造血干细胞联合骨髓间充质干细胞移植实验效果观察

Effect of cotransplantation of mesenchymal stem cells and hematopoietic stem cell on hematopoietic reconstitution

目的：对骨髓问充质干细胞（MSC）在造血干细胞移植后造血重建中的作用进行探讨。方法：分离培养水囊引产4～5个月龄胎儿来源的骨髓MSC和脐血CD34^＋造血干／祖细胞（HSC），作为供体细胞。23只重症联合免疫缺陷（scm）小鼠作为受体，随机分成4组，^137Cs全身照射350cGy后，在4h内从尾静脉输入供体细胞。①阴性对照组：5只，不输入任何细胞。②阳性对照组：6只，单纯输入HSC5×10^6ml^-1。③实验Ⅰ组：6只，输入HSC5×10^6ml^-1和MSC5×10^6ml^-1。④实验Ⅱ组：6只，输入HSC5×10^6ml^-1和MSC5×10^6ml^-1输注后观察各组小鼠的存活情况；每周计数外周血单个核细胞（MNC）总数；移植后第7周，PCR检测外周血人体细胞DNA，并用流式细胞仪测定供体骨髓人源细胞嵌合比例。结果：阴性和阳性对照组小鼠均于10d内死亡，骨髓病理切片提示小鼠死于骨髓衰竭。实验Ⅰ、Ⅱ组小鼠均保持良好的生存状态，皮肤黏膜无溃烂、出血，毛发无缺损。实验Ⅰ、Ⅱ组外周血象逐渐回升，第3周，实验Ⅱ组外周血MNC数量（2．63±0．32）×10^9L^-1，高于实验Ⅰ组的（1．33±0．21）×10^9L^-1（P〈0．01），第5周即恢复正常水平。第7周，实验Ⅰ、Ⅱ组小鼠外周血均检测出供体细胞；实验Ⅱ组人源细胞嵌合比例为（7．41±0．79）％，高于实验Ⅰ组（5．11±0．07）％（P〈0．05）。结论：SCID小鼠致死剂量照射后．联合输入HSC和MSC可明显促进造血干细胞移植后造血重建，且不增加移植物抗宿主病的发生率。

Aim： To explore the effect of cotransplantation of mesenchymal stem cells （MSC） and hematopoietic stem cell（HSC） on hematopoitie reeonstitution. Methods： Human bone marrow derived M SC and HSC were used as the donor. A total of 23 SCID mice, lethally irradiated, were allocated into 4 groups. Group A （ n = 5） ,transplated nothing. Group B （n =6） ,transplated HSC 5 × 10^6 ml^-1. Group C（n =6） ,transplated HSC 5 × 10^6 ml^-1and MSC 5 × 10^6 ml^-1. Group D （n = 6） , transplated HSC 5 × 10^6 ml^-1 and MSC 5 × 10^8 ml^-1. The hematopoietic reconstitution was observed by detecting the number of MNC in the peripheral blood of each mouse every week. At the 7th week PCR and FACS were performed to determine the human-derived cells in mice bone marrow and the level of their chimerism. Results： All the mice in Group A and B died of hematopoietic failure within 10 days after the transplantation. While the mice in group C and D survived, At the 7th week, human-derived cells were found in peripheral blood of Group C and D, and the ratio of chimerism in the bone marrow of Group D were （ 7.41 ± 0. 79 ） , higher than that of Group C （ 5.11 ±0. 07, P 〈 0. 05 ）. Conclusion ： Cotransplantation of MSC and HSC after lethally irradiation could significantly reconstitute hematopoiesis of SCID mice, and reduce treatment related mortality.