刚地弓形虫排泄分泌抗原刺激NK细胞过继转输对小鼠B16F10黑色素瘤生长的抑制作用

目的观察刚地弓形虫排泄分泌抗原(TgESA)刺激后的NK细胞对小鼠B16F10黑色素瘤生长的作用。方法取弓形虫RH株体外培养12h后,从培养上清收集获得弓形虫ESA。取10只C57BL/6小鼠随机分为2组,每组5只,每只小鼠右腋窝皮下接种B16F10黑素瘤细胞2×10^5个。接种后第7天,随机取1组小鼠腹腔注射100μlTgESA,另1组注射等量的PBS。接种后第14天,无痛处死小鼠,无菌条件下制备小鼠脾细胞悬液,分离2组荷瘤鼠的NK细胞,分别记为NKB16F10和NKESA,用于后续过继转输实验。取30只C57BL/6小鼠随机分为对照组、NKB16F10组和NKESA组,每组10只。3组小鼠均右腋窝皮下接种B16F10细胞2×10^5个/鼠。其中NKB16F10组和NKESA组小鼠同时分别尾静脉注射2×10^5个NKB16F10和NKESA。观察瘤体生长情况以及各组小鼠死亡数和死亡时间,共观察35 d。结果 NK细胞过继转输后,NKd、组和“心間。组小鼠平均出瘤时间分别为(14.70 ±0.95)、(12.60 ± 0.70) d,均晚于对照组的(8.50 ± 0.85) d (P < 0.05).3组小鼠出瘤后,瘤体均不断增长,至第35天,NKB16F10组和NKESA组的平均瘤体面积分别为(686.53 ± 17.84)和(577.79 ± 49.70) mm^2,均小于对照组的(787.84 ± 19.94) mm^2 (P< 0.05 ).对照组、NKB16F10组和NKESA组小鼠分别于接种B16F10细胞后第24、27和30天开始出现死亡,至第35天,存活的小鼠数分别为3、4和6只。结论弓形虫ESA刺激的NK细胞过继转输小鼠后可较为明显地抑制B16F10黑色素瘤瘤体的生长。

Therapeutic effect of NK cells stimulated by Toxoplasma gondii excretory/secretory antigens on B16F10 melanoma in mice

Objective To observe the therapeutic effect of natural killer (NK) cells stimulated by Toxoplasma gondii excretory/secretory antigens (7gESA) on the growth of B16F10 melanoma in mice. Methods 7gESA was obtained from the culture of T. gondii tachyzoites in vitro for 12 hours. The tumor-bearing animal models were established by direct injection of melanoma B16F10 cells into the right axillary of ten mice (2×10^5/mouse). Five of the melanoma-inoculated mice were intraperitoneally treated with TgESA (100μl/mouse) 7 days after inoculation of B16F10 cells, the rest 5 mice were treated with PBS only as control. One week after the treatment, all mice were euthanized, and the NK cells were isolated from the splenocytes of two groups (named NKBi6fio and NKexa). Total 30 naive C57BL/6 mice were randomly divided into 3 groups with 10 mice each group. Each mouse was inoculated with 2 × 105 B16F10 cells subcutaneously into axillary. One week after melanoma inoculation, one group of mice were passive transferred with 2 × 10^5 of NKBi6fio cells through tail vein injection (group of NKBESA)- Another group of mice were transferred with the same number of NKESA cells (group of NKpxa). The third group of mice were given with PBS only as control. After passive transfer, the tumor growth, mortality and survival rate in the transferred mice were observed and recorded for 35 days. Results The visible time of tumor in mice of NKesa group (14.70 ± 0.95) d] was significantly longer than that in NKBi6fio group (12.60 ± 0.70) d] and in control group (8.50 ± 0.85) d](P < 0.05). The tumor size in NKESA group (577.79 ± 49.70) mm^2] was also significantly smaller than that in NKBi6Fio group (686.53 ± 17.84) mm^2] and in control group (787.84 ± 19.94) mm^2] at D35. The delayed appearance of tumor and smaller tumor size in group of NKB]6F10 was also significant compared to control group. Mice began to die at D24 for control group, D27 for NKB|6F10 group and D30 for NKjsa group after tumor inoculation, and the number of mice remain alive was 3, 4 and 6 respectively at terminal time D35 with significant difference between NKESA group and control group. Conclusion Passive transfer of NK cells stimulated by TgESA conferred therapeutic effect on B16F10 melanoma in recipient mice.