异基因造血干细胞移植中嵌合体的STR-PCR定量检测

目的:研究异基因造血干细胞移植后供者单个核细胞、CD3+T细胞、CD15+粒细胞植入情况。方法:采集22例清髓性外周血干细胞移植患者移植前及移植后不同时间的骨髓标本,应用免疫磁珠进行CD3+T细胞、CD15+粒细胞分选,常规提取DNA,PCR扩增后,进行毛细管电泳,确定基因位点及峰面积,计算供者嵌合率。结果:22例患者均在中位时间+12d(+9d～+15d)造血恢复。骨髓单个核细胞在+7d时供者嵌合率为70%,+14d供者嵌合率均超过90%,+21d和+28d供者嵌合率均超过95%以上。CD15+粒细胞在+7d时嵌合率为40%,在+14d、+21d和+28d供者嵌合率>95%。CD3+T细胞在+7d、+14d、+21d和+28d供者嵌合率分别为65%、80%、84%和90.9%。移植后早期完全嵌合组急性GVHD发生率高于混合嵌合组。结论:在+28d3群细胞均可达到供者完全嵌合状态。定量检测对判断早期植入、预测复发,指导临床干预治疗具有重要意义。

A quantitative detection of graft chimerism following allogeneic hematopoietic stem cell transplantation by PCR-STR

Aim:To analyze the graft chimerism of mononuclear cell(MNC),CD3+ T lymphocytes and CD15+ granulocytes chimerism following allogeneic hematopoietic stem cell transplantation(allo-HSCT).Methods:Twenty-two patients underwent allo-HSCT.CD3+ T lymphocytes and CD15+ granulocytes from bone marrow of donors and recipients before and after transplantation in different phases were sorted by immunomagnetic beads.DNA were extracted and then amplified by PCR,and geneloci,peak area were addressed with capillary electrophoresis to assay the chimerism.Results:The median time of haematogenesis recovery was 12(9～15) days post transplant in all 22 patients.Graft chimerism of MNC was 70% at 7 day post transplant,reaching 90% on 14 days,then achieving 95% at 21 and 28 days post transplant.The graft chimerism of granulocyte was 40% at 7 days post transplant,and all patients got a more than 95% chimerism at 14,21 and 28 days post transplant.The graft chimerism of T lymphocyte was 65%,80%,84% and 90.9% at 7,14,21 and 28 days post transplant,respectively.The incidence of acute graft versus host disease(aGVHD) in complete donor chimerism(CDC) group was higher than that in mix-chimerism(MC) in earlier period after transplant.Conclusion:It is concluded that MNC,CD3+ T lymphocyte,and CD15+ granulocyte chimerism could reach a CDC at 28 days post transplant following allo-HSCT.Sequential and quantitative detection of chimerism may be significant to evaluate engraftment,predict disease relapse,and provide a basis for early intervention treatment.