碱性成纤维细胞生长因子增强重组人骨形态发生蛋白-2诱导成骨能力的剂量依赖性

Dose-dependence of bone induction ability of recombinant human bone morphogenetic protein - 2 en-hanced by basic fibroblast growth factor

目的 对碱性成纤维细胞生长因子（basic fibroblast growth factor,bFGF）增强重组人骨形态发生蛋白-2（recombinant human bone morphogenetic protein-2,rhBMP-2）诱导成骨能力的剂量依赖性进行研究。方法280只BALB/c小鼠随机分为8组,每组35只,实验组rhBMP-2的剂量均为0.5mg,bFGF的剂量分别为0、2、10、40、80、300、500活性单位（IU）;设立单纯牛松质骨载体组为对照组。按实验设计,分别将rhBMP-2混悬液及bFGF-PVP溶液与牛松质骨载体充分混匀,负压下真空抽吸,冻干。将复合植骨材料植入小鼠右侧股部肌袋内,各组分别于术后第3、5、7、9、14、21d六个时间点取材,进行组织学、X线分析以及钙含量测定,观察各组的诱导成骨情况。结果（1）bFGF剂量为10～80IU时,间充质细胞增殖、分化,软骨细胞及新生骨形成均早于其它各组,成骨量多于其它各组,组织钙含量明显高于其它各组（P<0.05）,提示此剂量bFGF使rhBMP-2的诱导成骨能力明显增强。（2）bFGF剂量为0.2、300 IU时,各组在间充质细胞增殖、分化,软骨细胞及新生骨形成以及组织钙含量方面差异均无显著性意义（P>0.05）,提示此剂量bhBMP-2的诱导成骨能力无明显影响。（3）bFGF剂量为500IU时,术后第21d,仅1例标本可见散在的极少量新骨形成。

Objective To study the dose-dependence of bone induction of recombinant human bone morphogenetic protein - 2(rhBMP - 2) enhanced by basic fibroblast growth factor(bFGF) . Methods BALB/c mice ( n = 280) were divided into eight groups at random, 35 in each group; rhBMP - 2 was mixed with 0. 1 mol/L PBS(pH=7. 4) and then homogenized into suspension, bFGF and polyvinylpyrrolidone (PVP) were mixed with 0. 1 mol/L PBS and then made into solution with viscosity. The suspension and solution were mixed thoroughly with the bovine cancellous bone as the vector under vacuum and lyophilized. Composites were implanted in the right thigh pouches of the mice receiving respectively rhBMP -2 (0. 5 mg) and bFGF(0, 2, 10, 40, 80, 300, 500 IU ) served as the experimental subjects and those receiving vectors only served as the control. In order to investigate osteoinduction and osteogenesis, histological analysis was performed 3, 5, 7, 9, 14 and 21 days after the implantation. Radiographs of new bone in thigh pouches were taken and ealcium content was measured 21 days after the implantation. Results 1) Proliferation and differentiation of mesenchymal cells, chondrogenesis and osteogenesis induced by rhBMP - 2/ bFGF (with doses from 10to80IU) appeared earlier than those induced by other groups; the amount of osteogenesis was greater than that in other groups; calcium content was higher than those in other groups ( P < 0. 05) . These results indicate that bFGF enhanced obviously the osteoinduction and osteogenesis by rhBMP - 2. 2) No significant difference was seen on proliferation and differentiation of mesenchymal cells, chondrogenesis, osteogenesis and calcium content( P < 0. 05) in groups rhBMP - 2/ bFGF(with doses 0, 2, 300 IU). These results suggested that no effect was seen with 0, 2 or 300 IU of bFGF. 3) Only a little new bone induced by rhBMP - 2/ bFGF(with dose 500 IU) in just one sample was seen; calcium content in group rhBMP - 2/bFGF(with dose 500 IU) was lower than that in other groups 21 days after the im-plantation ( P < 0. 05). These results indicate that bFGF with dose of 500 IU had a powerful inhibitory effect. 4) No new bone was seen in group control 21 days after the implantation. Conclusion The dose-dependent effect is clearly seen in the bone induction of rhBMP - 2 enhanced by bFGF .