摘要
目的探讨溶瘤Ⅱ型单纯疱疹病毒(oHSV2)诱导结肠癌肝转移小鼠模型产生的脾淋巴细胞对结肠癌肺转移瘤生长的影响。方法选择18只6周龄BALB/c雌性小鼠,取对数生长期小鼠结肠癌细胞株CT26细胞,分别接种于小鼠右侧背部(2×10^(5)/只)和脾(1×10^(5)/只),肿瘤细胞通过脾静脉血行转移至肝,构建CT26结肠癌肝转移瘤模型;采用随机数字表法分为oHSV2组和磷酸盐缓冲液(PBS)组,每组9只;oHSV2组采用100μl oHSV2(感染复数为1)进行皮下瘤内多点注射,PBS组采用100μl PBS进行皮下瘤内多点注射;隔1 d注射1次,共6次;采用Kaplan-Meier法分析小鼠存活情况,观察肿瘤生长情况;荷瘤后第20天处死两组结肠癌肝转移模型小鼠,分离其脾淋巴细胞。摸索结肠癌肺转移瘤CT26细胞株最佳接种数量和最适观察时间点;选择9只正常的6周龄BALB/c雌性小鼠,采用随机数字表法分为实验组、阴性对照组和空白对照组,每组3只。实验组小鼠尾静脉同时接种oHSV2诱导结肠癌肝转移小鼠模型产生的脾淋巴细胞(4×10^(7)/只)和CT26细胞(2×10^(5)/只),阴性对照组小鼠尾静脉同时接种相同周龄正常小鼠的脾淋巴细胞(4×10^(7)/只)和CT26细胞(2×10^(5)/只),空白对照组小鼠尾静脉只注射CT26细胞(2×10^(5)/只);荷瘤后第10天全部处死3组小鼠,观察肺转移瘤的生长,小鼠存活情况,观察小鼠各组织病理形态改变。结果结肠癌肝转移小鼠模型中,oHSV2组9只小鼠中7只未发现肝转移灶,2只出现1~2个长径<2 mm肝转移灶;PBS组9只小鼠均出现多发性肝转移灶,肿瘤长径1~10 mm。oHSV2组小鼠总生存优于PBS组小鼠(P<0.001)。结肠癌肺转移瘤小鼠模型中,小鼠尾静脉接种肿瘤细胞的最佳细胞数量为2×10^(5)/只,最佳观察时间点为尾静脉注射后第10天;荷瘤24 d后阴性对照组和空白对照组小鼠均全部死亡,实验组小鼠在第60天仍全部存活,3组间总生存差异有统计学意义(P=0.007);HE染色结果显示,实验组小鼠肺脏组织未见明确肿瘤细胞,阴性对照组和空白对照组的肺脏组织有广泛弥漫的肿瘤细胞。结论oHSV2诱导结肠癌肝转移小鼠模型产生的脾淋巴细胞能够有效抑制CT26小鼠结肠癌肺转移瘤的生长。
Objective To study the effect of splenic lymphocytes isolated from mouse models of colorectal carcinoma with liver metastases induced by oncolytic herpes simplex virus typeⅡ(oHSV2)on the growth of pulmonary metastases of colorectal carcinoma.Methods A total of 186-week-old BALB/c female mice were selected,colorectal carcinoma cell line CT26 of mice in logarithmic phase was inoculated at the right back(2×10^(5) per mouse)and spleen(1×10^(5) per mouse)of mice,and tumor cells had hematogenous metastasis to liver through splenic vein.CT26 colorectal carcinoma with liver metastases model was constructed.All mice were respectively divided into oHSV2 group and phosphatic buffered saline(PBS)group,9 mice in each group according to the random number table method.Mice in oHSV2 group were treated with subcutaneous intratumoral multi-point injection of 100μl oHSV2(the multiplicity of infection was 1)for 6 cycles,while mice in PBS group were treated with subcutaneous intratumoral multi-point injection of 100μl PBS for 6 cycles in total,once injection every other day;the survival of mice was analyzed by using Kaplan-Meier method and tumor growth was observed.The mice of both groups in mouse models of colorectal carcinoma with liver metastases were killed on day 20 and their splenic lymphocytes were isolated.After investigation of the most suitable inoculation number and the optimal observation time of colorectal carcinoma with pulmonary metastases CT26 cell lines,96-week-old BALB/c female mice were divided into the experimental group,the negative control group and the blank control group according to the random number table method,with 3 mice in each group.Mice in the experimental group were injected with splenic lymphocytes(4×10^(7) per mouse)and CT26 cells(2×10^(5) per mouse)isolated from mouse models induced by oHSV2 via the tail vein,mice in the negative control group were injected with splenic lymphocytes(4×10^(7) per mouse)and CT26 cells(2×10^(5) per mouse)isolated from normal mice with same weeks old via the tail vein,and mice in the blank control group were injected with only CT26 cells(2×10^(5) per mouse)via the tail vein.The above 3 groups were executed on day 10 after inoculation,and tumor growth,histopathological changes of mice were also observed;the survival of mice was analyzed by using Kaplan-Meier method.Results In models of colorectal carcinoma with liver metastases,liver metastases lesions were not detected in 7 mice and 2 mice had 1-2 liver metastases lesions with long diameter less than 2 mm of oHSV2 group;in PBS group,9 mice all had multiple liver metastases lesions with tumor long diameter ranging from 1 to 10 mm.And mice in oHSV2 group survived much longer than that of mice in PBS group(P<0.001).In models of pulmonary metastases,the optimal number of CT26 cells in mouse tail vein was 2×10^(5) per mouse;the best observation time point was day 10 after tail vein injection.On day 24 after inoculation,all mice in the negative control group and the blank control group died,while mice in the experimental group all survived on day 60,and the difference of the overall survival in the above 3 groups was statistically significant(P=0.007).HE staining results showed that the lung tissues of the experimental group did not show clear tumor cells,whereas the lung tissues of the negative control group and the blank control group showed extensive diffuse tumor cells.Conclusions Splenic lymphocytes produced by oHSV2 induction in mouse models of colorectal carcinoma with liver metastases can effectively inhibit the development of pulmonary metastases in colorectal carcinoma CT26 cell of mice.
作者
邹灵婕
张文
宋艳
刘尚梅
Zou Lingjie;Zhang Wen;Song Yan;Liu Shangmei(Department of Pathology,National Cancer Center/National Clinical Research Center for Cancer/Cancer Hospital,Chinese Academy of Medical Sciences and Peking Union Medical College,Beijing 100021,China;Department of Immunology,National Cancer Center/National Clinical Research Center for Cancer/Cancer Hospital,Chinese Academy of Medical Sciences and Peking Union Medical College,Beijing 100021,China)
出处
《肿瘤研究与临床》
CAS
2022年第5期321-325,共5页
Cancer Research and Clinic
基金
中国医学科学院医学与健康科技创新工程项目(2017-I2M-3-020)。
关键词
溶瘤病毒
结肠肿瘤
转移瘤
淋巴细胞
Oncolytic viruses
Colonic neoplasms
Neoplasm metastasis
Lymphocytes