The Role And Mechanism Of Androgen And Androgen Receptor In The Regulation Of Bone Mineral Density And Bone Biomechanical Properties In High Fat Diet Mice By Exercise

Purpose Obesity induced by high-fat diet could reduce bone mineral density and damage bone microstructure,thus lead to osteoporosis.Androgen play an important role in male bone growth,peak bone mass acquisition and bone mass maintenance,mainly via binding to androgen receptor(AR).The decrease of androgen is closely related to the occurrence of osteoporosis.However,the role and mechanisms with regard to androgens and androgen receptors in the regulation of bone mineral density and biomechanical properties in high-fat diet mice by exercise remain unknown.Our previous study has found that the serum level of andrgoen in obesity male rats were decreased significantly,and the present study established androgen deficiency and AR blockade mice models by castration and AR antagonist flutamide embedding,respectively.Then those above mice were fed with high-fat diet in combination with or without a 6-week exercise intervention to explore the role and mechanisms of androgen and AR in exercise-regulated bone mineral density and bone biomechanical properties of high-fat diet mice,which would provide appropriate exercise protocol and theoretical basis for preventing and treating osteoporosis caused by high-fat diet.Methods Fourty-two C57BL/6 male mice(6 weeks of age)were randomly divided into normal diet group(C,n=6)and high fat diet(HFD,n=36)group.The mice in the normal diet group weren't exerted any intervention,whereas mice in high-fat diet group were further divided into 6 groups: high-fat diet control group(HFD,n = 6),high-fat diet+ exercise group(HFD + E,n = 6),high fat diet+ castration group(HFD + Cas,n = 6),high fat diet+ castration + exercise group(HFD + Cas + E,n = 6),high fat diet+ flutamide group(HFD + F,n = 6),high fat diet+ flutamide + exercise group(HFD + F + E,n = 6)after feeding with high-fat diet.Then two weeks after castration and flutamide embedding,mice in HFD + E,HFD + Cas + E,HFD + F + E groups performed moderate-intensity treadmill intervention duration for 6 weeks.All Mice were anesthetized and sacrificed at 36 h after training finished,and the bilateral tibia and femur were collected.The wet weight and length of bone were measured by electronic scales and vernier caliper,respectively.The three-dimensional space structure was reconstructed by Micro-CT,and the results of BMD(bone mineral density),BV/TV(bone volume/total volume),and Tb.N(trabecular number),Tb.Th(trabecular thickness),Con.D(connectivity density),SMI(structure model index),Tb.Sp(trabecular separation),Ct.Th(cortical thickness)were calculated.Biomechanical properties of bone such as maximum load,fracture load,bending strength and elastic modulus were detected by three-point bending method,the levels of AR,IGF-1,IGF-1R protein expression were determined by Western blot.Results 1.Effects of exercise on bone mineral density and biomechanical properties in high-fat diet mice(1)Exercise increased the wet weight of bone in mice with high-fat diet,whereas had no effect on the length of bone Compared with C group,the wet weight of femur and tibia in HFD group wassignificantly reduced(p<0.01,p<0.01);compared with HFD group,the wet weight of femur and tibia in HFD+E group was remarkably increased(p<0.01,p<0.05).For the length of femur and tibia,there was no significant difference among the groups.(2)Exercise improved cancellous and cortical bone structure in high-fat diet mice Compare with C group,cancellous bone BV/TV,Tb.N,and Con.D in HFD group were significantly reduced(p<0.01,p<0.01,p<0.05),SMI and Tb.Sp significantly increased(p<0.05,p<0.01);compared with HFD group,BV/TV,Tb.N and Con.D were significantly increased(p<0.01,p<0.01,p<0.01),and Tb.Sp significantly decreased(p<0.01).As for cortical bone,there was no difference for BV/TV and Ct.Th between C and HFD group;compared with HFD group,BV/TV and Ct.Th in HFD+E group were significantly increased(p<0.05),p<0.05).