MicroRNA regulation of osteoblast physiology and glucocorticoid signaling

Endogenous glucocorticoids are critical for normal bone physiology. However, glucocorticoid excess, due to systemic administration or syndromes such as Cushing’s disease, causes osteopenia and metabolic disorder. Overall, glucocorticoid receptor signaling must be tightly controlled for optimal bone health. We identified microRNA-433 (miR-433) as negative regulator of glucocorticoid signaling in the osteoblast lineage. In vitro, inhibition of miR-433 activity made mesenchymal cells more responsive to glucocorticoids and increased glucocorticoid receptor residence in the nucleus, suggesting that miR-433 may target mechanisms designed to limit the responsiveness of cells to glucocorticoid signaling. In addition to effects on glucocorticoid signaling, miR-433 is a negative regulator of osteoblastic differentiation. miR-433 decreases as osteoblast differentiation progresses, and inhibition of miR-433 activity increases osteoblastic maker gene expression. To better understand the function of miR-433 in bone, we generated transgenic mice expressing a miR-433 competitive inhibitor (tough decoy) in osteoblastic cells. Calvarial bone from the miR-433 decoy mice has increased mRNA for osteocalcin and the direct miR-433 target Runx2. Further, miR-433 decoy mice display increased trabecular and cortical bone thickness due to increased bone formation, although molecular mechanisms remain to be identified. We propose to test the hypothesis that miR-433 targets genes and pathways critical for osteoblastogenesis and for limiting glucocorticoid receptor signaling. In Aim 1, we will comprehensively characterize the skeletal phenotype of both male and female miR-433 decoy mice in maturity and aging. We will also identify miR-433 targets using a non-biased approach, to better understand how miR-433 regulates osteoblast biology. In Aim 2, we will determine the mechanisms by which miR-433 limits glucocorticoid responsiveness at a molecular level, as well as determining the impact of miR- 433 on the response of bone to exogenous glucocorticoids excess. Overall Impact: miR-433 is a novel negative of both glucocorticoid responsiveness and osteoblast differentiation. Glucocorticoid excess is the most common secondary cause of osteopenia, and tissue sensitivity to glucocorticoids is regulated by multiple mechanisms. Understanding the interaction between osteoblastogenesis and glucocorticoid signaling is critical for the design of novel strategies to limit the adverse effects of glucocorticoid excess on the skeleton.