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P Bovolenta

Publications and source records attributed to P Bovolenta.

At least 19 recordsLinked to original sources
PubMed2005

Comparative analysis of gnathostome Otx gene expression patterns in the developing eye: implications for the functional evolution of the multigene family.

We have performed a detailed analysis of the expression pattern of the three gnathostome Otx classes in order to gain new insights into their functional evolution. Expression patterns were examined in the developing eye of a chondrichthyan, the dogfish, and an amniote, the chick, and compared with the capacity of paralogous proteins to induce a pigmented phenotype in cultured retina cells in cooperation with the bHLH-leucine zipper protein Mitf. This analysis indicates that each Otx class is characterized by highly specific and conserved expression features in the presumptive RPE, where Otx1 and Otx2, but not Otx5, are transcribed at optic vesicle stages, in the differentiating neural retina, where Otx2 and Otx5 show a conserved dynamic expression pattern, and in the forming ciliary process, a major site of Otx1 expression. Furthermore, the paralogous proteins of the dogfish and the mouse do not display any significant difference in their capacity to induce a pigmented phenotype, suggesting a functional equivalency in the specification and differentiation of the RPE. These data indicate that specific functions selectively involving each Otx orthology class were fixed prior to the gnathostome radiation and highlight the prominent role of regulatory changes in the functional diversification of the multigene family.

Animals↗
PubMed2001

Bmp4 mediates apoptotic cell death in the developing chick eye.

The bone morphogenetic protein (BMP) expression in vertebrates suggests a reiterative function of these molecules during eye development. However, genetic analysis in mice has provided only partial information. Using the chick embryo as a model system, we have analyzed possible additional functions of BMP4 during optic cup formation. Here we describe the expression pattern of Bmp4 and Bmp7 and we show that, in contrast to the mouse, the prospective lens placode ectoderm expresses high levels of Bmp4 but no Bmp7. After optic vesicle invagination, Bmp4 is expressed in the prospective dorsal neural retina, where BmprIA, BmprII, and Smad1, components of the BMP4 signal transduction pathway, are also expressed. In toto terminal deoxynucleotidyl transferase-mediated biotinylated UTP nick end-labeling analysis shows that the dorsal optic cup is the site of a spatiotemporally restricted apoptosis, which parallels the expression not only of Bmp4 but also of Msx1 and Msx2, genes implicated in BMP4-mediated apoptosis. The use of optic vesicle cultures as well as in ovo local addition of BMP4 and its antagonist Noggin proves that the local activity of BMP4 is responsible for programmed cell death in the dorsal optic cup. In addition, we show that Noggin is able to reduce the rate of cell proliferation in the dorsal part of the optic cup whereas BMP4 increases the number of BrdU-positive cells in retina cultures. These results provide evidence that BMP4 contributes to eye development by promoting cell proliferation and programmed cell death.

Animals↗
PubMed2001

Otx genes are required for tissue specification in the developing eye.

Patterning of the vertebrate eye appears to be controlled by the mutual regulation and the progressive restriction of the expression domains of a number of genes initially co-expressed within the eye anlage. Previous data suggest that both Otx1 and Otx2 might contribute to the establishment of the different eye territories. Here, we have analysed the ocular phenotype of mice carrying different functional copies of Otx1 and Otx2 and we show that these genes are required in a dose-dependent manner for the normal development of the eye. Thus, all Otx1(-/-); Otx2(+/-) and 30% of Otx1(+/-); Otx2(+/-) genotypes presented consistent and profound ocular malformation, including lens, pigment epithelium, neural retina and optic stalk defects. During embryonic development, optic vesicle infolding was severely altered and the expression of pigment epithelium-specific genes, such as Mitf or tyrosinase, was lost. Lack of pigment epithelium specification was associated with an expansion of the prospective neural retina and optic stalk territories, as determined by the expression of Pax6, Six3 and Pax2. Later in development the presumptive pigment epithelium region acquired features of mature neural retina, including the generation of Islet1-positive neurones. Furthermore, in Otx1(-/-); Otx2(+/-) mice neural retina cell proliferation, cell differentiation and apoptotic cell death were also severely affected. Based on these findings we propose a model in which Otx gene products are required for the determination and differentiation of the pigment epithelium, co-operating with other eye patterning genes in the determination of the specialised tissues that will constitute the mature vertebrate eye.

