犬骨髓成年多能祖细胞诱导分化血管内皮细胞的实验研究

Differentiation of vascular endothelial cells from canine bone marrow multipotent adult progenitor cells

目的 内皮细胞移植对损伤组织的修复治疗至关重要 ,本研究旨在探讨骨髓成年多能干细胞(MAPCs)体内外诱导分化血管内皮细胞的可行性。方法 采用Percoll密度梯度法分离培养MAPCs。应用 10ng ml血管内皮细胞生长因子 (VEGF)对骨髓MAPCs进行体外诱导分化 2～ 3周 ,使其定向分化成为血管内皮细胞。采用细胞形态学、细胞免疫组化以确定诱导分化的效果。将标记BrdU的MAPCs自体移植于犬心肌内 ,观察局部微环境对于骨髓MAPCs的分化能力。结果 应用 10ng mlVEGF孵育骨髓MAPCs 2～ 3周 ,可见MAPCs分化为血管内皮细胞 :细胞形态呈鹅卵石样 ;形成血管样结构 ;细胞vWF免疫染色阳性。移植于心肌内的MAPCs在体内形成新生血管 ,血管内皮BrdU染色与vWF染色均阳性。结论 骨髓MAPCs可在体外VEGF诱导下或在体内微环境作用下分化为成熟血管内皮细胞。骨髓MAPCs可为损伤组织的移植修复治疗提供内皮细胞资源。

Objective To study the f ea sibility of in vitro and in vivo differentiation of vascular endothelial cells (EC) from canine bone marrow multipotent adult progenitor cells (MAPCs)in order to provide an ideal cell source for the restoration therapy of damaged ti ssue. Methods MAPCs isolated from canine bone marrow using Percol l density gradient solution were grown in DMEM medium supplemented with 10 ng/ml vascular endothelial growth factor (VEGF) for 2-3 weeks. The morphology and spe cial antigen of differentiated cells were observed by microscopy and immunohisto chemical staining. In vivo differentiation of endothelial cells from bone ma rrow MAPCs labeled with bromodeoxyuridine (BrdU) was identified by morphology an d immunohistochemical staining for observing the effects of physiological enviro nment on differentiation of endothelial cells. Results After 2-3 weeks of incubation with 10 ng/ml VEGF, the cells formed a monolayer with a typical EC morphology, vascular structure, and vWF positive. In vivo, MAP Cs labeled with BrdU could differentiate, in response to the effect of local mic r oenvironment, into endothelial cells that contributed to angiogenesis in myocard i um. Conclusion Multipotent adult progenitor cells isolated from c anine bone marrow could differentiate into vascular endothelial cells by VEGF in duction in vitro or under the influence of physiological environment in vi vo, which may provide an important source of endothelial cells for cellular tr ansplantation therapy of damaged tissue.