Suppression of wild-type p53 and JunB fusion gene to the growth of transplanted tumor of hepatocellular carcinoma in nude mice

Abstract

Objective To explore the suppressive effects of wt-p53 and JunB fusion gene to transplanted tumor of hepatocellular carcinoma in nude mice. Methods Hepatocellular carcinoma HepG2 cells were transfected by wt-p53/junB fusion gene plasmid and lipidosome Lipofectmine 2000 with green fluorescent protein (GFP) (fusion gene vector), empty plasmid and Lipofectmine 2000 with GFP (empty vector) and pure Lipofectmine 2000. The positive clone was selected by using G418 antibiotics selection culture medium and RT-PCR was performed to identify the positive clone. Afterwards, 18 BALB/c nude mice were randomly divided into experimental group, negative control group and blank control group with 6 mice in each group using computer random number table method. The nude mice in 3 groups were inoculated with HepG2 cells transfected by fusion gene vector, empty vector and pure HepG2 cells respectively and 2×107 tumor cell suspension was injected subcutaneously on the right side of nude mice's back. The nude mice were killed 28 days after injection and the gross tumor volume, tumor weight and tumor control rate were calculated. The gross tumor volume and tumor weight were compared using one-way analysis of variance and pairwise comparison was performed using LSD-t test. Results The transfecting rate of HepG2 cells was 40% 24 hours after transfected by fusion gene vector and empty vector. After 10 days selection by using G418 antibiotics selection culture medium, HepG2 cells transfected by fusion gene vector and empty vector acquired large single cell positive clone and fusion gene stably-expressed HepG2 cells was gained after the cultivation was amplified. HepG2 cells transfected by fusion gene vector expressed higher level of p53 and JunB gene compared with the HepG2 cells transfected by empty vector through gel electrophoresis. The gene relative amount of p53 in the experimental group was 6.09 and JunB was 0.13, while in the negative control group were 0.42 and 0.04. The transplanted tumors were located subcutaneously on the backside of nude mice of 3 groups. The tumors of mice in the experimental group were smaller with a few millimeters in diameter. The tumors of mice in the negative control group and blank control group obviously uplifted on the mice back with the diameter between ten to dozens millimeters. The transplanted tumors were cauliflower-like, had smooth surface without coating and obvious interface with surrounding tissue. The tumors were hard-texture without adhesion with deep tissue. The surfaces of abdominal organs were smooth and no obvious metastasis was observed. The gross tumor volume of mice in the experimental group and negative control group were (47±21)mm3, (384±166)mm3 respectively and there was significant difference between two groups (LSD-t=4.638, P<0.05). And the tumor weight in two groups were(2.6±0.5)g and (10.1±0.6)g respectively and there was also significant difference (LSD-t=19.560, P<0.05). There was significant difference in the gross tumor volume and tumor weight between the experimental group and blank control group [(292±103)mm3, (9.6±0.7)g] (LSD-t=5.740, 27.225; P<0.05). The tumor control rate of experimental group was 84%. Conclusion The growth of transplanted tumor of hepatocellular carcinoma in nude mice can be obviously inhibited by fusion gene of wt-p53 and JunB. Key words: Gene fusion; Transfection; Mice, inbred BALB C; Neoplasm transplantation