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Currently, no effective therapies exist for fibrodysplasia ossificans progressiva (FOP), a rare congenital syndrome in which heterotopic bone is formed in soft tissues owing to dysregulated activity of the bone morphogenetic protein (BMP) receptor kinase ALK2 (also known as ACVR1). From a screen of known biologically active compounds, we identified saracatinib as a potent ALK2 kinase inhibitor. In enzymatic and cell-based assays, saracatinib preferentially inhibited ALK2, compared with other receptors of the BMP/TGF-β signaling pathway, and induced dorsalization in zebrafish embryos consistent with BMP antagonism. We further tested the efficacy of saracatinib using an inducible ACVR1Q207D-transgenic mouse line, which provides a model of heterotopic ossification (HO), as well as an inducible ACVR1R206H-knockin mouse, which serves as a genetically and physiologically faithful FOP model. In both models, saracatinib was well tolerated and potently inhibited the development of HO, even when administered transiently following soft tissue injury. Together, these data suggest that saracatinib is an efficacious clinical candidate for repositioning in FOP treatment, offering an accelerated path to clinical proof-of-efficacy studies and potentially significant benefits to individuals with this devastating condition.

Activin A functions in BMP signaling in two ways: it either engages ACVR1B to activate Smad2/3 signaling or binds ACVR1 to form a non-signaling complex (NSC). Although the former property has been studied extensively, the roles of the NSC remain unexplored. The genetic disorder fibrodysplasia ossificans progressiva (FOP) provides a unique window into ACVR1/Activin A signaling because in that disease Activin can either signal through FOP-mutant ACVR1 or form NSCs with wild-type ACVR1. To explore the role of the NSC, we generated 'agonist-only' Activin A muteins that activate ACVR1B but cannot form the NSC with ACVR1. Using one of these muteins, we demonstrate that failure to form the NSC in FOP results in more severe disease pathology. These results provide the first evidence for a biological role for the NSC in vivo and pave the way for further exploration of the NSC's physiological role in corresponding knock-in mice., Competing Interests: SA, RC, CC, AD, HK, QZ, AR, RL, XW, JM, JJ, KN, NR, JL, AM, AE, VI The author is an employee of Regeneron Pharmaceuticals, Inc. Regeneron is currently developing a monoclonal antibody that neutralizes Activin A (REGN2477) as a potential therapy in fibrodysplasia ossificans progressiva (see https://clinicaltrials.gov/ct2/show/NCT03188666). EG, EM, TT No competing interests declared, (© 2020, Aykul et al.)

Patients with anterior cruciate ligament (ACL) rupture are two times as likely to develop posttraumatic osteoarthritis (PTOA). Annually, there are ∼900,000 knee injuries in the United States, which account for ∼12% of all osteoarthritis (OA) cases. PTOA leads to reduced physical activity, deconditioning of the musculoskeletal system, and in severe cases requires joint replacement to restore function. Therefore, treatments that would prevent cartilage degradation post-injury would provide attractive alternatives to surgery. Sclerostin (Sost), a Wnt antagonist and a potent negative regulator of bone formation, has recently been implicated in regulating chondrocyte function in OA. To determine whether elevated levels of Sost play a protective role in PTOA, we examined the progression of OA using a noninvasive tibial compression overload model in SOST transgenic (SOST TG ) and knockout (Sost -/- ) mice. Here we report that SOST TG mice develop moderate OA and display significantly less advanced PTOA phenotype at 16 weeks post-injury compared with wild-type (WT) controls and Sost -/- . In addition, SOST TG built ∼50% and ∼65% less osteophyte volume than WT and Sost -/- , respectively. Quantification of metalloproteinase (MMP) activity showed that SOST TG had ∼2-fold less MMP activation than WT or Sost -/- , and this was supported by a significant reduction in MMP2/3 protein levels, suggesting that elevated levels of SOST inhibit the activity of proteolytic enzymes known to degrade articular cartilage matrix. Furthermore, intra-articular administration of recombinant Sost protein, immediately post-injury, also significantly decreased MMP activity levels relative to PBS-treated controls, and Sost activation in response to injury was TNFα and NF-κB dependent. These results provide in vivo evidence that sclerostin functions as a protective molecule immediately after joint injury to prevent cartilage degradation. © 2018 The Authors. Journal of Bone and Mineral Research Published by Wiley Periodicals Inc., (© 2018 The Authors. Journal of Bone and Mineral Research Published by Wiley Periodicals Inc.)

