The Roles of P2Y(2) Purinergic Receptors in Osteoblasts and Mechanotransduction

We previously demonstrated, using osteoblastic MC3T3-E1 cells, that P2Y(2) purinergic receptors are involved in osteoblast mechanotransduction. In this study, our objective was to further investigate, using a knockout mouse model, the roles of P2Y(2) receptors in bone mechanobiology. We first examined bone structure with micro-CT and measured bone mechanical properties with three point bending experiments in both wild type mice and P2Y(2) knockout mice. We found that bones from P2Y(2) knockout mice have significantly decreased bone volume, bone thickness, bone stiffness and bone ultimate breaking force at 17 week old age. In order to elucidate the mechanisms by which P2Y(2) receptors contribute to bone biology, we examined differentiation and mineralization of bone marrow cells from wild type and P2Y(2) knockout mice. We found that P2Y(2) receptor deficiency reduces the differentiation and mineralization of bone marrow cells. Next, we compared the response of primary osteoblasts, from both wild type and P2Y(2) knockout mice, to ATP and mechanical stimulation (oscillatory fluid flow), and found that osteoblasts from wild type mice have a stronger response, in terms of ERK1/2 phosphorylation, to both ATP and fluid flow, relative to P2Y(2) knockout mice. However, we did not detect any difference in ATP release in response to fluid flow between wild type and P2Y(2) knock out osteoblasts. Our findings suggest that P2Y(2) receptors play important roles in bone marrow cell differentiation and mineralization as well as in bone cell mechanotransduction, leading to an osteopenic phenotype in P2Y(2) knockout mice.