TGF-β_1影响钛表面成骨细胞增殖分化的体外研究

The effects of TGF-β_1 on the proliferation and differentiation of osteoblasts located on titanium disc in vitro

目的动态观测TGF-β1和钛片表面微形态对胎鼠成骨细胞增殖分化的影响以及两者之间的相关关系。方法将传代后的成骨细胞接种于不同处理(机械打磨、喷砂处理TPS)钛片表面,加入外源性TGF-β1(浓度为10ng/ml)后MTT法检测细胞增殖,并检测细胞内ALP和细胞外OC的合成分泌状况;设置玻片对照组。结果外源性TGF-β1后对各组钛片表面的成骨细胞增殖均有不同程度的抑制作用,细胞接种后期抑制作用更明显。细胞接种早期,ALP活性和OC分泌量各实验组和对照组与未加因子相比有明显增加(P<0.05)。细胞接种后期,成骨细胞ALP合成在机械组与未加因子相比明显减少(P<0.05),喷砂组和TPS组则仍然增加;OC分泌量在机械组和对照组变化不明显,而喷砂组和TPS组OC分泌与未加因子相比明显增加(P<0.05)。结论浓度为10ng/ml的外源性TGF-β1对不同钛片表面成骨细胞增殖有一定抑制作用。外源性TGF-β1和粗糙钛片表面对成骨细胞分化有一定协同促进作用。

Objective To investigate the consequential changes of proliferation and differentiation of fetal rat osteoblasts incubated with TGF - β1, and to make a primary understanding of the relationship between TGF - β1 and different surface treatment of titanium during osteoblast growth. Methods The primary osteoblasts separated from calvarium of 10 fetal rats were cultured and a finite cell line was established. Then cells were seeded on the different surfaces of Ti disc（Smooth surface,Sandblasted surface and TPS surface）and TGF- β1 with a dose of 10nm/ml was administrated to osteoblasts incubated on the different surfaces of Ti disc. At the different time points after addition of TGF- β1, cell proliferation activity was evaluated with MTT colorimetric assay. At the different time points ofter addition of TGF- β1 ,ALP activity of cellular layer and OC content of medium were tested to evaluate the differentiational status of osteoblasts. Osteoblasts were cultured on the surface of glass in the same condition as control group and tested as same as the experiment groups. Results The results of MTT assay showed that TGF - β1 inhibited proliferation of osteoblasts in all groups in a time- dependent manner. In the early phase,TGF -β1 had significantly decreased proliferation acivity of osteoblasts on the smooth surfaces or under control conditions on glass. Osteoblast proliferation of all groups was inhibited obviously compared with non - addition of TGF-β1, at 7 days. The present study showed thet TGF- β1, added to the cultured cells in vitro in the early phase, had signifcantly increased ALP activity of cells on all surfaces. Compared with non - addition of TGF-β1,cell ALP activity was inhibited on the smooth surfaces while promoted on the sandblasted surfaces and TPS surfaces as time went by. It was also found that TGF- β1 had significantly increased OC content of osteoblasts of the experiment groups（except TPS） in the early phase. When osteoblasts had matured, OC content between addition and non- addition of TGF - β1 on the smooth surface showed no difference while OC content was obviously increased on the rough surfaces （ P 〈0.05）.Conclusion Extraneous TGF- β1 with a dose of 10ng/ml inhibited proliferation acivity of osteoblasts on the different surfaces of Ti disc and the effect was time - depemdent. Extaneous TGF-β1 levels affected osteoblast differentiation.