Loss of Sostdc1, a growth factor paralogous to Sost, causes the formation of ectopic incisors, fused molars, abnormal hair follicles, and resistance to kidney disease. Sostdc1 is expressed in the periosteum, a source of osteoblasts, fibroblasts and mesenchymal progenitor cells, which are critically important for fracture repair. Here, we investigated the role of Sostdc1 in bone metabolism and fracture repair. Mice lacking Sostdc1 (Sostdc1(-/-)) had a low bone mass phenotype associated with loss of trabecular bone in both lumbar vertebrae and in the appendicular skeleton. In contrast, Sostdc1(-/-) cortical bone measurements revealed larger bones with higher BMD, suggesting that Sostdc1 exerts differential effects on cortical and trabecular bone. Mid-diaphyseal femoral fractures induced in Sostdc1(-/-) mice showed that the periosteal population normally positive for Sostdc1 rapidly expands during periosteal thickening and these cells migrate into the fracture callus at 3days post fracture. Quantitative analysis of mesenchymal stem cell (MSC) and osteoblast populations determined that MSCs express Sostdc1, and that Sostdc1(-/-) 5day calluses harbor >2-fold more MSCs than fractured wildtype controls. Histologically a fraction of Sostdc1-positive cells also expressed nestin and α-smooth muscle actin, suggesting that Sostdc1 marks a population of osteochondral progenitor cells that actively participate in callus formation and bone repair. Elevated numbers of MSCs in D5 calluses resulted in a larger, more vascularized cartilage callus at day 7, and a more rapid turnover of cartilage with significantly more remodeled bone and a thicker cortical shell at 21days post fracture. These data support accelerated or enhanced bone formation/remodeling of the callus in Sostdc1(-/-) mice, suggesting that Sostdc1 may promote and maintain mesenchymal stem cell quiescence in the periosteum., (Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.)

Bone morphogenetic proteins (BMPs) are highly conserved signaling molecules that are part of the transforming growth factor (TGF)-beta superfamily, and function in the patterning and morphogenesis of many organs including development of the dentition. The functions of the BMPs are controlled by certain classes of molecules that are recognized as BMP antagonists that inhibit BMP binding to their cognate receptors. In this study we tested the hypothesis that USAG-1 (uterine sensitization-associated gene-1) suppresses deciduous incisors by inhibition of BMP-7 function. We learned that USAG-1 and BMP-7 were expressed within odontogenic epithelium as well as mesenchyme during the late bud and early cap stages of tooth development. USAG-1 is a BMP antagonist, and also modulates Wnt signaling. USAG-1 abrogation rescued apoptotic elimination of odontogenic mesenchymal cells. BMP signaling in the rudimentary maxillary incisor, assessed by expressions of Msx1 and Dlx2 and the phosphorylation of Smad protein, was significantly enhanced. Using explant culture and subsequent subrenal capsule transplantation of E15 USAG-1 mutant maxillary incisor tooth primordia supplemented with BMP-7 demonstrated in USAG-1+/- as well as USAG-1-/- rescue and supernumerary tooth development. Based upon these results, we conclude that USAG-1 functions as an antagonist of BMP-7 in this model system. These results further suggest that the phenotypes of USAG-1 and BMP-7 mutant mice reported provide opportunities for regenerative medicine and dentistry.

WNT signaling is critical in most aspects of skeletal development and homeostasis, and antagonists of WNT signaling are emerging as key regulatory proteins with great promise as therapeutic agents for bone disorders. Here we show that Sost and its paralog Sostdc1 emerged through ancestral genome duplication and their expression patterns have diverged to delineate non-overlapping domains in most organ systems including musculoskeletal, cardiovascular, nervous, digestive, reproductive and respiratory. In the developing limb, Sost and Sostdc1 display dynamic expression patterns with Sost being restricted to the distal ectoderm and Sostdc1 to the proximal ectoderm and the mesenchyme. While Sostdc1(-/-) mice lack any obvious limb or skeletal defects, Sost(-/-) mice recapitulate the hand defects described for Sclerosteosis patients. However, elevated WNT signaling in Sost(-/-); Sostdc1(-/-) mice causes misregulation of SHH signaling, ectopic activation of Sox9 in the digit 1 field and preaxial polydactyly in a Gli1- and Gli3-dependent manner. In addition, we show that the syndactyly documented in Sclerosteosis is present in both Sost(-/-) and Sost(-/-); Sostdc1(-/-) mice, and is driven by misregulation of Fgf8 in the AER, a region lacking Sost and Sostdc1 expression. This study highlights the complexity of WNT signaling in skeletal biology and disease and emphasizes how redundant mechanism and non-cell autonomous effects can synergize to unveil new intricate phenotypes caused by elevated WNT signaling., (© 2013 The Authors. Published by Elsevier Inc. All rights reserved.)

