脐带来源间充质干细胞对人B细胞系细胞的调控  

Regulation effects of umbilical cord derived mesenchymal stem cells on human B cell lines

作　　者：徐祎欣[1] 樊竑冶[1] 董冠军[1] 刘飞[1] 任德善 侯亚义[1] 

机构地区：[1]南京大学医学院和生物医药技术国家重点实验室,南京210093

出　　处：《中国免疫学杂志》2014年第1期40-47,共8页Chinese Journal of Immunology

基　　金：江苏省教育厅2011年高校科研成果产业化推进项目(JHB2011-1);第53批中国博士后科学基金面上资助项目(2013M531331)资助

摘　　要：目的:探讨脐带来源的MSC对B细胞生长、活化、功能、信号通路的影响,为阐明MSC对B细胞的调控以及作用机制提供理论基础。方法:以人非Hodgkin淋巴瘤B细胞系Daudi、Namalwa和Raji为研究模型,在CpG刺激和不刺激下,与脐带MSC分别共培养3天后,收集悬浮的B细胞,进行细胞计数,PI染色检测细胞周期,Annexin V染色检测凋亡;流式细胞仪检测表面免疫球蛋白、MHC分子和共刺激分子的表达,Transwell实验证明细胞接触的依赖性。实时定量PCR检测与MSC共培养6、12、24、48 h后细胞因子的mRNA水平。Western blot检测MAPK、JAK-STAT、PI3K信号通路蛋白的激活情况,抑制剂证明信号通路对细胞功能的影响。结果:MSC可阻滞Namalwa和Raji于G0/G1期,对3种B细胞系增殖和凋亡没有显著影响;显著抑制Daudi、Namalwa表面IgM的表达和3种B细胞表面CD86的表达,且抑制效应不依赖细胞接触和COX2的作用;显著上调Daudi内CCL2、COX2、IL-8的mRNA水平。但不影响TNF-α;显著激活3种细胞株上的ERK信号,ERK抑制剂可减弱MSC对Daudi表面分子和CCL2的调控。结论:MSC可阻滞B细胞系周期,通过分泌可溶性因子抑制B细胞系表面分子的活化、促进细胞因子的产生,ERK信号通路参与了MSC对B细胞系上述功能的调控,提示MSC抑制B细胞的免疫效应从而在治疗免疫疾病上发挥重要作用。Objective：To investigate the effects of mesenchymal stem cells （MSC） on the cell growth, activations, functions and signal pathways of B cells, and provide the theoretical basis to elucidate the regulation mechanisms of MSC on B cells. Methods： We used three human non-Hodgkin＇s lymphoma B cell lines Dandi, Namalwa, Raji as the research models. B cell lines were co-cul- tured with umbilical cord derived MSC for three days in the presence or absence of CpG, respectively, then collected suspension cells and detected the cell count, cell cycle （by PI staining）, apoptosis （by Annexin-V staining）, surface expression of immunoglobulin and co-stimulation molecules （by flow cytometry）, the dependence of eell-eontaet （by transwell assay）. Real-time PCR was used to determine the mRNA levels of eytokines after 6,12,24,48 hours of culture with MSC. Western blot was used to inspect the protein activations of MAPK, JAK-STAT, PI3K signal pathways, and the inhibitor was used to verify the influence of signal pathway on cell functions. Results： MSC arrested cell cycles of Namalwa and Raji cells on GO/G1 phase, but had no significant effect on the apoptosis of three B cell lines. MSC remarkably inhibited the expressions of CD86 （all three cell lines） and surface IgM （Dandi and Namalwa） on B cells, and the suppression was independent on cell-contact, cell-ratio and Cox2. MSC continuously and tremendously up-regulated the mRNA expressions of CCL2, Cox2, IL-8, but not TNF-α in Daudi. The cell signal results showed a sustained phosphorylation of ERK in all three cell lines by MSC, and treatments of ERK inhibitor impaired the suppression of surface molecules and the up-regulation of CCL2 on Daudi cells. Conclusion： MSC could arrest the cell cycle of B cell lines, inhibit the activation of surface molecules and promote the expression of eytokines on human B cell lines through secreting soluble factors. ERK signal involved in these regulations of B cell functions. Above all, it implied that MSC suppr

关 键 词：间充质干细胞 B细胞 CPG ERK 

分 类 号：R392.12[医药卫生—免疫学]