增强型生物活性玻璃一胶原复合支架材料的成骨效能研究

Enhanced bioactive glass/collagen composite scaffold promotes osteogenesis

目的 探讨增强型生物活性玻璃一胶原复合支架材料的体内外成骨效能。方法取第3代BMSCs种植于支架材料（支架组），以等量细胞常规培养作为非支架组，培养1、3、5、7、9、11d采用阿尔玛蓝法动态检测细胞的增殖率。取第3代BMSCs种植于支架材料（体外实验组），以等量细胞常规培养作为体外对照组，培养4、7d采用实时定量逆转录聚合酶链式反应（qRT—PCR）检测细胞骨形态发生蛋白一2（BMP一2）、碱性磷酸酶（ALP）、I型胶原的mRNA表达。裸鼠皮下植入复合成骨样细胞的支架材料（体内实验组），取正常骨组织作为体内对照组，6周后以X线片、qRT—PCR、组织学染色评估成骨情况。结果培养7～11d支架组细胞增殖率显著高于非支架组，差异均有统计学意义（P〈0．05）。体外实验组培养7dBMP一2、ALP、I型胶原的mRNA表达显著高于体外对照组，差异均有统计学意义（P〈0．05）。体内实验组x线片示植入区域有密度增高影，支架材料形成白色硬性组织；BMP一2、I型胶原、骨钙素、骨桥蛋白的mRNA表达较体内对照组均增高，差异有统计学意义（P〈0．05）；组织学染色示支架材料大部分降解，新生骨形成明显。结论增强型生物活性玻璃一胶原复合支架材料具有良好的生物相容性．体内外均具有成骨效应。

Objective To study the in vivo and vitro biocompatibility and osteogenetic capacity of enhanced hioactive glass/collagen composite scaffold. Methods Bone marrow stromal cells （BMSCs） were collected and induced to osteoblast-like cells. The growth rate of BMSCs was detected and compared progressively through Alamar Blue. The RNAs of the cells were collected and detected for bone morphogenetic protein-2 （BMP-2）, alkaline phosphatase （ALP）, collagen I （Col- I） through qRT-PCR on the fourth and seventh days. Scaffolds with induced osteoblasts were embedded into 3 nude mice subcutaneously in vivo and detected after 6 weeks. X-ray, qRT-PCR and tissue staining were used to detect the mRNA expressions of BMP-2, Col I, osteocalcin （OCN） and osteopontin （OPN） and bone formation. Results SEM（scanning electronic microscopy） showed BMSCs attached to the scaffold tightly and viably and proliferated actively on the scaffold. The growth rate in the experimental group was significantly higher after 7 days （ P 〈 O. 05） than in the control group, qRT-PCR showed that the mRNA expressions of BMP-2, ALP and Col- I in the experimental group were significantly higher than in the control group on the seventh day （ P 〈 0.05） . X-ray showed that the dense images of embedded scaffolds were locally similar to those of normal bone after 6 weeks. qRT-PCR showed that the mRNA expressions of BMP-2, Col I, OCN and OPN in the experimental group were significantly higher than those of normal bone （ P 〈 0.05）. HE and Masson staining of the paraffin sections showed the scaffolds degraded generally and osteoblasts and chondrocytes proliferated abundantly and dis- tributed irregularly. Bone formation could be observed obviously. Conclusion Enhanced bioactive glass/collagen composite scaffolds have good biocompatibility and osteogenetic capacity in vitro and vivo.