Current endeavors in the type 2 diabetes (T2D) field include gaining a better understanding of extracellular signaling pathways that regulate pancreatic islet function. Recent data suggest that both Bmp and Wnt pathways are operative in pancreatic islets and play a positive role in insulin secretion and glucose homeostasis. Our laboratory found the dual Bmp and Wnt antagonist Sostdc1 to be upregulated in a mouse model of islet dysmorphogenesis and nonimmune-mediated lean diabetes. Because Bmp signaling has been proposed to enhance β-cell function, we evaluated the role of Sostdc1 in adult islet function using animals in which Sostdc1 was globally deleted. While Sostdc1-null animals exhibited no pancreas development phenotype, a subset of mutants exhibited enhanced insulin secretion and improved glucose homeostasis compared with control animals after 12-wk exposure to high-fat diet. Loss of Sostdc1 in the setting of metabolic stress results in altered expression of Bmp-responsive genes in islets but did not affect expression of Wnt target genes, suggesting that Sostdc1 primarily regulates the Bmp pathway in the murine pancreas. Furthermore, our data indicate that removal of Sostdc1 enhances the downregulation of the closely related Bmp inhibitors Ctgf and Gremlin in islets after 8-wk exposure to high-fat diet. These data imply that Sostdc1 regulates expression of these inhibitors and provide a means by which Sostdc1-null animals show enhanced insulin secretion and glucose homeostasis. Our studies provide insights into Bmp pathway regulation in the endocrine pancreas and reveal new avenues for improving β-cell function under metabolic stress.

Sclerostin is expressed by osteocytes and has catabolic effects on bone. It has been shown to antagonize bone morphogenetic protein (BMP) and/or Wnt activity, although at present the underlying mechanisms are unclear. Consistent with previous findings, Sclerostin opposed direct Wnt3a-induced but not direct BMP7-induced responses when both ligand and antagonist were provided exogenously to cells. However, we found that when both proteins are expressed in the same cell, sclerostin can antagonize BMP signaling directly by inhibiting BMP7 secretion. Sclerostin interacts with both the BMP7 mature domain and pro-domain, leading to intracellular retention and proteasomal degradation of BMP7. Analysis of sclerostin knock-out mice revealed an inhibitory action of sclerostin on Wnt signaling in both osteoblasts and osteocytes in cortical and cancellous bones. BMP7 signaling was predominantly inhibited by sclerostin in osteocytes of the calcaneus and the cortical bone of the tibia. Our results suggest that sclerostin exerts its potent bone catabolic effects by antagonizing Wnt signaling in a paracrine and autocrine manner and antagonizing BMP signaling selectively in the osteocytes that synthesize simultaneously both sclerostin and BMP7 proteins.

The glomerular basement membrane (GBM) is a key component of the filtering unit in the kidney. Mutations involving any of the collagen IV genes (COL4A3, COL4A4, and COL4A5) affect GBM assembly and cause Alport syndrome, a progressive hereditary kidney disease with no definitive therapy. Previously, we have demonstrated that the bone morphogenetic protein (BMP) antagonist uterine sensitization-associated gene-1 (USAG-1) negatively regulates the renoprotective action of BMP-7 in a mouse model of tubular injury during acute renal failure. Here, we investigated the role of USAG-1 in renal function in Col4a3-/- mice, which model Alport syndrome. Ablation of Usag1 in Col4a3-/- mice led to substantial attenuation of disease progression, normalization of GBM ultrastructure, preservation of renal function, and extension of life span. Immunohistochemical analysis revealed that USAG-1 and BMP-7 colocalized in the macula densa in the distal tubules, lying in direct contact with glomerular mesangial cells. Furthermore, in cultured mesangial cells, BMP-7 attenuated and USAG-1 enhanced the expression of MMP-12, a protease that may contribute to GBM degradation. These data suggest that the pathogenetic role of USAG-1 in Col4a3-/- mice might involve crosstalk between kidney tubules and the glomerulus and that inhibition of USAG-1 may be a promising therapeutic approach for the treatment of Alport syndrome.

