Role of FGFs in the control of programmed cell death during limb development

We have investigated the role of FGFs in the control of programmed cell death during limb development by analyzing the effects of increasing and blocking FGF signaling in the avian limb bud. BMPs are currently considered as the signals responsible for cell death. Here we show that FGF signaling is also necessary for apoptosis and that the establishment of the areas of cell death is regulated by the convergence of FGF- and BMP-mediated signaling pathways. As previously demonstrated, cell death is inhibited for short intervals (12 hours) after administration of FGFs. However, this initial inhibition is followed (24 hours) by a dramatic increase in cell death, which can be abolished by treatments with a BMP antagonist (Noggin or Gremlin). Conversely, blockage of FGF signaling by applying a specific FGF-inhibitor (SU5402) into the interdigital regions inhibits both physiological cell death and that mediated by exogenous BMPs. Furthermore, FGF receptors 1, 2 and 3 are expressed in the autopodial mesoderm during the regression of the interdigital tissue, and the expression of FGFR3 in the interdigital regions is regulated by FGFs and BMPs in the same fashion as apopotosis. Together our findings indicate that, in the absence of FGF signaling BMPs are not sufficient to trigger apoptosis in the developing limb. Although we provide evidence for a positive influence of FGFs on BMP gene expression, the physiological implication of FGFs in apoptosis appears to result from their requirement for the expression of genes of the apoptotic cascade. We have identified MSX2 and Snail as candidate genes associated with apoptosis the expression of which requires the combined action of FGFs and BMPs.     

The role of bone morphogenetic proteins in endochondral bone formation

Bone morphogenetic proteins (BMPs) were originally identified as proteins capable of inducing endochondral bone formation when implanted at extraskeletal sites. BMPs have diverse biological activities during early embryogenesis and various aspects of organogenesis. BMPs bind to BMP receptors on the cell surface, and these signals are transduced intracellularly by Smad proteins. BMP signal pathways can be inhibited by both extra- and intracellular mechanisms. As for skeletal development, genetic studies suggest that BMPs are skeletal mesoderm inducers. Recent studies of tissue-specific activation and inactivation of BMP signals have revealed that BMP signals control proliferation and differentiation of chondrocytes, differentiation of osteoblasts and bone quality. These findings may contribute not only to understanding of bone biology and pathology, but also to improvement of the clinical efficacy of BMPs.     

Smad8 is expressed in the anterior necrotic zone: evidence for a role of bone morphogenetic proteins/SMAD signaling in the activation of a molecular cascade that culminates in cell death

Bone morphogenetic proteins (BMPs) play a crucial role in programmed cell death (PCD), a biological process required for the sculpturing of the embryonic limbs. However, it is unknown if BMP signaling directly promotes cell death, or if it induces a molecular cascade that culminates in cell death. Given that Smad8, which encodes one component of BMP signaling, is expressed during the regression of interdigital tissue and responds to BMPs, we presumed that it may be expressed in other cell death areas during chick limb development such as the anterior and posterior necrotic zones (ANZ and PNZ). The present study found that the Smad8 expression pattern in the anterior mesoderm of the hindlimb is very similar to that observed in limbs stained to detect cell death. Also, BMPs and retinoic acid, which act as apoptosis-promoting factors, induced expression of Smad8 before the onset of cell death, while sonic hedgehog protein, acting as a survival factor, inhibited Smad8 expression in the ANZ. However, although there was correlation between Smad8 expression patterns and PCD in the ANZ, phosphorylated forms of SMAD1/5/8 and TUNEL staining did not co-localize in dying cells. Interestingly, a short pulse of BMP was sufficient to trigger cell death. On the other hand, most dying cells were located in the avascular region, while many cells expressing Smad8 were located in the vascular region of the ANZ. These results suggest that BMPs mediated by SMAD signaling activate a molecular cascade that culminates in PCD.     

