青蒿琥酯诱导凋亡的U937细胞负载树突状细胞后的免疫应答_医学论文.docVIP

青蒿琥酯诱导凋亡的U937细胞负载树突状细胞后的免疫应答_医学论文 青蒿琥酯诱导凋亡的U937细胞负载树突状细胞后的免疫应答_医学论文 作者：郑智茵，王建峰，胡致平，沈建平，高瑞兰，叶宝东，林圣云，沈一平【摘要】 本研究探讨凋亡白血病细胞负载的树突状细胞（dendritic cells, DC）能否诱导特异性细胞毒T淋巴细胞反应。 用青蒿琥酯诱导U937细胞凋亡, 从正常人外周血单个核细胞诱生DC，将凋亡U937细胞负载DC, 并添加TNF诱导DC成熟, 然后与自体T淋巴细胞共育,并联合 IL以诱生细胞毒性T淋巴细胞（CTL）；应用流式细胞术分析DC和T细胞免疫表型, 采用Dextran内吞试验检测DC的抗原摄取能力；用ELISA法检测IL的产生；采用MTT法检测凋亡U937细胞负载及未负载DC诱导的自体T淋巴细胞对不同的靶细胞（U937细胞和NB4细胞）的杀伤效应。结果表明：青蒿琥酯1 μg/ml作用于U937细胞48小时后, U937细胞的凋亡率达51.52%, DC在未成熟阶段时吞噬Dextran的能力最强，负载凋亡U937细胞之后并不能促使未成熟DC（iDC）分化为成熟DC（mDC）。凋亡U937细胞负载后的iDC在TNF的作用下能够成为mDC。与未负载的mDC相比, 凋亡U937细胞负载后的mDC（mDC）分泌IL水平明显增高, 而且由其所激发和扩增的T细胞以CD8+的T细胞为主。细胞毒性实验显示: mDC诱导的T细胞的对U937细胞的杀伤显著超过对NB4细胞的杀伤。结论： mDC能有效地诱导特异性抗白血病效应。 【关键词】 白血病 Immune Response of Dendritic Cells Capturing Antigens from Apoptotic U937 Cells Induced by Artesunate Abstract The objective of study was to investigate whether U937 cellsm peripheral blood mononuclear cells of health donors were loaded with apoptotic U937 cells, and induced to maturation in the presence of TNFα. Matured DCs were cocultured with autologous Tlymphocytes, and combined with IL2 in order to induce the leukemiaspecific CTL. The phenotypes of DCs and T lymphocytes were tested by flow cytometry. The ability of DC capturing antigens was measured by DextranFITC endocytosis. The IL12p70 level was assayed by ELISA kit. The proliferation of CTL and CTL activity were measured by MTT assay. The results showed that the apoptotic rate of the U937 cells was 51.2% when U937 cells were induced by 1 μg/ml ART for 48 hours in vitro. DCs had the most powerful ability of endocytosis in its immature phase. Apoptotic U937 cells could not induce the features of DC maturation, and apoptotic U937 cellpulsed immature DCs could be matured with TNFα. The IL12p70 level secreded by apoptotic U937 cellloaded mature DCs（mDC） was higher than that of nonation of autologous T lymphocytes cocultured with mDCApoU937 was significantly remarkable and the content of CD8+ CTL was significantly higher in comp