Individuals with congenital renal hypoplasia display a defect in the growth of nephrons during development. Many genes that affect the initial induction of nephrons have been identified, but little is known about the regulation of postinductive stages of kidney development. In the absence of the growth factor bone morphogenic protein 7 (BMP7), kidney development arrests after induction of a small number of nephrons. The role of BMP7 after induction, however, has not been fully investigated. Here, we generated a podocyte-specific conditional knockout of BMP7 (Bmp7(flox/flox);Nphs2-Cre(+) [BMP7 CKO]) to study the role of podocyte-derived BMP7 in nephron maturation. By postnatal day 4, 65% of BMP7 CKO mice had hypoplastic kidneys, but glomeruli demonstrated normal patterns of laminin and collagen IV subunit expression. Developing proximal tubules, however, were reduced in number and demonstrated impaired cellular proliferation. We examined signaling pathways downstream of BMP7; the level of cortical phosphorylated Smad1, 5, and 8 was unchanged in BMP CKO kidneys, but phosphorylated p38 mitogen-activated protein kinase was significantly decreased. In addition, beta-catenin was reduced in BMP7 CKO kidneys, and its localization to intracellular vesicles suggested that it had been targeted for degradation. In summary, these results define a BMP7-mediated regulatory axis between glomeruli and proximal tubules during kidney development.

Twisted gastrulation (Tsg) is a secreted glycoprotein that binds bone morphogenetic proteins (BMP)-2 and -4 and can display both BMP agonist and antagonist functions. Tsg promotes BMP-mediated endochondral ossification, but its activity in adult bone is not known. We created tsg null mice and examined the consequences of the tsg deletion on the skeleton in vivo and on osteoblast function in vitro. Analysis of the skeletal phenotype of 4-week-old tsg null mice revealed a 40% decrease in trabecular bone volume, but osteoblast and osteoclast number, and bone formation and resorption were not affected. The phenotype was transient, and at 7 weeks of age tsg null mice were not different from control wild-type mice. The decreased trabecular bone is congruent with a defect in endochondral bone formation. In osteoblasts isolated from tsg null mice, tsg gene inactivation decreased the BMP-2 stimulatory effects on osteocalcin expression and alkaline phosphatase activity, indicating that in the bone microenvironment endogenous Tsg enhances BMP activity. Accordingly, tsg null cells displayed impaired BMP signaling. These results were confirmed by Tsg down-regulation in primary osteoblasts from wild-type mice using RNA interference. In conclusion, endogenous Tsg is required for normal BMP activity in osteoblastic cells in vitro, but it plays a minor role in the regulation of adult bone homeostasis in vivo.

Neurogenesis from endogenous progenitor cells in the adult forebrain ventricular wall may be induced by the local viral overexpression of cognate neuronal differentiation agents, in particular BDNF. Here, we show that the overexpression of noggin, by acting to inhibit glial differentiation by subependymal progenitor cells, can potentiate adenoviral BDNF-mediated recruitment of new neurons to the adult rat neostriatum. The new neurons survive at least 2 months after their genesis in the subependymal zone and are recruited primarily as GABAergic DARPP-32+ medium spiny neurons in the caudate-putamen. The new medium spiny neurons successfully project to the globus pallidus, their usual developmental target, extending processes over several millimeters of the normal adult striatum. Thus, concurrent suppression of subependymal glial differentiation and promotion of neuronal differentiation can mobilize endogenous subependymal progenitor cells to achieve substantial neuronal addition to otherwise non-neurogenic regions of the adult brain.

During skull development, the cranial connective tissue framework undergoes intramembranous ossification to form skull bones (calvaria). As the calvarial bones advance to envelop the brain, fibrous sutures form between the calvarial plates. Expansion of the brain is coupled with calvarial growth through a series of tissue interactions within the cranial suture complex. Craniosynostosis, or premature cranial suture fusion, results in an abnormal skull shape, blindness and mental retardation. Recent studies have demonstrated that gain-of-function mutations in fibroblast growth factor receptors (fgfr) are associated with syndromic forms of craniosynostosis. Noggin, an antagonist of bone morphogenetic proteins (BMPs), is required for embryonic neural tube, somites and skeleton patterning. Here we show that noggin is expressed postnatally in the suture mesenchyme of patent, but not fusing, cranial sutures, and that noggin expression is suppressed by FGF2 and syndromic fgfr signalling. Since noggin misexpression prevents cranial suture fusion in vitro and in vivo, we suggest that syndromic fgfr-mediated craniosynostoses may be the result of inappropriate downregulation of noggin expression.

