Inhibition Of Bone Morphogenetic Protein Signaling Research Articles

Bone morphogenetic protein signaling inhibitor improves differentiation and function of 3D muscle construct fabricated using C2C12

Functional tissue-engineered artificial skeletal muscle tissue has great potential for pharmacological and academic applications. This study demonstrates an in vitro tissue engineering system to construct functional artificial skeletal muscle tissues using self-organization and signal inhibitors. To induce efficient self-organization, we optimized the substrate stiffness and extracellular matrix (ECM) coatings. We modified the tissue morphology to be ring-shaped under optimized self-organization conditions. A bone morphogenetic protein (BMP) inhibitor was added to improve overall myogenic differentiation. This supplementation enhanced the myogenic differentiation ratio and myotube hypertrophy in two-dimensional cell cultures. Finally, we found that myotube hypertrophy was enhanced by a combination of self-organization with ring-shaped tissue and a BMP inhibitor. BMP inhibitor treatment significantly improved myogenic marker expression and contractile force generation in the self-organized tissue. These observations indicated that this procedure may provide a novel and functional artificial skeletal muscle for pharmacological studies.

Inhibition of BMP signaling pathway induced senescence and calcification in anaplastic meningioma

PurposeMeningiomas are the most common type of brain tumors and are generally benign, but malignant atypical meningiomas and anaplastic meningiomas frequently recur with poor prognosis. The metabolism of meningiomas is little known, so few effective treatment options other than surgery and radiation are available, and the targets for treatment of recurrence are not well defined. The Aim of this paper is to find the therapeutic target.MethodsThe effects of bone morphogenetic protein (BMP) signal inhibitor (K02288) and upstream regulator Gremlin2 (GREM2) on meningioma’s growth and senescence were examined. In brief, we examined as follows: 1) Proliferation assay by inhibiting BMP signaling. 2) Comprehensive analysis of forced expression GREM2.3) Correlation between GREM2 mRNA expression and proliferation marker in 87 of our clinical samples. 4) Enrichment analysis between GREM2 high/low expressed groups using RNA-seq data (42 cases) from the public database GREIN. 5) Changes in metabolites and senescence markers associated with BMP signal suppression.ResultsInhibitors of BMP receptor (BMPR1A) and forced expression of GREM2 shifted tryptophan metabolism from kynurenine/quinolinic acid production to serotonin production in malignant meningiomas, reduced NAD + /NADH production, decreased gene cluster expression involved in oxidative phosphorylation, and caused decrease in ATP. Finally, malignant meningiomas underwent cellular senescence, decreased proliferation, and eventually formed psammoma bodies. Reanalyzed RNA-seq data of clinical samples obtained from GREIN showed that increased expression of GREM2 decreased the expression of genes involved in oxidative phosphorylation, similar to our experimental results.ConclusionsThe GREM2-BMPR1A-tryptophan metabolic pathway in meningiomas is a potential new therapeutic target.

A83-01 and DMH1 effects in the zebrafish spermatogonial niche: Unraveling the roles of TGF-β and BMP signaling in the Fsh-mediated spermatogonial fate

