Molecular biological analyses of cranio-maxillo-facial bone metabolism and application of the gene therapy

人骨髓间充质干细胞(HMSCs)在骨形态发生蛋白-2(BMP-2)和碱性成纤维细胞生长因子(BFGF)的作用下具有高度的增殖和细胞周期调节作用。BrDU、增殖细胞核抗原(增殖细胞核抗原)和PTTG(垂体瘤转化基因)等细胞周期基因分别在全周期、GO/G1期和G2/M期均有较好的表达。超微结构分析显示，在BMP-2作用下，hMSCs通过改变细胞核(常染色质变为异染色质)和细胞质(平滑内质网变为颗粒状内质网)的形态而分化。在使用hMSCs和合适的支架(如天然明胶)的再生医学中，伴随着BMP-2和bFGF2的启动，可以加速颅骨的再生。在含有地塞米松、抗坏血酸和β-甘油磷酸的成骨细胞培养上清液和细胞中观察到碱性磷酸酶(ALP)的高表达水平。移植后4周，人骨髓间充质干细胞和成骨细胞因子免疫组织化学显示人碱性磷酸酶和骨钙素有明显表达，骨量增加。这些结果可能解释了hMSCs在成骨细胞谱系的再生医学中的应用。未来，该方案可能与基因治疗联合用于治疗严重骨代谢异常的hMSCs。

The human modesenchymal stem cells,(hMSCs) are highly proliferative and cell-cycle regulated by applications of bone morphogenetic protein-2 (BMP-2) and basic fibroblast growth factor (bFGF). The gene expressions of the cell cycles such as BrDU, PCNA (proliferating cell nuclear antigen) and PTTG (pituitary tumor transforming gene) are well demonstrated at entire, GO/G1 and G2/M phase, respectively. Ultrastrucural analyses revealed that hMSCs are differentiated upon BMP-2 administration by changing the nuclear (of euchromatin to heterochromatin) and cytoplasmic (smooth endoplasmic reticulum to granular endoplasmic reticulum) morphology. In regenerative medicine using hMSCs and the proper scaffold such as a natural gelatin, there are accelerated cranial bone regeneration together with BMP-2 and bFGF_initiations. High expression levels of alkaline phosphatase (ALP) in osteogenic medium culture, which contains dexamethasone, ascorbic acid, and beta-glycerophosphate, observed in both supernatant and cells. In 4 weeks after transplanting hMSCs and osteogenic cytokines demonstrated the significant immunohistochemical expressions of human ALP and osteocalcin as well as increased bone mass. These results may explain that hMSCs are useful in regenerative medicine in osteogenic lineage. In future, this regimen may be used in conjunction with gene therapy on hMSCs for severe abnormal bone metabolism.