| Objective: The bone fracture cicatrization is a very complex wound repair process , with the influences and regulation of many factors,in which nervous system's control action holds a very important status. Studies found that nerve fibers played a regulatory role by the release of a variety of neurotransmitters, but the specific adjustment mechanism is not entirely clear. It is frequent to see children with metaphyseal fractures complicating with peripheral nerve injury in clinic, but the influence of peripheral nerve on metaphyseal fracture healing and its control mechanism needs to be further studied. In this experiment models of tibial metaphyseal bone defect with denervation were made, and the differences in healing of metaphysis between the experiment group and control group were compared with after a variety of physical examination and observation under microscope. Then the effect of neuropeptide on the healing of tibial metaphysis and its possible regulatory mechanism were explored, so to provide some theoretical basis for promoting children's metaphyseal fracture healing by improving neurological function. Methods: 40 healthy china white rabbits whose ages were 4-6 weeks, and body weights were 1.0-1.5kg were selected. Under the aseptic condition, vertical cut was marked in front of the left hind legs knee joint at the level of shinbone tubercle. The metaphysic of shinbone near-end and the epiphyseal plate that was like white thin line were exposed fully. Power drill whose diameter was 3mm was used to drill hole in epiphyseal plate parallel direction in the position 0.5cm under epiphyseal plate from shinbone inner side, to make a bone defect 1cm long approximately. What should be paid attention to was that don't injury the epiphyseal plate and the articular cartilage. Because the tibial metaphyseal damage of bone defect type was stable, it was not necessary to fix the injured legs inside and outside. All the young rabbits were randomly divided into two groups: the tibial metaphyseal damage group without innervation (experimental group) and that with innervation (control group). The sciatic nerve and femoral nerve of the left side of hind legs of young rabbit in experimental group were cut off. Sciatic nerve and femoral nerve were remained in the control group. Then all the experimental animals were put into the cage after the operation, and fed with unified standard feed unification. We should pay attention to the temperature and the prevention of wound infection. 5 rabbits in both groups were selected randomly and killed by air embolism to do radiology examination in 1, 2, 4, 8 weeks after the operation. Growth bone callus and connection of bone bridges in the bone defect of animals in each group were observed. Tibia in both side were gotten and the soft tissue around the bone were stripped to observe the growth of callus. The healing callus in the injury metaphyseal bone was selected and examined with light microscope to do tissue analysis to evaluate the healing of bone fracture and callus quality and quantity.Results: Result of X-ray examination: There was no significance in the result of X-ray inspections between the two groups in 1 week after the operation, and anomalous transparent area with uneven density in metaphyseal bone could be found. The osteotylus quantity of the two groups: experimental group:0.09±0.02,control group:0.07±0.01,P>0.05,it was no significance。The stripped high density shade occurred in tibial metaphysis in 2 weeks after the operation in each group. Compared with control group, the quantity of bony callus in experimental group was more, but the density was lower and uneven. The osteotylus quantity of the two groups: experimental group:0.19±0.01,control group:0.16±0.01,P<0.01,The difference between the two groups was significant(P<0.01).The same result could be found in each group in 4 weeks after the operation. The osteotylus quantity of the two groups: experimental group:0.31±0.02,control group:0.28±0. 02,P<0.05 , The difference between the two groups was significant(P<0.05).X-ray results showed the massive over-growth osteotylus in experimental group in 8 weeks after the operation, the osteotylus highlight in the surface of bone, osteotylus models the shape to be mature gradually in the control group. The osteotylus quantity of the two groups: experimental group:0.35±0.03,control group:0.26±0.02,P<0.01,The difference between the two groups was significant(P<0.01). With the visual observation of the roughly specimen in tibial metaphysis, the osteotylus quantity of the two groups are of no obvious differences in 1, 2 week after the operation. The osteotylus quantity was more in experimental group than that in the control group in 4 weeks after the operation, the osteotylus filled the bone damage, and was higher than the bone surface. The osteotylus was obviously higher in the bone surface in 8 weeks after the operation in experimental group, assumes the excessive growth, but the bad intensity. The bone damage place of tibial metaphysic is filled with osteotylus in the control group, smooth, close to normal bone tissue. Specimen carries on the histology observation through the light microscope, massive fiber structure prolifera- tion is presented in experimental group the 1st week after the operation, few tiny newborn trabecular bone forms; Hyperplasia of fiber structure is active in the control group. Abundant thick newborn bone trabeculae form, the osteoblast is obvious its circumference. Histological score result of the two groups 1 week after the operation: 2.4±0.6, 3.0±0.7. The histological score difference of two groups is non-statistical significance in 1 week after operation (P>0.05). After the 2nd week of the operation, the newborn trabeculae of experimental group forms increasingly, but the osteotylus grows sparsely, and is low-density, disarranged, and the osteoclast are seen. Many knitting bone form in the control group with high maturity, the osteotylus is compact, along the line of force arrangement, assumes the stockade shape, external callus joins forces the bridge joint at some place obviously. Histological score result of the two groups 2 weeks after the operation: 4.3±0.8, 5.5±0.7,The histological score difference of two groups have statistics significance in 2 weeks after operation(P<0.05). Woven bone forms 4 weeks after the technique in experimental groups with low maturity, the osteotylus are more, but relatively sparse, Trabecular bone is narrow, Osteoi is thick. There are still many osteoclast. Woven bone in control group are further increaseingly mature and fuses gradually, new bone connect with each other, the bone damage department is stuffed, with rule arrangement, born trabeculae is basically wide and compact, the gap between born trabeculae reduces. Histological score result of the two groups 4 weeks after the operation: 5.5±0.6, 6.7±0.8. The histological score difference of two groups have statistics significance in 4 weeks after operation (P<0.05). The later period of bone fracture cicatrization in 8 weeks after the operation, the osteotylus grow overly in the experimental group, the maturity is low, the arrangement is disorderand sparse. Osteotylus connect into film in control group, arrange rulely along the line of force, the structure is close to normal bone tissue, the proportion of the osseous callus in the newborn callus's is higher obviously than in the experimental group, the fiber structure quantity are relatively massive in the group without the control of the innervation in each phase spot . The histological score difference of two groups have statistics significance in 8 weeks after operation. SP immunohisto- chemical staining observeing one week after the experiment in the experimental group, we can see that some SP expression in inosteotylus, in a scattered distribution points, a yellow staining. SP-positive expression can be seen in bone fracture callus tissue in control group, focalling distribution along the vascular, a brown staining. Only a very low level of SP expression can be seen in 2, 4 weeks after the experiment in inosteotylus of the experimental group, a large number of SP-positive nerve fibers distribution present in marginal woven bone in control group, and increase gradually, with significantly enhancing dark-brown stain. SP expression level is still very low 8 weeks after the experiment in experimental group,there is still more positive SP expression in control group, but significantly decreased compared to the point four weeks after the experiment,and was stained brown.Conclusions: 1. In the bone fracture cicatrization process,the distribution of Substance P is concerned with metaphyseal fracture restoration, Substance p plays an important action, especially on the healing of metaphyseal injury. 2. The peripheral nervous system has a regulative function on the metabolism and shape formulation of metaphysis bone, so to help for the normal growth of ossify. 3. The concentration of Substance P in fracture influence the bony callus quantity and quality and biomechanics intension, the diminution of Substance P result in excess bony callus,but the mechanism need to be futher studied.4. The metaphysis fracture complicating with peripheral nerve damage should be treated as soon as possible, not only the fracture,but also the injured nerve.It is necessary to repair the nerve.
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