Role And Mechanism Of Osteocyte Neuropeptide Y In Chronic Stress-induced Bone Loss

Objectives:The objectives of this study were to explore the role and mechanism of osteocyte neuropeptide Y in chronic stress-induced bone loss.Methods:Chronic unpredictable mild stress(CUMS)model was constructed using 10-week-old C57BL/6 mice.The behavioral changes of mice after CUMS intervention were determined by the open-field and elevated crossmaze experiments.Micro-computed tomography(Micro-CT)and biomechanics were used to evaluate bone volume and bone strength in the femoral of mice.The levels of osteoblast marker osteocalcin and bone resorption marker type I collagen cross-linked C-terminal peptide in serum and the levels of norepinephrine,acetylcholine and NPY in bone cortical homogenates were measured by enzyme-linked immunosorbent assay.Immunofluorescence staining was performed to assess the level of NPY protein in mouse brain tissue.Semi-quantitative PCR was performed to calculate the expression levels of NPY receptor genes in bone cortex and bone marrow.The effects of osteocyte derived culture media(OCY-CM),NPY and Y1 R antagonist BIBO3304 on osteogenic differentiation of bone mesenchymal stem cells(BMSCs)were determined by alizarin red S staining.Results:(1)Compared with normal controls,mice in the CUMS intervention group exhibited anxiety-like behavior,significantly decreased femoral bone mass and bone strength,and significantly increased levels of the autonomic neurotransmitters norepinephrine and acetylcholine and NPY in bone cortical homogenates;osteogenic and osteoclastic activities were significantly increased in mice after 2 weeks of CUMS intervention and significantly decreased after 4 and 8 weeks of intervention.(2)OCY-CM and NPY inhibited osteogenic differentiation of BMSCs,and the Y1 R antagonist BIBO3304 partially counteracted the inhibitory effect of NPY and OCY-CM on osteogenic differentiation of BMSCs.(3)Bone marrow cavity injection of NPY resulted in bone loss,and BIBO3304 was able to reverse the above phenotype.(4)The autonomic modulator γ-glutamine increased bone mass and improved bone microarchitecture in mice after CUMS intervention.Conclusion:Osteocyte NPY secretion increased after CUMS intervention,NPY inhibited osteogenic differentiation of BMSCs via Y1 R,and the autonomic modulator γ-glutamine increased bone mass and improved bone microarchitecture in mice after CUMS intervention.