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Bone-specific Overexpression of NPY Modulates Osteogenic Differentiation.

Our previous work showed that neuropeptide Y (NPY) is expressed by osteoblast lineage cells and when administered in vitro can inhibit osteoblast lineage differentiation. We hypothesize that NPY, generated by mature osteoblast lineage cells, can control and modulate differentiation of osteoprogenitor cells. Therefore, in this study we have evaluated the effects of bone-specific overexpression by generating Col2.3NPY-ires GFPtopaz transgenic mice. As described in previous studies, this transgenic approach will direct NPY overexpression to mature osteoblasts and osteocytes. Real-time PCR analysis of bone-derived RNA showed increased expression of NPY in samples derived from Col2.3NPY mice compared to wild type. To characterize the Col2.3NPY mice phenotype in vivo, trabecular and cortical bone morphometry was measured in three-month-old mice using μCT analysis. Lower trabecular and cortical bone mass and morphometric parameters were observed in both male and female mice, exhibiting 30-35% lower trabecular volume and 7-9% lower cortical volume, without differences in linear bone growth. Gene expression analysis in Col2.3NPY mice shows increased expression of OC when compared to wild type. No difference was detected in expression levels for BSP and DMP-1. Our preliminary data on dynamic histomorphometry has shown reduced mineral apposition rate in Col2.3NPY mice. This apparent discrepancy in bone mass and gene expression will be addressed by evaluating cellular parameters of bone lineage using histomorphometry and evaluation of the effects of NPY overexpression on proliferation and maturation of the osteoblast lineage cells in vivo. We have utilized primary culture to assess the effects of NPY overexpresion. Markers of mature osteoblast lineage cells (BSP, OC, and DMP-1) in BMSCs derived from Col2.3NPY mice were enhanced compared to wild type. There was no difference in ALP activity or mineralization in MSC cultures derived from NPY versus wild type mice. In contrast to stromal cells, calvarial osteoblast derived from Col2.3NPY mice show a decrease in mineralization colonies. This difference between two culture models could be a result of the presence of more mature NPY producing cells driven by Col2.3 at the early days of calvarial osteoblast culture compared to BMSC. Further studies are required to clarify the bone phenotype of Col2.3NPY mice and to understand the mechanism by which bone derived NPY affects osteoprogenitor differentiation.

neuropeptide Y; Col2.3NPY; osteoprogenitor cells

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