PubMed Health⌕ Search

Biomedical subjects

B Dastugue

Publications and source records attributed to B Dastugue.

At least 37 records · Page 2Linked to original sources
PubMed2000

Esterification of vitamin A by the human placenta involves villous mesenchymal fibroblasts.

Vitamin A (retinol) and its active derivatives (retinoic acids) are essential for growth and development of the mammalian fetus. Maternally derived retinol must pass the placenta to reach the developing fetus. Despite its apparent importance, little is known concerning placental transfer and metabolism of retinol, and particularly of placental production and storage of retinyl esters. To elucidate this metabolic pathway, we incubated, in the presence of retinol, 1) human full-term placental explants and 2) primary cultures of major cells types contributing to placental function: trophoblasts and villous mesenchymal fibroblasts. We used HPLC to determine the types and concentrations of retinyl esters produced by these explants and cells. About 14% of total cellular retinol in placental explants was esterified. The most abundant esters were myristate and palmitate. Primary cell cultures showed that fibroblasts efficiently produced retinyl esters, but trophoblasts did not. In both types of experiments, no retinyl esters were detected in the culture medium, suggesting that retinyl esters were produced for storage purpose. These results suggest that villous mesenchymal fibroblasts are primary sites of retinol esterification and storage in the placenta.

Adult↗
PubMed1999

Neuroblastoma B104 cell line as a model for analysis of neurite outgrowth and neuronal aggregation induced by Reissner's fiber material.

The subcommissural organ (SCO) is a specialized ependymal structure of the brain that secretes glycoproteins into the cerebrospinal fluid (CSF), which condense to form a thread-like structure - Reissner's fiber (RF). The effects of soluble material released by RF were examined on neuroblastoma B104 cells grown in serum-free medium, using "low-density" and "high-density" culture systems. In the presence of soluble RF material, low-density cultures were suitable for analysis of the enhanced neurite outgrowth of B104 cells, while high-density cultures allowed the increased B104 cell aggregation to be examined. RF-induced neuronal aggregation and neuritic outgrowth were restricted to a perimeter around the RF. This standardized cell culture system reproduced in part the effects observed previously with primary cortical and spinal cord cell cultures and may serve the analysis of the mechanisms leading to aggregation and neurite outgrowth. In the present study, we analyzed variations in the rate of neural cell adhesion molecules, such as N-CAM and N-cadherin, induced by soluble RF material in high-density cultures.

Animals↗
PubMed1999

SCO-spondin is evolutionarily conserved in the central nervous system of the chordate phylum.

Bovine SCO-spondin was shown to be a brain-secreted glycoprotein specifically expressed in the subcommissural organ, an ependymal differentiation located in the roof of the Sylvian aqueduct. Also, SCO-spondin makes part of Reissner's fiber, a phylogenetically and ontogenetically conserved structure present in the central canal of the spinal cord of chordates. This secretion is a large multidomain protein probably involved in axonal growth and/or guidance. As Reissner's fiber is highly conserved in the chordate central nervous system, we sought genes orthologous to the bovine SCO-spondin gene by Southern blot analysis in several members of the chordate phylum: urochordates, cephalochordates, cyclostomes, and lower and higher vertebrates, including humans. In addition, conserved glycoproteins present in the subcommissural organ and Reissner's fiber were revealed by immunohistochemistry using antibodies raised against bovine Reissner's fiber. Variation in the sites of Reissner's fiber production according to chordate subphylum, presence of this structure in the spinal cord, and conservation of the SCO-spondin gene are discussed in the context of chordate central nervous system development. These results indicate that SCO-spondin is an ancient ependymal secretion, making part of Reissner's fiber, that may have had an important function during the evolution of the central nervous system in chordates, including that of the spinal cord.

Animals↗
PubMed1999

Proteolipoprotein gene analysis in 82 patients with sporadic Pelizaeus-Merzbacher Disease: duplications, the major cause of the disease, originate more frequently in male germ cells, but point mutations do not. The Clinical European Network on Brain Dysmyelinating Disease.

Pelizaeus-Merzbacher Disease (PMD) is an X-linked developmental defect of myelination affecting the central nervous system and segregating with the proteolipoprotein (PLP) locus. Investigating 82 strictly selected sporadic cases of PMD, we found PLP mutations in 77%; complete PLP-gene duplications were the most frequent abnormality (62%), whereas point mutations in coding or splice-site regions of the gene were involved less frequently (38%). We analyzed the maternal status of 56 cases to determine the origin of both types of PLP mutation, since this is relevant to genetic counseling. In the 22 point mutations, 68% of mothers were heterozygous for the mutation, a value identical to the two-thirds of carrier mothers that would be expected if there were an equal mutation rate in male and female germ cells. In sharp contrast, among the 34 duplicated cases, 91% of mothers were carriers, a value significantly (chi2=9. 20, P<.01) in favor of a male bias, with an estimation of the male/female mutation frequency (k) of 9.3. Moreover, we observed the occurrence of de novo mutations between parental and grandparental generations in 17 three-generation families, which allowed a direct estimation of the k value (k=11). Again, a significant male mutation imbalance was observed only for the duplications. The mechanism responsible for this strong male bias in the duplications may involve an unequal sister chromatid exchange, since two deletion events, responsible for mild clinical manifestations, have been reported in PLP-related diseases.

Cystic Fibrosis Transmembrane Conductance Regulato↗
PubMed1999

Screening for cystic fibrosis transmembrane conductance regulator gene mutations in men included in an intracytoplasmic sperm injection programme.

