The Experimental Study Of Metformin On Steroid-induced Osteonecrosis Of The Femoral Head In Rats

Objective:To investigate the effect of metformin on the prevention of steroid-induced osteonecrosis of the femoral head,metformin was given at the same time when steroid-indduced osteonecrosis of the femoral head was induced in model rats,and provided evidence for prevention of osteonecrosis of the femoral head possiblely caused by long-term or higher doses of glucocortic oid.Methods:60 healthy SD rats were randomly divided into three groups,including the group A?blank control group?,group B?modle group?and group C?intervention group?,20 rats in each group.Rats in group B and group C were injected with prednisolone acetate 24.5mg/kg once a week for 12 weeks.Rats in group C were given 0.2g/kg.d metformin for 12 weeks,rats in group A and group B were given 2mL physiological saline daily,which lasted for 12 weeks.During the experiment time,all the animals were given an intramuscular injection of gentamicin sulphate?50,000U/rat?weekly for infection prophylaxis.At the 12th week of experiment,blood samples about 2mL were taken from every rats by eyeball removal,the blood samples were all centrigugated,and the serums above were collected carefully.The levels of N.terminal propep tide of type I collagen?PINP?,bone specific alkaline phosphatase?BALP?,and beta-crosslaps??-CTx?in serum were tested by enzyme-linked immun osorbent assay?ELISA?.¹⁸F-NaF Micro-PET images were taken in 10 rats from every group after blood collection,and assessed the metabolism of the femoral head by comp aring the standardized uptake value?SUV?,then the rats were killed by spinal dislocation method and remove the bilateral femoral heads were removed.Observing and comparing changes in the general morphology of the specimen,using 10%formalin solution fixed specimen,10 femoral heads selected from each group for Micro-CT scanning and Calculating the microstructural paramete-rs?BV/TV?BS/BV?Tb.Th?Tb.N?Tb.Sp and CWT?.All specimens were made pathological sectionswere after fixation and decalcification,and HE staining was performed for histomorphological observation.Data were expressed as mean±standard deviation????±s?,and analyzed by One-Way ANOVA test using SPSS22.0 software.P<0.05 means statistical significance.Results:1.Serum bone turnover marker testPINP:The average concentration of PINP in group B was?336.31±38.66?ng/mL,which is higher,compared with that in group A?258.45±57.34?ng/mL,there was a significant difference?P=0.000<0.05?,but it was statistically lower than group C?381.76±33.08?ng/mL,there was a significant difference?P<0.05?.And there was a statistically significant difference between the group A and group C?P=0.000<0.05?.BALP:The average concentration of BALP in group B was?3.83±0.26?ug/L,which is higher,compared with that in group A?3.56±0.16?ug/L,there was a significant difference?P=0.000<0.05?,but it was statistically lower than group C?4.21±0.23?ug/L,there was a significant difference?P=0.000<0.05?.And there was a statistically significant difference between group A and group C?P=0.003<0.05?.?-CTx:The average concentration of?-CTx in group B was?46.08±4.35?pg/mL,which is higher,compared with that in group A?35.96±4.84?pg/mL,there was a significant difference?P=0.000<0.05?,and it was statistically higher than that in group C?38.94±4.69?pg/mL,there was a significant difference?P=0.000<0.05?.And there was a statistically significant difference between group A and group C?P=0.003<0.05?.Group C was slightly higher than that in group A,but there was no statistically significant difference between the two groups?P=0.1>0.05?.2.Micro-PET/CT imaging resultsSUVmax:The average value of SUVmax in group A was?3.52±0.25?,which is lower compared with that in group B?5.14±0.27?,there was a significant difference?P=0.000<0.05?,and it was statistically lower than that in group C?6.69±0.36?,there was a significant difference?P=0.000<0.05?.Group C was higher than that in group B,there was statistically significant difference between the two groups?P=0.000<0.05?.The average value of SUVmin in group A was?2.88±0.27?,which is lower compared with that in group B?3.72±0.28?,there was a significant difference?P=0.000<0.05?,and it was statistically lower than that in group C?4.28±0.24?,there was a significant difference?P=0.000<0.05?.Group C was higher than that in group B,there was statistically significant difference between the two groups?P=0.000<0.05?.The average value of SUVmean in group A was?3.19±0.21?,which is lower compared with that in group B?4.32±0.20?,there was a significant difference?P=0.000<0.05?,and it was statistically lower than that in group C?5.22±0.32?,there was a significant difference?P=0.000<0.05?.Group C was higher than that in group B,there was statistically significant difference between the two groups?P=0.000<0.05?.Micro-CT scan:The trabeculars in group A were arranged neatly,thick,and the gap was small.In group B,the trabeculars became thinner,the gap became wider,the number decreased,and trabecular fracture became more.The trabecular in group C was thicker than that in group B,the gap was smaller than that in group B,and the number was more than that in the group B,and there was no obvious trabecular fracture.The BS/BV in group C was smaller than that in group B,there was a significant difference?P=0.001<0.05?.The Tb.Th in group C was higher than that in group B,there was a significant difference?P=0.01<0.05?.The Tb.N in group C was more than that in group B,there was a significant difference?P=0.000<0.05?.And the Tb.N in group B was less than that in group A,there was a significant difference?P=0.000<0.05?.The Tb.N in group C was less than that in group A,but there was no significant difference?P=0.748>0.05?.There was no significant difference in BV/T,Tb.Sp and CWT among the groups?P>0.05?.3.Morphological histological observationGeneral morphology observation:The femoral heads cartilage in group A was intact,smooth,and there was no deformation collapse.The femoral heads in group B had dark surface and poor luster.There were small areas of superficial cartilage destruction,and some femoral heads were deformed to varying degrees.In group C,the surface of femoral headswas relatively smooth,part of the femoral heads became dark and had poor gloss,but there were no deformation and collapse.Histological observation:In group A,the trabeculars were arranged regularly,the bone cells were clear,the nucleus was in the middle,and occasional empty bone lacunae were found.The medullary hematopoietic cells were abundant,there were a few adipocytes,and there were more osteoblasts around the trabecular bone.In group B,The trabeculars were arranged sparsely and thinly.The structure was in disorder and there were more trabecular fractures.The necrotic bone cells and empty bone lacunae were significantly increased.The fat cells covered the bone marrow cavity.In group C the structure of trabecular was relatively complete and regular.There were few empty bone lacunae and scattered osteoblasts around the lacunae.The number of empty lacunae cells was significantly less than that in group B?P<0.05?.Conclusion.1.Metformin can increase serum bone formation markers PINN and BALP and reduces serum bone resorption markers of?-CTx in the model rats of steroid-induced osteonecrosis of the femoral head.2.Metformin can increase the number and thickness of trabecular and reduces bone surface area/bone volume value in the model rats of steroid-induced osteonecrosis of the femoral head.3.Metformin can reduce the rate of empty bone lacunae in the model rats of steroid-induced osteonecrosis of the femoral head.Summary:Metformin has preventive effects on steroid-induced osteonecrosis of the femoral head.