Skeletal homeostasis is determined by systemic hormones and local factors. Bone morphogenetic proteins (BMP) are unique because they induce the differentiation of mesenchymal cells toward cells of the osteoblastic lineage and also enhance the differentiated function of the osteoblast. However, the activity of BMPs needs to be tempered by intracellular and extracellular antagonists. BMPs bind to specific receptors and signal by phosphorylating the cytoplasmic proteins mothers against decapentaplegic (Smad) 1 and 5, which form heterodimers with Smad 4, and after nuclear translocation regulate transcription. BMP antagonists can be categorized as pseudoreceptors that compete with signaling receptors, inhibitory Smads that block signaling, intracellular binding proteins that bind Smad 1 and 5, and factors that induce ubiquitination and proteolysis of signaling Smads. In addition, a large number of extracellular proteins that bind BMPs and prevent their binding to signaling receptors have emerged. They are the components of the Spemann organizer, noggin, chordin, and follistatin, members of the Dan/Cerberus family, and twisted gastrulation. The antagonists tend to be specific for BMPs and are regulated by BMPs, indicating the existence and need of local feedback mechanisms to temper BMP cellular activities.

Background: The activity of bone morphogenetic proteins (BMPs) is regulated extracellularly by several families of secreted, negatively-acting factors. These BMP antagonists participate in the control of a diverse range of embryonic processes, such as establishment of the dorsal-ventral axis, neural induction, and formation of joints in the developing skeletal system. The ongoing process of neurogenesis in the adult brain also requires inhibition of BMP ligand activity. To date, the three-dimensional structures of these antagonists as well as the nature of their interaction with ligand have remained unknown. Toward that end, we have determined the crystal structure of the antagonist Noggin bound to BMP-7., Methods: The complex of the two homodimeric proteins was preformed, isolated by size exclusion chromatography, and crystallized at neutral pH. To probe the molecular interface of the complex and to quantitate the activity of a human mutant form, variant Noggin proteins were produced and their binding affinities were measured in vitro. The correlation between binding affinity and biological activity was examined with Noggin-soaked beads implanted in the developing chick limb bud., Results and Conclusions: The structure of the complex reveals that Noggin inhibits BMP signaling by blocking the binding sites of both types of receptors (Type I and Type II), mimicking their modes of binding. The affinity of Noggin variants for BMP-7 correlated well with the inhibition of BMP-induced chondrogenesis in the chick limb bud, confirming that Noggin acts by sequestering the ligand in an inactive state. Interestingly, the scaffold of Noggin was found to contain a cystine knot topology and protein fold similar to that of BMPs, indicating that ligand and antagonist may have evolved from a common ancestral gene.

The interplay between bone morphogenetic proteins (BMPs) and their antagonists governs developmental and cellular processes as diverse as establishment of the embryonic dorsal-ventral axis, induction of neural tissue, formation of joints in the skeletal system and neurogenesis in the adult brain. So far, the three-dimensional structures of BMP antagonists and the structural basis for inactivation have remained unknown. Here we report the crystal structure of the antagonist Noggin bound to BMP-7, which shows that Noggin inhibits BMP signalling by blocking the molecular interfaces of the binding epitopes for both type I and type II receptors. The BMP-7-binding affinity of site-specific variants of Noggin is correlated with alterations in bone formation and apoptosis in chick limb development, showing that Noggin functions by sequestering its ligand in an inactive complex. The scaffold of Noggin contains a cystine (the oxidized form of cysteine) knot topology similar to that of BMPs; thus, ligand and antagonist seem to have evolved from a common ancestral gene.

Bone morphogenetic proteins (BMPs) are expressed broadly and regulate a diverse array of developmental events in vivo. Essential to many of these functions is the establishment of activity gradients of BMP, which provide positional information that influences cell fates. Secreted polypeptides, such as Noggin, bind BMPs and inhibit their function by preventing interaction with receptors on the cell surface. These BMP antagonists are assumed to be diffusible and therefore potentially important in the establishment of BMP activity gradients in vivo. Nothing is known, however, about the potential interactions between Noggin and components of the cell surface or extracellular matrix that might limit its diffusion. We have found that Noggin binds strongly to heparin in vitro, and to heparan sulfate proteoglycans on the surface of cultured cells. Noggin is detected only on the surface of cells that express heparan sulfate, can be specifically displaced from cells by heparin, and can be directly cross-linked to a cell surface proteoglycan in culture. Heparan sulfate-bound Noggin remains functional and can bind BMP4 at the plasma membrane. A Noggin mutant with a deletion in a putative heparin binding domain has reduced binding to heparin and does not bind to the cell surface but has preserved BMP binding and antagonist functions. Our results imply that interactions between Noggin and heparan sulfate proteoglycans in vivo regulate diffusion and therefore the formation of gradients of BMP activity.

