| Fracture healing is an extremely complex biological repair processes, and it is affected by many factors. How to promote fracture healing has been one of the important topics for many scientists. It is the nervous system that has a regulatory role for human physiological and pathological processes. In normal bone developing and bone repairing process, there are a large number of nerve fibers in bone tissue. In the fracture healing process, callus formation and reconstruction are carried out under the regulation of the nervous system, and it is necessary that the normal fracture healing process needs complete innervation. Nervous system regulation of fracture healing includes both central and peripheral, and these two respects are interrelated and mutually each other. In clinical practice it is often observed that fracture patients who accompanied by nervous (central nervous system/peripheral nerve system, CNS/PNS) injury can often be seen in abnormal growth of callus, and even appear in heterotopic ossification in the muscle. This phenomenon has led to people's thinking, and also aroused the interest of many scholars. As early as 1880 Variot and Remy had found nerve fibers exist in the bone tissue of humans and animals by tissue staining method. Since then domestic and foreign scholars had conducted a series of studies in this field, and many hypotheses about the mechanism of nerve in the fracture healing had been also put forward. But the exact mechanism has remained unclear. Microstructure of fracture healing process can well reflect the changes of cellular process in bone formation and construction. It is an important tool for the research of bone formation and building process. Understanding the relationship between nerve tissue injury and fracture healing, we can have a more clear knowledge for the neural control of bone tissue. And there is a potential application prospects for the clinical treatment of nonunion, heterotopic ossification and osteoporosis.Objective Through the animal model establishment of Wistar rats femoral shaft fracture combined with nerve(CNS/PNS)injury, at different time nodes in the early stage of fracture healing, we measure the new allus on X-ray imaging and tissue biopsy. And to explore the differences and/or similarities of callus generation between the states of nerve loss and normal innervation in Wistar rats femoral shaft fracture healing, and also the differences and/or similarities between CNS loss and PNS loss.Methods The experiment was conducted from September 2008 to October 2009 at the the the Animal Center of Second Military Medical University.3-month-old male Wistar rats,120 (Chinese Academy of Sciences, Shanghai Experimental Animal Center), weighing 274-295 (284.81±4.04) g. The rats were randomly divided into three groups, and 40 rats each group. The groups were simple femoral shaft fracture group (SFF), ipsilateral femoral shaft fracture+sciatic nerve transection group (NIR), and femoral shaft fracture+T10 spinal cord transection group (SCI). The rats were made intraperitoneal anesthesia with 10% chloral hydrate (300mg/kg).①Modeling established for SFF group: The rats was fixed on the anatomy board in prone position. Take a 1cm long vertical incisionin in inner of patella of right lower limb. Longitudinal cut off the tendon tissue of the quadriceps, dislocate the patella to the medialr, inserted a diameter of 1mm intramedullary nail (10, injection needles, the needle had been cut off) in marrow cavity from the femoral concaveto the trochanteric. Pinch-off mid-femoral shaft, resulting in transverse fracture, pay attention not to cut-off the nail.②Modeling established for NIR group:The establishment of femoral shaft fracture modeling was same with the SFF group. Separated and found the sciatic nerve through the septum between the femur fore lintermuscular and lateral intermuscular,and the sciatic nerve was cut out about lmm.③Modeling established for SCI group:The establishment of femoral shaft fracture modeling was same with the SFF group. We took median dorsal incision, cut open the skin and subcutaneous tissue, peelied spinous muscle djacent to spinous prosess, exposed T10 lamina, excision T10 spinous prosess and lamina, revealed T10 spinal cord dura, the spinal cord was transection with a ship knife, cut about 2mm. And to facilitate hemostasis, wound was covered by gelatin sponge. Postoperativly, rats were single-cage reared, natural light, the cycle timing foot feed, and unlimited water drinking. In order to urination, the bladder of rats were given artificial massage every 6 hours until the automatic restoration of bladder voiding. And the padding were kept dry,which were replaced every 2 to 3 days. Respectively, the rats were killed after 2 and 4 weeks. Then the following data should be collected, such as X-ray bone callus width measurement, wet weighing of femur specimens weight, bone callus HE staining, immunohistochemical staining of bone callus, picric acid-fuchsin stain of non-decalcified sections of gravity, and static bone histomorphometry measurements.Results①The observation of microscope morphological 2 weeks later:SFF group, mainly osteoblasts, less newborn cartilage, dense bone callus, and less osteoid; NIR group, consisting mainly of cartilage cells, cell osteoporosis, little osteoblasts callus sparse and more osteoid; SCI group:rendering the transition state of new cartilage cells to mature chondrocytes, cartilage cells mainly, and low density callus. The observation of microscope morphological 4 weeks later:SFF group, osteoblasts increased, arranged more closely, there were scattered osteoclasts, newborn nutrition blood vessel had been formed, trabecular bone density, cis-arranged line of force laws, cortical bone could be seen scattered in the bone lacunae; NIR group, osteoblasts arranged looser compared with SFF group, osteoclasts disappeared, new nutrient vessels were more, and bone trabecular sparse. SCI group, osteoblasts scattered and no osteoclasts were found, bone trabeculae were arranged irregularly, sparsly, more lacunae.②The data were statisticsed by SPSS 11.0 statistical software. The comparison of the femur X-ray callus width measurement after 2 and 4 weeks postoperation. There were statistical different(P<0.05) between SFF group and NIR groups, SFF group and SCI group. While there were no statistical different (P> 0.05)between NIR group and SCI group; The comparison of the femur wet weight after 2 and 4 weeks postoperation. There were no statistical different(P>0.05) between SFF group and NIR groups, SFF group and SCI group. While there were statistical different (P< 0.05)between NIR group and SCI group.Conclusions Although callus formation is a necessary condition for fracture healing, the quality of fracture healing is neither the amount of callus quantity nor the size of callus volume, and it is the bio-mechanical strength of callus that truly reflects the quality of fracture healing. This study showed that different types of nerve injury on fracture healing process had different effects. Both acceleration of fracture healing process for the peripheral nerve cut-off, and delayed fracture healing process for the spinal cord cut-off,the early callus are pathologic. Therefore, a complete nerve is an indispensable condition of fracture healing. |
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