骨骼肌来源和骨骼肌筋膜来源的间充质干细胞成软骨差异的研究

Experimental research of differences between source of skeletal muscle and skeletal muscle fascia source of mesenchymal stem cells into cartilage

目的观察骨骼肌来源和骨骼肌筋膜来源的间充质干细胞成软骨能力的差异，以期找到合适的种子细胞达到抑制甚至逆转骨性关节炎软骨细胞退变。 方法取膝关节置换的患者的髌下骨骼肌组织及骨骼肌筋膜组织，酶消化法原代培养至第3代。波形蛋白（Vimentin）免疫荧光鉴定两种细胞，然后两种细胞进行三维立体培养（3D培养）来模拟体内细胞的生长环境并石蜡切片，阿尔新蓝（Alcian blue）染色研究两种细胞的成软骨能力差异。同时通过实时荧光定量聚合酶链反应（FQ-PCR）检测成软骨及成骨标志物的基因表达量差异。结果骨骼肌干细胞和骨骼肌筋膜干细胞爬片Vimentin免疫荧光结果，可以看到骨骼肌干细胞Vimentin免疫荧光的信号很弱，而骨骼肌筋膜干细胞的Vimentin免疫荧光呈阳性表现；两种细胞培养第14及第21天在3D培养条件下Alcian blue染色的情况是染色均有加深，但骨骼肌筋膜干细胞更加明显；骨骼肌筋膜干细胞的成软骨标志物性别决定区Y框蛋白9（SOX9）、Ⅱ型胶原（CollagenⅡ）表达量明显高于骨骼肌干细胞。结论骨骼肌筋膜来源间充质干细胞属于成纤维类细胞，而骨骼肌来源间充质干细胞属于成肌细胞。骨骼肌筋膜来源间充质干细胞成软骨能力较骨骼肌来源间充质干细胞更强，相反骨骼肌来源间充质干细胞的成骨能力更强。

ObjectiveTo observe the difference of cartilage ability of skeletal muscle and skeletal muscle fascia-derived mesenchymal stem cells in order to find suitable seed cells to inhibit or even reverse osteoarthritic degeneration.MethodsThe patients underwent patellofemoral skeletal muscle tissue and skeletal muscle fascia tissue of patients undergoing knee arthroplasty, and the primary culture was carried out to the 3rd generation. Vimentin immunofluorescence was used to identify two kinds of cells, and then two kinds of cells were cultured in three-dimensional culture （3D culture） to simulate the growth environment of cells and paraffin sections and Alcian blue to study the differences in cartilage capacity. At the same time, the difference of gene expression between cartilage and osteogenic markers was detected by real-time fluorescent quantitative polymerase chain reaction （FQ-PCR）.ResultsClimb piece of Vimentin immunofluorescence of mesenchymal stem cells more than that of keletal muscle fascia stem cells; Alcian blue stain of keletal muscle fascia stem cells deepen that of mesenchymal stem cells; expression quantity of sex determining region Y-box 9 （SOX9）, Collagen Ⅱ of keletal muscle fascia stem cells larger than that of mesenchymal stem cells.ConclusionSkeletal muscle fascia-derived mesenchymal stem cells belonging to fibroblasts, and skeletal muscle source mesenchymal stem cells belong to myoblasts. Skeletal fascia-derived mesenchymal stem cells become more capable of cartilage than mesenchymal stem cells in skeletal muscle, whereas skeletal muscle mesenchymal stem cells have stronger osteogenic ability.