Loss of Sostdc1, a growth factor paralogous to Sost, causes the formation of ectopic incisors, fused molars, abnormal hair follicles, and resistance to kidney disease. Sostdc1 is expressed in the periosteum, a source of osteoblasts, fibroblasts and mesenchymal progenitor cells, which are critically important for fracture repair. Here, we investigated the role of Sostdc1 in bone metabolism and fracture repair. Mice lacking Sostdc1 (Sostdc1(-/-)) had a low bone mass phenotype associated with loss of trabecular bone in both lumbar vertebrae and in the appendicular skeleton. In contrast, Sostdc1(-/-) cortical bone measurements revealed larger bones with higher BMD, suggesting that Sostdc1 exerts differential effects on cortical and trabecular bone. Mid-diaphyseal femoral fractures induced in Sostdc1(-/-) mice showed that the periosteal population normally positive for Sostdc1 rapidly expands during periosteal thickening and these cells migrate into the fracture callus at 3days post fracture. Quantitative analysis of mesenchymal stem cell (MSC) and osteoblast populations determined that MSCs express Sostdc1, and that Sostdc1(-/-) 5day calluses harbor >2-fold more MSCs than fractured wildtype controls. Histologically a fraction of Sostdc1-positive cells also expressed nestin and α-smooth muscle actin, suggesting that Sostdc1 marks a population of osteochondral progenitor cells that actively participate in callus formation and bone repair. Elevated numbers of MSCs in D5 calluses resulted in a larger, more vascularized cartilage callus at day 7, and a more rapid turnover of cartilage with significantly more remodeled bone and a thicker cortical shell at 21days post fracture. These data support accelerated or enhanced bone formation/remodeling of the callus in Sostdc1(-/-) mice, suggesting that Sostdc1 may promote and maintain mesenchymal stem cell quiescence in the periosteum.

Fibrodysplasia ossificans progressiva (FOP) is a rare genetic disorder characterized by episodic yet cumulative heterotopic ossification (HO) of skeletal muscles, tendons, ligaments, and fascia. FOP arises from missense mutations in Activin Receptor type I (ACVR1), a type I bone morphogenetic protein (BMP) receptor. Although initial findings implicated constitutive activity of FOP-variant ACVR1 (ACVR1FOP) and/or hyperactivation by BMPs, it was later shown that HO in FOP requires activation of ACVR1FOP by Activin A. Inhibition of Activin A completely prevents HO in FOP mice, indicating that Activin A is an obligate driver of HO in FOP, and excluding a key role for BMPs in this process. This discovery led to the clinical development of garetosmab, an investigational antibody that blocks Activin A. In a phase 2 trial, garetosmab inhibited new heterotopic bone lesion formation in FOP patients. In contrast, antibodies to ACVR1 activate ACVR1FOP and promote HO in FOP mice. Beyond their potential clinical relevance, these findings have enhanced our understanding of FOP's pathophysiology, leading to the identification of fibroadipogenic progenitors as the cells that form HO, and the discovery of non-signaling complexes between Activin A and wild type ACVR1 and their role in tempering HO, and are also starting to inform biological processes beyond FOP. [ABSTRACT FROM AUTHOR]

Group 2 innate lymphoid cells (ILC2s) are type 2 cytokine-producing cells that have important roles in helminth infection and allergic inflammation. ILC2s are tissue-resident cells, and their phenotypes and roles are regulated by tissue-specific environmental factors. While the role of ILC2s in the lung, intestine and bone marrow has been elucidated in many studies, their role in adipose tissues is still unclear. Here, we report on the role of ILC2-derived bone morphogenetic protein 7 (BMP7) in adipocyte differentiation and lipid accumulation. Co-culture of fat-derived ILC2s with pluripotent mesenchymal C3H10T1/2 cells and committed white preadipocyte 3T3-L1 cells resulted in their differentiation to adipocytes and induced lipid accumulation. Co-culture experiments using BMP7-deficient ILC2s revealed that BMP7, produced by ILC2s, induces differentiation into brown adipocytes. Our results demonstrate that BMP7, produced by ILC2s, affects adipocyte differentiation, particularly in brown adipocytes. [ABSTRACT FROM AUTHOR]

While osteo- and chondro-inductive activities of recombinant human bone morphogenetic protein 7 are well established, evaluation of the role of endogenous BMP7 in skeletal homeostasis has been hampered by perinatal lethality in BMP7 knockout mice. Here, we examined physiological roles of endogenous BMP7 in joint homeostasis and showed that proteoglycan contents in articular cartilage were significantly reduced in the absence of BMP7. Loss of BMP7 did not affect survival of articular cartilage cells, but resulted in reduced expression of aggrecan and enhanced expression of matrix metalloproteinase 13. We also found extensive synovial hyperplasia and enhanced expression of Activin A. These findings suggest that locally produced BMP7 is prerequisite for postnatal synovial joint homeostasis and may be involved in osteoarthritic changes in adults. [ABSTRACT FROM AUTHOR]