Transforming growth factor-β (TGF-β) and bone morphogenetic protein (BMP) signaling has fundamental roles in the regulation of the stem cell niche for both embryonic and adult stem cells. In zebrafish, male germ stem cell niche is regulated by follicle-stimulating hormone (Fsh) through different members of the TGF-β superfamily. On the other hand, the specific roles of TGF-β and BMP signaling pathways are unknown in the zebrafish male germ stem cell niche. Considering this lack of information, the present study aimed to investigate the pharmacological inhibition of TGF-β (A83-01) and BMP (DMH1) signaling pathways in the presence of recombinant zebrafish Fsh using testicular explants. We also reanalyzed single cell-RNA sequencing (sc-RNA-seq) dataset from adult zebrafish testes to identify the testicular cellular sites of smad expression, and to understand the physiological significance of the changes in smad transcript levels after inhibition of TGF-β or BMP pathways. Our results showed that A83-01 potentiated the pro-stimulatory effects of Fsh on spermatogonial differentiation leading to an increase in the proportion area occupied by differentiated spermatogonia with concomitant reduction of type A undifferentiated (Aund) spermatogonia. In agreement, expression analysis showed lower mRNA levels for the pluripotency gene pou5f3, and increased expression of dazl (marker of type B spermatogonia and spermatocyte) and igf3 (pro-stimulatory growth factor) following the co-treatment with TGF-β inhibitor and Fsh. Contrariwise, the inhibition of BMP signaling nullified the pro-stimulatory effects of Fsh, resulting in a reduction of differentiated spermatogonia and increased proportion area occupied by type Aund spermatogonia. Supporting this evidence, BMP signaling inhibition increased the mRNA levels of pluripotency genes nanog and pou5f3, and decreased dazl levels when compared to control. The sc-RNA-seq data unveiled a distinctive pattern of smad expression among testicular cells, primarily observed in spermatogonia (smad 2, 3a, 3b, 8), spermatocytes (smad 2, 3a, 8), Sertoli cells (smad 1, 3a, 3b), and Leydig cells (smad 1, 2). This finding supports the notion that inhibition of TGF-β and BMP signaling pathways may predominantly impact cellular components within the spermatogonial niche, namely spermatogonia, Sertoli, and Leydig cells. In conclusion, our study demonstrated that TGF-β and BMP signaling pathways exert antagonistic roles in the zebrafish germ stem cell niche. The members of the TGF-β subfamily are mainly involved in maintaining the undifferentiated state of spermatogonia, while the BMP subfamily promotes spermatogonial differentiation. Therefore, in the complex regulation of the germ stem cell niche by Fsh, members of the BMP subfamily (pro-differentiation) should be more predominant in the niche than those belonging to the TGF-β (anti-differentiation). Overall, these findings are not only relevant for understanding the regulation of germ stem cell niche but may also be useful for expanding in vitro the number of undifferentiated spermatogonia more efficiently than using recombinant hormones or growth factors.

SMAD1 Is Dispensable for CDX2 Induction but Required for the Repression of Ectopic Small-Intestinal Gene Expression in Human-Pluripotent-Stem-Cell-Derived Colonic Organoids

The generation of gastrointestinal tissues from human pluripotent stem cells has provided unprecedented insight into the molecular mechanisms that drive the patterning of the primitive gut tube. Previous work has identified bone-morphogenetic-protein (BMP) signaling as an important mediator of mid/hindgut versus foregut and hindgut versus midgut cell fate choice. Inhibition of BMP signaling during gut tube morphogenesis inhibits the expression of the pan-intestinal transcription factor CDX2. Treatment of CDX2+ mid/hindgut cultures with BMP patterns them into hindgut, which gives rise to colonic organoids (HCOs). While the role for BMP signaling is clear, the molecular mechanisms through which BMP signaling patterns the mid/hindgut and colon remain unclear. BMPs bind to BMP receptors, activating a signaling cascade that results in the activation of SMADs, which function as transcription factors. We hypothesized that one of these factors, SMAD1, would be necessary for establishing the CDX2 domain and the colon domain. Unexpectedly, endoderm derived from SMAD1-deficient induced pluripotent stem cells was capable of inducing CDX2 in response to WNT and FGF signaling. In addition, CDX2+ gut tube cultures could activate posterior HOX genes in response to BMP. However, examination of HCOs following cytodifferentiation revealed that SMAD1-deficient HCOs ectopically expressed small-intestinal markers despite expressing posterior HOX genes. These results indicate that there is redundancy of SMADs during early hindgut patterning but that SMAD1 is required for the inhibition of small-intestinal gene expression in HCOs.

Characterization of erythroferrone structural domains relevant to its iron-regulatory function

