| Objectives This study was designed to investigate the effect of insulin treatment on femoral fracture healing in type2 diabetes mellitus rats with osteoporosis.Methods Fifty Sprague-Dawley female rats, aged 8-weeks-old and weighing 212~236 g were used in our study.After one week of acclimatization,the rats were randomly divided into 5 groups with 10 rats per group.Control group(F; sham+ fracture), ovariectomy group(OF; Ovx+fracture), type2 diabetes mellitus group(DF; T2DM+Ovx), type2 diabetes mellitus and ovariectomy group(DOF; T2DM+Ovx+Fracture), insulin treatment group(DOFI; T2DM+Ovx+Fracture+Insulin). Combination of high-fat diet-fed and low-dose streptozotocin injection to establish rat model of type 2 diabetes mellitus, the insulin sensitivity index and non-fasting blood glucose were measured to ensure the successful establishment of T2 DM model. Insulin(2~4 IU/day/rat) was administered to group DOFI from the 5th week, while the rats in groups F and DOF were administered with the same amount of distilled water. The postmenopausal osteoporosis model was established by ovariectomy bilaterall, after the success of modeling,a closed femoral fracture surgery were performed at the right hind limbs with internal fixation.The X-Ray photograph were taken post-operation to make sure the classification of femoral fracture can meet the requirements of our experiment.After 3-weeks of surgery,the X-Ray radiographs were taken again,than the animal were sacrificed and samples were obtained. Body weight and blood glucose levels were monitored closely in the experiment; ELISA was used to detect the serum osteocalcin concentration;The X-ray film of the fracture site was taken to evaluate the healing condition;Measurement of bone mineral density in fracture site by dual energy X-ray absorptiometry;HE staining was used to observe the morphological changes of the fracture site; Immuohistochemistry was used to assess OCN expression in the healing fracture tissues.Results 1 At the beginning of the study,the baseline weight of all rats was similar.At the end of the study,group DF and group DOF were lighter than group F(P<0.05). Group OF and group DOFI were heavier than group F(P<0.05).2 The non-fasting blood glucose levels of diabetic rats(DF,DOF,DOFI) was found to be increased when compared to the non-diabetic rats(P<0.05),after the treatment of insulin,the blood glucose levels of group DOFI tended to me normal.3 High-fat diet group presented lower insulin sensitivity than that in regular chow-fed-rats(P<0.05). 4 Serum levels of osteocalcin was measured to provide an evaluation of bone formation activity after fracture under insulin treatment. The serum osteocalcin concentration of group F was significantly higher than that other groups(P<0.05),Insulin increased OCN levels by 28.7% compared with group DOF(P<0.05),the serum osteocalcin concentration of group DOF was lower than that of group OF and group DF,but there is no statistical significance.5 The radiographic score of group F was higher than the other groups(P<0.05),the radiographic score of group DOFI was higher than group DOF(P<0.05),the radiographic score of group DOF was lower than group OF and group DF,but the difference was not statistically significant. 6 Histological observation showed that the fracture healing in group F was better than others.Compared with group DOF,group DOFI has a high degree of bone healing,good continuity of callus, more bony callus, little cartilage cells but no cartilage island,the trabecular bone was more mature,the formation of lamellar bone was found. 7 Immunohistochemical staining showed that the DOFI group compared with DOF group, positive expression of osteocalcin is more obvious.Conclusions 1 The femoral fracture healing was impaired in the diabetic model group(DF); The femoral fracture healing was impaired in the ovariectomized group(OF);The femoral fracture healing was impaired in the diabetes mellitus combined with osteoporosis group(DOF).2 The impaied fracture healing was improved by insulin in rats with type 2 diabetes mellitus and osteoporosis. |
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