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The activity of interleukine 2(IL-2) in culture supernatants of lymphokine-activated killer(LAK)cellsand tumor infiltrating lymphocytes (TIL)as well as cytotoxicity of LAK cells on cultured leukemic cells were detected by MTT colorimetry. The results showed that high activity of IL-2 in culture supernatants of LAK and TIL cells was found as well as it could be used for culture of IL-2 edpendent cell lines and clinic experiment. The significant cytotoxicity of LAK cells on... The activity of interleukine 2(IL-2) in culture supernatants of lymphokine-activated killer(LAK)cellsand tumor infiltrating lymphocytes (TIL)as well as cytotoxicity of LAK cells on cultured leukemic cells were detected by MTT colorimetry. The results showed that high activity of IL-2 in culture supernatants of LAK and TIL cells was found as well as it could be used for culture of IL-2 edpendent cell lines and clinic experiment. The significant cytotoxicity of LAK cells on leukemic cell lines could be found in vitro ,which was consistent with the ratio of effector cells to target cells. The number of living leukemic cells is consistently related with the concentration of formazan metabolite of MTT. It suggested that the numbers of living cells and cytotoxicity of LAK cells could be correctly estimated by the determina-tion of OD value. 本文采用MTT比色法测定了LAK和TIL细胞培养上清IL-2活性及LAK细胞介导对体外培养的白血病细胞株的杀伤活性。结果表明LAK细胞和TIL细胞培养上清有较高的IL-2活性,可用于支持培养IL-2依赖株的生长;LAK细胞对体外培养的白血病细胞株有明显的杀伤作用。其杀伤效应随效靶比例增加而增高;同时亦证明:活的白血病细胞数与MTT甲赞形成产物呈直线相关性,提示通过MTT法测定OD值能较好地反映活细胞数目及细胞毒效应。 Tumor-infiltrating lymphocytes (TIL) isolated from two renal cell carcinoma were cultured in vitro in the presence of rh IL-2.The results showed that TIL was expanded 32-203 times and exhibited specific strong cytotoxic activity against autologous fresh renal tumor target cells (up to 53% and 640%).Phenotype characterization of the preparations showed that the CD3,CD4 and CD8 phenotype expressions of non-induced TIL cells did not change in the process of cultivation. Whereas,the... Tumor-infiltrating lymphocytes (TIL) isolated from two renal cell carcinoma were cultured in vitro in the presence of rh IL-2.The results showed that TIL was expanded 32-203 times and exhibited specific strong cytotoxic activity against autologous fresh renal tumor target cells (up to 53% and 640%).Phenotype characterization of the preparations showed that the CD3,CD4 and CD8 phenotype expressions of non-induced TIL cells did not change in the process of cultivation. Whereas,the CD3 expression and the CD/CD8ratio increased significantly to 95% and 1.65 respectively in the culture of activated TIL at 32th day. 对两例原发性肾癌患者手术切除肿瘤组织中肿瘤浸润性淋巴细胞(TIL)进行了体外分离与培养。结果表明:TIL体外扩增倍数分别达32~203倍,对自体肿瘤靶细胞的最高杀伤活性达53%和64%,且呈现一定的靶细胞特异性。免疫组化分析结果:未经激活的TIL细胞其膜抗原(CD3,CD4,CD8)的表达动态变化不大,但经IL-2激活的TIL细胞随着培养无数的增加,其CD3细胞数比例及CD4/CD8比值上升明显,在培养至32天时分别达95%和1.65。 In order to investigate the functions of tumor cells and tumorinfiltrating lymphocytes,IL-2 DNA liposome were directly injected into B16F10 melanoma mass; 10 days later,Northern blots analysis confirmed IL-2 mRNA expression both in tumor cells and TIL. IL2 could be detected in the supernatants of freshly isolated tumor cells after G418 resistant selection. Higher expression of MHC class Ⅰ(H-2b) molecules on tumor cells and much more LAF-1 molecules on TIL cells could be observed.... In order to investigate the functions of tumor cells and tumorinfiltrating lymphocytes,IL-2 DNA liposome were directly injected into B16F10 melanoma mass; 10 days later,Northern blots analysis confirmed IL-2 mRNA expression both in tumor cells and TIL. IL2 could be detected in the supernatants of freshly isolated tumor cells after G418 resistant selection. Higher expression of MHC class Ⅰ(H-2b) molecules on tumor cells and much more LAF-1 molecules on TIL cells could be observed. The cytotoxicity of TIL to B16F10 cells was augmented significantly after intratumoral injection of IL-2 DNA liposome.These results show that liposome can mediated IL-2 gene transfer into tumor cells and TIL cells in situ,then increase the immunogenicity of the tumor cells and induce local tumor specific immune responses. 将脂质体包裹的IL-2基因直接注射至B16F10黑色素瘤瘤体内,研究肿瘤细胞和肿瘤浸润性淋巴细胞(TIL)的功能变化。10d后,Northernblot鉴定显示瘤体内注射IL-2基因后肿瘤细胞及TIL中IL-2mRNA表达阳性;经G418筛选后的肿瘤细胞培养上清中可检测出IL-2;肿瘤细胞表面表达了较高水平的MHCⅠ类抗原(H-2K ̄bD ̄b);TIL表面粘附分子LAF-1表达也明显高于对照组,其杀伤B16细胞的活性明显增强。结果表明瘤体内注射脂质体包裹的IL-2基因后,可在肿瘤原位将基因转移至肿瘤细胞及TIL中,提高肿瘤细胞的MHCⅠ类抗原的表达,增强肿瘤细胞的免疫原性,并通过提高TIL粘附分子的表达,共同促进TIL的肿瘤特异性杀伤功能。
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