骨髓间质干细胞复合多孔磷酸钙陶瓷修复兔颅骨缺损的研究

目的探讨以骨髓间质干细胞(MSCs)作为种子细胞、β-磷酸三钙(β-TCP)多孔生物陶瓷作为支架材料构建组织工程化人工骨，修复兔实验性颅骨标准缺损的可行性。方法选择5月龄新西兰大白兔34只，制作颅骨标准缺损后分成3组。A组(n=20)：分离培养兔同胎异体MSCs。体外扩增后接种到预制的β-TCP多孔生物陶瓷材料上，细胞-材料复合体经体外孵育后，无菌条件下植入颅骨缺损；B组(n=10)：采用单纯β-TCP材料修复兔颅骨缺损；C组(n=4)：兔实验性颅骨标准缺损区未做骨修复。术后6周和12周分别处死动物、取材，进行缺损区大体、组织学、组织化学和免疫组织化学分析。结果颅骨缺损处A组术后6周表面肉眼可见骨样组织形成，组织学提示材料部分降解，未降解吸收的材料孔洞内广泛分布着新生骨组织，成骨细胞外基质丰富，I型胶原染色阳性；术后12周，支架材料几乎完全降解，缺损区被新生骨组织所取代。B组术后6周，可见从缺损区边缘有新生骨组织向支架材料内长入，支架材料部分吸收；术后12周，可见从缺损区边缘长入到支架材料内的新生骨组织逐渐增多，但材料的中心部位未发现新生骨形成。C组术后12周，仅见少量骨组织从缺损区边缘向缺损区内长入，缺损中央大部分区域未得到修复。结论体外培养扩增的兔MSCs在不添加外源性生长因子的情况下，与pTCP复合植入后可以在体内诱导、分化为成骨细胞，并能够对标准的颅骨缺损进行有效的修复，可进一步推广应用。

REPAIR OF CRANIAL DEFECTS WITH BONE MARROW DERIVED MESENCHYMAL STEM CELLS AND β-TCP SCAFFOLD IN RABBITS

OBJECTIVE: To determine whether culture expanded bone marrow derived mesenchymal stem cells (MSCs) in combination with beta-tricalcium phosphate(beta-TCP) can repair critical cranial defects in New Zealand rabbits. METHODS: In group A(n = 20), MSCs from homogeneous rabbits were isolated and expanded in vitro and then implanted onto the pre-molded porous beta-TCP. The MSCs-beta-TCP complexes were implanted into rabbit critical cranial defects. In group B (n = 10), The defects were repaired with beta-TCP only. In group C(n = 4), the defects were left un-repaired. Samples were extracted 6 and 12 weeks after operation for histological, histochemical and immunohistochemical analysis. RESULTS: In group A, bone-like tissue formation could be seen on the surface of the implants. Microscopic analysis demonstrated certain degradation of beta-TCP and extensive new bone filling in rich extracellular matrix after 6 weeks. The cells were stained positively for type I collagen. After 12 weeks, the bioceramics had almost completely degraded and abundant bone formation could be seen in the whole defects. In group B, marginal bone ingrowth was observed after 6 weeks and the number of osteoblasts increased significantly after 12 weeks. However, no new bone formation could be detected in the middle of the material. In group C, only a small quantity of new bone formation was found along the margin of defects. CONCLUSION: Transplantation of MSCs with beta-TCP can serve as an example of a cell-based treatment for bone regeneration in skeletal defects.