| Objective 1.To assay the serum IGF-1R level of PDO patients,preliminarily exploring the underlying mechanism of PDO.2.To compare the difference between PDO rat model and DO rat model,and to explore roles of IGF-1R/?-catenin signaling pathway on the imbalance of boneglucose metabolism in PDO.3.To confirm the therapeutic effect of Psoralea carylifolia L.on PDO and to explore the regulatory action of IGF-1R/?-catenin signaling pathway in this process.Methods 1.Clinical researchTwenty serum samples of PDO patients and postmenopausal normal women respectively who were admitted to the Spine Surgery Department of The First Affilitated Hospital of Guangzhou University of Chinese Medicine,were collected.The serum IGF-1R levels were assayed by ELISA.2.Animal research(1)Establishment of rat DO model and underlying mechanism explorationSD female rats were diveded into 4 groups: SHAM group,OP group,DM group DO group.OP group and DO group rats were bilaterally ovariectomized,small mass of celiac fat was taken out from SHAM group rats.After 2 weeks,DM group and DO group rats were fed with high-sugar/fat chow for 4 weeks,then were injected with STZ.3 and 4 days later,the fasting blood glucose was tested consecutively.12 weeks(6 weeks after STZ injection),16 weeks,20 weeks and 24 weeks after ovariectomy,rats in all group were respectively sacrificed and the following parameters were tested:(1)(Areal)Bone mineral density(BMD),bone mineral content(BMC)and bone area(Area)of L1-3 vertebrae were measured by dual-energy x-ray absorptiometry(DXA).(2)Compression strength of L1 vertebrae by compression testing.(3)volume BMD(vBMD),relative bone volume to tissue volume(BV/TV),trabecular number(Tb.N),trabecular space(Tb.Sp),trabecular thickness(Tb.Th)of L2 vertebrae scanned by micro-CT.(4)Histomorphology of L3 vertebrae.(5)Relative expression of IGF-1R?GSK3???-catenin mRNA of L4 vertebrae by qPCR.(6)Examination of proteins expression of IGF-1R?GSK3???-catenin by western blot(2)Treatment of postmenopausal DO rats with Psoralea carylifolia L.and the mechanism explorationThe remaining DO rats were divided into 4 groups: model group(PDO),metformin group(Me),high-dosage Psoralea carylifolia L.group(P-H)and low-dosage Psoralea carylifolia L.group(P-L).Me group rats were administered orally with metformin,P-H group with high-dosage Psoralea carylifolia L.,P-L group with low-dosage Psoralea carylifolia L.and DO group with distilled water.2 milliliters of liquid were equally given to all group rats.Parameters as “(1)(1)-(5)” were tested.Results 1.Clinical researchSerum IGF-1R level of PDO patients was significantly higher than that of control group(P<0.05).2.Animal research(1)Comparison of DO models and exploration on the pathogenesis of PDO(1)Establishment of DM modelAfter peritoneal STZ injection,rats in both DM group and DO group had developed DM(with diabetic morbidities of 72% and 64% respectively).(2)Establishment of the secondary OP(namely DO)Six weeks after STZ injection(at W12),BMD of lumbar vertebraes of rats in DM group was decreased with no significance compared with both SHAM group and OP group(P>0.05),and that in DO group was significantly increased compared with SHAM,OP and DM groups(P<0.01 and P<0.05).DM group showed insignificantly lower compression strength than SHAM did(P>0.05),while DO group showed notably lower compression strength than SHAM did(P<0.01),with no remarkable difference between DO and DM groups(P>0.05).(3)IGF-1R expression in rat lumbar vertebraesAt W20,expression of IGF-1R mRNA in both DM and DO groups was profoundly upregulated compared with in SHAM group(P<0.05 and P<0.01).At W24,expression of IGF-1R mRNA in DO group was profoundly upregulated compared with in both SHAM and DM groups(P<0.05 and P<0.01).At W20,expression of proteins including p-IGF-1R and p-GSK3?in DO group were strikingly upregulated in comparison with in SHAM group and p-GSK3? in DO group was strikingly upregulated,with upregulation trend of p-IGF-1R and p-?-catenin at W24.(2)The therapeutic effect of Psoralea carylifolia L.on rat PDO and exploration on its related mechanism(1)Blood glucose of all groupsAt W4,blood glucose of rats in Me group was notably lowered(P<0.01),and in both Me and P-H groups blood glucose was was notably lowered at W8 and at W12(P<0.01 and P<0.05),compared with in PDO group.(2)Improvement on OP in all groupAt W12,BMD,BMC and AREA in both Me and P-H groups were significantly increased in contrast to in PDO group(P<0.05).Compression strength was significantly enhanced in P-H group(P<0.05)but insignificantly in either Me group or P-L group compared with PDO group.vBMD and Tb.Th in P-H group were significantly increased compared with in DO group(P<0.05 and P<0.01),and vBMD in P-H group was significantly increased compared with in Me group(P<0.05).PDO group showed sparse and thin trabecula,far distance and abundant lipid droplets between trabecula.Me group and P-H group showed increased and thickened trabecula,decreased trabecula separation and lipid droplets compared with PDO group.P-L group showed similar morphology with trabecula slightly thickened compared with PDO group.(3)Expression of IGF-1R mRNA,GSK3? mRNA and ?-catenin mRNA in lumbar vertebraes of all groupsAt W12,expression of IGF-1R and GSK3? mRNA in both P-H and P-L groups trend to be downregulated,and ?-catenin mRNA in P-H group trend to be upregulated slightly,and OCN mRNA in P-H group remarkably upregulated(P<0.05)in comparison with in PDO group.Conclusion 1.Clinical researchIGF-1R may be the key mediator in the onset of PDO.2.Animal researchSTZ injection solely and STZ injection combined with ovariectomy are capable of establishing rats DO model(mimicking DO and PDO state respectively),with the latter inducing DO model more rapidly.IGF-1R/?-catenin signaling pathway may be a factor in the imbalance between glucose-bone metabolism in PDO.Psoralea carylifolia L.could cure PDO through improving blood glucose,enhancing bone mass and bone strength,and improving micorstructure probably via regulating IGF-1R/?-catenin signaling pathway. |
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