Controlling of bone morphogenetic protein signaling

Bone morphogenetic proteins (BMPs) are multifunctional growth factors and play crucial roles during embryonic development, skeletal development, and cell fate determination. Their signals are transduced from cell membrane to the nucleus through intracellular signaling mediators. At present, different signaling pathways have been identified, and elaborate of network of regulators involved in the signaling control. The aim of the present review is to describe the recent understanding of BMPs signaling with emphasis on the regulation of its signal transduction at extracellular level, intracellular level, Smad-interacting factors in the nucleus, and Smad-independent signaling pathways, respectively.     

BMP signaling in the control of skin development and hair follicle growth

Bone morphogenetic proteins (BMPs), their antagonists, and BMP receptors are involved in controlling a large number of biological functions including cell proliferation, differentiation, cell fate decision, and apoptosis in many different types of cells and tissues during embryonic development and postnatal life. BMPs exert their biological effects via using BMP-Smad and BMP-MAPK intracellular pathways. The magnitude and specificity of BMP signaling are regulated by a large number of modulators operating on several levels (extracellular, cytoplasmic, nuclear). In developing and postnatal skin, BMPs, their receptors, and BMP antagonists show stringent spatio-temporal expressions patterns to achieve proper regulation of cell proliferation and differentiation in the epidermis and in the hair follicle. Genetic studies assert an essential role for BMP signaling in the control of cell differentiation and apoptosis in developing epidermis, as well as in the regulation of key steps of hair follicle development (initiation, cell fate decision, cell lineage differentiation). In postnatal hair follicles, BMP signaling plays an important role in controlling the initiation of the growth phase and is also involved in the regulation of apoptosis-driven hair follicle involution. However, additional efforts are required to fully understand the mechanisms and targets involved in the realization of BMP effects on distinct cell population in the skin and hair follicle. Progress in this area of research will hopefully lead to the development of new therapeutic approaches for using BMPs and BMP antagonists in the treatment of skin and hair growth disorders.     

Zebrafish bmp4 functions during late gastrulation to specify ventroposterior cell fates

Bone morphogenetic proteins (BMPs) are key mediators of dorsoventral patterning in vertebrates and are required for the induction of ventral fates in fish and frogs. A widely accepted model of dorsoventral patterning postulates that a morphogenetic BMP activity gradient patterns cell fates along the dorsoventral axis. Recent work in zebrafish suggests that the role of BMP signaling changes over time, with BMPs required for global dorsoventral patterning during early gastrulation and for tail patterning during late gastrulation and early somitogenesis. Key questions remain about the late phase, including which BMP ligands are required and how the functions of BMPs differ during the early and late gastrula stages. In a screen for dominant enhancers of mutations in the homeobox genes vox and vent, which function in parallel to bmp signaling, we identified an insertion mutation in bmp4. We then performed a reverse genetic screen to isolate a null allele of bmp4. We report the characterization of these two alleles and demonstrate that BMP4 is required during the later phase of BMP signaling for the specification of ventroposterior cell fates. Our results indicate that different bmp genes are essential at different stages. In addition, we present genetic evidence supporting a role for a morphogenetic BMP gradient in establishing mesodermal fates during the later phase of BMP signaling.     

BMP/Smad信号通路与哺乳动物卵泡发生

BMPs属于TGF-b超家族成员, 在调节哺乳动物的生长、细胞增殖和分化等方面有很广泛的生物学功能。越来越多的证据显示, BMPs在雌性哺乳动物生殖, 尤其在卵泡发生过程中发挥重要作用。Smads蛋白是BMP家族细胞内信号转导分子, 可将BMPs胞外信号从细胞膜传递入细胞核。文章对BMPs、BMP/Smad信号通路和BMP如何被调节进行概述, 并重点对BMP/Smad信号通路在卵泡发生过程中所起的调控作用进行综述。     

BMPs and BMPRs in chicken ovary and effects of BMP-4 and -7 on granulosa cell proliferation and progesterone production in vitro