Animals↗
PubMed2001

Control of retinal ganglion cell axon growth: a new role for Sonic hedgehog.

Retinal ganglion cell (RGC) axons grow towards the diencephalic ventral midline during embryogenesis guided by cues whose nature is largely unknown. We provide in vitro and in vivo evidence for a novel role of Sonic hedgehog (SHH) as a negative regulator of growth cone movement. SHH suppresses both the number and the length of neurites emerging from the chick retina but not from neural tube or dorsal root ganglia explants, without interfering with their rate of proliferation and differentiation. Similarly, retroviral-mediated ectopic expression of Shh along the chick visual pathway greatly interferes the growth of RGC axons. Upon SHH addition to grown neurites, the intracellular level of cAMP decreases, suggesting that the dampening of growth cone extension mediated by SHH may involve interaction with its receptor Patched which is expressed by RGC. Based on these findings, we propose that Shh expression at the chiasm border defines a constrained pathway within the ventral midline which serves to guide the progression of RGC axons.

Animals↗
PubMed2000

Nervous system proteoglycans as modulators of neurite outgrowth.

The proteoglycans are multifunctional macromolecules composed of a core polypeptide and a variable number of glycosaminoglycan chains. The structural diversity and complexities of proteoglycan expression in the developing and adult Nervous System underlies the variety of biological functions that these molecules fulfill. Thus, in the Nervous System, proteoglycans regulate the structural organisation of the extracellular matrix, modulate growth factor activities and cellular adhesive and motility events, such as cell migration and axon outgrowth. This review summarises the evidences indicating that proteoglycans have an important role as modulators of neurite outgrowth and neuronal polarity. Special emphasis will be placed on those studies that have shown that proteoglycans of certain subtypes inhibit neurite extension either during the development and/or the regeneration of the vertebrate Central Nervous System.

Animals↗
PubMed2000

Early and dynamic expression of cSfrp1 during chick embryo development.

Secreted frizzled related proteins (SFRPs) are a new class of signalling molecules that appear to antagonise the activity of the Wnt proteins. Here we report the dynamic expression pattern of cSfrp1, a new member of this family, at early stages of chick embryo development. cSfrp1 transcripts are first detected at pre-streak stages throughout the chick blastula but, during early primitive streak formation, expression is restricted to the anterior primitive streak and later to the blastoderm anterior to the Hensen' s node. This pattern of expression overlaps with that of Otx2 and is complementary to that of cWnt8c. During neural plate formation cSfrp1 mRNAs are abundantly localized only to the anterior domain of the embryo but, as neural tube closes, the expression extends caudally. Later, the main sites of expression in the neural tissue are the telencephalic vesicles, the epiphysis, the developing eyes and the ventral hindbrain and neural tube. Additionally, cSfrp1 transcripts were found in the axial and lateral mesoderm, the otic placode, the trigeminal ganglia, the mesoderm of the branchial arches, the developing limb buds, as well as in the mesodermal component of the developing kidney.

Animals↗
PubMed1999

Genomic cloning and characterization of the human homeobox gene SIX6 reveals a cluster of SIX genes in chromosome 14 and associates SIX6 hemizygosity with bilateral anophthalmia and pituitary anomalies.

The Drosophila gene sine oculis (so), a nuclear homeoprotein that is required for eye development, has several homologues in vertebrates (the SIX gene family). Among them, SIX3 is considered to be the functional orthologue of so because it is strongly expressed in the developing eye. However, embryonic SIX3 expression is not limited to the eye field, and SIX3 has been found to be mutated in some patients with holoprosencephaly type 2 (HPE2), suggesting that SIX3 has wide implications in head development. We report here the cloning and characterization of SIX6, a novel human SIX gene that is the homologue of the chick Six6(Optx2) gene. SIX6 is closely related to SIX3 and is expressed in the developing and adult human retina. Data from chick and mouse suggest that the human SIX6 gene is also expressed in the hypothalamic and the pituitary regions. SIX6 spans 2567 bp of genomic DNA and is split in two exons that are transcribed into a 1393-nucleotide-long mRNA. Chromosomal mapping of SIX6 revealed that it is closely linked to SIX1 and SIX4 in human chromosome 14q22.3-q23, which provides clues about the origin and evolution of the vertebrate SIX family. Recently three independent reports have associated interstitial deletions at 14q22.3-q23 with bilateral anophthalmia and pituitary anomalies. Genomic analyses of one of these cases demonstrated SIX6 hemizygosity, strongly suggesting that SIX6 haploinsufficiency is responsible for these developmental disorders.

