上海交通大学学报（医学版）

上海交通大学学报(医学版) ›› 2020, Vol. 40 ›› Issue (06): 728-735.doi: 10.3969/j.issn.1674-8115.2020.06.005

• 论著·基础研究 • 上一篇    下一篇

OX40/FcγR多基因人源化小鼠模型的构建及在激动型OX40抗体研究中应用的验证

刘明东*，刘小波*，赵英杰，张 燕，张慧慧，李福彬   

  1. 上海交通大学基础医学院免疫学与微生物学系，上海市免疫学研究所，上海200025
  • 出版日期:2020-06-28 发布日期:2020-06-28
  • 通讯作者: 李福彬，电子信箱：fubin.li@sjtu.edu.cn。
  • 作者简介:刘明东（1992—），男，硕士生；电子信箱：kai123lmd@163.com。刘小波（1982—），男，博士后；电子信箱：iamliuxiaobo@163.com。*为共同第一作者。
  • 基金资助:
    国家自然科学基金（31422020，31700806，31600704）；上海市科学技术委员会项目（18140902600，19431902900，15ZR1436400）；上海高水平地方高校创新团队（SSMU-2DCX20180100）。

Establishment and validation of OX40/FcγR-humanized mice for the study of agonistic anti-OX40 antibody

LIU Ming-dong*, LIU Xiao-bo*, ZHAO Ying-jie, ZHANG Yan, ZHANG Hui-hui, LI Fu-bin   

  1. Department of Immunology and Microbiology, Shanghai Jiao Tong University College of Basic Medicine Sciences; Shanghai Institute of Immunology, Shanghai 200025, China
  • Online:2020-06-28 Published:2020-06-28
  • Supported by:
    National Natural Science Foundation of China (31422020, 31700806, 31600704); Shanghai Science and Technology Commission Project (18140902600, 19431902900, 15ZR1436400); Innovative Research Team of High-level Local Universities in Shanghai (SSMU-2DCX20180100).

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摘要： 目的·建立一种能够快速获得可用于激动型抗体活性评估的OX40/FcγR [肿瘤坏死因子受体超家族成员4（tumor necrosis factor receptor superfamily member 4，OX40） /Fcγ受体（Fcγ receptor，FcγR） ]多基因人源化小鼠模型的方法。方法·将表达人源OX40分子的小鼠骨髓细胞与表达人源FcγR的小鼠骨髓细胞等比例混合后，通过尾静脉注射到经过辐照的野生型小鼠体内，构建骨髓嵌合小鼠。通过流式细胞术检测骨髓嵌合小鼠中OX40/FcγR人源分子的表达情况，确认免疫系统的重建效率。对构建成功的骨髓嵌合小鼠，使用流式细胞术评估人类抗人OX40单克隆抗体对CD4+、CD8+ T细胞的免疫激活活性。2组间比较采用t检验，多组间比较采用单因素方差分析。结果·流式细胞术的结果显示，骨髓嵌合小鼠脾脏、外周血和淋巴结中的B淋巴细胞、髓系细胞均可以表达高水平的人源FcγR分子（均P<0.05）；骨髓嵌合小鼠T淋巴细胞在体外激活后有明显的人源OX40分子的表达（P<0.05）。使用人类激动型抗人OX40抗体对骨髓嵌合小鼠进行处理显示，人类激动型抗人OX40抗体可以明显增强小鼠体内T细胞的γ-干扰素（interferon γ，IFN-γ）表达量，和CD4+ T细胞上OX40分子的表达(均P<0.05)。结论·以表达人源OX40分子的小鼠与表达人源FcγR的小鼠的骨髓细胞为基础构建的骨髓嵌合小鼠模型能够同时表达人源OX40和FcγR分子，可用于评估人类激动型抗人OX40抗体的体内活性。

关键词: 肿瘤坏死因子受体超家族成员4, Fcγ受体, 骨髓嵌合小鼠, 多基因人源化小鼠

Abstract:

Objective · To establish a rapid method to evaluate the activity of agonistic antibody using OX40 (tumor necrosis factor receptor superfamily member 4)/FcγR (Fcγ receptor)-humanized mice. Methods · Bone marrow cells from OX40-humanized mice and FcγR-humanized mice were collected and mixed with equal ratio. Then the mixed bone marrow cells were administrated into irradiated wild-type mice through the tail veins. The reconstruction efficiency of the immune system was confirmed by detecting the expression of hOX40 and hFcγR in the immune cells of chimera mice. After the chimera mice were generated successfully, they were used to evaluate the immunostimulatory activity of anti-hOX40 antibodies to CD4+ or CD8+ T cells. The results of flow cytometry were statistically analyzed. The unpaired t-test was used to compare the means between the two groups, and one-way ANOVA was used to compare the means between multiple groups. Results · Flow cytometry analysis showed that wild-type recipient mice were efficiently reconstituted with hFcγR expressing cells and hOX40 expressing cells to generate OX40/FcγR-humanized bone marrow chimera mice. In these mice, B cells and myeloid cells expressed hFcγRs (P<0.05), and T cells expressed hOX40 upon in vitro stimulation (P<0.05). When these mice were used to evaluate the immunostimulatory activity of anti-hOX40 antibody, significant expressions of IFN-γ and hOX40 were observed (P<0.05). Conclusion · OX40/FcγR-humanized bone marrow chimera mice are generated based on hFcγR expressing cells and hOX40 expressing cells, suggesting a rapid method to build a mouse model with both hFcγR and hOX40 expression. These mice are suitable for evaluating the immunostimulatory activity of agonistic human anti-hOX40 antibodies.

Key words: tumor necrosis factor receptor superfamily member 4 (OX40), Fcγ receptor (FcγR), bone marrow chimera mice, polygenic humanized mice

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