脂肪来源干细胞修复骨骼肌放射损伤的病理学研究

Influence of adipose-derived stem cell transplantation on irradiation-induced pathological changes in skeletal muscles of rabbit

目的 观察CM-Dil标记的脂肪来源干细胞（adipose-derived stem cells,ASCs）植入放射损伤骨骼肌后的病理学改变,探讨ASCs对兔骨骼肌放射损伤的修复影响.方法 64只新西兰兔单侧臀部予9 MeV电子线单次照射80 Gy后采用随机数字表法分为ASCs组和磷酸盐缓冲液（PBS）组,每组32只,照射24 h后照射侧分别肌肉注射1 ml含5×107/ml标记后ASCs的PBS悬液和1 mlPBS缓冲液,未照射一侧作为各组正常对照.分别于照射后1、4、8和26周收集各组实验动物肌肉标本.观察标记后ASCs在骨骼肌组织中的分布及迁移,分析骨骼肌损伤病理学分级,观察4和26周时肌组织超微病理结构改变.结果 标记后的ASCs能在损伤部位迁移.ASCs组受照射骨骼肌组织病理损伤分级较PBS组在照射后1、4、8、26周显著减低（U=11.5、12.0、11.0,P＜0.05）.ASCs组26周时再生肌细胞数（27.01土9.36）显著高于PBS组（5.23 土4.23）（t=15.12,P＜0.05）.ASCs组4和26周肌原纤维损伤程度较PBS组轻,26周ASCs组肌细胞肌间隙可见肌原纤维状结构增生.结论 ASCs能减轻受照射骨骼肌的病理组织学损伤,促进肌卫星细胞的代偿增生、再生肌组织,可能是修复骨骼肌放射损伤的部分机制.

Objective To observe the pathological changes of irradiated skeletal muscles in a rabbit model after CM-Dil fluorescence-labeled adipose-derived stem cells （ASCs） transplantation and to explore the influence of ASCs on the pathological repair of radiation-induced skeletal muscle injury.Method Unilateral hips of 64 rabbits were randomly divided into ASCs group （n =32） and PBS group （n =32） and they were irradiated with a single dose of 80 Gy of 9 MeV electrons beam from a linear accelerator.The non-irradiated sides with normal skeletal muscle were used as controls.24 hours after irradiation,1 ml of the CM-Dil fluorescence-labeled ASCs （5 × 107） were locally transplanted into the irradiated side of the skeletal muscle,while PBS injection was applied as a control.1,4,8 and 26 week after irradiation,8 animals of each group were randomly executed and the tissue samples were collected.The distribution and migration of ASCs labeled by CM-Dil were observed in the frozen sections using the fluorescence microscope.The pathological changes and histological severity scoring of skeletal muscles were determined in HE staining sections.The ultrastructural changes of the muscular tissues were observed using a transmission electron microscope.Results By fluorescence tracing,migration of ASCs along the needle passage was observed in the tissues with irradiation injury.For the ASCs group,remarkable decrease of the histological severity scores was noted at 4,8,and 26 week after irradiation compared with those of PBS control group（U =11.5,12.0,11.0,P 〈 0.05）.The number of centronucleated regenerating myofibers in the ASCs group was significantly higher than that in the PBS group 26 week after irradiation（27.01 土9.36 vs.5.23 土 4.23,P 〈 0.05）.Interestingly,26 week after the irradiation,myofibrils-like structures were identified in the irradiated muscle cells after ASCs transplantation.Conclusions ASCs transplantation could repair the radiation-induced skeletal muscle injury perhaps by promoting the compensatory hyperplasia of muscle satellite cells as well as myogenic differentiation.