Iron delivery to the plasma is closely coupled to erythropoiesis, the production of red blood cells, as this process consumes most of the circulating plasma iron. In response to hemorrhage and other erythropoietic stresses, increased erythropoietin stimulates the production of the hormone erythroferrone (ERFE) by erythrocyte precursors (erythroblasts) developing in erythropoietic tissues. ERFE acts on the liver to inhibit bone morphogenetic protein (BMP) signaling and thereby decrease hepcidin production. Decreased circulating hepcidin concentrations then allow the release of iron from stores and increase iron absorption from the diet. Guided by evolutionary analysis and Alphafold2 protein complex modeling, we used targeted ERFE mutations, deletions, and synthetic ERFE segments together with cell-based bioassays and surface plasmon resonance to probe the structural features required for bioactivity and BMP binding. We define the ERFE active domain and multiple structural features that act together to entrap BMP ligands. In particular, the hydrophobic helical segment 81 to 86 and specifically the highly conserved tryptophan W82 in the N-terminal region are essential for ERFE bioactivity and Alphafold2 modeling places W82 between two tryptophans in its ligands BMP2, BMP6, and the BMP2/6 heterodimer, an interaction similar to those that bind BMPs to their cognate receptors. Finally, we identify the cationic region 96-107 and the globular TNFα-like domain 186-354 as structural determinants of ERFE multimerization that increase the avidity of ERFE for BMP ligands. Collectively, our results provide further insight into the ERFE-mediated inhibition of BMP signaling in response to erythropoietic stress.

Abstract 2844: BMP inhibition overcomes chemoresistance of prostate cancer

Abstract Chemoresistance is a major therapeutic challenge to prostate cancer and its underlying molecular mechanism is poorly understood. Previously, it has been suggested that bone morphogenetic protein (BMP) signaling is down-regulated during the prostate cancer progression from the early androgen-sensitive stage to the metastatic castration-resistant stage. However, no literature reports are available for BMP signaling in the more advanced-chemoresistant prostate cancer. In this study, we found the expression levels of the BMP type I receptor members, Activin-like kinase-2 (ALK2) and Activin-like kinase-3 (ALK3), were significantly higher in the chemoresistant prostate cancer cells than those in the chemosensitive prostate cancer cells. In addition, the phospho-Smad1/5/9 proteins, the pivotal intracellular effectors of the BMP signaling, were notably elevated in the chemoresistant prostate cancer cells over the chemosensitive prostate cancer cells, indicating that BMP signaling is highly activated in the chemoresistant prostate cancer cells. We also found that BMP signaling inhibition with either DMH1 or the knockdown of ALK2/ALK3 sensitized chemoresistant prostate cancer cells to the chemotherapy drug docetaxel in a dose dependent manner. Our further study indicates that DMH1 suppressed the migration and invasion of chemoresistant prostate cancer cells in vitro, and attenuated chemoresistant prostate tumor growth in the mouse xenograft model in vivo. In addition, we showed that DMH1 disrupted the sphere formation in DU145-TxR and PC3-TxR cells, suppressed the expression of marker genes of the cancer stem cells (CSCs), and inhibited the efflux activity of multidrug resistance P-glycoprotein (P-gp1). In conclusion, our study demonstrates that BMP signaling is associated with prostate cancer chemoresistance and BMP signaling inhibition effectively overcomes the cancer chemoresistance potentially through the disruption of CSCs’ stemness and P-gp1 efflux activity. Citation Format: Jijun Hao, Chen Xie, Zhijun Wang, Yong Bar, Jose Aguilar, Austin Kyan, Li Zhing. BMP inhibition overcomes chemoresistance of prostate cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2844.

Intrinsic BMP inhibitor Gremlin regulates alveolar epithelial type II cell proliferation and differentiation

Type 1 alveolar epithelial cells (AT1s) and type 2 alveolar epithelial cells (AT2s) regulate the structural integrity and function of alveoli. AT1s mediate gas exchange, whereas AT2s serve multiple functions, including surfactant secretion and alveolar repair through proliferation and differentiation into AT1s as progenitors. However, mechanisms regulating AT2 proliferation and differentiation remain unclear. Here we demonstrate that Gremlin, an intrinsic inhibitor of bone morphogenetic protein (BMP), induces AT2 proliferation and differentiation. Transient overexpression of Gremlin in rat lungs by adenovirus vector delivery suppressed BMP signaling, induced proliferation of AT2s and the production of Bmp2, which in turn led to the recovery of BMP signaling and induced AT2 differentiation into AT1s. Bleomycin-induced lung injury upregulated Gremlin and showed a similar time course of biomarker expression comparable to the adenovirus model. TGF-β and IL-1β induced Gremlin expression in fibroblasts. Taken together, our findings implicate that Gremlin expression during lung injury leads to precisely timed inhibition of BMP signaling and activates AT2s, leading to alveolar repair.

Exosomal miR-128-3p reversed fibrinogen-mediated inhibition of oligodendrocyte progenitor cell differentiation and remyelination after cerebral ischemia.

