Experimental Study Of HGF And TGF-β₁of Gastrocnemius During Distraction Osteogenesis Of Rabbit Tibia

ObjectiveTo establish the rabbit model of tibial lengthening using the monolateral external fixation. To observe the structural change of gastrocnemius and changes of HGF and TGF-β1expresssion during rabbit tibia distraction osteogenesis, so as to understand the ability of adaptive remodeling and change rule of gastrocnemius and discuss the remodeling molecular mechanism.MethodsForty-seven rabbits were randomly divided into the experimental group and the control group. The osteotomy was done in the control group (A group) but they wouldn’t be distracted and be killed after7days of the operation. According to the distraction degree, the experimental group was divided in three groups, the1cm group (B group),2cm group (C group),3cm (D group). Every group was subdivided into five groups based the mineralized times after the lengthening process, including the0day group (A1,B1,C1group), the7days group (A2,B2,C2group), the14days group (A3,B3,C3group), the28days group (A4,B4,C4group), the56days group (A5,B5,C5group). Every subgroup included three rabbits. The experimental group wad performed the tibial osteotomy fixed with the monlateral external fixator and began distracted after5days of the operation in a rate of lmm/d for twice a day. The X-ray films were taken first, then the rabbits were killed at different time. To observe the histological morphology and ultrastructure changes at each consolidation time point by means of light microscope (HE staining). The extended gastrocnemius of different groups were performed immunohistochemical staining to determine HGF and TGF-β1expression during different time.Result1. It can be seen by X-ray that callus images gradually changed clearly following the consolidation period, the amount of callus increased comparing with the previous callus images, the cortical bone and medullary cavity. At last, the callus union was acquired. It also can be seen that the amount of callus in A group is much more than B group and C group at same consolidation period. The amount of callus in B group is better than C group.2. The structural changes were observed by light microscope including the muscle fibres’degradation, muscle plasma crumbling degeneration, muscle nuclei internalization. During the consolidation period, muscle repaired itself gradually, until the late stage of consolidation period (56days), the structure of gastrocnemius returns to normal. Following lengthening degree increased, the histological damage of gastrocnemius tend to increase. The endomysial fibrosis and nuclear retraction phenomenon increased significantly. When the distraction is up to30%, this phenomenon is more significant and muscle damage may be irreversible.3. The HGF and TGF-β in control group was a little but negative; at consolidation0day the expression of HGF was strong positive; at consolidation7days was also active, but less than the0day; from the consolidation14days, the expression of HGF began decline; at the consolidation28days, the expression of HGF further decline, at the consolidation56days, the expression of HGF was the same as the control group. The expression of TGF-β1was less than the control group; then it began increase and at the consolidation14days, it was strong positive. Then the expression of TGF-β1began decline, at the consolidation56days, it became normal.Conclusion1. During slow limb lengthening with a degree, the subclinical peripheral muscle injury is reversible. The natural repair of the damage muscle has begun as soon as the muscle was injury. When beyond this distraction range, it may induce irreversible damage.2. During the process of tibia distraction osteogenesis, the rule of expression change of HGF and TGF-β1was in accordance with the rule of structure change of gastrocnemius. The result may cause that HGF and TGF-β1mediates the functional remodeling of gastrocnemius.