Bone morphogenetic proteins (BMPs) and their receptors (BMPRs) are now known to have important roles in mammalian ovarian folliculogenesis. This study determined the expression of the mRNA encoding for BMPs and their receptors in the chicken ovary and explored possible roles for them. The expression of the mRNA for BMP-2, -4, -6, -7, and BMPR-IA, -IB, and -II was determined and quantified by a semiquantitative RT-PCR. The mRNAs for all the BMPs and receptors determined were present in both the granulosa (G) and theca (T) cells of the F1, F2, and F3 follicles. All BMP mRNAs increased in G cells with follicular development, whereas only BMP-7 mRNA had this trend in the T cells. BMP-2, -4, and -6 mRNAs in T were similar between follicles. BMPR-IA mRNA was similar in F2G and F3G but lower in F1G. BMPR-IB mRNA was similar in G of all follicles, and BMPR-II mRNA increased with development. In the T, each receptor subtype showed equal distribution between follicles. mRNA levels for BMPR-IB and -II were higher in G than in T, suggesting that the G is a major target for BMPs. BMP-4 and -7 stimulated basal, IGF-I-, and gonadotropin-stimulated progesterone production by cultured G cells, with differential responses between cells from the F1 and F3/4. This suggests involvement in follicular differentiation. BMP-4 and -7 reversed the inhibitory effects of transforming growth factor (TGF)-alpha on basal and gonadotropin-stimulated G cell progesterone production, with greater effect in the F1 than in the F3/4. This effect suggests an important role for BMPs interacting with TGF-alpha in modulating the effects of gonadotropins and IGF-I on follicular differentiation. Finally, BMP-7 stimulated G cell proliferation, but BMP-4 inhibited TGF-alpha+ IGF-I- and/or FSH-stimulated G cell proliferation, suggesting a role in the control of follicular growth during development. These effects of BMP-4 and -7 on the G cell function showed relationships with the expression levels of the BMPs and the BMPR-II.     

BMP signalling in early Xenopus development

Bone morphogenetic proteins (BMPs) are typically members of the transforming growth factor β (TGF-β) family with diverse roles in embryonic development. At least five genes with homology to BMPs are expressed during Xenopus development, along with their receptors and intracellular signalling pathways. The evidence suggests that BMPs have roles to play in both mesoderm induction and dorsoventral patterning. Studies in Xenopus have also identified a number of inhibitory binding proteins for the classical BMPs, encoded by genes such as chordin and noggin. These proteins appear to be responsible for establishing a morphogen gradient of BMP4 activity, which specifies different dorsoventral fates in early gastrulae. An emerging theme is that inhibition of BMP signalling is an important mechanism regulating cell fate decisions in early development. BioEssays 21:751–760, 1999. © 1999 John Wiley & Sons, Inc.     

Bone morphogenetic proteins and their receptors in the eye

The human genome encodes at least 42 different members of the transforming growth factor-beta superfamily of growth factors. Bone morphogenetic proteins (BMPs) are the largest subfamily of proteins within the transforming growth factor-beta superfamily and are involved in numerous cellular functions including development, morphogenesis, cell proliferation, apoptosis, and extracellular matrix synthesis. This article first reviews BMPs and BMP receptors, BMP signaling pathways, and mechanisms controlling BMP signaling. Second, we review BMP and BMP receptor expression during embryonic ocular development/ differentiation and in adult ocular tissues. Lastly, future research directions with respect to BMP, BMP receptors, and ocular tissues are suggested.     

Suppression of tumor necrosis factor-mediated apoptosis by nuclear factor kappaB-independent bone morphogenetic protein/Smad signaling

