不同途径输注造血干细胞对小鼠移植效果的比较

Comparison of Efficacies of Allogeneic Hematopoietic Stem Cell Transplantations between Different Routes of Administration in Mice

为比较骨髓腔内(IBM)或静脉(IV)等不同途径输注小鼠造血干细胞后在受体内的分布特点及对移植效果的影响,将C57BL/6胎鼠及新生鼠外周血(FNPB)单个核细胞(MNC)移植经亚致死量60Coγ射线辐照的BALB/c鼠。受鼠随机分为6组:单侧IBM组;双侧IBM组;IV组;双侧IBM+IV组;照射对照组;空白对照组。用组织冰冻切片和流式细胞术动态了解CFSE标记的FNPBMNC在受体内的分布变化,观察移植后受鼠生存状况、植入水平、造血恢复及GVHD情况。结果显示:IBM注射的供体FNPBMNC主要积聚于注射侧骨髓腔内,少量FNPBMNC可经血循环二次归巢,肺部滞留很少。单/双侧IBM组输注的FNPBMNC总数无明显差异,但经双侧IBM输入FN-PBMNC渗漏至外周血或其它组织器官的比例更少;IBM各组造血重建速度均优于IV组,尤以双侧IBM组最快,其外周血象和造血干祖细胞集落产率在移植后第21天已接近正常水平,单侧IBM组、双侧IBM+IV组次之;IBM组各时点植入水平均明显高于IV组,双侧IBM组注射侧胫骨植入水平与单侧IBM组无明显差异,二次移植中双侧IBM组的植入水平明显高于IV组;IBM移植组90天存活率≥80%,IV组仅为50%。结论:双侧IBM有利于更多造血干细胞归巢,促进植入和造血重建,提高IBM途径移植的效果。

When hematopoietic stem cells（HSCs） were administrated by intravenous infusion （Ⅳ） , most of them were trapped in some nonhematopoietic organs as like lungs that had abundant blood capillaries. Only a small fraction of injected cells could home to the bone marrow, which reduced the engraftment of HSCs. The purpose of intra-bone marrow （IBM） transplantation was to facilitate the homing of HSCs directly. Based on the established murine model for allogeneic umbilical cord blood transplantation （UCBT） by IBM injection, the objective of this study was to compare the distribution of fetal and neonatal peripheral blood （ FNPB ） mononuclear cells （MNC） in vivo and the efficacy of HSCT by different routes of administration in mice. BALB/c recipient mice exposed to sublethal dose ^60Coγ-ray were transplanted with FNPBMNCs from C57BL/6 mice. Recipient mice were divided into six groups at random： unilateral-IBM group; bilateral-IBM group; Ⅳ group; bilateral-IBM ＋ Ⅳ group; irradiated control group and normal group. The distribution of CFSE-labeled FNPBMNCs in the recipients was observed in frozen sections of different organs or by flow cytometry. The survival rate, engraftment level, recovery of hematopoietic function and GVHD of recipient mice were studied. The results showed that infused by IBM route, FNPBMNCs mainly accumulated in the bone marrow （BM） cavity of the injected side tibia. Some of them could enter the BM of noninjected bones via blood circulation and few were trapped in the lung. Though same amount of FNPBMNCs were injected into recipient mice of unilateral and bilateral-IBM group, less cells could leak into peripheral blood or other tissues when transplanted by bilateral-IBM route. Therefore, in term of accelerating hemopoietic recovery, the injection of IBM route was better than Ⅳ route, especially bilateral IBM injection of HSCs, which neared the normal level of peripheral blood cells and colony-forming units of bone marrow nucleated cells at day 21 after transplantation, followed by unilateral-IBM group and bilateral-IBM ＋ Ⅳ group. The percentages of H-2Db cell subsets in the three IBM groups were much higher than that in Ⅳ group. There was no significant difference of the engraftment level in the injected side tibia between the unilateral and bilateral-IBM group. When secondary transplantation was performed, the engraftment level in bilateral-IBM group was still much high- er than that in Ⅳ group. At day 90, the survival rates of IBM groups were all ≥ 80%, while that of Ⅳ group was only 50%. It is concluded that bilateral-IBM route can facilitate the homing of more HSCs, accelerate the engraftment of HSCs and hematopoietic reconstitution, which promoted the efficacy of IBM-HSCT.