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目的研究脾内注射携带人白细胞介素2(hIL-2)与鼠白细胞介素12(mIL1-2)融合基因的逆转录病毒包装细胞株对大鼠诱发肝细胞癌的生长抑制作用。方法构建携带hIL-2和mIL-12融合基因的逆转录病毒载体GCIL1-2EIL-2PN,转染包装细胞PA317后,分别在大鼠肝癌诱发后90天(早期治疗组)及105天(晚期治疗组)进行脾内注射治疗,观察大鼠生存时间及毒性反应。ELISA法检测大鼠血清IL-12和IL-2浓度,放射性活度测定法检测脾脏NK细胞活性。结果IL-12+IL-2联合基因治疗组中,早期治疗及晚期治疗的生存时间分别为188.1±14.2天及168.5±13.6天;IL-12基因治疗组分别为168.2±13.4天及149.1±13.8天(与联合基因治疗组相比,P<0.01);IL-2基因治疗组分别为145.9±11.3天及131.1±10.5天(P<0.01)。3个治疗组中,早期治疗的平均生存时间均长于晚期治疗(P<0.01)。IL-12+IL-2联合基因治疗组中,早期治疗后长期存活率(≥240天)为25%(5/20),治疗后5天肝癌组织中浸润的淋巴细胞明显增多,治疗后2个月血清hIL-2及mIL-12仍维持在较高水平。各基因治疗组NK细胞活性较对照组显著增高(P<0.01)。IL-12+IL-2联合基因组NK细胞活性明显高于单基因治疗组(P<0.01)。结论脾内直接注射携带hIL-2和(或)mIL-12基因的逆转录包装细胞株可显著提高NK细胞的活性,抑制?
Objective To investigate the inhibitory effect of retrovirus packaging cell line transfected with human interleukin 2 (hIL-2) and murine interleukin-12 (mIL1-2) fusion gene on the growth of rat hepatocellular carcinoma induced by spleen injection. Methods The retroviral vector GCIL1-2EIL-2PN carrying the fusion gene of hIL-2 and mIL-12 was constructed and transfected into PA317 cells. The cells were treated for 90 days (early treatment group) and 105 days (late treatment group) Group) for spleen injection therapy, to observe the survival time and toxicity in rats. Serum levels of IL-12 and IL-2 were detected by ELISA, and activity of NK cells in spleen was measured by radioactivity assay. Results The survival time of early treatment and late treatment of IL-12 + IL-2 combined gene therapy group was 188.1 ± 14.2 days and 168.5 ± 13.6 days, respectively. The IL-12 gene therapy group was 168.2 ± 13.4 days and 149.1 ± 13.8 Day (P <0.01 compared with the combined gene therapy group); IL-2 gene therapy group was 145.9 ± 11.3 days and 131.1 ± 10.5 days (P <0.01), respectively. Among the three treatment groups, the average survival time of early treatment was longer than that of advanced treatment (P <0.01). Long-term survival (≥240 days) after initial treatment was 25% (5/20) in IL-12 + IL-2 combined gene therapy group. The number of infiltrating lymphocytes in HCC was significantly increased 5 days after treatment Month serum hIL-2 and mIL-12 remained at a high level. The NK cell activity of each gene therapy group was significantly higher than that of the control group (P <0.01). IL-12 + IL-2 combined with genomic NK cell activity was significantly higher than the single-gene therapy group (P <0.01). Conclusion The direct injection of the retroviral packaging cell line carrying hIL-2 and / or mIL-12 gene in spleen can significantly increase the activity of NK cells and inhibit it.