To investigate the role of exosomal miR-128-3p in promoting fibrinogen-mediated inhibition of oligodendrocyte progenitor cell (OPC) differentiation and the therapeutic potential of exosomal miR-128-3p in cerebral ischemia. Mouse models of middle cerebral artery occlusion (MCAO) were established as described previously. MCAO was treated with fibrinogen and exosomes by stereotactically injecting into the left stratum. Mouse cortical OPCs were used for mRNA and miRNA sequencing analysis. Exosomes were isolated from neural stem cells (NSCs) of mice. Fibrinogen deposition suppressed remyelination after MCAO and inhibited OPC differentiation by activating ACVR1, the bone morphogenetic protein (BMP) signaling type I receptor. In vitro, miR-sequencing and verification studies revealed that miR-128-3p is associated with BMP signaling mediated by ACVR1. Additionally, transfer of NSC-derived exosomal miR-128-3p to OPCs significantly increased myelin basic protein expression and inhibited BMP signaling. Furthermore, NSC-derived exosomal miR-128-3p protected against fibrinogen-induced demyelination related to BMP signaling, reduced the infarct volume, and improved neurological function after MCAO. Fibrinogen deposition inhibits remyelination after ischemic damage and NSC-derived exosomal miR-128-3p promotes OPC differentiation into OLs by suppressing BMP signaling, indicating that NSC-derived exosomal miR-128-3p represents a potential therapeutic target for ischemic stroke.

Identification of the Time Period during Which BMP Signaling Regulates Proliferation of Neural Progenitor Cells in Zebrafish.

Bone morphogenetic protein (BMP) signaling regulates neural induction, neuronal specification, and neuronal differentiation. However, the role of BMP signaling in neural progenitors remains unclear. This is because interruption of BMP signaling before or during neural induction causes severe effects on subsequent neural developmental processes. To examine the role of BMP signaling in the development of neural progenitors in zebrafish, we bypassed the effect of BMP signaling on neural induction and suppressed BMP signaling at different time points during gastrulation using a temporally controlled transgenic line carrying a dominant-negative form of Bmp receptor type 1aa and a chemical inhibitor of BMP signaling, DMH1. Inhibiting BMP signaling from 8 hpf could bypass BMP regulation on neural induction, induce the number of proliferating neural progenitors, and reduce the number of neuronal precursors. Inhibiting BMP signaling upregulates the expression of the Notch downstream gene hairy/E(spl)-related 2 (her2). Inhibiting Notch signaling or knocking down the Her2 function reduced neural progenitor proliferation, whereas inactivating BMP signaling in Notch-Her2 deficient background restored the number of proliferating neural progenitors. These results reveal the time window for the proliferation of neural progenitors during zebrafish development and a fine balance between BMP and Notch signaling in regulating the proliferation of neural progenitor cells.

