SMAD1/5 signaling in osteoclasts regulates bone formation via coupling factors

Abstract

Bone remodeling occurs via coupling between bone resorption by osteoclasts and bone formation by osteoblasts. The mechanisms that regulate osteoclast signals to osteoblasts are not well understood. Published studies have reported that BMP signaling in osteoclasts regulate osteoclast coupling targets. To investigate the necessity of canonical BMP signaling on osteoclast differentiation and coupling, we mated Smad1fl/fl; Smad5fl/fl mice to c-Fms-Cre mice. We analyzed male mice at 3 months of age to determine the skeletal phenotype of the Smad1fl/fl; Smad5fl/fl;c-Fms-Cre (SMAD1/5 cKO) mice. There was a 1.2-fold decrease in trabecular BV/TV in SMAD1/5 cKO. Analyses of osteoclast serum markers in SMAD1/5 cKO mice, showed a significant increase in CTX-1 (1.5 fold) and TRAP ELISA (3 fold) compared to control mice. In these same mice, there was a 1.3-fold increase in cortical thickness. Consistent with the increase in cortical thickness, we found a 3-fold increase in osteoblast activity as measured by P1NIP ELISA assay from SMAD1/5 cKO mice. To explain the changes in cortical thickness and P1NP activity, we determined conditioned media from SMAD1/5 cKO osteoclast cultures enhanced mineralization of an osteoblast cell line and coupling factors expressed by osteoclasts that regulate osteoblast activity Wnt1 (4.5-fold increase), Gja1 (3-fold increase) and Sphk1 (1.5-fold increase) were all upregulated in osteoclasts from SMAD1/5 cKO compared to control osteoclasts. Lastly osteoclasts treated with dorsomorphin, a chemical inhibitor of SMAD1/5 signaling, demonstrates an increase in Wnt1 and Gja1 expression similar to the SMAD1/5 cKO mice. Previous studies demonstrated that TGF-β signaling in osteoclasts leads to increases in WNT1 expression by osteoclasts. Therefore, our data suggest that TGF-β and BMP signaling pathways in osteoclasts could act in an antagonistic fashion to regulate osteoblast activity through WNT1 and other coupling factors.

Disruption of BMP Signaling Prevents Hyperthyroidism-Induced Bone Loss in Male Mice

“…(64) Vice versa, BMP signaling in osteoclasts also appears to alter osteoblast function as inhibited SMAD1/5 signaling in osteoclasts has been shown to increase osteoblast activity via osteoclast-to-osteoblast coupling factors such as WNT1, gap junction alpha-1 protein, and sphingosine kinase 1. (65) Finally, recent reports indicate tight connections between BMP and Wnt signaling. (40,66) in vitro blockade of the BMP pathway reversed T 3 -stimulated Dkk1 expression but did not alter the expression of Wnt target genes Axin2 nor Opg, suggesting additional mechanisms might obstruct Wnt signaling in osteoblasts.…”

The Bone Morphogenetic Protein Pathway: The Osteoclastic Perspective

“…Despite the comprehensive knowledge about BMP signaling in osteoblasts, its role in osteoclast formation has long been underrated. Several studies report on the endogenous expression of several BMP ligands (BMP1, BMP2, BMP4, BMP6, BMP7), SMAD proteins (SMAD1/5, SMAD4), and BMP receptors (BMPR1A, BMPR1B, BMPR2) in osteoclasts or osteoclast-like cell lines ( Anderson et al, 2000 ; Garimella et al, 2008 ; Jensen et al, 2010 ; Broege et al, 2013 ; Tasca et al, 2015 , 2018 ).…”

“…Downstream of receptors, in vitro genetic ablation of SMAD1/5 or SMAD4 in osteoclast precursors led to the formation of fewer and smaller multinucleated osteoclasts and to a reduction of resorption pits and resorbed areas ( Tasca et al, 2015 ). Correspondingly, mice with a conditional Smad1/5 knockout in osteoclast precursors (Smad1 fl/fl ;Smad5 fl/fl ;c-Fms-Cre mice, 12-week-old, male) showed mild bone gain due to reduced bone resorption and stimulated bone formation ( Tasca et al, 2018 ). In contrast, Smad4 deletion in mature osteoclasts (Smad4 fl/fl ;Ctsk-Cre mice, 8-week-old, female) increased osteoclast formation and bone resorption leading to an osteopenic phenotype, however, caused by disrupted TGFβ signaling and independent of the BMP pathway ( Morita et al, 2016 ).…”

“…Vice versa, osteoclast-specific Bmpr1a knockout enhanced bone formation suggesting an important role of BMPR1A within osteoblast and osteoclast coupling ( Okamoto et al, 2011 ). Furthermore, osteoblasts can be recruited by osteoclasts to sites of active bone remodeling, mediated through coupling factors such as WNT1 ( Weivoda et al, 2016 ; Tasca et al, 2018 ), Gap junction alpha-1 protein (GJA1) ( Tasca et al, 2018 ), and sphingosine kinase 1 (SPHK1) ( Ryu et al, 2006 ; Pederson et al, 2008 ; Tasca et al, 2018 ). A recent study proposed that especially SMAD1/5-dependent signaling in osteoclasts might regulate bone formation since mRNA levels of aforementioned coupling factors were upregulated in osteoclasts with deleted SMAD1/5 ( Tasca et al, 2018 ).…”

Activin-A Induces Fewer, but Larger Osteoclasts From Monocytes in Both Healthy Controls and Fibrodysplasia Ossificans Progressiva Patients

“…In contrast to the generally accepted role of BMPs in osteogenesis, several studies have shown that BMP signaling is also important for osteoclast formation. Inhibition of this signaling, either via deletion of BMPR2 (33) or by inhibiting SMAD 4 or SMAD 1/5 (34,35) reduces DC-STAMP expression and inhibits osteoclastogenesis. Similarly, the inhibitory effect of Activin-A on BMP signaling could be the cause of the inhibited osteoclastogenesis seen in this study.…”