Novel Human Osteoblast Effector Gene EPDR1 Demonstrates Differing Outcomes after Knock-down or Knock-out in Murine In vitro Osteoblastogenesis and During Murine Bone Modeling

Abstract

EPDR1 is a recently characterized human osteoblast effector gene previously identified by intersecting BMD GWAS data with functional genomics in differentiating human mesenchymal stem cells (MSC). Loss of EPDR1 function biases hMSC towards adipogenic differentiation via metabolic and immunologic reprogramming. However, the role of EPDR1 in murine cells and bone development is not known. We generated Epdr1¬fl/fl mice to investigate its role in osteoblastic and adipogenic differentiation of mouse MSC (mMSC) and to dissect the effect of global loss of EPDR1 function during bone development using whole body knock-out EIIA-Cre mediated genetic recombination. Adenovirus mediated recombination of Epdr1fl/fl mMSC led to a decrease in alkaline phosphatase expression and mineralization similar to our published results with hMSC. The Epdr1 recombined cells showed a general reduction in Runx2, Alpl, Sp7, Ibsp, and Bglap expression suggesting a requirement of EPDR1 function for osteoblastogenesis. However, unlike results observed in hMSC upon EPDR1 silencing, mMSC adipogenesis decreased with Epdr1 recombination and was accompanied by a modest reduction in Pparg gene expression. Next, we crossed the Epdr1fl/fl mice with EIIA-Cre to generate a global Epdr1 knock-out mice. These mice appeared normal at birth and showed no developmental abnormalities. Intriguingly, the overall body weight of Epdr1 knockout male mice showed a reduction at 6 months of age that could be associated with reduction in the amount of body fat depots. At the cellular level, , mMSC harvested from 1-month old Epdr1-/- showed a minor reduction in mineralization capability, but these differences were not observed with cells harvested from 3- and 6-months old mice. Correspondingly, bone parameters of the 6 months old Epdr1fl/fl and Epdr1-/- mice using microCT imaging of the femurs showed no major genotype differences. Accordingly, gene expression analysis using the tibial diaphysis and metaphysis from the same mice groups did not establish genotype or sex-specific reduction on bone metabolism. We are currently processing microCT images from the 1-and 3-month-old femur samples, but do not anticipate significant changes. Collectively, these results indicate a compensatory mechanism could exist upon global Epdr1 knock-out in the murine model, and warrants future investigations using conditional loss of Epdr1 function to understand osteoblast and adipocyte cell lineage specific roles in vivo.