Liu Guangpeng, Zhao Li, Cui Lei, Liu Wei, Cao Yilin
National Tissue Engineering Research and Development Center, Shanghai 200235, People's Republic of China.
Biomed Mater. 2007 Jun;2(2):78-86. doi: 10.1088/1748-6041/2/2/004. Epub 2007 Mar 14.
In this study we investigated not only the cellular proliferation and osteogenic differentiation of human bone marrow stromal cells (hBMSCs) on the novel beta-tricalcium phosphate (beta-TCP) scaffolds in vitro but also bone formation by ectopic implantation in athymic mice in vivo. The interconnected porous beta-TCP scaffolds with pores of 300-500 microm in size were prepared by the polymeric sponge method. beta-TCP scaffolds with the dimension of 3 mm x 3 mm x 3 mm were combined with hBMSCs, and incubated with (+) or without (-) osteogenic medium in vitro. Cell proliferation and osteogenic differentiation on the scaffolds were evaluated by scanning electron microscopy (SEM) observation, MTT assay, alkaline phosphatase (ALP) activity and osteocalcin (OCN) content measurement. SEM observation showed that hBMSCs attached well on the scaffolds and proliferated rapidly. No significant difference in the MTT assay could be detected between the two groups, but the ALP activity and OCN content of scaffolds (+) were much higher than those of the scaffolds (-) (p < 0.05). These results indicated that the novel porous beta-TCP scaffolds can support the proliferation and subsequent osteogenic differentiation of hBMSCs in vitro. After being cultured in vitro for 14 days, the scaffolds (+) and (-) were implanted into subcutaneous sites of athymic mice. In beta-TCP scaffolds (+), woven bone formed after 4 weeks of implantation and osteogenesis progressed with time. Furthermore, tissue-engineered bone could be found at 8 weeks, and remodeled lamellar bone was also observed at 12 weeks. However, no bone formation could be found in beta-TCP scaffolds (-) at each time point checked. The above findings illustrate that the novel porous beta-TCP scaffolds developed in this work have prominent osteoconductive activity and the potential for applications in bone tissue engineering.
在本研究中，我们不仅在体外研究了人骨髓基质细胞（hBMSCs）在新型β-磷酸三钙（β-TCP）支架上的细胞增殖和成骨分化，还通过在无胸腺小鼠体内异位植入研究了骨形成情况。采用聚合物海绵法制备了孔径为300 - 500微米的相互连通的多孔β-TCP支架。将尺寸为3毫米×3毫米×3毫米的β-TCP支架与hBMSCs结合，并在体外于有（+）或无（-）成骨培养基的条件下孵育。通过扫描电子显微镜（SEM）观察、MTT法、碱性磷酸酶（ALP）活性和骨钙素（OCN）含量测定来评估支架上的细胞增殖和成骨分化。SEM观察显示hBMSCs在支架上附着良好且增殖迅速。两组间MTT法检测无显著差异，但（+）支架的ALP活性和OCN含量远高于（-）支架（p < 0.05）。这些结果表明新型多孔β-TCP支架能够在体外支持hBMSCs的增殖及随后的成骨分化。体外培养14天后，将（+）和（-）支架植入无胸腺小鼠的皮下部位。在（+）β-TCP支架中，植入4周后形成编织骨，且骨生成随时间进展。此外，8周时可发现组织工程骨，12周时还观察到重塑的板层骨。然而，在每个检查时间点，（-）β-TCP支架中均未发现骨形成。上述研究结果表明，本研究中开发的新型多孔β-TCP支架具有显著的骨传导活性及在骨组织工程中的应用潜力。
