同种记忆性CD8^+T细胞与移植物抗宿主病相关性的研究

Study on the Relationship Between Memory CD8^+T Cell and GVHD

目的用同种异基因骨髓移植诱导小鼠急性移植物抗宿主病(GVHD)模型,研究GVHD与同种记忆性CD8^+T细胞的关系。方法供鼠C3H.SW(H-2D^b,CD45.2^+)和受鼠B6/SJL(H-2D^b,CD45.1^+),两者MHC抗原均为H-2D^b,但供受鼠之间miHAs有差异。小鼠致死剂量辐照后移植供鼠T^-BM+ CD8^+T细胞,构建CD8^+T细胞依赖的GVHD模型。观察受鼠体重和生存率变化;流式细胞仪检测受鼠肝脏和脾脏中供鼠来源CD8^+T细胞的浸润数、分泌INF-γ的效应CD8^+T细胞数和与凋亡相关的annexin V分子在CD8^+T细胞的表达情况。结果建立的CD8^+T细胞依赖的致死性急性GVHD小鼠模型,移植后小鼠体重显著下降并有大幅波动,其变化与分泌INF-γ的CD8^+T细胞数量成反比。移植后28d小鼠逐渐出现死亡,70d时死亡率达58%(7/12)。随着GVHD病程的进展,供鼠CD8^+T细胞表现出典型的记忆性T细胞应答的过程,即扩增、凋亡紧缩和残存T细胞的再扩增;分泌INF-γ的效应性CD8^+T细胞也表现出同样的变化;细胞表面凋亡相关分子的表达显示CD8^+T细胞的减少与凋亡的发生有关。结论在模拟目前临床病人采用的同种异基因骨髓移植配型方式的同种异基因骨髓移植小鼠模型中,供者来源的效应性记忆性CD8^+T细胞数量的波动与记忆性T细胞产生过程中数量的变化关系极为密切、受鼠的体重变化与效应性CD8^+T细胞浸润数量的波动成反比的现象说明,在GVHD发生过程中有抗原特异性记忆性CD8^+T细胞形成。

Objective To study the relationship between memory CD8^T cell and GVHD by using a mouse model of CD8^T cell-mediated acute graft versus host disease（GVHD）. Methods C3H.SW （H-2Db, CD45.2~） mice were used as donors and B6/SJL（H-2Db, CD45.1） mice as recipients, both of them are MHC-identical but miHA-mismatched. Before transplantation, the B6 recipients received ^137γ, ray （9.5 Gy） TBI administered in three times from a ^137Cs source, then the T cell-depleted bone marrow （T BM） cells and CD8^T cells from C3H.SW donors were injected immediately via tail vein. The weights of recipient mice and dead rate were calculated after transplantation. Results The quantitative changes in the livers and spleens of recipient mice of the donor CD8^＋T cells（CD45.2^＋）, donor CD8^＋T cells that produced IFN-γ and the apoptosis related molecular annexin V on donor CDS^T cells were analyzed by Flow Cytometry. In the miHA-mismatched CDS^T cell-mediated acute GVHD mouse model, the recipient mice began to die 28 days following transplantation and the dead rate was 58% until 70 days. The analysis showed that in parallel to GVHD development, donor CD8^Tcells andCD8^T cells secreting high levels of IFN-γ peaked in the livers and spleens of recipient mice by day 14, considerably declined by day 28, increased again by day 42 after transplantation. The decline in donor CD8^T cells was accompa- nied with increased annexin V-positive donor CD8^T cells. These results suggest that a typical memory T cell response, as manifested by T cell expansion, contraction, and re-expansion, develops in recipients during the GVHD process, and a small population of reactive CD8^T cells survives after the death phase of effect cells. Conclusions The MHC-identical but miHA mismatched mouse model is analogous to human clinical miHA-mismatched bone marrow transplantation. There is the relation of the donorCD8^Tcells to GVHD persistence.