Amino Acid Sequence↗
PubMed1999

Genomic cloning, structure, expression pattern, and chromosomal location of the human SIX3 gene.

The Drosophila gene sine oculis (so) is a nuclear homeoprotein that is required for eye development. Homologous genes to so, denoted SIX genes, have been found in vertebrates. Among the SIX genes, SIX3 is considered to be the functional homologue of so. To provide insight into the potential implications of SIX3 in human ocular malformations, we have cloned and characterized the human SIX3 gene. In human eye, SIX3 produces a 3-kb transcript that codes for a 332-amino-acid polypeptide that is virtually identical to its mouse and chick homologues. Expression of SIX3 was detected in human embryos as early as 5-7 weeks of gestation and found to be maintained in the eye throughout the entire period of fetal development. At 20 weeks of gestation, expression of SIX3 in the human retina was detected in the ganglion cells and in cells of the inner nuclear layer. The human SIX3 gene spans 4.4 kb of genomic DNA and is split in two exons separated by a 1659-bp intron. SIX3 was mapped to human chromosome 2p16-p21, between the genetic markers D2S119 and D2S288. Interestingly, the map position of human SIX3 overlaps the locations of two dominant disorders with ocular phenotypes that have been assigned to this chromosomal region, holoprosencephaly type 2 and Malattia Leventinese.

Amino Acid Sequence↗
PubMed1999

Expression of the beta-chain of the complement regulator C4b-binding protein in human ovary.

Human C4b-binding protein (C4BP) is an important regulator of the complement system that also binds and inactivates the anticoagulant vitamin K-dependent protein S. These two activities are performed by two distinct polypeptides of 70 kDa and 45 kDa known as alpha and beta chains, respectively. C4BP is present in plasma in various isoforms with different alpha/beta composition. We report here that C4BPbeta, but not C4BPalpha, is expressed in adult human ovary. Expression of C4BPbeta was detected in all ovarian biopsies analyzed (n = 15), independently of age and phase of the menstrual cycle. In situ hybridization and immunostaining analyses on cryostat sections demonstrated expression of C4BPbeta in both regressing corpus luteum and corpus albicans, but not in the follicles, the corpus luteum, the ovary stroma or the vascular cells. In addition, we noted that the expression pattern of the C4BPbeta mRNA resembles that described for the connective tissue that invades the degenerating corpus luteum and causes a progressive fibrosis that gradually converts it into a scar, the corpus albicans. RT-PCR and immunostaining analyses of primary cultures derived from human ovaries demonstrated the presence of fibroblast-like cells that express C4BPbeta. As a whole, these data suggest a role for the C4BPbeta in human ovary during the healing and scar resorption processes that leads to the formation of the corpus albicans and its replacement by ovarian stroma.

Adult↗
PubMed1999

Six9 (Optx2), a new member of the six gene family of transcription factors, is expressed at early stages of vertebrate ocular and pituitary development.

The Drosophila gene sine oculis (so) is a nuclear homeoprotein, which is required for eye development. Several homologues of so have been found in vertebrates. We report here a detailed expression analysis in chick and mouse of Six9 (Optx2), a novel gene of the Six/sine oculis family closely related to Six3. Six9 (Optx2) is first expressed at presomitic stages in the head-fold, both in the neural plate and in the underlying axial mesoderm. Thereafter, Six9 (Optx2) is strongly expressed in the presumptive and differentiating neural retina and ventral optic stalk, in the olfactory placodes, in the hypothalamus and in the pituitary gland. This expression pattern largely overlaps with that of Six3, but several differences exist between the expression domain of the two genes. At presomitic stages, the posterior boundary of Six3 expression is at the same axial level both in the prechordal plate and in the overlying neural plate. In contrast, Six9 (Optx2) expression in the prechordal plate extends more caudal to that of the neural plate, occupying a more restricted V-shaped territory. Similarly, during the early events of eye patterning, Six3 is first expressed in the entire optic vesicle and lens placode. Only later does its expression become confined to the prospective and differentiating neural retina. Conversely, Six9 (Optx2) is never observed in the lens placode of either chick and mouse, and from early stages of optic vesicle development, Six9 (Optx2) transcripts are restrained to the prospective ventral neural retina and optic stalks.