The activation of nuclear factor kappaB (NF-kappa B) plays a pivotal role in the regulation of tumor necrosis factor (TNF)-mediated apoptosis. However, little is known about the regulation of TNF-mediated apoptosis by other signaling pathways or growth factors. Here, unexpectedly, we found that bone morphogenetic protein (BMP)-2 and BMP-4 inhibited TNF-mediated apoptosis by inhibition of caspase-8 activation in C2C12 cells, a pluripotent mesenchymal cell line that has the potential to differentiate into osteoblasts depending on BMP stimulation. Utilizing both a trans-dominant IkappaBalpha inhibitor of NF-kappaB expressed in C2C12 cells and IkappaB kinase beta-deficient embryonic mouse fibroblast, we show that BMP-mediated survival was independent of NF-kappaB activation. Rather, the antiapoptotic activity of BMPs functioned through the Smad signaling pathway. Thus, these findings provide the first report of a BMP/Smad signaling pathway that can inhibit TNF-mediated apoptosis, independent of the prosurvival activity of NF-kappaB. Our results suggest that BMPs not only stimulate osteoblast differentiation but can also promote cell survival during the induction of bone formation, offering new insight into the biological functions of BMPs.     

Bmp6 and Bmp7 are required for cushion formation and septation in the developing mouse heart.

The mature heart valves and septa are derived from the cardiac cushions which initially form as local outgrowths of mesenchymal cells within the outflow tract and atrioventricular regions. Endocardial cells respond to signals from the overlying myocardium and undergo an epithelial-to-mesenchymal transformation to invade the intervening extracellular matrix. The molecules that can induce and maintain these cell populations are not known, but many candidates, including several TGFbetas and BMPs, have been proposed based on their expression patterns and activities in other systems. In the present study, we describe the expression of Bmp6 and Bmp7 in overlapping and adjacent sites, including the cardiac cushions during mouse embryonic development. Previous analyses demonstrate that neither of these BMPs is required during cardiogenesis, but analysis of Bmp6;Bmp7 double mutants uncovers a marked delay in the formation of the outflow tract endocardial cushions. A proportion of Bmp6;Bmp7 mutants also display defects in valve morphogenesis and chamber septation, and the embryos die between 10.5 and 15.5 dpc due to cardiac insufficiency. These data provide the first genetic evidence that BMPs are involved in the formation of the cardiac cushions.     

Combinatorial gene therapy for bone regeneration: cooperative interactions between adenovirus vectors expressing bone morphogenetic proteins 2, 4, and 7

Bone morphogenetic proteins (BMPs) have demonstrated effectiveness as bone regeneration agents whether delivered as recombinant proteins or via gene therapy. Current gene therapy approaches use vectors expressing single BMPs. In contrast, multiple BMPs are coordinately expressed during bone development and fracture healing. Furthermore, BMPs likely exist in vivo as heterodimeric molecules having enhanced biological activity. In the present study, we test the hypothesis that gene therapy-based bone regeneration can be enhanced by expressing combinations of BMPs. For in vitro studies, mesenchymal cell lines were transduced with individual adenoviruses containing BMP2, 4, or 7 cDNA under control of a CMV promoter (AdBMP2, 4, 7) or virus combinations. Significantly, combined transduction with AdBMP2 plus AdBMP7 or AdBMP4 plus AdBMP7 resulted in a synergistic stimulation of osteoblast differentiation. This synergy is best explained by formation of BMP2/7 and 4/7 heterodimers. To test in vivo biological activity, fibroblasts were transduced with specific virus combinations and implanted into C57BL6 mice. Consistent with in vitro results, strong synergy was observed using combined AdBMP2/BMP7 treatment, which induced twofold to threefold more bone than would be predicted based on the activity of individual AdBMPs. These studies show that dramatic enhancement of osteogenesis can be achieved using gene therapy to express specific combinations of interacting regenerative molecules.     

Shaping BMP morphogen gradients through enzyme-substrate interactions

Bone morphogenetic proteins (BMPs) regulate dorsal/ventral (D/V) patterning across the animal kingdom; however, the biochemical properties of certain pathway components can vary according to species-specific developmental requirements. For example, Tolloid (Tld)-like metalloproteases cleave vertebrate BMP-binding proteins called Chordins constitutively, while the Drosophila Chordin ortholog, Short gastrulation (Sog), is only cleaved efficiently when bound to BMPs. We identified Sog characteristics responsible for making its cleavage dependent on BMP binding. "Chordin-like" variants that are processed independently of BMPs changed the steep BMP gradient found in Drosophila embryos to a shallower profile, analogous to that observed in some vertebrate embryos. This change ultimately affected cell fate allocation and tissue size and resulted in increased variability of patterning. Thus, the acquisition of BMP-dependent Sog processing during evolution appears to facilitate long-range ligand diffusion and formation of a robust morphogen gradient, enabling the bistable BMP signaling outputs required for early Drosophila patterning.     

