双荧光小鼠异基因骨髓移植模型的建立和应用  

Establishment and Applications of Double-Fluorescent Protein Allo-Transplantation Mice Model

作　　者：叶艳艳[1] 杨默[1] 袁乐中 江汕[1] 易正山[1] 沈克锋[1] 麦青[1] 黄皓[1] 江千里[1] 

机构地区：[1]南方医科大学南方医院血液科,广东广州510515

出　　处：《中国实验血液学杂志》2015年第5期1451-1454,共4页Journal of Experimental Hematology

基　　金：国家自然科学基金项目(30901367;31070866);南方医院国家基金匹配基金(G2011021;2010037)

摘　　要：目的:建立红荧光蛋白(mRFP)→绿荧光蛋白(eGFP)转基因小鼠异基因骨髓移植模型,结合半固体脱钙法观察移植后骨髓中供者细胞的分布及供受者细胞间的相互关系。方法:C57BL/6雌性eGFP转基因小鼠清髓性放疗后,经尾静脉输注(5×106)个供者FVB雄性mRFP转基因小鼠的骨髓细胞。给放疗对照组输注等量的PBS。观察移植后受鼠的一般情况、植入水平、造血恢复、移植物抗宿主病(GVHD)的发生率和生存期。在受鼠造血恢复的过程中,取股骨半固体脱钙,用共聚焦显微镜等观察骨髓中红绿细胞的分布、形态和相互作用关系。结果:eGFP受鼠的造血恢复时间为(20±3.07)d,外周血中RFP+细胞比例为(93.94±1.59)%;1个月内4/10只受鼠有aGVHD表现;半固体脱钙后股骨的质密结构被逐步替代,在共聚焦显微镜下可见骨髓腔中mRFP+细胞主要位于骨小梁周围且被大量的GFP+细胞所包绕;三维构建后可清楚地观察到骨髓微环境中红绿荧光细胞的相互作用。结论:成功建立了双荧光小鼠异基因骨髓移植模型,结合半固体脱钙法可清晰地观察到移植后骨髓中供者细胞的分布及供受者细胞间的相互关系,为临床观察移植后供者细胞的分布、归巢等相关研究提供了基础。Objective: To establish allo-transplantation model by using mRFP + to eGFP + transgenic mice and to observe the distribution of donor cells and donor-recipient cellular interaction in the bone marrow after semi-solid decalcification( SSD). Methods: After myeloablative irradiation,C57 BL /6 female eGFP + transgenic mice were infused with( 5 × 106) bone marrow cells from FVB male donor mice through tail vein. The control group was infused with PBS.Then the general conditions,engraftment level,hematopoietic recovery,incidence of GVHD and survival of recipients were evaluated after transplantation. In the recovery process,SSD was used to treat the femora before observing the cells distribution,morphology and interaction by confocal microscopy directly or after making frozen section. Results: WBC of recipient eGFP + mice was recovered on( 20 ± 3. 07) d,( 93. 94 ± 1. 59) % in peripheral cells were RFP + cells( n =10),GVHD happened in 4 of 10 mice within 1 month. During SSD,the hard components were replaced gradually and RFP + cells could be seen mainly in the bone trabecula and surrounded by eGFP + cells under confocal microscope,their interactions could be further observed clearly in bone marrow microenvironment in three-dimensional reconstruction.Conclusion: The double fluorescent allo-transplantation mouse model successfully established,by means of our novel protocol named SSD,the donor and recipient cell location and their interaction can be visually observed,which provides the basis for clinical studies on the distribution and homing of donor cells,and some related explorations after transplantation.

关 键 词：荧光小鼠 异基因骨髓移植模型 半固体脱钙 

分 类 号：R457.7[医药卫生—治疗学]