降钙素基因相关肽联合重组人骨形态发生蛋白对兔成骨样细胞增殖和碱性磷酸酶活性的影响

目的研究降钙素基因相关肽(CGRP)及重组人骨形成蛋白-2(rhBMP-2)在促进兔骨髓来源成骨样细胞增殖和分化方面是否有协同作用。方法取经诱导第三代兔骨髓基质细胞获得的兔成骨样细胞以2×10~6/ml的浓度接种于96孔板,分为6组,用含有不同条件的培养基进行培养。包括①A组:空白对照组,②B组:0.05 ng/ml CGRP,③C组:5 ng/ml CGRP,④D组:100 ng/ml rhBMP-2,⑤E组:5 ng/ml CGRP+100ng/ml rhBMP- 2,⑥F组:0.05ng/ml CGRP+100 ng/ml rhBMP-2。用四唑盐比色法(MTT)法测定细胞增殖情况和碱性磷酸酶( ALP)染色法检测其对ALP活性的影响。结果①100ng/ml rhBMP-2组、0.05ng/ml CGRP+100ng/ml rhBMP-2组、5ng/ml CGRP+100ng/ml rhBMP-2组细胞增殖OD值比对照组显著增强(P<0.05),其中0.05ng/ml CGRP+100ng/ml rhBMP-2组最强(P<0.01)。②100ng/ml rhBMP-2组、0.05ng/ml CGRP+100ng/ml rhBMP-2组、5ng/ml CGRP+100ng/ml rhBMP-2组,3组之间的ALP表达无明显差异(P>0.05).但3组与对照组相比显著增强( P<0.05)。结论CGRP和rhBMP-2合用对兔骨髓来源成骨样细胞的增殖有一定的协同作用,但对ALP无协同作用,与单用rhBMP-2的效果相当。

The Effects of CGRP and rhBMP-2 on the Proliferation and ALP Activity of Rabbit Osteoblast-like Cells

Objective To study the synergistic effects of calcitonin gene-related peptide(CGRP)and recombinant human bone morphogenetic protein-2(rhBMP-2)on the generation and activity of rabbit osteoblast-like cells obtained from rabbit bone marrow stromal cells by induced mineralization.Method The osteoblast-like cells were obtained from rabbit bone marrow stromal cells by induced mineralization and subcultured to the 3rd passage used for the experiment.The cells were seeded separately into 96 well plates with approximately 2×10~6cells/well and incubated in different mediums.The samples were derided into 6 groups:①Group A,blank group,②Group B,0.05 ng/ml CGRP,③Group C,5 ng/ml CGRP,④Group D,100 ng/ml rhBMP-2,⑤Group E,5 ng/ml CGRP+100ng/ml rhBMP-2,⑥GroupF,0.05ng/ml CGRP+100 ng/ml rhBMP-2.The proliferation of osteoblast-like cells were measured by MTT and the activity of ALP studied by staining method.Result The optical density(OD)values of cells proliferation were higher in 100ng/ml rhBMP,0.05ng/ml CGRP+100ng/ml rhBMP-2 and 5ng/ml CGRP+100ng/ml rhBMP-2 groups than in normal control(p<0.05),with the highest in 0.5ng/mlCGRP+100ng/ml rhBMP-2,and also higher in first three groups than in normal control for ALP activity(p<0.05), but no obvious differences between first three groups(p>0.05).Conclusion The combination of rhBMP-2 and CGRP has synergistic effect on the proliferation of rabbit osteoblast-like cells,but not for ALP.