Animals↗
PubMed1999

cSix4, a member of the six gene family of transcription factors, is expressed during placode and somite development.

We describe the expression pattern of cSix4, a chick homologue of the murine Six4/AREC3 gene. cSix4 transcripts are detected at gastrula stages in the blastoderm surrounding the developing axial midline. As the neural plate begins to form cSix4 mRNA is detected in a crescent-shaped band, which surrounds the anterior developing neural plate and corresponds to the presumptive placode region. This expression is maintained in all the placodes (olfactory, optic, neural and otic) as they develop but with different characteristics. Further, abundant expression of cSix4 was localised to the paraxial mesoderm and the entire developing somites, becoming restricted first to their dorsal portion, then to the dermomyotome and finally to the myotome. cSix4 expression is maintained in the developing and adult muscular tissue. Additional sites of cSix4 expression are the presumptive and developing limb buds, the notochord, trigeminal ganglia, cells of the spinal cord, particularly the motor neurones, the dorsal root ganglia, the neural retina, as well as the epithelial component of the developing kidney.

Animals↗
PubMed1998

A neurite outgrowth-inhibitory proteoglycan expressed during development is similar to that isolated from adult brain after isomorphic injury.

The expression of proteoglycans (PGs) in the mammalian central nervous system (CNS) appears to be strictly regulated both during development and after damage to the mammalian CNS. Recently, we have isolated from membranes of injured adult brain a neurite outgrowth-inhibitory proteoglycan (IMP), the activity of which could be specifically counteracted by a monoclonal antibody (mAB) against the PG. We described in this report the characterization of perinatal membrane proteoglycan (PMP), a heparan-sulfate/chondroitin-sulfate-containing PG expressed during brain development. Its maximal expression was observed around postnatal day 3, decreasing strongly in normal adult tissue. This PG was purified and characterized using mABs generated against IMP. The comparison of PMP and IMP properties indicates that the two PGs are highly related and share expression patterns, biochemical characteristics, and the ability to inhibit neurite initiation in culture. However, IMP and PMP displayed a distinct effect on neurite elongation, which may be explained by their differences in glycosilation pattern. The data presented in this report support the idea that proteoglycans expressed during CNS development are re-expressed following injury.

Animals↗
PubMed1998

Expression pattern of cSix3, a member of the Six/sine oculis family of transcription factors.

We describe the expression pattern of cSix3, a chick homologue of the murine Six3. cSix3 transcripts are expressed from presomitic stages in the most anterior portion of the neural plate. As the neural tube folds and the optic vesicles evaginate, cSix3 is expressed in the optic vesicle and the rostroventral forebrain. At later stages, cSix3 is found in most of the structures derived from the anterior neural plate, i.e. olfactory epithelium, septum, adenohypophysis, hypothalamus and preoptic areas. During eye development, cSix3 expression is first found in the entire optic vesicle and the overlying ectoderm but soon becomes restricted to the prospective neural retina and to the lens placode. In the developing neural retina, cSix3 is expressed in the entire undifferentiated neuroepithelium but is rapidly downregulated, first in the postmitotic photoreceptors and later in the majority of retinal ganglion cells.

Animals↗
PubMed1997

Implication of OTX2 in pigment epithelium determination and neural retina differentiation.

