Li Xiulan, Zhang Yang, Shi Yijian, Wang Zengli
Tianjin Institute of Orthopedics, Tianjin, P.R. China.
Zhongguo Xiu Fu Chong Jian Wai Ke Za Zhi. 2006 Feb;20(2):107-11.
To explore the regulator factor of osteogenesis induced by the fibroblast in vitro so as to provide enough seeding cells for the bone tissue engineering.
The fibroblasts were isolated and purified from granulation of New Zealand rabbits, and they were incubated in the media of fibronectin (FN) 10, 20, 40, 60 and 80 microg/ml, respectively, in the experimental groups 1 - 5, but there was no FN in the control group. The markers for osteogenic features were investigated by fibroblast morphogenesis, calcium nodules formation ratios, labeling of tetracycline fluorescence, labeling of 3H-TdR, determination of osteocalcin, and labeling of 3H-proline within 2 weeks.
The morphological changes of the fibroblasts were manifested as transference from a long spindle to a round or multiple form, shifted nucleus increased in number, confluence and formed multilayered structure. There was a piling-up of calcium crystals that were gradually merged into foggy substances. The foggy substances increased and formed nodules. The calcium nodules formation ratios were as follows: 15.35% +/- 3.45% in the control group, and 53.73% +/- 9.49%, 75.21% +/- 9.80%, 98.34% +/- 15.20%, 61.83% +/- 10.04%, and 45.11% +/- 8.70% in the experimental groups 1-5, respectively. There was a significant difference between the control group and the 5 experimental groups at 14 days (P < 0.05), and a significant difference between the experimental group 3 and the other experimental groups at 14 days (P < 0.05). The histochemical study on the nodules with the specific labeling of tetracycline fluorescence indicated that the nodules were composed of new bones.
Fibronectin can stimulate the fibroblast to proliferate, secrete osteocalcin, and synthesize collagen fibrils. Fibronectin, in an optimal dose of 40-60 microg/ml, is capable of inducing the fibroblast to form the bone.
探讨成纤维细胞在体外诱导成骨的调节因子，为骨组织工程提供充足的种子细胞。
从新西兰兔肉芽组织中分离纯化成纤维细胞，实验组1 - 5分别置于含10、20、40、60和80μg/ml纤连蛋白（FN）的培养基中培养，对照组培养基中不含FN。在2周内通过成纤维细胞形态发生、钙结节形成率、四环素荧光标记、3H-TdR标记、骨钙素测定及3H-脯氨酸标记等方法研究成骨特征标志物。
成纤维细胞形态变化表现为从长梭形转变为圆形或多形，核移位数量增加，汇合并形成多层结构。有钙晶体堆积，逐渐融合成雾状物质。雾状物质增多并形成结节。钙结节形成率如下：对照组为15.35%±3.45%，实验组1 - 5分别为53.73%±9.49%、75.21%±9.80%、98.34%±15.20%、61.83%±10.04%和45.11%±8.70%。14天时对照组与5个实验组间差异有统计学意义（P < 0.05），14天时实验组3与其他实验组间差异有统计学意义（P < 0.05）。对结节进行四环素荧光特异性标记的组织化学研究表明，结节由新骨组成。
纤连蛋白可刺激成纤维细胞增殖、分泌骨钙素并合成胶原纤维。最佳剂量为40 - 60μg/ml的纤连蛋白能够诱导成纤维细胞形成骨。
