Change in hematopoietic function of lethal dose irradiated mice models following skeletal muscle satellite cells transplantation

背景:骨骼肌卫星细胞是一种具有增殖、分化潜能的肌源性干细胞.近年,国外已有报道骨骼肌细胞具可被某种特定微环境因子激活分化为造血干细胞,从而具有造血重建的潜能.目的:初步验证肌源性的成体干细胞-骨骼肌卫星细胞分化为造血干细胞的能力.设计:验证性动物实验.单位:天津中医药大学基础医学院组胚教研室.材料:选用雌性昆明种成年小鼠65只,体质量25～28 g.出生5 d昆明种乳鼠5只,均由北京大学医学部实验动物中心提供.方法:实验于2001-08/2003-08在北京大学医学部人体解剖学与组织胚胎学系细胞培养室完成.胶原酶、胰蛋白酶消化分离5只乳鼠骨骼肌卫星细胞,分离5只成年昆明种小鼠骨髓单个核细胞.取成年雌性小鼠60只作为受体,用60Co γ8.0 Gy照射,然后随机分为4组:对照组不做任何处理;培养液输注组小鼠尾静脉注射DMEM,F-12培养液;卫星细胞输注组小鼠尾静脉注射卫星细胞悬液0.3 mL(细胞浓度1×109 L-1);骨髓细胞输注组小鼠尾静脉注射骨髓细胞悬液0.3 mL(细胞浓度1×109 L-1).主要观察指标:①观察各组小鼠14 d存活率.②照射后14 d处死存活下来的受体小鼠:肉眼计数脾脏表面结节数;对骨髓单个核细胞涂片进行Wright-Girmesa染色.结果:①脾集落形成单位的测定:照射后14 d,卫星细胞注射组小鼠脾结节数与骨髓细胞注射组比较,差异无显著性意义(P＞0.05).②骨髓单个核细胞组织学鉴定结果:卫星细胞输注组与骨髓细胞输注组骨髓单个核细胞涂片中有相对较多的早期造血细胞出现,其形态符合早期造血细胞的生物学特性.③被照射鼠的存活情况:对照组和培养液输注组小鼠在被照射后9～13 d全部死亡.照射后14 d,卫星细胞输注组和骨髓细胞输注组分别有8和13只鼠存活.结论:骨骼肌卫星细胞具有诱导分化为造血干细胞的功能.BACKGROUND: Skeletal muscle satellite cells are muscle-derived stem cells with proliferation and differentiation potential. Recently, foreign researches have reported that skeletal muscle satellite cells can be activated by some definite microenvironmental factors and differentiate into hematopoietic stem cells and thereby they will have the potential of hematopoietic reconstruction.OBJECTIVE: To initially validate the potential of adult muscle-derived stem cells- skeletal muscle satellite cells differentiating into hematopoietic stem cells.DESIGN: Validation animal experiment.SETTING: Department of Histology and Embryology, College of Basic Medical Science, Tianjin University of Traditional Chinese Medicine.MATERIALS: Sixty-five male Kunming adult mice, weighing 25-28 g, were involved in this study. Five Kunming neonate rats, aged 5 days, were provided by the Laboratory Animal Center, Department of Medicine, Peking University.METHODS: This experiment was carried out in the Laboratory for Cell Culture, Department of Human Anatomy and Histo-embryology, School of Basic Medical Sciences, Peking University Health Science Center between August 2001 and August 2003. Skeletal muscle satellite cells of 5 neonate rats were isolated by collagenase and trypsin digestion. Bone marrow mononuclear cells of 5 adult Kunming mice were isolated. Sixty adult female mice were used as recipients, irradiated with 60Coγ 8.0 Gy and then randomized into 4 groups: control group, in which, the mice were untouched; culture fluid infusion group, in which, the mice were injected with DMEM/F-12 medium through caudal vein; satellite cell infusion group, in which, the mice were injected with 0.3 mL satellite cell suspension through caudal vein (cell concentration 1×109 L-1); bone marrow-derived cell infusion group, in which, the mice were injected with 0.3 mL bone marrow-derived cell suspension (cell concentration 1×109 L-1) through caudal vein.MAIN OUTCOME MEASURES:①The survival rate of 14-day-old mice in each group. ②The surviving recipient mice were euthanized 14 days after irradiation, and tubercles on the surface of spleen were counted by naked observation; Bone marrow mononuclear cell smear was stained by Wright-Gimesa.RESULTS:① Determination of colony forming unit-spleen (CFU-S): No significant difference in the number of spleen tubercles of mice existed between satellite cell infusion group and bone marrow-derived cell infusion group 14 days after irradiation (P＞0.05). ②Histological identification of bone marrow-derived mononuclear cells: Many hematopoietic cells appeared at the early stage in the bone marrow-derived mononuclear cell smears between satellite cell infusion group and bone marrow-derived cell infusion group. Their morphology meets the biological characteristics of hematopoietic cells at the early stage. ③ The survival condition of irradiated mice: All the mice in the control group and culture fluid infusion group died 9 to 13 days after irradiation. In contrast, 8 mice from the satellite cell infusion group and 13 the bone marrow-derived cell infusion group survived 14 days after irradiation.CONCLUSION: Skeletal muscle satellite cells have the function of differentiating into hematopoietic stem cells.中国科技核心期刊(ISTIC)中国科学引文数据库(CSCD)0428618-86201