气液交互法培养小鼠来源的肿瘤类器官的建立及优化

Establishment and optimization of air-liquid interface method cultured mouse-derived tumor organoids

目的构建一种不需要人工重建就可以保留免疫微环境中各类免疫细胞的小鼠来源的肿瘤类器官的气液交互法。方法利用气液交互法建立并培养小鼠来源的肿瘤类器官;用苏木精-伊红(HE)染色分析肿瘤类器官的组织学形态;用流式细胞测量术分析肿瘤类器官中各类免疫细胞的比例;用白细胞介素-2(IL-2)维持肿瘤类器官中T淋巴细胞的数量。结果小鼠CT26结肠癌、Lewis肺癌和MC38结肠癌这3种肿瘤类型的肿瘤类器官都分别保留了原始肿瘤的组织形态;与小鼠CT26结肠癌原始肿瘤组织相比,肿瘤类器官中CD4^(+)T细胞、CD8^(+)T细胞和B细胞的比例基本保持不变,巨噬细胞比例从66.7%减少到44.3%,树突状细胞的比例从66.5%减少到7.25%;与小鼠Lewis肺癌原始肿瘤组织相比,肿瘤类器官中CD4^(+)T细胞、巨噬细胞、树突状细胞和B细胞的比例基本保持不变,CD8^(+)T细胞的比例从5.6%减少到1.95%;与小鼠MC38结肠癌肿瘤组织相比,肿瘤类器官中CD4^(+)T细胞、CD8^(+)T细胞和B细胞的比例基本保持不变,巨噬细胞的比例从33.5%减少到17%,树突状细胞的比例从46.8%减少到1.2%;IL-2可以增加3种肿瘤类型来源的肿瘤类器官中肿瘤浸润T淋巴细胞的数量。结论小鼠肿瘤类器官保留了原始肿瘤的组织学形态,一定程度上保留了各类免疫细胞。IL-2可以增加T淋巴细胞的数量。

Objective To construct a mouse-derived tumor organoid that can retain various types of immune cells in the immune microenvironment without artificial reconstruction.Methods The mouse-derived tumor organoids were cultured and constructed by air-liquid interface method.The histological morphology of the tumor organoids was analyzed by hematoxylin and eosin(HE).The proportion of various types of immune cells in the tumor organoids was analyzed by flow cytometry.The interleukin-2(IL-2)was used to maintain the number of T lymphocytes in tumor organoids.Results The tumor organoids from three tumor types of mouse CT26 colon cancer,Lewis lung cancer and MC38 colon cancer all retained the histomorphology of the original tumor.Compared with mouse CT26 colon cancer tumor tissues,the proportion of CD4^(+)T cells,CD8^(+)T cells and B cellsin tumor organoids didn't show significant change and the proportion of macrophages decreased from 66.7%to 44.3%.The proportion of dendritic cells decreased from 66.5%to 7.25%.Compared with mouse Lewis lung cancer tumor tissue,the proportion of CD4^(+)T cells,macrophages,dendritic cells and B cells in tumor organoids remained unchanged and the proportion of CD8^(+)T cells decreased from 5.6%to 1.95%.Compared with mouse MC38 colon cancer tumor tissue,the proportion of CD4^(+)T cells,CD8^(+)T cells and B cells in tumor organoids remained unchanged,the proportion of macrophages decreased from 33.5%to 17%,and the proportion of dendritic cells decreased from 46.8%to 1.2%.IL-2 could increase the tumor-infiltrating T lymphocytes in tumor organoids derived from three tumor types.Conclusions Mouse tumor organoids retain the histological morphology of the original tumor and the various types of immune cells are retained in a certain extent.IL-2 can increase the quantity of T lymphocytes.