摘要
目的探讨大鼠骨髓间充质干细胞BMSCs(bone marrow mesenchymal stem cells)的分离、培养、扩增方法及外源性转化生长因子TGF-β1(transforming growth factor-1)诱导BMSCs向肌成纤维细胞定向分化的可能性,为放射性诱导的颌骨纤维化提供体外理论依据。方法取大鼠股骨的骨髓,全髓培养法分离培养骨髓间充质干细胞;培养至第3代时分别用+/-TGF-β1的2组诱导培养基定向分化诱导;诱导2周后显微镜下观察细胞形态变化,Real-time PCR检测细胞的基因表达情况;细胞爬片行免疫组化鉴定α-SMA及胶原纤维的表达。结果大鼠股骨骨髓组织中可分离出骨髓间充质干细胞,体外生长形态类似成纤维细胞,呈梭型、纺锤型,定向诱导后TGF-β1组细胞可表现出肌成纤维细胞的特性(α-SMA+),同时伴有COLⅠ、COLⅢ、α-SMA基因的高表达。结论从大鼠股骨骨髓组织中可分离出骨髓间充质干细胞,TGF-β1能定向诱导BMSCs向肌成纤维细胞方向分化,为放射诱导的颌骨纤维萎缩机制提供了体外理论依据。
Objective To investigate the methods of isolation, cultivation and amplification of Bone marrow mesenchymal stem cells （BMSCs） and the feasibility of inducing BMSCs to differentiate to myofibroblast with exogenous Transforming growth factor-1 （TGF-β1） in vitro to provide the theory for jaw fibrosis induced by radiation . Methods The Bone marrow mesenehymal stem cells （BMSCs） were obtained from rats＇ Bone marrow of femur,and the isolation and cultivation were processed by all cultivation;The passage m cells were ex posed to two kinds of defined mediums for phenotype differentiation. After two weeks of induction, the morphological change of cells was observed in Microscope; Real-time PCR was used to detect cells＇ gene expression;the identification of α-SMA and collagenous fiber was pursued by imnmnohistoehemistry. Results Bone mesenchymal stem cells could be isolated from bone marrow tissue of the rat femoral; the morphology of spindle and shuttle was similar to fibroblast; after directional induction by TGF-β1 ,the cells showed the characteristics of Myofihroblasts （α-SMA ＋ ） accompanied with the increase of COLI, COL Ⅲ and α-SMA. Conclusions Bone marrow mesenchymal stem cells,which have stable biological characteristics,can be isolated from the rat femoral bone marrow tissue; TGF-β1 can induce BM SCs to differentiate to Myofibroblast directly,which provides the theory for fibrosis of jaws induced by radiation in vitro.
出处
《口腔医学》
CAS
2014年第1期1-4,共4页
Stomatology
基金
国家自然科学基金(81271112)
上海市教委"曙光计划"(10SG19)