Regulation of Parietal Cell Homeostasis by Bone Morphogenetic Protein Signaling

<h3>Background and Aims</h3> Loss of bone morphogenetic protein (BMP) signaling in the stomach, achieved by transgenic expression of the BMP inhibitor noggin (<i>H</i>⁺<i>/K</i>⁺<i>-Nog</i> mice), causes parietal cell (PC) loss, spasmolytic polypeptide-expressing metaplasia, a marker of preneoplasia, and activation of cell proliferation. We examined if specific inhibition of BMP signaling in PCs leads to aberrations in epithelial homeostasis. <h3>Methods</h3> Mice with floxed alleles of BMP receptor 1a (<i>Bmpr1a</i>^flox/flox mice) were crossed to <i>H</i>⁺<i>/K</i>⁺<i>-Cre</i> mice to generate <i>H</i>⁺<i>/K</i>⁺<i>-Cre;Bmpr1a</i>^flox/flox mice. Morphology of the mucosa was analyzed by hematoxylin and eosin staining. Distribution of H⁺/K⁺-ATPase-, IF-, and Ki-67-positive cells was analyzed by immunostaining. Expression of pit and neck cell mucins was determined by staining with the lectins Ulex Europaeus Agglutinin 1 and Griffonia (Bandeiraea) simplicifolia lectin II, respectively. Isolation of PCs from control and <i>Nog</i>-expressing mice was achieved by crossing <i>H</i>⁺<i>/K</i>⁺<i>-Nog</i> mice to <i>Rosa26-tdTomato</i> (Tom) mice to generate <i>H</i>⁺<i>/K</i>⁺<i>-Nog;Rosa26-tdTom</i> mice. <i>H</i>⁺<i>/K</i>⁺<i>-Cre</i> mice were then crossed to <i>H</i>⁺<i>/K</i>⁺<i>-Nog;Rosa26-tdTom</i> mice to generate <i>H</i>⁺<i>/K</i>⁺<i>-Cre;H</i>⁺<i>/K</i>⁺<i>-Nog;Rosa26-tdTom</i> mice. Tom-labeled PCs were purified by flow cytometry. Changes in PC transcripts were measured by RNA-Seq. <h3>Results</h3> Six-month-old <i>H</i>⁺<i>/K</i>⁺<i>-Cre;Bmpr1a</i>^flox/flox mice exhibited increased epithelial cell proliferation, presence of transitional cells showing colocalization of IF with both Griffonia (Bandeiraea) simplicifolia lectin II-binding mucins and the H⁺/K⁺-ATPase, and expansion of Ulex Europaeus Agglutinin 1-positive cells. PC transcripts from Nog-expressing mice demonstrated induction of markers of Spasmolytic Polypeptide-Expressing Metaplasia. <h3>Conclusion</h3> PC-specific loss of BMP signaling alters the homeostasis of the gastric epithelium leading to the development of metaplasia.

GTPase-activating protein TBC1D5 coordinates with retromer to constrain synaptic growth by inhibiting BMP signaling

Formation and plasticity of neural circuits rely on precise regulation of synaptic growth. At Drosophila neuromuscular junction (NMJ), Bone Morphogenetic Protein (BMP) signaling is critical for many aspects of synapse formation and function. The evolutionarily conserved retromer complex and its associated GTPase-activating protein TBC1D5 are critical regulators of membrane trafficking and cellular signaling. However, their functions in regulating the formation of NMJ are less understood. Here, we report that TBC1D5 is required for inhibition of synaptic growth, and loss of TBC1D5 leads to abnormal presynaptic terminal development, including excessive satellite boutons and branch formation. Ultrastructure analysis reveals that the size of synaptic vesicles and the density of subsynaptic reticulum are increased in TBC1D5 mutant boutons. Disruption of interactions of TBC1D5 with Rab7 and retromer phenocopies the loss of TBC1D5. Unexpectedly, we find that TBC1D5 is functionally linked to Rab6, in addition to Rab7, to regulate synaptic growth. Mechanistically, we show that loss of TBC1D5 leads to upregulated BMP signaling by increasing the protein level of BMP type II receptor Wishful Thinking (Wit) at NMJ. Overall, our data establish that TBC1D5 in coordination with retromer constrains synaptic growth by regulating Rab7 activity, which negatively regulates BMP signaling through inhibiting Wit level.

TXNIP Suppresses the Osteochondrogenic Switch of Vascular Smooth Muscle Cells in Atherosclerosis.

The osteochondrogenic switch of vascular smooth muscle cells (VSMCs) is a pivotal cellular process in atherosclerotic calcification. However, the exact molecular mechanism of the osteochondrogenic transition of VSMCs remains to be elucidated. Here, we explore the regulatory role of TXNIP (thioredoxin-interacting protein) in the phenotypical transitioning of VSMCs toward osteochondrogenic cells responsible for atherosclerotic calcification. The atherosclerotic phenotypes of Txnip-/- mice were analyzed in combination with single-cell RNA-sequencing. The atherosclerotic phenotypes of Tagln-Cre; Txnipflox/flox mice (smooth muscle cell-specific Txnip ablation model), and the mice transplanted with the bone marrow of Txnip-/- mice were analyzed. Public single-cell RNA-sequencing dataset (GSE159677) was reanalyzed to define the gene expression of TXNIP in human calcified atherosclerotic plaques. The effect of TXNIP suppression on the osteochondrogenic phenotypic changes in primary aortic VSMCs was analyzed. Atherosclerotic lesions of Txnip-/- mice presented significantly increased calcification and deposition of collagen content. Subsequent single-cell RNA-sequencing analysis identified the modulated VSMC and osteochondrogenic clusters, which were VSMC-derived populations. The osteochondrogenic cluster was markedly expanded in Txnip-/- mice. The pathway analysis of the VSMC-derived cells revealed enrichment of bone- and cartilage-formation-related pathways and bone morphogenetic protein signaling in Txnip-/- mice. Reanalyzing public single-cell RNA-sequencing dataset revealed that TXNIP was downregulated in the modulated VSMC and osteochondrogenic clusters of human calcified atherosclerotic lesions. Tagln-Cre; Txnipflox/flox mice recapitulated the calcification and collagen-rich atherosclerotic phenotypes of Txnip-/- mice, whereas the hematopoietic deficiency of TXNIP did not affect the lesion phenotype. Suppression of TXNIP in cultured VSMCs accelerates osteodifferentiation and upregulates bone morphogenetic protein signaling. Treatment with the bone morphogenetic protein signaling inhibitor K02288 abrogated the effect of TXNIP suppression on osteodifferentiation. Our results suggest that TXNIP is a novel regulator of atherosclerotic calcification by suppressing bone morphogenetic protein signaling to inhibit the transition of VSMCs toward an osteochondrogenic phenotype.