(3)Exercise increased bone biomechanical properties in high-fat diet mice Compared with C group,Tv BMD in HFD group was significantly reduced(p<0.01);compared with HFD group,Tv BMD,Max.Load,Fracture Load and Yield stress in HFD+E group were significantly increased(p<0.01,p<0.05,p<0.05,p<0.05);for Elastic Modulus,there was no difference among groups.2.The role and mechanism of androgens in exercise-regulated bone mineral density and bone biomechanical properties in high-fat diet mice(1)Androgen defiency partially attenuated the effects of exercise on the wet weight of bone Compared with HFD group,the wet weight of femur and tibia in HFD+ Cas group was significantly reduced(p<0.05,p<0.05);compared with HFD + Cas group,the wet weight of the femur and tibia in HFD + Cas + E group was significantly increased(p<0.01,p<0.05);Compared with HFD+E group,the wet weight of tibial in HFD + Cas + E mice was significantly reduced(p<0.05).(2)Androgen deficiency attenuated the improvement of exercise on cancellous bone and cortical bone structure in high-fat diet mice Compared with HFD group,cancellous bone BV/TV,Tb.N,and Con.D in HFD+ Cas group were significantly reduced(p<0.01,p<0.01,p<0.01),and Tb.Sp was significantly increased(p<0.01).Compared with HFD +E group,BV / TV,Tb.N,and Con.D in HFD+Cas+E group mice were significantly reduced(p<0.01,p<0.01,p<0.01),and Tb.Sp was significantly increased(p<0.01);there was no significant difference for Tb.Th and SMI between groups.For cortical bone,compared with HFD group,Ct.Th of mice in HFD +Cas group were significantly reduced(p<0.01);compared with HFD+E group,BV/TV and Ct.Th in HFD+ Cas+ E group were significantly reduced(p<0.01,p<0.01).(3)Androgen deficiency reduced the improvement of exercise on bone biomechanical properties in high-fat diet mice Compared with HFD group,Tv BMD in HFD +Cas group was significantly decreased(p<0.01);compared with HFD+ Cas group,the Tv BMD of mice in HFD + Cas+E group was significantly increased(p<0.01);compared with mice in HFD+E group,Tv BMD,maximum load,fracture load were significantly reduced in mice inHFD + Cas +E group(p<0.01,p<0.05,p<0.05).(4)Mechanisms of androgens in exercise-regulated bone mineral density and biomechanical properties in high-fat diet mice-AR mediated IGF-1 and IGF-1R pathways? Compared with HFD group mice,the expression levels of AR,IGF-1and IGF-1R protein in HFD + E group were significantly increased(p<0.01,p<0.05,p<0.05);compared with HFD +E group,AR,the protein expression levels of IGF-1 and IGF-1R in HFD +Cas+E group were significantly reduced(p<0.05,p<0.05,p<0.01).3.The role and mechanism of androgen receptor in exercise-improved bone mineral density and bone biomechanical properties in high-fat diet mice(1)AR blockade attenuated exercise-induced increase in bone wet weight in high-fat diet mice Compared with HFD group,the wet weight of femur in HFD +F group was significantly reduced(p<0.01);compared with HFD +F group,the wet weight of femur and tibia in HFD+F+E group was significantly increased(p<0.01,p<0.05);compared with HFD +E group,the wet weight of femur and tibia was significantly reduced in HFD +F +E group.(2)AR blockade reduced the improvement of exercise on cancellous bone and cortical bone structure in high-fat diet mice Compared with HFD group,cancellous bone Con.D in HFD +F group was significantly reduced(p<0.01),SMI was significantly increased(p<0.01);compared with HFD +E group,the BV/TV,Tb.N and Con.D were significantly decreased(p<0.01,p<0.01,p<0.01),SMI and Tb.Sp were significantly increased(p<0.01,p<0.01)in HFD +F +E group.For cortical bone,there was no significant difference among groups.(3)AR blockade attenuated the improvement of exercise on bone mineral density(but not bone biomechanics)Compared with HFD+ E group,the Tv BMD of mice in HFD +F +E group was significantly reduced(p<0.01),and there was no significant difference for maximum load,fracture load,yield stress and elastic modulus between two groups.(4)Mechanisms of AR in exercise-regulated bone mineral density and bone biomechanical properties in high-fat diet mice-IGF-1 and IGF-1R Compared with HFD group,the levels of AR,IGF-1 and IGF-1R protein expression in HFD +E group were remarkably increased(p<0.05,p<0.05,p<0.05);compared with HFD +E group,the expression levels of AR and IGF-1R protein in the HFD + F+ E group were significantly reduced(p<0.05,p<0.05).Conclusions 1.Moderate intensity exercise could improve cancellous and cortical bones structure,increase bone mineral density and biomechanical properties.2.Androgen and androgen receptor play vital roles in exercise-induced improvement on bone mineral density and biomechanical mice properties in high-fat diet mice,most likely by upregulating bone IGF-1 /IGF-1R protein levels.