摘要
用反转录PCR的方法,从BALB/c小鼠脾细胞中扩增出B7-1cDNA后,插入pcDNA3质粒中构建成小鼠B7-1cDNA的真核表达载体pCD-mB7.1,经酶切鉴定和序列分析证实此表达载体中插入的B7-1cDNA的序列与文献报道一致.通过脂质体介导将pCD-mB7.1导入B7-1的小鼠黑色素瘤细胞系B16(F0)中,经RT-PCR和RNA斑点杂交初步证实B7-1在肿瘤细胞中获得了稳定有效的表达.同源小鼠脾淋巴细胞与肿瘤细胞混合培养后采用LDH释放改良法测定淋巴细胞特异杀伤活性,结果显示,与野生型和模拟转染的B16细胞相比,B7-1基因转染的B16细胞能较有效诱导淋巴细胞产生针对野生型B16细胞的特异杀伤活性(p<0.02).这说明,将B7-1基因导入肿瘤细胞表达能提高其免疫原性。
Recent studies suggested that expression of B7 1 in tumor cells is effective at inducing antitumor immune responses. In this research, mouse B7 1 cDNA was amplified by RT PCR from spleen cells of BALB/c mice, and inserted into a eukaryotic expression vector, pcDNA3. Then the inserter was sequenced, the results were corresponed to the data from Freeman et al. The recombinant, named pCD mB7.1, was transfected into B7 1 negative melanoma B16(F0) cells by lipofectin mediated gene transfer. The mouse B7 1 expression in modified B16 cells (B16 mB7.1) were detected within at least six months by RT PCR and RNA molecular hybrid consequently. Non adherent splenocytes from non immunized C57BL/6 mice were incubated with non replicating B16 wt, B16 neo as control, or B16 mB7.1 respectively. Then the lymphocytes were tested for cytotoxic activity against B16 wt cells. The results suggested that the costimulation signal of B7 1 is required for the activation of T lymphocytes, the expression of B7 1 by tumor cells is effective at inducing antitumor immune responses.
出处
《武汉大学学报(自然科学版)》
CSCD
1999年第2期225-228,共4页
Journal of Wuhan University(Natural Science Edition)
基金
国家教委基金