The Effects Of High Fat-fed/STZ Combined With Ovariectomy On Femoral Fracture Healing In Rats

Objectives The purpose of this study is to investigate the effect of type 2 diabetic with estrogen deficiency on fracture healing by establishing femoral fracture model of Hight Fat-fed/STZwith ovariectomy in rats.Methods Forty 2-month-old female SPF grade Sprague Dawley(SD) rats, Adaptive feeding after a week, they were randomly divided into four groups with 10 animals each,namely: 1) Fracture + control group(Sham+F) with normal diet, they were received intraperitoneal injection of citrate buffer at third weeks and exposed the abdominal cavity without removing the ovaries at fourth weeks, right femoral fracture was performed at tenth weeks; 2) Hight Fat-fed/STZ+ fracture group(HFD/STZ + F group) with high-fat diet, they were received intraperitoneal injection of STZ solution at third weeks and exposed the abdominal cavity without removing the ovaries at fourth weeks, right femoral fracture was performed at tenth weeks; 3) Ovariectomy + Fracture group(OVX+F) with normal diet, they were received intraperitoneal injection of citrate buffer at third weeks and ovariectomy was performed at fourth weeks, right femoral fracture was performed at tenth weeks; 4) Hight Fat-fed/STZ + Ovariectomy + fracture group(HFD/STZ + OVX +F group)with high-fat diet, they were received intraperitoneal injection of STZ solution at third weeks and ovariectomy was performed at fourth weeks, right femoral fracture was performed at tenth weeks; HFD/STZ + F and HFD/STZ + OVX + F rats were tested random blood glucose by tail vei blood at a week after injection of STZ solution, glucose value greater than 16.7mmol/L was considered as a successful model of type 2 diabetes,the weight and random blood glucose of rats in each group were monitored regularly. The rats with Ovariectomy or Fracture were got intramuscular injection of penicillin to prevent infection at 30 minutes before each surgery and 2 days after each surgery. The rats of each group were immediately taken X-ray to check whether there is a successful model after fracture surgery, All rats were taken X-ray photography again to observe the fracture healing and further to carry out the image score at 3 weeks after fracture surgery. All rats were killed after animal experiment, femoral specimens were decalcified for 12 weeks and embedded in paraffin, sliced, stained with Saffranin O stained, histological grading, Col-Ⅰimmunohistochemical staining, IRS score was used to express the positive protein expression of immunohistochemistry, The data were analyzed by SPSS20.0 software.Results 1 Weight results: at eighth weeks, the weight of rats in HFD/STZ+F group was significantly lower than Sham+F group(p<0.05), the weight of rats in OVX +F group was significantly higher than Sham+F group(p<0.05), the weight of rats in HFD/STZ+F group and HFD/STZ+OVX+F group were significantly lower than OVX +F group(p<0.05), the weight of rats in HFD/STZ+OVX+F group was significantly higher than HFD/STZ+F group(p<0.05); At twelfth weeks, the weight of rats in OVX +F group was significantly higher than Sham+F group(p<0.05), the weight of rats in HFD/STZ+F group and HFD/STZ+OVX+F group were significantly lower than OVX +F group and Sham+F group respectively( p<0.05). 2 Blood glucose results: at fourth, eighth and twelfth weeks, the blood glucose of rats in HFD/STZ+F group and HFD/STZ+OVX+F group were significantly higher than OVX +F group and Sham+F group respectively(p<0.05).3 Radiographic findings: 1) score of fracture healing: the score of fracture healing in HFD/STZ+F group(1.44±0.52), OVX+F group(1.64±0.20) and HFD/STZ+OVX+F group(1.22±0.44) were significantly lower than Sham+F group (2.49±0.53) respectively( p<0.05), which indicated that the fracture healing in HFD/STZ+F group, OVX+F group and HFD/STZ+OVX+F group was poor. 2) Area of callus: the area of callus in HFD/STZ+F group(1058.63±301.14) and HFD/STZ+OVX+F group(1375.13±390.60) were significantly higher than OVX+F group(690.38±288.12)and Sham+F group(611.25±57.53) respectively( p<0.05), the area of callus in HFD/STZ+OVX+F group(1375.13±390.60) was significantly higher than HFD/STZ+F group(1058.63±301.14)( p<0.05). 4 Histological results: 1) Saffranin O stained: the union rates in HFD/STZ+F group(12.5%), OVX+F group(20%) and HFD/STZ+OVX+F group(0) were significantly lower than Sham+F group(70%) respectively( p<0.05),which indicated that the fracture healing in HFD/STZ+F group, OVX+F group and HFD/STZ+OVX+F group was poor. 2) Immunohistochemical results: the IRS score of expression of the positive protein immunohistochemistry in HFD/STZ+F group, OVX+F group and HFD/STZ+OVX+F group were significantly lower than Sham+F group, which indicated that the fracture healing in HFD/STZ+F group, OVX+F group and HFD/STZ+OVX+F group was poor.Conclusions 1 The healing of femoral fractures in rats induced by high fat and STZ was delayed, which indicated that type 2 diabetes has a negative effect on fracture healing; 2The delayed healing of femoral fractures in ovariectomized rats, which suggests that estrogen deficiency has a negative effect on fracture healing; 3 The healing of femoral fractures in rats induced by fat-fed/STZ combined with ovariectomy was delayed, which indicated that type 2 diabetes combined with estrogen deficiency delayed fracture healing and shown a more serious tend of negative effect on fracture healing than the single model.