摘要
目的:构建BALB/c小鼠原位肝癌模型,分离肿瘤小鼠体内的CD4^+CD25^+调节性T细胞(Tregs)和树突状细胞(DC),研究Tregs对DC功能的影响。方法:采用肝癌细胞悬液直接接种BALB/c小鼠肝脏建立原位肝癌模型。模型制作成功后25d处死动物,无菌取出脾脏,用免疫磁珠分选法分离纯化肝癌小鼠脾脏中的Tregs、CD4^+CD25^-效应T细胞和DC。Tregs和效应T细胞共培养,CCK8法检测Tregs对效应T细胞增殖情况的影响。DC和Tregs直接接触共培养,用流式细胞仪检测DC表面协同刺激分子CD80/CD86表达的变化,ELISA法检测培养上清液中TNF-α、IL-12的分泌。结果:成功构建了小鼠原位肝癌模型,并分离出高纯度的Tregs、效应T细胞和DC。Tregs在体外能抑制CD4^+CD25^-效应T细胞的增殖。DC和Tregs共培养时,Tregs会下调DC表面协同刺激分子CD80/CD86的表达,并抑制DC分泌IL-12和TNF-α。结论:肝癌小鼠模型中Tregs能抑制DC的功能,从而抑制机体的抗肿瘤免疫,针对Tregs进行肿瘤免疫治疗是肿瘤治疗的新策略。
Objective： To investigate how CD4~＋CD25~＋ regulatory T cells（Tregs）impacted on dendritic cells which were separated from BALB/c mice with hepatocellular carcinoma. Methods： BALB/c mice with hepatocellular carcinoma were established in laboratory. Twenty-five days after the models, the tumor-bearing mice were sacrificed and fresh spleen tissues were separated. Tregs, CD4~＋CD25^-T cells and dendritic cells were separated from the mice＇s spleen and purified by immuno-magnetic beads according to the manufacturers＇ instructions. Tregs were co-cultured with CD4~＋CD25^-T cells and the proliferation experiment was measured with CCK8. Tregs were directly co-cultured with dendritic cells and then the expression of CD80/CD86 on dendritic cells was detected by flow cytometry. The levels of TNF-α and IL-12 in supernatants were detected by ELISA. Results： We successfully established the tumor model of hepatocellular carcinoma and obtained high purity of Tregs, CD4~＋CD25^-T cells and dendritic cells. Tregs could suppress the proliferation of CD4~＋CD25^-T cells in vitro. Tregs could down-regulate the expression of CD80/CD86 on dendritic cells and inhibit the expression of TNF-α and IL-12. Conclusion： Tregs play an important role in the establishment and persistence of tumor immune suppression. Tregs can inhibit the function of dendritic cells in tumor-bearing mice. So Tregs may be a promising therapeutic target for cancer.
作者
杜勇
黄智铭
林秀清
陈新
DU Yong;HUANG Zhiming;LIN Xiuqing;CHEN Xin(Department of Pediatrics, the First Affliated Hospital of Wen-zhou Medical University, Wenzhou, 325015;Department of Gastroenterology and Hepatology, the First Affli-ated Hospital of Wenzhou Medical University, Wenzhou, 325015)
出处
《温州医科大学学报》
CAS
2017年第10期718-722,共5页
Journal of Wenzhou Medical University
基金
浙江省自然科学基金资助项目(Y2090660)
