造血干细胞移植后患者间充质干细胞增殖功能的研究

Expanding Capacity of Mesenchymal Stem Cells from Patients after Hematopoietic Stem Cell Transplantation

为了研究造血干细胞移植(hematopoietic stem cell transplantation,HSCT)后造血植活患者中骨髓源的间充质干细胞(mesenchymal stem cells,MSCs)的增殖能力,本研究选择造血干细胞移植后临床植活的34例患者作为研究对象,抽取34例患者及30例正常健康供者骨髓,进行微环境成纤维细胞集落形成单位(colony-formingunit-fibroblast,CFU-F)集落培养,同时对MSC进行培养传代,记录MSC的融合时间及传代总数;利用流式细胞术(flowcytometry,FCM)检测患者MSC表面抗原的表达;将此34例患者培养结果与30例正常对照结果进行比较统计。另外对其中骨髓加外周血加第二供者脐血移植的10例患者与骨髓加外周血移植的19例患者的MSC体外扩增指标进行比较统计。结果表明:34例患者中有31例(91.1%)患者的MSC原代培养能达到贴壁融合,有1例(2.9%)达到部分融合,2例(5.9)未达融合;与正常对照相比,患者的MSCs融合时间明显延长,传代总数明显减少,CFU-F数明显降低,即其MSC的体外扩增能力明显受损。骨髓加外周血加第二供者脐血移植的10例患者与骨髓加外周血移植的19例患者相比,MSC达到融合的时间短,体外传代总数多,CFU-F计数高。结论:移植后患者的MSC呈明显受损状态,加用第二供者脐血细胞能部分改善患者MSC体外增殖功能。

The aim of this study was to investigate the expanding capacity of bone marrow-derived mesenchymal stem cells （MSCs） in 34 patients who received a marrow and/or peripheral blood stem cells transplant （SCT）. Marrow samples were obtained from iliac crest aspirates of healthy individuals （normal controls） and patients for the isolation, purification, and expansion of MSCs. The different passage MSCs of patients were analyzed by flow cytometry （FCM） to reveal the cell surface antigen expression. The expanding function of MSCs from patients after SCT, which might be affected by cytotoxic therapy in the conditioning regimen, colony-forming unit-fibroblast （CFU-F）, confluent time, and passage number of the culture were measured, and then compared with those in the normal controls. At the same time, the numbers of colony-forming unit of hematopoietic progenitor were detected and compared with normal controls. In addition, CFU-F, confluent time, and passage number of MSCs were compared between group of BMSCs plus PBSCs co-transplanted patients and group of BMSCs plus PBSCs with the second donor umbilical cord blood cells （UBCs） cotransplanted patients. The results indicated that a confluent monolayer of stroma cells was generated in 31 out of the 34 cases （91. 1% ）, a subconfluent monolayer was generated in one case （ 2.9% ）, no adherent stromal layer was generated in 2 cases （5.9%）. As compared with the normal controls, the time generating a confluent layer of stroma cells from primary MSCs of patients was longer significantly, and the passage number and CFU-F of MSCs of patients in vitro was less than that of normal controls significantly. Compared with the group of BMSCs plus PBSCs co-transplanted patients, the confluent time of MSCs in group of BMSCs plus PBMSCs with UBCs co-transplanted patients was shorter, the passage number and CFU-F count in this group were higher. It is concluded that the MSCs of patients after HSCT are damaged, and the co-transplant of BMSCs and PBSCs plus UBCs can partially improve in vitro expanding capacity of MSCs from patients.