Therapeutical Effects Of Geraniin On Experimental Osteoporosis Fracture In Rabbits Based On Wnt/?-catenin Signal Pathway

[Objective]To study the therapeutic effect of geraniin on Osteoporotic fracture(OPF)in rabbits.To investigate the regulating effects of geranim on ?-catenin,LEF1,Frizzled2,Dkk1,Wnt5a in Wnt/?-catenin signaling pathway of OPF rabbit bone tissue.[Methods]1.36 rabbits were randomly divided into 6 groups:sham operation group,OPF model group,geraniin at 20,10 and 5 mg/kg groups and simvastatin at 5 mg/kg group.2.Rabbit OPF model was duplicated:The model group rabbits were resected bilaterally with ovaries and the oviducts were ligated.After bone mineral density(BMD)were measured to confirm the successful duplication of osteoporosis model,bone saw wire was used to excision the bone of the middle segment of radius about 3 mm to duplicate the fracture model.The bone in middle segment of radius was exposed but not excised in the sham group.The OPF model was identified by X-rays.3.After the OPF model was successfully replicated,the corresponding drugs were given by gavage for 1 time/day according to the above groups.After 8 weeks of continuous administration,the samples were taken and set aside.All animals were subcutaneously injected with tetracycline(30 mg/kg)on the 11th day before beingexecuted as the first fluorescent marker,and calcein(6 mg/kg)was subcutaneously injected on the 4th day before being executed as the second fluorescent marker,the interval between the two fluorescence labeling was 7 days.4'.The fractured radius was scanned by PET/CT machine and the Region of interest(RIO)was reconstructed in three dimensions.The stereometric indicators of bone trabecular were analyzed,which including bone volume/total volume(BV/TV),trabecular thickness(Tb.Th),trabecular number(Tb.N),bone surface/bone volume(BS/BV),trabecular separation(Tb.Sp)and so on.5.The fourth lumbar vertebrae of rabbits in each group were made into non-decalcified bone slices,and the laser confocal microscope was used to take photos and calculate the Mineral addition rate(MAR).6.The paraffin sections of rabbit femur were made into decalcified sections,and the expressions of ?-catenin,LEF1,Frizzled2,Dkk1 and Wnt5a protein in femur were detected by Immunohistochemical(IHC)assay using DAB staining method.[Results]1.The results of bone mineral density and X-ray showed that OPF model was successfully established.2.PET/CT of radial fracture showed that after 8 weeks of administration,20 mg/kg,10 mg/kg,5 mg/kg geraniin group and 5 mg/kg simvastatin group could significantly increase BV/TV,Tb.Th,Tb.N and decrease BS/BV,Tb.Sp compared with the model group.3.MAR results of the fourth lumbar spine showed that after 8 weeks of administration,20 mg/kg,10 mg/kg,5 mg/kg geraniin group and 5 mg/kg simvastatin group could significantly increase MAR compared with the model group.4.Immunohistochemical staining results showed that geraniin could up-regulate the protein expression of p-catenin,Frizzled2,LEF1 and down-regulate the protein expression of Dkk1,Wnt5a with a dose-effect relationship.[Conclusion]1.Geraniin has obvious therapeutic effect on rabbit OPF,which can improve bone microstructure,promote bone formation and increase MAR,and has dose-effect relationship.2.Geraniin significantly up-regulates the protein expression of ?-catenin,Frizzled2,LEF1 and down-regulates the protein expression of Dkk1,Wnt5a,which can promote fracture healing by regulating Wnt/?-catenin signaling pathway.