Ubiquitin Ligase Smurf1 Mediates Tumor Necrosis Factor-induced Systemic Bone Loss by Promoting Proteasomal Degradation of Bone Morphogenetic Signaling Proteins

Abstract

Chronic inflammatory disorders, such as rheumatoid arthritis, are often accompanied by systemic bone loss, which is thought to occur through inflammatory cytokine-mediated stimulation of osteoclast resorption and inhibition of osteoblast function. However, the mechanisms involved in osteoblast inhibition remain poorly understood. Here we test the hypothesis that increased Smad ubiquitin regulatory factor 1 (Smurf1)-mediated degradation of the bone morphogenetic protein pathway signaling proteins mediates reduced bone formation in inflammatory disorders. Osteoblasts derived from bone marrow or long bone samples of adult tumor necrosis factor (TNF) transgenic (TNF-Tg) mice were used in this study. TNF decreased the steady-state levels of Smad1 and Runx2 protein similarly to those in long bones of TNF-Tg mice. In the presence of the proteasome inhibitor MG132, TNF increased accumulation of ubiquitinated Smad1 protein. TNF administration over calvarial bones caused decreases in Smad1 and Runx2 protein levels and mRNA expression of osteoblast marker genes in wild-type, but not in Smurf1 ؊/؊ mice. Vertebral bone volume and strength of TNF-Tg/Smurf1 ؊/؊ mice were examined by a combination of micro-CT, bone histomorphometry, and biomechanical testing and compared with those from TNF-Tg littermates. TNF-Tg mice had significantly decreased bone volume and biomechanical properties, which were partially rescued in TNF-Tg/Smurf1 ؊/؊ mice. We conclude that in chronic inflammatory disorders where TNF is increased, TNF induces the expression of ubiquitin ligase Smurf1 and promotes ubiquitination and proteasomal degradation of Smad1 and Runx2, leading to systemic bone loss. Inhibition of ubiquitin-mediated Smad1 and Runx2 degradation in osteoblasts could help to treat inflammation-induced osteoporosis.Osteoporosis and fragility fractures are common and preventable complications of rheumatoid arthritis (RA).2 For example, one study has reported that 53.3% of RA patients had osteoporosis and 19.3% had vertebral fractures, rates that are much higher than those of the general population (1). These complications are thought to occur through inflammatory stimulation of osteoclast bone resorption and inhibition of osteoblast function (2) mediated by cytokines, such as TNF (3-5). TNF and other cytokines are overproduced by various cells in the inflamed joints of RA patients, which lead to severe local erosion of cartilage and bone, as well as periarticular osteopenia and systemic osteoporosis.TNF is a strong inhibitor of osteoblast functions in vitro, but the molecular mechanisms that mediate the inhibitory effects of TNF on osteoblasts have not been fully investigated. TNF inhibits the recruitment of osteoblast progenitors, reduces expression of genes produced by mature osteoblasts, and promotes osteoblast apoptosis through the NF-B pathway (6 -10). It decreases the expression and DNA binding activity of runt-related transcription factor 2 (Runx2), which is partially through suppression of Runx2 gene transcription and destabilization o...

Vitamin K2 stimulates osteoblastogenesis and suppresses osteoclastogenesis by suppressing NF-κB activation

“…We have previously demonstrated that one mechanism by which TNF· suppresses osteoblast differentiation is through NF-κB-mediated antagonism of TGFß-and BMP-2-induced SMAD signaling in differentiating osteoblasts (17). These data are consistent with the demonstration that NF-κB signaling up-regulates SMAD7, an inhibitor of SMAD activation in osteoblastic cells (18) and that TNF· promotes systemic bone loss by promoting proteasomal degradation of bone morphogenetic signaling proteins through up-regulation of SMAD ubiquitination regulatory factor 1 (Smurf1) (19). Recently, a direct inhibitory action of NF-κB on bone formation has been ratified in vivo where time-and stage-specific inhibition of the inhibitor of κB kinase (IKK) in differentiated osteoblasts increased trabecular bone mass and bone mineral density (BMD) and ameliorated ovariectomy-induced bone loss by promoting a compensatory increase in bone formation (20).…”

Cyclic AMP Response Element-binding Protein H (CREBH) Mediates the Inhibitory Actions of Tumor Necrosis Factor α in Osteoblast Differentiation by Stimulating Smad1 Degradation

“…TNF␣ suppresses BMP2-induced osteoblast differentiation and increases the expression of Smurf1, leading to the subsequent degradation of Smad1 and Runx2, which are critical mediators of BMP2 signaling (25,43). Smurf1 interacts directly with Smad1 and Runx2, and stimulates the degradation of these proteins in ubiquitin-and proteasome-dependent manners (44).…”

Glucocorticoids and Tumor Necrosis Factor α Increase Oxidative Stress and Suppress Wnt Protein Signaling in Osteoblasts

“…In addition, there is evidence that TNF␣ suppresses RUNX2 and osterix as well as bone morphogenetic protein-induced osteoblast differentiation (2,(67)(68)(69). Hence, these alternative mechanisms may contribute to the attenuating effects of Dex and TNF␣ on osteoblastogenesis.…”