Transmembrane anterior posterior transformation 1 regulates BMP signaling and modulates the protein stability of SMAD1/5

The bone morphogenetic protein (BMP) signaling pathway plays pivotal roles in various biological processes during embryogenesis and adult homeostasis. Transmembrane anterior posterior transformation 1 (TAPT1) is an evolutionarily conserved protein involved in murine axial skeletal patterning. Genetic defects in TAPT1 result in complex lethal osteochondrodysplasia. However, the specific cellular activity of TAPT1 is not clear. Herein, we report that TAPT1 inhibits BMP signaling and destabilizes the SMAD1/5 protein by facilitating its interaction with SMURF1 E3 ubiquitin ligase, which leads to SMAD1/5 proteasomal degradation. In addition, we found that the activation of BMP signaling facilitates the redistribution of TAPT1 and promotes its association with SMAD1. TAPT1-deficient murine C2C12 myoblasts or C3H/10T1/2 mesenchymal stem cells exhibit elevated SMAD1/5/9 protein levels, which amplifies BMP activation, in turn leading to a boost in the transdifferentiation or differentiation processing of these distinct TAPT1-deficient cell lines changing into mature osteoblasts. Furthermore, the enhancing effect of TAPT1 deficiency on osteogenic differentiation of C3H/10T1/2cells was observed in an invivo ectopic bone formation model. Importantly, a subset of TAPT1 mutations identified in humans with lethal skeletal dysplasia exhibited gain-of-function activity on SMAD1 protein levels. Thus, this finding elucidates the role of TAPT1 in the regulation of SMAD1/5 protein stability for controlling BMP signaling.

Multifaceted Functions of TWSG1: From Embryogenesis to Cancer Development.

Bone morphogenetic proteins (BMPs) play an important role in development. Twisted gastrulation BMP signaling modulator 1 (TWSG1) was initially identified as a regulator of the dorsoventral axis formation in Drosophila. The mechanism of BMP signaling modulation by TWSG1 is complex. TWSG1 inhibits BMP signaling by binding to BMP ligands including BMP4, whereas it enhances signaling by interacting with Chordin, a BMP antagonist. Therefore, TWSG1 can act as both a BMP agonist and antagonist. TWSG1 has various functions ranging from embryogenesis to cancer progression. TWSG1 knockout mice showed neural, craniofacial, and mammary defects. TWSG1 also regulated erythropoiesis and thymocyte development. Furthermore, the relationship between TWSG1 and cancer has been elucidated. Allelic loss of TWSG1 was detected in colorectal cancer. TWSG1 expression was upregulated in papillary thyroid carcinoma and glioblastoma but downregulated in gastric and endometrial cancers. TWSG1 suppressed BMP7-enhanced sphere formation and migration in endometrial cancer cells, indicating its tumor-suppressive role. Further studies are required to clarify the TWSG1 function and its association with BMP signaling in cancer development. Finally, TWSG1 is abundantly expressed in human and mouse ovaries and sustains follicular growth in rodent ovaries. Thus, TWSG1 has various functions ranging from fertility to cancer. Therefore, TWSG1 signaling modulation may be beneficial in treating specific diseases such as cancer.

