Endocrine Abstracts

Osteocalcin (OC) is the main non-collagenous protein of the bone and it has a regulatory role in mineralization. Extra-skeletal roles have been recently hypothesized for OC (e.g., modulation of glucose-induced insulin secretion by interacting with GPRC6A in β-cells). However, OC-mediated GPRC6A activation has been demonstrated only in rodents and heterologous systems. In this study we aimed at evaluating the dose-dependent response to OC and the eventual activation of the GPCR-dependent pathways in a physiologic-like model of human pancreatic β-cells. 1.1B4 cell line, derived from electrofusion of PANC-1 cell line and primary human β-cells, were treated for 5 and 30 min with different concentrations of OC (20 to 100 ng/ml) either alone or in combination with 10 μM NPS2143, a non-competitive antagonist of GPRC6A. mRNA and protein expression were evaluated in order to assess the expression of GPRC6A and the activation profile of Erk and Akt pathways. As for human β-cells in vivo, which physiologically express nearly undetectable levels of GPRC6A, 1.1B4 cells displayed undetectable levels of GPRC6A protein. On the contrary, GPRC6A mRNA was slightly detectable. A slight increase (1.5 fold) of pErk/tErk was induced by OC 40 ng/ml at 30 min. Instead, pAkt was dose-dependently induced within 5 min with a 2-fold peak with OC 80 ng/ml; at 30 min a residual activation was evident for OC 20 and 40 ng/ml. NPS2143 did not affect the activation profile of Erk and Akt if not for a slight peak activation of both pathways with OC 60 ng/ml, instead of 40 ng/ml for Erk and 80 ng/ml for Akt. In summary: i) GPRC6A is not expressed (or it is expressed at very low levels) by human pancreatic β-cells; ii) the common OC signaling pathways, identified in heterologous expression systems, are only slightly induced in human pancreatic β-cells; iii) the inhibition of GPRC6A do not affect the activation profile of Erk and Akt and, hence, GPRC6A may not be involved in physiological OC signaling in human β-cells.