Objective: To investigate the expression of transforming growth factor beta1 (TGF-beta1) and insulin-like growth factor I (IGF-I) in new bone after low frequency micro movement.
Methods: Fifteen female sheep from Shandong province were involved in the study and their bilateral tibias transversely osteotomized in the middle shafts with a defect of 2 mm. The hind limbs were fixed with unilateral external fixators connected to a controlled micro movement device. Ten days after osteotomy, one hind limb of each sheep randomly was selected to perform micro movement at an amplitude of 0. 25 mm and a frequency of 1 Hertz, 30 min a day for 4 weeks (micro movement group). The other hind limb served as the control group. Five sheep were sacrificed at 3, 4 and 6 weeks after osteotomy, respectively, and specimens were harvested for detecting the expression of TGF-beta1 and IGF-I by immunohistochemistry and RT-PCR.
Results: Immunohistochemistry: In the third postoperative week in the micro movement group, the expression of TGF-beta1 was detected in different areas of new chondrocytes at the margin of callus, mainly in proliferating area, and IGF-I expressed in osteoblasts at the margin of endochondral ossification area, calcified and mature chondrocytes and osteocytes. There was seldom expression of IGF-I and little expression of TGF-beta1 in the corresponding area in the control one. In the 4th postoperative week in the micro movement group, the expression of TGF-beta1 diminished gradually with the mature of new bone and be located in extracellular matrix and osteoblasts around ossified areas; The expression of IGF-I reached the peak and be located mainly in osteoblasts of new bone surface, maturing osteocytes and calcifying osteoid. But there was little expression of them in the control group. In the sixth postoperative week in the micro movement group, there was a little expression of IGF-I expression but little expression of TGF-beta1; there was nearly no expression of them in the control group. In the micro movement group, the absorbance values of TGF-beta1 at 3 and 4 weeks and of IGF-I at 3, 4 and 6 weeks were significantly higher than those in control group (P<0. 05). RT-PCR: In the third and fourth postoperative weeks in the micro movement group, there was higher expression of mRNA of TGF-beta1 and TGF-I than those in control group; in the sixth postoperative week, the expression diminished gradually, but was higher than that in control group. The absorbance values of TGF-beta1 at 3 and 4 weeks and IGF-I at 3, 4 and 6 weeks were significantly higher than those of control group (P<0. 05).
Conclusion: Low frequency and controlled micro movement in the early stage of the fracture healing can promote the expression of TGF-beta1 and IGF-I. They worked together to regulate the process of the endochondral ossification, while in the late stage the differentiation of osteocytes and mineralization of osteoid were regulated mainly by IGF-I, which played an important role in regulating the cell biological behavior during micro movement.