Role of ecto-NTPDases on UDP-sensitive P2Y6receptor activation during osteogenic differentiation of primary bone marrow stromal cells from postmenopausal women

This study aimed at investigating the expression and function of uracil nucleotide-sensitive receptors (P2Y2, P2Y4, and P2Y6) on osteogenic differentiation of human bone marrow stromal cells (BMSCs) in culture. Bone marrow specimens were obtained from postmenopausal female patients (68 ± 5 years old, n = 18) undergoing total hip arthroplasty. UTP and UDP (100 µM) facilitated osteogenic differentiation of the cells measured as increases in alkaline phosphatase (ALP) activity, without affecting cell proliferation. Uracil nucleotides concentration-dependently increased [Ca2+]i in BMSCs; their effects became less evident with time (7 > 21 days) of the cells in culture. Selective activation of P2Y6 receptors with the stable UDP analog, PSB 0474, mimicked the effects of both UTP and UDP, whereas UTPγS was devoid of effect. Selective blockade of P2Y6 receptors with MRS 2578 prevented [Ca2+]i rises and osteogenic differentiation caused by UDP at all culture time points. BMSCs are immunoreactive against P2Y2, P2Y4, and P2Y6 receptors. While the expression of P2Y6 receptors remained fairly constant (7∼21 days), P2Y2 and P2Y4 became evident only in less proliferative and more differentiated cultures (7