Podocyte injury is the first step in the progression of glomerulosclerosis. Previous studies have demonstrated the beneficial effect of bone morphogenetic protein 7 (Bmp7) in podocyte injury and the existence of native Bmp signaling in podocytes. Local activity of Bmp7 is controlled by cell-type specific Bmp antagonists, which inhibit the binding of Bmp7 to its receptors. Here we show that the product of Twisted gastrulation (Twsg1), a Bmp antagonist, is the central negative regulator of Bmp function in podocytes and that Twsg1 null mice are resistant to podocyte injury. Twsg1 was the most abundant Bmp antagonist in murine cultured podocytes. The administration of Bmp induced podocyte differentiation through Smad signaling, whereas the simultaneous administration of Twsg1 antagonized the effect. The administration of Bmp also inhibited podocyte proliferation, whereas simultaneous administration of Twsg1 antagonized the effect. Twsg1 was expressed in the glomerular parietal cells (PECs) and distal nephron of the healthy kidney, and additionally in damaged glomerular cells in a murine model of podocyte injury. Twsg1 null mice exhibited milder hypoalbuminemia and hyperlipidemia, and milder histological changes while maintaining the expression of podocyte markers during podocyte injury model. Taken together, our results show that Twsg1 plays a critical role in the modulation of protective action of Bmp7 on podocytes, and that inhibition of Twsg1 is a promising means of development of novel treatment for podocyte injury. [ABSTRACT FROM AUTHOR]

The number of nephrons, the functional units of the kidney, varies among individuals. A low nephron number at birth is associated with a risk of hypertension and the progression of renal insufficiency. The molecular mechanisms determining nephron number during embryogenesis have not yet been clarified. Germline knockout of bone morphogenetic protein 7 (Bmp7) results in massive apoptosis of the kidney progenitor cells and defects in early stages of nephrogenesis. This phenotype has precluded analysis of Bmp7 function in the later stage of nephrogenesis. In this study, utilization of conditional null allele of Bmp7 in combination with systemic inducible Cre deleter mice enabled us to analyze Bmp7 function at desired time points during kidney development, and to discover the novel function of Bmp7 to inhibit the precocious differentiation of the progenitor cells to nephron. Systemic knockout of Bmp7 in vivo after the initiation of kidney development results in the precocious differentiation of the kidney progenitor cells to nephron, in addition to the prominent apoptosis of progenitor cells. We also confirmed that in vitro knockout of Bmp7 in kidney explant culture results in the accelerated differentiation of progenitor population. Finally we utilized colony-forming assays and demonstrated that Bmp7 inhibits epithelialization and differentiation of the kidney progenitor cells. These results indicate that the function of Bmp7 to inhibit the precocious differentiation of the progenitor cells together with its anti-apoptotic effect on progenitor cells coordinately maintains renal progenitor pool in undifferentiated status, and determines the nephron number at birth. [ABSTRACT FROM AUTHOR]

Elongation of the digit rays resulting in the formation of a defined number of phalanges is a process poorly understood in mammals, whereas in the chicken distal mesenchymal bone morphogenetic protein (BMP) signaling in the so-called phalanx-forming region (PFR) or digit crescent (DC) seems to be involved. The human brachydactylies (BDs) are inheritable conditions characterized by variable degrees of digit shortening, thus providing an ideal model to analyze the development and elongation of phalanges. We used a mouse model for BDB1 (Ror2W749X/W749X) lacking middle phalanges and show that a signaling center corresponding to the chick PFR exists in the mouse, which is diminished in BDB1 mice. This resulted in a strongly impaired elongation of the digit condensations due to reduced chondrogenic commitment of undifferentiated distal mesenchymal cells. We further show that a similar BMP-based mechanism accounts for digit shortening in a mouse model for the closely related condition BDA1 (IhhE95K/E95K), altogether indicating the functional significance of the PFR in mammals. Genetic interaction experiments as well as pathway analysis in BDB1 mice suggest that Indian hedgehog and WNT/β-catenin signaling, which we show is inhibited by receptor tyrosine kinase-like orphan receptor 2 (ROR2) in distal limb mesenchyme, are acting upstream of BMP signaling in the PFR. [ABSTRACT FROM AUTHOR]

