论文部分内容阅读
目的:观察表达CD40L的小鼠结肠癌colon26瘤苗体内外对DC活性的影响。方法:以脂质体法将CD40L基因转染colon26细胞获得稳定表达CD40L的colon26/CD40L瘤苗。colon26/CD40L细胞与小鼠骨髓来源DC共培养,流式细胞术检测DC表型变化,RT-PCR法检测细胞因子基因P19、P35、P40、IL-18和IFN-γ的表达,ELISA法检测共培养上清中IL-12、IL-23和IFN-γ的水平。BALB/c小鼠皮下注射colon26细胞制备荷瘤小鼠模型,colon26/CD40L致敏DC治疗荷结肠癌小鼠,ELISA法检测荷瘤小鼠外周血中IL-12、IL-23、IFN-γ、IL-10和TGF-β的水平,H-E染色观察肝、脾和肿瘤组织的病理学变化。结果:成功获得高表达CD40L的丝裂霉素(MMC)处理的colon26/CD40L瘤苗。DC与colon26/CD40L瘤苗共培养后,DC表面共刺激分子CD80、CD86、MHCⅠ和MHCⅡ表达增高(P<0.01);共培养体系中可检测到P19、P35、P40、IL-18和IFN-γmRNA的表达,而在其他组未检测到上述基因的表达;共培养细胞上清中有较高水平的IL-12、IL-23和IFN-γ(P<0.01)。co-lon26/CD40L瘤苗致敏DC治疗组与colon26/CD40L瘤苗治疗组、DC治疗组比较,小鼠移植瘤的体积和质量明显减小和减少(P<0.05);小鼠外周血IL-12、IL-23和IFN-γ明显升高(P<0.01),IL-10和TGF-β明显降低(P<0.01);小鼠移植瘤组织病理变化显著减轻。结论:表达CD40L的丝裂霉素(MMC)处理的结肠癌瘤苗可促进DC的成熟,刺激DC分泌细胞因子,增强DC体内外活性,从而增强治疗体系的抗肿瘤免疫效应。
OBJECTIVE: To observe the effect of colon26 colon of CD40L-expressing mouse colon carcinoma on DC activity in vitro and in vivo. Methods: Colon26 / CD40L cell line stably expressing CD40L was obtained by transfecting CD40L gene into colon26 cells by liposome. Colon26 / CD40L cells were co-cultured with mouse bone marrow-derived DCs, the DC phenotypes were detected by flow cytometry, the expression of cytokines P19, P35, P40, IL-18 and IFN-γ were detected by RT- Cultures supernatants IL-12, IL-23 and IFN-γ levels. BALB / c mice were injected subcutaneously with colon26 cells to prepare tumor-bearing mouse model. Colon26 / CD40L-sensitized DCs were used to treat colon cancer mice. The levels of IL-12, IL-23 and IFN- , IL-10 and TGF-β, and the pathological changes of liver, spleen and tumor were observed by HE staining. Results: The colon26 / CD40L vaccine treated with mitomycin (MMC) highly expressing CD40L was successfully obtained. After coculturing DCs with colon26 / CD40L tumor cells, the expressions of CD80, CD86, MHCⅠ and MHCⅡ on the surface of DCs were increased (P <0.01); P19, P35, P40, IL- γ mRNA expression was detected in the other groups, while the levels of IL-12, IL-23 and IFN-γ in the co-cultured cells were higher than those in other groups (P <0.01). The volume and the quality of the xenografts of mice inoculated with co-lon26 / CD40L vaccine were significantly decreased and decreased compared with those in the colon26 / CD40L and DC treated groups (P <0.05) -12, IL-23 and IFN-γ were significantly increased (P <0.01), while IL-10 and TGF-β were significantly decreased (P <0.01). The pathological changes in the transplanted tumor of mice were significantly reduced. CONCLUSION: MMC vaccine expressing CD40L can promote the maturation of DC, stimulate the secretion of cytokines by DC, enhance the activity of DC in vitro and in vivo, and enhance the anti-tumor immune effect of the therapeutic system.