K562白血病株树突状细胞诱导及其抗肿瘤功能的体外研究

Study on the anti-tumor function of K562 leukemia dendritic cell induced by VPA in vitro

目的将K562细胞诱导分化为树状突细胞(DC),并以K562-DC作为刺激细胞诱导细胞毒T淋巴细胞(CTL)反应,观察其体外特异性抗肿瘤效应,对白血病细胞来源DC用于白血病免疫治疗进行初步探讨。方法(1)低浓度丙戊酸钠将K562细胞诱导分化为DC(K562-DC)。(2)DC的鉴定:①形态学:倒置显微镜(Olympus)下观察细胞形态并摄相。②免疫表型:PE结合的鼠抗人CD1a、HLADR、CD83、CD80及CD86单抗用流式细胞仪分析检测细胞表面标志。③功能鉴定:采用同种异体混合淋巴细胞反应。(3)MTT杀伤实验检测DC诱导特异性细胞毒性T淋巴细胞的杀伤活性。结果(1)丙戊酸钠(VPA)培养第7天时,细胞均匀分散,变形,体积增大,数量增多,细胞表面可见多个突起,且具有刺状突起的细胞数量较前增多。(2)通过流式细胞术检测诱导的K562DC表面分子的表达,结果发现丙戊酸钠(VPA)诱导的K562DC各表面标志的表达,与K562细胞相比均明显上调(P<0.05)。(3)丙戊酸钠(VPA)诱导的K562DC具有较强的诱导CTL杀伤肿瘤细胞的能力且随效靶比例增高而增强,并显著高于未负载肿瘤可溶性抗原的DC及单纯T细胞组(P<0.05)。结论以K562细胞为靶细胞,观察VPA对K562细胞诱导向树突状细胞分化作用。进一步的实验证实,VPA可以诱导K562细胞向树突状细胞分化,而且诱导的树突状细胞具有较强的抗原呈递功能和诱导特异性细胞毒性T淋巴细杀伤活性作用。

Objective To investigate the function of dendritic cell （DC） derived from the K562 cells in stimulating the reaction of T-lymph cell （CTL） and its specific effect on tumor resistance, and to evaluate the efficiency of leukemia cell derived DC for the leukemia immunity treatment. Methods （1） K562 cells were induced into DC by lower dose of VPA . （2）Appraisement of DC ① Morphology ： Observing the cells under the converted Olympus and taking pictures. ② Immunophenotype： Using flow cytometer （FCM） to analyze and examine PE combined monoclonal antibody ： CD1a,HLA DR,CD83 ,CD80 and CD86. ③Function appraisal： Adopt homogeneity variant mixed lymphocyte reaction. （3） Use the method of MTT to examine the aetivity of DC in inducing CTL. Results （1） On the seventh day, theeells deformed and dispersed equally, meanwhile, their solidity expands and their quantity manifolds. Many tubers appear on the cell surface and the cells with thorn increased than before. （2） The expression of the specific molecules increased in K562 derived DC compared with K562 cells by using FCM examiantion. （3） Revulsived K562DC has stronger power in inducing CTL to execute tumor cells, the higher of the proportion, the stronger of its function; and. this power is notably higher than unmixed T cell group and DC which has no tumor soluble antigen. Conclusion K562 cells can be induced and differentiated into DC by lower dose of VPA, and these K562 derived DCs have powerful antigen submit function and effect in inducing CTL to kill the tumor cells. words]