A Bone Morphogenetic Protein Signaling Inhibitor, LDN193189, Converts Ischemia-Induced Multipotent Stem Cells into Neural Stem/Progenitor Cell-Like Cells.

Stem cell therapy is used to restore neurological function in stroke patients. We have previously reported that ischemia-induced multipotent stem cells (iSCs), which are likely derived from brain pericytes, develop in poststroke human and mouse brains. Although we have demonstrated that iSCs can differentiate into neural lineage cells, the factors responsible for inducing this differentiation remain unclear. In this study, we found that LDN193189, a bone morphogenetic protein (BMP) inhibitor, caused irreversible changes in the shape of iSCs. In addition, compared with iSCs incubated without LDN193189, the iSCs incubated with LDN193189 (LDN-iSCs) showed upregulated expression of neural lineage-related genes and proteins, including those expressed in neural stem/progenitor cells (NSPCs), and downregulated expression of mesenchymal and pericytic-related genes and proteins. Moreover, microarray analysis revealed that LDN-iSCs and NSPCs had similar gene expression profiles. Furthermore, LDN-iSCs differentiated into electrophysiologically functional neurons. These results indicate that LDN193189 induces NSPC-like cells from iSCs, suggesting that bioactive molecules regulating BMP signaling are potential targets for promoting neurogenesis from iSCs in the pathological brain, such as during ischemic stroke. We believe that our findings will bring us one step closer to the clinical application of iSCs.

Chromatin remodeler Znhit1 controls bone morphogenetic protein signaling in embryonic lung tissue branching

Branching morphogenesis is a key process essential for lung and other organ development in which cellular and tissue architecture branch out to maximize surface area. While this process is known to be regulated by differential gene expression of ligands and receptors, how chromatin remodeling regulates this process remains unclear. Znhit1 (zinc finger HIT-type containing 1), acting as a chromatin remodeler, has previously been shown to control the deposition of the histone variant H2A.Z. Here, we demonstrate that Znhit1 also plays an important role in regulating lung branching. Using Znhit1 conditional KO mice, we show that Znhit1 deficiency in the embryonic lung epithelium leads to failure of branching morphogenesis and neonatal lethality, which is accompanied by reduced cell proliferation and increased cell apoptosis of the epithelium. The results from the transcriptome and the chromatin immunoprecipitation assay reveal that this is partially regulated by the derepression of Bmp4, encoding bone morphogenetic protein (BMP) 4, which is a direct target of H2A.Z. Furthermore, we show that inhibition of BMP signaling by the protein inhibitor Noggin rescues the lung branching defects of Znhit1 mutants ex vivo. Taken together, our study identifies the critical role of Znhit1/H2A.Z in embryonic lung morphogenesis via the regulation of BMP signaling.

Bone morphogenetic protein pathway responses and alterations of osteogenesis in metastatic prostate cancers.

Prostate cancer is a common cancer in men that annually results in more than 33 000 US deaths. Mortality from prostate cancer is largely from metastatic disease, reflecting on the great strides in the last century of treatments in care for the localized disease. Metastatic castrate resistant prostate cancer (mCRPC) will commonly travel to the bone, creating unique bone pathology that requires nuanced treatments in those sites with surgical, radio and chemotherapeutic interventions. The bone morphogenetic protein (BMP) pathway has been historically studied in the capacity to regulate the osteogenic nature of new bone. New mineralized bone generation is a frequent and common observation in mCRPC and referred to as blastic bone lesions. Less common are bone destructive lesions that are termed lytic. We queried the cancer genome atlas (TCGA) prostate cancer databases for the expression of the BMP pathway and found that distinct gene expression of the ligands, soluble antagonists, receptors, and intracellular mediators were altered in localized versus metastatic disease. Human prostate cancer cell lines have an innate ability to promote blastic- or lytic-like bone lesions and we hypothesized that inhibiting BMP signaling in these cell lines would result in a distinct change in osteogenesis gene expression with BMP inhibition. We found unique and common changes by comparing these cell lines response and unique BMP pathway alterations. We treated human PCa cell lines with distinct bone pathologic phenotypes with the BMP inhibitor DMH1 and found distinct osteogenesis responses. We analyzed distinct sites of metastatic PCa in the TCGA and found that BMP signaling was selectively altered in commons sites such as lymph node, bone and liver compared to primary tumors. Overall we conclude that BMPs in metastatic prostate cancer are important signals and functional mediators of diverse processes that have potential for individualized precision oncology in mCRPC.

