Differential Effect Of Type 1 Diabetes On Trabecular And Cortical Bone And Its Mechanism

Objective: To study and explore the differential mechanism of trabecular bone and cortical bone loss in type 1 diabetic mice.Methods: The C57BL/6J mice were randomly divided into diabetic group and wild type group.Mice in diabetic group were intraperitoneally injected STZ 50 μg/(kg·d)for 5 days to establish type 1 diabetes mellitus model,while those in control group received same volume of saline.X-ray radiology,Micro-CT,HE staining and bone histomorphometry were used to observe the bone mass and osteoblast activity changes.Real-time PCR and immunohistochemical staining was used to determine the expression levels of β-catenin and other canonical Wnt target genes,such as Low-density lipoprotein receptor-related protein 6(LRP-6),lymphoid enhancer factor(LEF-1),T cell factor(TCF),and osteoblast target genes alkaline phosphatase(ALP),osteocalcin(OCN),osterix(Osx),runt-related transcription factor 2(RunX2).Results: X-ray radiology,Micro-CT and HE staining showed that the decrease of bone mass in the distal femur of the T1 DM group was the most obvious,however the decrease of bone mass in the distal end of the femur and mid-end femur were relatively small.Micro-CT analysis of femur revealed the differing extent of bone loss in the trabecular and cortical bones.The trabecular parameters [bone volume per total volume(BV/TV),trabecular thickness(Tb.Th),and trabecular number(Tb.N)] were significantly lower in T1 DM mice than those in control mice [(0.16±0.01)vs.(1.27±0.08),(0.04±0.01)vs.(0.09±0.01),(1.90±0.27)vs.(4.53±0.45);P<0.01],while the corresponding cortical parameters [bone area per tissue area(BA/TA)and cortical thickness(CT.Th)] decreased to a much smaller extent [(0.35±0.05)vs.(0.40±0.02),(0.17±0.01)vs.(0.18±0.01);P<0.05] in T1 DM mice.Also,osteoblast surface per bone surface(Ob.S/BS),mineralizing surface(MS/BS),mineral appositional rate(MAR),the reduction of osteoblast number per tissue area(N.Ob/T.Ar)and bone formation rate(BFR)were also greater in the femoral distal metaphysis [(12.15±0.46)vs.(21.69±0.37),(30.19±1.02)vs.(38.02±0.70),(0.74±0.06)vs.(1.13±0.10),(0.25±0.02)vs.(0.48±0.04),P=0.000] than in the femoral diaphysis [(12.80±0.13)vs.(16.41±0.29),(36.16±1.22)vs.(42.21±0.54),(0.67±0.34)vs.(0.83±0.03),(0.27±0.02)vs.(0.36±0.04);P=0.000].The expression levels of β-catenin,other canonical Wnt target genes including LRP-6,TCF,LEF-1 and osteoblast target genes ALP,OCN,OSX,RunX2 were examined in the trabecular and cortical bones respectively.All these genes were downregulated in both trabecular and cortical bones in T1 DM mice,and the decrease in the expression level of LRP-6,β-catenin,TCF,LEF-1 and osteoblast target genes ALP,OCN,OSX,RunX2 was more striking in the trabecular bone [(0.18±0.13)vs.(1.00±0.13),(0.23±0.20)vs.(1.00±0.15),(0.29±0.23)vs.(1.00±0.23),(0.22±0.17)vs.(1.00±0.12),(0.52±0.10)vs.(1.00±0.00),(0.52±0.11)vs.(1.00±0.10),(0.42±0.01)vs.(1.00±0.10),(0.34±0.12)vs.(1.00±0.08);P=0.000] than in the cortical bone [(0.74±0.65)vs.(1.00±0.07),(0.63±0.45)vs.(1.00±0.09),(0.75±0.58)vs.(1.00±0.08),(0.70±063)vs.(1.00±0.05),(0.67±0.19)vs.(1.00±0.23),(0.73±0.08)vs.(1.00±0.23),(0.63±0.08)vs.(1.00±0.39),(0.62±0.08)vs.(1.00±0.11),P<0.01].Immunohistochemical results shows the decrease in the expression level of LRP-6,β-catenin,TCF,LEF-1 and osteoblast target genes ALP,OCN,OSX,RunX2 was more striking in the trabecular bone[(9.17±1.18)vs.(19.13±1.83),(8.92±0.92)vs.(20.72±1.32),(7.36±0.45)vs.(19.43±1.31),(8.46±0.61)vs.(8.46±0.61),(4.88±0.67)vs.(6.25±0.56),(9.17±0.42)vs.(11.06±0.67);P<0.01] than in the cortical bone [(8.46±0.61)vs.(11.45±1.57),(4.88±0.67)vs.(6.25±0.56),(9.17±0.42)vs.(11.06±0.67),(8.16±1.05)vs.10.98±1.09),(7.51±0.99)vs.(9.62±1.20),(6.92±0.86)vs.(10.34±1.22),P<0.01].Conclusion: In T1 DM mice,the reduction in the cancellous bone mass is greater than that in cortical bone mass,which might be caused by the greater downregulation of the Wnt/β-catenin signaling pathway in cancellous bone than in cortical bone.