异位高表达OCT4的人骨髓间充质干细胞对T淋巴细胞功能的影响  

Effect of Human Bone Marrow Mesenchymal Stem Cells with Ectopic High OCT4 Expression on T Lymphocyte Function

作　　者：郭晓萍[1] 陈燕飞 陈萍 潘劲 应培挺 赵宁[1] 汤永民[1] GUO Xiao-Ping;CHEN Yan-Fei;CHEN Ping;PAN Jin;YING Pei-Ting;ZHAO Ning;TANG Yong-Min(Division/Center of Pediatric Hematology-Oncology,Children’s Hospital,Zhejiang University School of Medicine,The Pediatric Leukemia Diagnostic and Therapeutic Technology Research Center of Zhejiang Province,National Clinical Research Center for Child Health,Hangzhou 310003,Zhejiang Province,China)

机构地区：[1]浙江大学医学院附属儿童医院血液肿瘤科/血液肿瘤中心,浙江省小儿白血病诊治技术研究中心,国家儿童健康与疾病临床医学研究中心,浙江杭州310003

出　　处：《中国实验血液学杂志》2023年第5期1523-1530,共8页Journal of Experimental Hematology

基　　金：浙江省自然科学基金一般项目(LY20H080007);浙江省小儿白血病诊治技术研究中心资助(JBZX-201904)。

摘　　要：目的:探讨异位高表达OCT4的人骨髓间充质干细胞(MSC)在体外对T细胞增殖、活化及分泌功能的影响。方法:分离健康儿童外周血单个核细胞,使用Anti-CD3/CD28单克隆抗体体外激活T淋巴细胞,白介素(IL)2体外刺激培养的T细胞1周。建立异位高表达OCT4的MSC(MSC-OCT4)与活化T细胞的体外共培养体系,收集共培养1周后的上清液,流式细胞仪测定Th1/Th2细胞因子(IL-2、IL-4、IL-6、IL-10、肿瘤坏死因子-α、γ干扰素)水平。收集共培养1周后的淋巴细胞,通过Countstar软件进行计数,用流式细胞仪测定T细胞及活化T细胞亚群的比例后,计算出绝对值,以均数±标准差表示。结果:与对照单独T细胞培养组相比,MSC及MSC-OCT4均能显著抑制CD3^(+)T细胞、CD3^(+)CD4^(+)T细胞和CD3^(+)CD8^(+)T细胞增殖。与MSC相比,MSC-OCT4能更好地抑制CD3^(+)CD8^(+)T细胞增殖(P=0.049),且主要抑制早期T细胞活化。与对照单独T细胞培养组相比,MSC及MSC-OCT4均能显著下调细胞因子IL-2和γ干扰素水平。与T细胞共培养1周后,MSC与MSC-OCT4组细胞因子IL-6水平显著增加。与对照MSC组相比,MSC-OCT4组在共培养1周后活细胞数更高(P=0.019),且能耐受更高浓度丝裂霉素C的抑制增殖作用。结论:MSC及MSC-OCT4在体外均能抑制IL-2刺激的T细胞增殖及活化,过表达OCT4后MSC能更强地抑制CD3^(+)CD8^(+)T细胞增殖,且具有更好的体外增殖能力,可能具有更好更持久地调节Th1/Th2造血因子平衡的作用。Objective:To explore the effect of human bone marrow mesenchymal stem cells(MSCs)with ectopic high OCT4 expression on T-cell proliferation,activation and secretion in vitro.Methods:Peripheral blood mononuclear cells were isolated from healthy children.Anti-CD3 and anti-CD28 monoclonal antibodies were used to activate T lymphocytes,which were stimulated by interleukin(IL)-2 for one week in vitro.Then MSCs with ectopic high OCT4 expression(MSC-OCT4)were co-cultured with activated T lymphocytes.After one week of co-culture,the supernatant was collected and the levels of Th1/Th2 cytokines[IL-2,IL-4,IL-6,IL-10,tumor necrosis factor(TNF)-αand interferon(IFN)-γ]were determined by flow cytometry.The lymphocytes after one week of co-culture were collected and counted by Countstar sof tware.After the proportions of activated/inactivated T cell subsets were determined by flow cytometry,the absolute lymphocyte counts were calculated and expressed as mean±standard deviation.Results:Compared with control T cell alone culture group,the proliferation of CD3^(+)T cells,CD3^(+)CD4^(+)T cells,and CD3^(+)CD8^(+)T cells were significantly inhibited in MSC group and MSC-OCT4 group.Compared with MSC,MSC-OCT4 could inhibit CD3^(+)CD8^(+)T cell proliferation better(P=0.049),and mainly inhibited early T cell activation.Compared with control T cell alone culture group,the levels of IL-2 and INF-γwere significantly down-regulated both in MSC group and MSC-OCT4 group.After co-culture with T cells for one week,the level of IL-6 significantly increased in MSC group and MSC-OCT4 group compared with that before co-culture.Compared with control MSC group,MSC-OCT4 group had higher viable cell numbers after 1 week of co-culture(P=0.019),and could resist the inhibition of proliferation by higher concentration of mitomycin C.Conclusion:Both MSC and MSC-OCT4 can inhibit the proliferation and activation of IL-2-stimulated T cells in vitro.After overexpression of OCT4,MSC has better proliferation ability in vitro and can inhibit the proliferation of

关 键 词：人骨髓间充质干细胞 转录因子 OCT4 免疫调节 细胞因子 

分 类 号：R392.4[医药卫生—免疫学] R457.7[医药卫生—基础医学]