TC-1-HPV16L1细胞模型与动物模型的建立与评价

Establishment and Evaluation of TC-1-HPV16L1 Cell Model and Animal Model

为了解决人乳头瘤病毒16型L1蛋白(HPV16L1)为主要靶点的疫苗缺乏肿瘤模型细胞来验证疫苗的免疫效果的问题,构建了一个细胞模型并可以利用此细胞在实验动物体内形成肿瘤,用于以HPV16L1为主要靶点疫苗的验证实验.利用这个模型细胞或肿瘤模型可以在体内外检测疫苗的免疫学性质和保护功效.首先,利用聚合酶链式反应(polymerase chain reaction,PCR)的方法获得目标基因HPV16L1的基因序列并连接到载体质粒中,将质粒转染到细胞TC-1后在杀稻瘟菌素抗性压力筛选下获得稳定表达HPV16L1的细胞株.对TC-1和TC-1-HPV16L1两种细胞的生长增殖和成瘤特性进行比较发现,外源基因的加入对细胞特性没有影响.通过体外杀伤实验证实细胞TC-1-HPV16L1能够被HPV16L1靶点疫苗免疫过的小鼠脾淋巴细胞杀伤.实验动物被疫苗免疫后,进行攻瘤实验发现疫苗只对TC-1-HPV16L1组具有保护作用.综上,在本研究中成功地构建了可以检测HPV16L1疫苗的抑瘤效果的模型细胞TC-1-HPV16L1,为HPV疫苗的研究提供了一个模型细胞.

The main purpose of this research is to construct a model cell which can form tumor in experimental animals, and the model cell and model animal can be used to study the vaccine of HPV16L1 as the main target in vitro and in vivo. First of all, HPV16L1 gene sequences were obtained by polymerase chain reaction ( PC R ) amplification and full-length open reading frame ( ORF) was constructed in-frame into the pcDNA 3. 1/blasticidin expression plasmid. The cell line of stable expression of HPV16L1 was obtained by transfection of recombinant plasmid into T C -1 cell line under the resistant screening of blasticidin. The addition of exogenous gene HPV16L1 did not cause significant changes in the characteristics of cells through the comparison of proliferation and tumorigenesis between TC -1 and TC - 1 - HPV16L1. T C - 1 - HPV16L1 can be killed by the spleen lymphocytes of mice i m m u n i z e d by H P V 1 6 L 1 target vaccine in vitro test. E xperiment of t umor challenge proved that the vaccine h a d protective effect o n T C - 1 - H P V 1 6 L 1 in animals.