Ym155 localizes to the mitochondria leading to mitochondria dysfunction and activation of AMPK that inhibits BMP signaling in lung cancer cells

The imidazolium compound Ym155 was first reported to be a survivin inhibitor. Ym155 potently induces cell death of many types of cancer cells in preclinical studies. However, in phase II clinical trials Ym155 failed to demonstrate a significant benefit. Studies have suggested that the cytotoxic effects of Ym155 in cancer cells are not mediated by the inhibition of survivin. Understanding the mechanism by which Ym155 induces cell death would provide important insight how to improve its efficacy as a cancer therapeutic. We demonstrate a novel mechanism by which Ym155 induces cell death by localizing to the mitochondria causing mitochondrial dysfunction. Our studies suggest that Ym155 binds mitochondrial DNA leading to a decrease in oxidative phosphorylation, decrease in TCA cycle intermediates, and an increase in mitochondrial permeability. Furthermore, we show that mitochondrial stress induced by Ym155 and other mitochondrial inhibitors activates AMP-activated kinase leading to the downregulation to bone morphogenetic protein (BMP) signaling. We provide first evidence that Ym155 initiates cell death by disrupting mitochondrial function.

Bone morphogenetic protein receptor inhibitors suppress the growth of glioblastoma cells.

Glioblastomas (GBMs) are aggressive brain tumors that are resistant to chemotherapy and radiation. Bone morphogenetic protein (BMP) ligand BMP4 is being examined as a potential therapeutic for GBMs because it induces differentiation of cancer stem cells (CSCs) to an astrocyte phenotype. ID1 is reported to promote self-renewal and inhibit CSC differentiation. In most cancers, ID1 is transcriptionally upregulated by BMP4 promoting invasion and stemness. This conflicting data bring into question whether BMP signaling is growth suppressive or growth promoting in GBMs. We utilized BMP inhibitors DMH1, JL5, and Ym155 to examine the role of BMP signaling on the growth of GBMs. DMH1 targets BMP type 1 receptors whereas JL5 inhibits both the type 1 and type 2 BMP receptors. Ym155 does not bind the BMP receptors but rather inhibits BMP signaling by inducing the degradation of BMPR2. We show that JL5, DMH1, and Ym155 decreased the expression of ID1 in SD2 and U87 cells. JL5 and Ym155 also decreased the expression of BMPR2 and its downstream target inhibitor of apoptosis protein XIAP. JL5 treatment resulted in significant cell death and suppressed self-renewal to a greater extent than that induced by BMP4 ligand. The lysosome inhibitor chloroquine increases the localization of BMPR2 to the plasma membrane enhancing JL5-induced downregulation of ID1 and cell death in SD2 cells. We show that BMP signaling is growth promoting in GBMs. These studies suggest the need for development of BMP inhibitors and evaluation as potential therapeutic for GBMs.

Inhibition of BMP signaling with LDN 193189 can influence bone marrow stromal cell fate but does not prevent hypertrophy during chondrogenesis.

SummaryBone morphogenetic protein (BMP) cascades are upregulated during bone marrow-derived stromal cell (BMSC) chondrogenesis, contributing to hypertrophy and preventing effective BMSC-mediated cartilage repair. Previous work demonstrated that a proprietary BMP inhibitor prevented BMSC hypertrophy, yielding stable cartilage tissue. Because of the significant therapeutic potential of a molecule capable of hypertrophy blockade, we evaluated the capacity of a commercially available BMP type I receptor inhibitor with similar properties, LDN 193189, to prevent BMSC hypertrophy. Using 14-day microtissue chondrogenic induction cultures we found that LDN 193189 permitted BMSC chondrogenesis but did not prevent hypertrophy. LDN 193189 was sufficiently potent to counter mineralization and adipogenesis in response to exogenous BMP-2 in osteogenic induction cultures. LDN 193189 did not modify BMSC behavior in adipogenic induction cultures. Although LDN 193189 is effective in countering BMP signaling in a manner that influences BMSC fate, this blockade is insufficient to prevent hypertrophy.