Effect Of Osteoblast Specific Knockout Wnt5a On Bone Loss In Disuse Osteoporotic Mice

Objectives Osteoblast-specific knockout Wnt5a mice were used to construct a tailsuspension-induced disuse osteoporosis model.Exploring the effect of the deletion of Wnt5a in osteoblast on bone mass and microstructure in the disuse osteoporotic mice model induced by tail-suspension.Methods Sixteen-week-old osteoblast-specific knockout Wnt5a female mice and their littermates born without Cre in the same batch were randomly divided into 4 groups,wild-type normal loading group(Wild-type Normal Loading,WTNL group),wild-type tail suspension group(Wild-type Tail Suspension,WTTS group),osteoblast-specific knockout Wnt5a normal loading group(Knockout Normal Loading,KONL),osteoblastspecific knockout Wnt5a tail suspension group(Knockout Tail Suspension,KOTS),5mice in each group.The mice in the WTTS group and KOTS group were established tail suspension model,and the others were bred normally.Body weight changes were observed during the modeling period.The mice of four groups were anesthetized and sacrificed three weeks after tail suspension modeling.The left tibia was used for MicroCT three-dimensional reconstruction and quantitative analysis of spongy bone parameters including bone mineral density(BMD)and trabecular bone volume(BV/TV),Trabecular thickness(Tb.Th),Trabecular number(Tb.N)and Trabecular separation(Tb.Sp),Structure Model Index(SMI);cortical bone parameters including cortical bone area(Ct.Ar),cortical thickness(Ct.Th),cortical bone porosity(Ct.Po).Three-point bending test was used to determine the biomechanical properties of the left femurs,which were reflected by maximum load and maximum stress.The controlateral femurs were detected by HE staining to analyze the histomorphology of their metaphysis.Real-time PCR was used to analyze the expression of relative m RNA of skulls originated from KO and WT mice.Collected results for statistical analysis.Results The results of Real-time PCR analysis showed that,compared with the WT group,the relative expression of Wnt5a the KO group was significantly decreased(P<0.0001).The HE staining and the three-dimensional reconstruction image of spongy bones showed that the microstructure of the cancellous bone of the WTNL group was normal,and the trabecular bones were arranged neatly and densely;the bone microstructure of the WTTS group degenerated abnormally;bone mass was significantly reduced in KONL group compared with WTNL group,same reduction was also found between KONL and KOTS group meanwhile the bone microstructure was abnormally changed.The three-dimensional reconstruction image of the cortical bone showed that the thickness of the cortical bone of the WTNL group was normal;compared with the WTNL group,the cortical bone of the WTTS group was significantly thinner;no significant change was observed in bone thickness among KONL,KOTS and WTNL group.Quantitative analysis of Micro-CT results showed that compared with the WTNL group,BV/TV、BMD、Tb.N、Tb.Th、Ct.Ar、and Ct.Th were significantly decreased in the WTTS group(P<0.05),Tb.Sp、Ct.Po、SMI increased significantly(P<0.05);Compared with KONL group,BV/TV,BMD,Tb.N of KOTS group were significantly decreased(P<0.05);SMI increased significantly;Compared with the WTNL group,BV/TV,BMD,Tb.N and Ct.Th in the KONL group were significantly reduced(P<0.05);compared with the WTTS group,Ct.Ar and Ct.Th in the KOTS group were increased significantly(P<0.05).The three-point bending of the femur showed that compared with the WTNL group,the maximum load and maximum stress of the WTTS group were decreased significantly(P<0.05);the maximum load and maximum stress of KOTS group was not statistically different from that of KONL group(P>0.05);Compared with the WTNL group,the maximum load and maximum stress of the KONL group decreased significantly(P<0.05);compared with the WTTS group,the maximum load and maximum stress of the KOTS group increased significantly(P<0.05).Conclusions 1 The WTTS group successfully established a disuse osteoporosis model through tail suspension,which induced bone loss,and the decline in cancellous bone mass was greater than that in cortical bone.2 Under normal physiological conditions,the osteoblast-specific Wnt5a knockout mice have significant loss of cancellous bone bone mass,abnormal changes in bone microstructure,and decreased bone strength compared with mice without knockout genes.3 Osteoblast-specific knockout of Wnt5a mice can partially delay the bone loss caused by tail suspension.Figure13;Table9;Reference104...