| Background:Approximately 5 to 10% of bone fractures cases are associated withdelayed union or non-union.Thus,how to enhance the fracture repair process,normalize the rate of healing,and decrease the likelihood of nonunion has garneredconsiderable interest from academia.The gold standard for attaining bony union in non-unions is autologous bonegraft.However,harvesting this graft is not without complications:multiplesurgical sites and incisions lead to a decrease in weight bearing status andfunction,and to an increase in blood loss,operative time,risk of fracture,riskof infection, and pain. Furthermore,autograft does not necessarily lead to fusion.These complications underscore the need to understand the molecular and cellularevents critical to bone formation,and to develop innovative strategies to enhancebone regeneration.Fracture repair can be envisioned as involving five distinguishable processes,including the immediate response to injury,intramembranous bone formation,chondrogenesis,endochondral bone formation leading to the reestablishment ofload bearing function, and bone remodeling. During fracture repair, a number ofgrowth factors, cytokines, and their cognate receptors are present at elevatedlevels in and around the fracture site.Many of these proteins are normallyexpressed in skeletal tissue, and others are released from associated inflammatorycells at the site of injury.The induction of these proteins is regulated bothspatially and temporally,suggesting that they play an active role in promotingfracture repair.Indeed several bone growth factors have been identified, such as BMPs,TGF-β,IGFs,FGFs and so on,and known to impact significantly on the differentiationof mesenchymal cells into osteogenic cells and subsequent bone formation.We all knew injury to muscles is very common while fracturing. Among the growth factors,IGF-1 is widely investigated because of their capacity of inducing new bone formation by the way of endochondral ossification and intramembranous ossification, and stimulating the proliferation and fusion of myoblasts. IGF-1 can stimulate muscle healing and have a considerable effect on the treatment of muscle injuries. Given the relatively short half-life of IGF - 1 in biologic systems, evaluation of the efficacy of IGF- 1 which is directly injected has been hampered by the time of delivery models, high cost, treatment repeated and dosing regimes used. With the development of gene engineering, transient expression is a desirable benefit and readily available, delivery of IGF - 1 using gene therapy strategies may circumvent some of the issues of time of delivery, localization, cost and sustained expression.Objective: To observe the effects of myoblasts carrying hIGF-1 gene injected into the C57BL/6J mice on the healing of tibial fracture.Methods: Forty-two eight-week-old C57BL/6J mice (22~25g, male) were used for this investigation .Closed fracture of tibiae were produced in left tibiae of the mice which were anesthetized with pentobarbital (30mg/kg) by intraperitoneal injection and then divided randomly into two groups:the experimental group and the control group. Experimental animals were injected with 3. 0×106 myoblasts by BrdU labeling, while control animals were injected with 0.3ml saline solution, seven mice in each group were executed on the 14th,21th,28th day after operation and the bone callus were stained by hematoxylin and eosin. quantitative analysis of the callus area and the relative amounts of bone, cartilage, and fibrous tissue in the callus of each section were calculated by computer program.At 2,4 weeks after fracture, evaluating cells surviving and the secretion of hIGF-1 were done by immunohistochemical staining of the skeletal muscle into which myoblasts carrying hIGF-1 gene and saline solution were injected, Mean gray value of the positive-stained cells was measured by a digital image processing software to evaluate the level of the expression of IGF - l;the anteroposterior and lateral radiographs of the left tibiae and transmission election microscopy(TEM) examination of the bone callus were taken.Results: The BrdU staining in mice transplanted with myoblast were positive and hIGF-1 was secreted, but the BrdU and hIGF-1 staining in mice transplanted without myoblast were negative. The values of protein expression of IGF - 1 in experimental group were significantly elevated compared with that in control group, but in experimental group at 2 weeks, the expression level of IGF-1 were not significantly higher than that in the experimental at 4 weeks (P>0. 05). Quantitative analysis of histological examination showed the amount of external callus in the experimental group were greater than that in the control group at each period, and the difference at 3,4 weeks demonstrated statistical significance ( P<0.05 ). The X-ray observation showed the experimental group demonstrated massive callus formation, and the difference at 2,4 weeks demonstrated statistical significance(P<0. 05). TEM observation showed the experimental group were better than the control in the quantity of generated cells in bone, the degree of active, the number and distribution of collagen, etc.Conclusions: 1,The myoblast carrying hIGF - 1 gene transplanted into the C57BL/6J mice can survive for a certain period of time, and can secrete hIGF-1; 2,Local injection of myoblasts carrying hIGF-1 gene may play some significance in the healing of tibial fracture in C57BL/6J mice; 3,Local injection of myoblasts carrying hIGF-1 gene may play some significance in the healing of tibial fracture in C57BL/6J mice by elevating the level of IGF-1 in serum and (or) around the fracture site through the type of action: paracrine and (or) endocrine.
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