The expression pattern of Otx2, a homeobox-containing gene, was analyzed from the beginning of eye morphogenesis until neural retina differentiation in chick embryos. Early on, Otx2 expression was diffuse throughout the optic vesicles but became restricted to their dorsal part when the vesicles contacted the surface ectoderm. As the optic cup forms, Otx2 was expressed only in the outer layer, which gives rise to the pigment epithelium. This early Otx2 expression pattern was complementary to that of PAX2, which localizes to the ventral half of the developing eye and optic stalk. Otx2 expression was always observed in the pigment epithelium at all stages analyzed but was extended to scattered cells located in the central portion of the neural retina around stage 22. The number of cells expressing Otx2 transcripts increased with time, following a central to peripheral gradient. Bromodeoxyuridine labeling in combination with immunohistochemistry with anti-OTX2 antiserum and different cell-specific markers were used to determine that OTX2-positive cells are postmitotic neuroblasts undergoing differentiation into several, if not all, of the distinct cell types present in the chick retina. These data indicate that Otx2 might have a double role in eye development. First, it might be necessary for the early specification and subsequent functioning of the pigment epithelium. Later, OTX2 expression might be involved in retina neurogenesis, defining a differentiation feature common to the distinct retinal cell classes.

Animals↗
PubMed1997

Neurite outgrowth inhibitor of gliotic brain tissue. Mode of action and cellular localization, studied with specific monoclonal antibodies.

Membranes from injured adult rat brain express a heparan/chondroitin sulphate proteoglycan that inhibits neurite outgrowth in vitro. We have developed monoclonal antibodies (Mabs) against this proteoglycan, two of which were characterized and used for the study of the inhibitor mode of action and localization in normal and injured adult brain. The antibodies recognized a molecule of apparent molecular weight 200 kDa in Western blots of injured brain membranes. One of the Mabs blocked both the inhibition of neurite outgrowth and the growth cone collapse activity, associated with the proteoglycan. In adult brain, inhibitor immunoreactivity was found predominantly in neurons but, after a lesion, it was associated mainly with reactive glial cells. The localization of neurite outgrowth inhibitors in reactive glia supports the idea that gliotic tissue is largely responsible for the failure of axonal regeneration in mammalian CNS.

Animals↗
PubMed1997

Control of early cell death by BDNF in the chick retina.

The developing chick retina undergoes at least two discrete periods of programmed cell death. The earlier period coincides with the main onset of neuron birth and migration (embryonic day 5-7), whereas the latter one corresponds to the well-documented process of retinal ganglion cell death following tectal innervation (embryonic day 10-14; Rager, G. H. (1980) Adv. Anat. Embryol. Cell Biol. 63, 1-92). In the early period, apoptosis is induced by nerve growth factor (NGF) acting via its p75 receptor (Frade, J. M., Rodríguez-Tébar, A. and Barde, Y.-A. (1996) Nature 383, 166-168). Here, we show that the application of brain-derived neurotrophic factor (BDNF) to chick embryos in ovo prevented retinal cell death in the early period, whereas exogenously applied NGF and neurotrophin-3 had no such effect. The addition of BDNF to embryos resulted in about 70% increase in the number of retinal ganglion cells in both E6 and E9 retinas relative to controls. BDNF is first expressed in both the pigment epithelium and neural retina of embryonic day 4 embryos, and at the same stage of development, its TrkB receptor is expressed in the neural retina. Our data indicate that early cell death is an important process in the neurogenesis of retinal ganglion cells and is regulated by locally produced BDNF.

Animals↗
PubMed1997

Vitronectin is expressed in the ventral region of the neural tube and promotes the differentiation of motor neurons.

The extracellular matrix protein vitronectin and its mRNA are present in the embryonic chick notochord, floor plate and in the ventral neural tube at the time position of motor neuron generation. When added to cultures of neural tube explants of developmental stage 9, vitronectin promotes the generation of motor neurons in the absence of either notochord or exogenously added Sonic hedgehog. Conversely, the neutralisation of endogenous vitronectin with antibodies inhibits over 90% motor neuron differentiation in co-cultured neural tube/notochord explants, neural tube explants cultured in the presence of Sonic hedgehog, and in committed (stage 13) neural tube explants. Furthermore, treatment of embryos with anti-vitronectin antibodies results in a substantial and specific reduction in the number of motor neurons generated in vivo. These results demonstrate that vitronectin stimulates the differentiation of motor neurons in vitro and in vivo. Since the treatment of stage 9 neural tube explants with Sonic hedgehog resulted in induction of vitronectin mRNA expression before the expression of floor plate markers, we conclude that vitronectin may act either as a downstream effector in the signalling cascade induced by Sonic hedgehog, or as a synergistic factor that increases Shh-induced motor neuron differentiation.

Amino Acid Sequence↗