of tumor size. Morphological observation revealed inflammatory cell infiltration in the tumors, massive necrosis and localization of tumor. Condusion It is difficult to make tumor vaccine for lymphoma by transferring the gene into the cells in vitro. Although adenovirus with lipofectin assistant attains higher transfection efficiency, it is not sufficient, the intratumoral injection may be considered as a safe and effective way of tumor vaccine administration.