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目的应用催产素(OT)体外诱导骨髓间充质干细胞(BMSCs)向心肌样细胞分化,探索BMSCs向心肌细胞分化的诱导方法。方法取SD大鼠四肢骨骨髓,分离培养BMSCs,应用OT定向诱导,相差显微镜观察细胞形态学变化,应用免疫细胞化学、激光扫描共焦显微镜技术检测结蛋白、α-横纹肌肌动蛋白、心肌特异性肌钙蛋白T(C-TnT)的表达,应用透射电镜对分化细胞进行鉴定。结果给予OT诱导后,BMCs形态和排列方式变化明显,可见诱导1周的细胞体积增大并延伸,多数细胞呈长梭形,平行排列生长。诱导4周的细胞密集呈片状生长,多数细胞呈杆状,并与邻近细胞相连形成类肌管结构。BMSCs体外经OT诱导后分化的细胞结蛋白、α-横纹肌肌动蛋白、C-TnT均表达阳性,其中α-横纹肌肌动蛋白和结蛋白的阳性率较高,分别为58.06%和33.24%,C-TnT阳性率较低为28.32%。激光扫描共焦扫描显微镜观察到分化细胞的胞质内α-横纹肌肌动蛋白和肌钙蛋白T均有阳性表达,分别呈现红色和绿色,当两者同时被观察时可见其共表达的部位呈黄色。透射电镜观察:分化细胞的胞质内可见到平行排列的肌丝,富含线粒体、糖原和核糖体。结论 OT可以诱导骨髓间充质干细胞定向分化为心肌样细胞。
OBJECTIVE: To induce the differentiation of bone marrow mesenchymal stem cells (BMSCs) into cardiomyocyte-like cells induced by oxytocin (OT) in vitro and explore the induction of BMSCs into cardiomyocytes. Methods BMSCs were isolated from bone of extremities of SD rats. BMSCs were isolated and cultured. OT morphological changes were observed by phase contrast microscopy. The expressions of desmin, α-striated muscle actin and cardiac myosin were detected by immunocytochemistry and laser scanning confocal microscopy Troponin T (C-TnT) expression, the use of transmission electron microscopy of differentiated cells were identified. Results After the induction of OT, the morphology and arrangement of BMCs changed obviously. The volume of cells in one week was increased and extended, and most of the cells were long fusiform and arranged in parallel. After 4 weeks of induction, the cells grew in the form of sheet-like growth. Most of the cells were rod-shaped and connected with adjacent cells to form myotube-like structures. The positive expression of BMSCs in vitro after induced by OT was positive, with the positive rates of α-striated actin and desmin being 58.06% and 33.24%, respectively. The positive rates of α-striated actin and C- The positive rate of C-TnT was 28.32%. Confocal laser scanning microscopy showed that the cytoplasmic α-striated actin and troponin T of the differentiated cells were positive, showing red and green, respectively, when the two were observed at the same time can be seen when its co-expression was yellow. Transmission electron microscopy: Differentiated cells can be seen in the cytoplasm arranged in parallel to the myofilament, rich in mitochondria, glycogen and ribosomes. Conclusion OT can induce bone marrow mesenchymal stem cells to differentiate into cardiomyocytes.