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目的研究激活的Notch1信号系统对体外培养的骨骺干细胞增殖与分化的调控作用。方法向体外培养的骨骺干细胞中分别加入Notch1激活剂rhNF-κB和抑制剂γ-secretaseinhibitorⅡ(MW167),空白对照加PBS缓冲液。用RT-PCR、免疫组化、MTT、流式细胞仪、碱性磷酸酶染色和Western印迹方法检测。结果骨骺干细胞中有Notch1和Jagged1表达;与对照组相比,rhNF-κB诱导组表达PCNA、胶原Ⅱ蛋白明显增多,促细胞增殖作用明显(P=0.027),并且进入分裂期(S期)的细胞增多(26.54%),而MW167诱导组碱性磷酸酶、CollagenⅩ蛋白表达明显增多,胶原Ⅱ蛋白及分化标志蛋白stathmin表达明显减少。结论当Notch信号系统被激活时,骨骺干细胞进行增殖;当Notch信号系统被抑制时,骨骺干细胞进行分化。
Objective To study the regulation of activated Notch1 signaling system on the proliferation and differentiation of epiphyseal stem cells in vitro. Methods Notch1 activator rhNF-κB and inhibitor γ-secretase inhibitor Ⅱ (MW167) were added into epiphysis stem cells cultured in vitro, and blank control and PBS buffer were added respectively. Detected by RT-PCR, immunohistochemistry, MTT, flow cytometry, alkaline phosphatase staining and Western blotting. Results Notch1 and Jagged1 were expressed in epiphyseal stem cells. Compared with the control group, the expression of PCNA was significantly increased in rhNF-κB induction group and the proliferation of collagen Ⅱ was significantly increased (P = 0.027) (26.54%). However, the expression of alkaline phosphatase and Collagen X in MW167 induced group was significantly increased, and the expression of collagen Ⅱ protein and stathmin protein was significantly decreased. Conclusion Epiphyseal stem cells proliferated when the Notch signaling system was activated. Epiphyseal stem cells differentiated when Notch signaling was inhibited.