Dynamics of BMP signaling in limb bud mesenchyme and polydactyly

Mutations in the Bone Morphogenetic Protein (BMP) pathway are associated with a range of defects in skeletal formation. Genetic analysis of BMP signaling requirements is complicated by the presence of three partially redundant BMPs that are required for multiple stages of limb development. We generated an inducible allele of a BMP inhibitor, Gremlin, which reduces BMP signaling. We show that BMPs act in a dose and time dependent manner in which early reduction of BMPs result in digit loss, while inhibiting overall BMP signaling between E10.5 and E11.5 allows polydactylous digit formation. During this period, inhibiting BMPs extends the duration of FGF signaling. Sox9 is initially expressed in normal digit ray domains but at reduced levels that correlate with the reduction in BMP signaling. The persistence of elevated FGF signaling likely promotes cell proliferation and survival, inhibiting the activation of Sox9 and secondarily, inhibiting the differentiation of Sox9-expressing chondrocytes. Our results provide new insights into the timing and clarify the mechanisms underlying BMP signaling during digit morphogenesis.     

Different Temporal Patterns in the Expressions of Bone Morphogenetic Proteins and Noggin During Astroglial Scar Formation After Ischemic Stroke

Bone morphogenetic proteins (BMPs) and their antagonists have roles in scar formation and regeneration after central nervous system injuries. However, temporal changes in their expression during astroglial scar formation in the ischemic brain are unknown. Here, we examined protein levels of BMP2, BMP7, and their antagonist noggin in the ischemic brain up to 4 weeks after experimental stroke in mice. BMP2 and BMP7 levels were increased from 1 to 4 weeks in the ischemic brain, and their expression was associated with astrogliosis. BMP7 expression was more intense and co-localized in reactive astrocytes in the ischemic subcortex at 1 week. Noggin expression began to increase after 2 weeks and was further increased at 4 weeks only in the ischemic subcortex, but the intensity was weak compared to the intensity of BMPs. Noggin was co-localized mainly in activated microglia. These findings show that expression of BMPs and noggin differed over time, in intensity and in types of cell, and suggest that BMPs and noggin have different roles in the processes of glial scar formation and neurorestoration in the ischemic brain.     

BMPs are mediators in tissue crosstalk of the regenerating musculoskeletal system

The musculoskeletal system is a tight network of many tissues. Coordinated interplay at a biochemical level between tissues is essential for development and repair. Traumatic injury usually affects several tissues and represents a large challenge in clinical settings. The current demand for potent growth factors in such applications thus accompanies the keen interest in molecular mechanisms and orchestration of tissue formation. Of special interest are multitasking growth factors that act as signals in a variety of cell types, both in a paracrine and in an autocrine manner, thereby inducing cell differentiation and coordinating not only tissue assembly at specific sites but also maturation and homeostasis. We concentrate here on bone morphogenetic proteins (BMPs), which are important crosstalk mediators known for their irreplaceable roles in vertebrate development. The molecular crosstalk during embryonic musculoskeletal tissue formation is recapitulated in adult repair. BMPs act at different levels from the initiation to maturation of newly formed tissue. Interestingly, this is influenced by the spatiotemporal expression of different BMPs, their receptors and co-factors at the site of repair. Thus, the regenerative potential of BMPs needs to be evaluated in the context of highly connected tissues such as muscle and bone and might indeed be different in more poorly connected tissues such as cartilage. This highlights the need for an understanding of BMP signaling across tissues in order to eventually improve BMP regenerative potential in clinical applications. In this review, the distinct members of the BMP family and their individual contribution to musculoskeletal tissue repair are summarized by focusing on their paracrine and autocrine functions.