The formation of the skeleton requires inductive signals that are balanced with their antagonists in a highly regulated negative feedback system. Inappropriate or excessive expression of BMPs (bone morphogenetic proteins) or their antagonists results in genetic disorders affecting the skeleton, such as fibrodysplasia ossificans progressiva. BMP signaling mediated through binding to its receptors is a critical step in the induction of abnormal ossification. Therefore, we hypothesized that engineering more effective inhibitors of this BMP-signaling process may lead to the development of therapies for such conditions. BMP4-induced heterotopic ossification was used as a model for testing the ability of the BMP antagonist Noggin to block de novo bone formation, either by local or systemic delivery. Since Noggin naturally acts locally, a Noggin mutein, hNOGΔB2, was engineered and was shown to circulate systemically, and its ability to block heterotopic ossification was tested in a mouse model with use of adenovirus-mediated somatic cell gene transfer. BMP4-induced heterotopic ossification can be prevented in vivo either by local delivery of wild-type Noggin or after somatic cell gene transfer of a Noggin mutein, hNOGΔB2. Furthermore, the data in the present study provide proof of concept that a naturally occurring factor can be engineered for systemic delivery toward a desirable pharmacological outcome. Clinical Relevance: Blocking bone formation is clinically relevant to disorders of heterotopic ossification in humans, such as fibrodysplasia ossificans progressiva. Furthermore, development of BMP antagonists as therapeutic agents may provide modalities for the treatment of other pathologic conditions that arise from aberrant expression of BMPs and/or from a lack of their antagonists. [ABSTRACT FROM AUTHOR]

Inductive signals from adjacent tissues initiate differentiation within the somite. In this study, we used mouse embryos mutant for the BMP antagonists noggin (Nog) and gremlin 1 (Grem1) to characterize the effects of BMP signaling on the specification of the sclerotome. We confirmed reduction of Pax1 and Pax9 expression in Nog mutants, but found that Nog;Grem1 double mutants completely fail to initiate sclerotome development. Furthermore, Nog mutants that also lack one allele of Grem1 exhibit a dramatic reduction in axial skeleton relative to animals mutant for Nog alone. By contrast, Pax3, Myf5 and Lbx1 expression indicates that dermomyotome induction occurs in Nog;Grem1 double mutants. Neither conditional Bmpr1a mutation nor treatment with the BMP type I receptor inhibitor dorsomorphin expands sclerotome marker expression, suggesting that BMP antagonists do not have an instructive function in sclerotome specification. Instead, we hypothesize that Nog- and Grem1-mediated inhibition of BMP is permissive for hedgehog (Hh) signal-mediated sclerotome specification. In support of this model, we found that culturing Nog;Grem1 double-mutant embryos with dorsomorphin restores sclerotome, whereas Pax1 expression in smoothened (Smo) mutants is not rescued, suggesting that inhibition of BMP is insufficient to induce sclerotome in the absence of Hh signaling. Confirming the dominant inhibitory effect of BMP signaling, Pax1 expression cannot be rescued in Nog;Grem1 double mutants by forced activation of Smo. We conclude that Nog and Grem1 cooperate to maintain a BMP signaling-free zone that is a crucial prerequisite for Hh-mediated sclerotome induction. [ABSTRACT FROM AUTHOR]

Gremlin is a glycoprotein that binds bone morphogenetic proteins (BMPs) 2,4, and 7, antagonizing their actions. Gremlin opposes BMP effects on osteoblastic differentiation and function in vitro and in vivo, and its overexpression causes osteopenia. To define the function of gremlin in the skeleton, we generated gremlin 1 (grem1) conditional null mice by mating mice where grem1 was flanked by loxP sequences with mice expressing the Cre recombinase under the control of the osteocalcin promoter. grem1 null male mice displayed increased trabecular bone volume due to enhanced osteoblastic activity, because mineral apposition and bone formation rates were increased. Osteoblast number and bone resorption were not altered. Marrow stromal cells from grem1 conditional null mice expressed higher levels of alkaline phosphatase activity. Gremlin down-regulation by RNA interference in ST-2 stromal and MC3T3 osteoblastic cells increased the BMP-2 stimulatory effect on alkaline phosphatase activity, on Smad 1/5/8 phosphorylation, and on the transactivation of the BMP/Smad reporter construct 12XSBE-Oc-pGL3. Gremlin down-regulation also enhanced osteocalcin and Runx-2 expression, Wnt 3a signaling, and activity in ST-2 cells, In conclusion, deletion of grem1 in the bone microenvironment results in sensitization of BMP signaling and activity and enhanced bone formation in vivo. [ABSTRACT FROM AUTHOR]