The present study was undertaken to evaluate the frequency and nature of mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene in infertile patients undergoing intracytoplasmic sperm injection. A total of 90 patients were screened for a panel of 10 mutations in the CFTR gene frequently involved in congenital absence of the vas deferens (CAVD); the patients included 14 with azoospermia and CAVD, 39 patients with azoospermia without CAVD (n = 39) and 37 patients with severe oligozoospermia. The length of the polymorphic polypyrimidine tract (allele 5T, 7T and 9T) in the intron 8/exon 9 splice-acceptor site was also determined. In 10 out of 14 patients with CAVD, CFTR mutations were found; nine patients had one DeltaISOdiaDeltaF508 mutation and one patient had two CFTR mutations (N1303K/R117H). Allele 5T was present in eight of these patients. In six patients, 5T was the non-DeltaISOdiaDeltaF508 allele and in two patients there was no known CFTR mutation. None of the CFTR mutations were observed in patients with azoospermia without CAVD or with severe oligozoospermia and the frequency of allele 5T was 3.6% (three out of 78 alleles) and 1.35% (one out of 74 alleles) respectively. Our observation suggests that the CFTR gene is not involved in either spermatogenesis or in the pathology of the genital tract, except for CAVD.

Adult↗
PubMed1999

Mobilization of two retroelements, ZAM and Idefix, in a novel unstable line of Drosophila melanogaster.

We describe a novel transposition system in a line of Drosophila melanogaster called RevI in which two retroelements are mobilized. These elements are the retroelement ZAM, recently described in the literature, and a novel element designated Idefix. Like ZAM, Idefix displays the structural features of a vertebrate retrovirus. Its three open reading frames encode predicted products resembling the products of the gag, pol, and env genes of retroviruses. In situ hybridization and Southern analyses performed on the RevI genome revealed the presence of some 20 copies of ZAM and Idefix, whereas ZAM is absent and Idefix is present in only four copies on the chromosomal arms of the original parental line. From RevI, a series of mutations affecting eye coloration has been recovered. The genetic and molecular analyses of these mutations have shown that most of them affected the white locus through three rounds of mutational events. The first mutational event was previously shown to be caused by a ZAM insertion 3 kb upstream of the transcription start site of white. It confers a red-brick phenotype to the orange eye coloration of the parental line. The second event results from the insertion of an Idefix copy 1.7 kb upstream of the transcription start site of the white gene, which modifies the red-brick phenotype to orange. This second mutational event was recovered as a recurrent specific mutation in 11 independent individuals. The third event results from an additional Idefix located 1.7 kb upstream of white that is responsible for the full reversion of the orange phenotype to red-brick. The fact that such mutations due to recurrent appearances of both ZAM and Idefix at the white locus result in such a variety of phenotypes brings to light a new molecular system in which the interference of mobile elements with the correct expression of the host gene can be tested.

Alleles↗
PubMed1999

The Krüppel-like core promoter binding protein gene is primarily expressed in placenta during mouse development.

The human core promoter binding protein (hCPBP) has been identified as a DNA-binding protein involved in the regulation of TATA box-less genes like those encoding the pregnancy-specific glycoproteins. Structurally, hCPBP contains three zinc fingers in the C-terminal domain, which is highly conserved in a number of proteins that constitute the Krüppel-like family of transcription factors. In the present work, we report the molecular cloning of the mouse CPBP (mCPBP) and its expression pattern during development as well as in adult tissues. The mouse cDNA encodes a protein of 283 amino acids that share 94.4% of identity with the hCPBP. The highest level of mCPBP transcript was detected in placenta, and its expression was lower in total embryos and in adult tissues. We also show by in situ hybridization that during embryonic development the mCPBP gene is mainly expressed in extra-embryonic structures throughout gestation; essentially no specific expression was detected in embryonic tissues. Our data demonstrate that CPBP transcript is enriched in the trophoblastic tissue and strongly suggest that its encoded polypeptide regulates target genes involved in placental development and pregnancy maintenance.

Amino Acid Sequence↗
PubMed1999

The integration machinery of ZAM, a retroelement from Drosophila melanogaster, acts as a sequence-specific endonuclease.

Retroviruses and retrotransposons insert into the host genome with no obvious sequence specificity. We examined the target sites of the retroelement ZAM by sequencing each host-ZAM junction in the genome of Drosophila melanogaster. Our overall data provide compelling evidence that ZAM integration machinery recognizes and leads to ZAM insertion into the sequence 5'-GCGCGCg-3'. This unique property of ZAM will facilitate the development of new tools to study the integration process of retroelements.

Animals↗
PubMed1999

Plasticity within the lateral somatic mesoderm of Drosophila embryos.

Each of 30 Drosophila larval somatic muscles has its individual shape, insertion sites and innervation. From the very beginning, the formation of individual muscles is controlled by a set of muscle identity genes. The four lateral transverse muscles (LT1-LT4) are thought to be specified by the combinatorial activity of Krüppel (Kr), apterous (ap) and muscle specific homeobox (msh) genes whilst the activity of the ladybird (lb) genes is required for proper formation of the neighbouring segmental border muscle (SBM). We have recently shown that ectopic expression of lb changes the identity of Kr-expressing lateral muscle precursors and recruits them to form enlarged or duplicated SBMs. Here we report that loss of msh function leads to a similar transformation resulting in the overproduction of SBMs. Inversely, in msh gain of function embryos, the prospective SBM myoblasts change their identity resulting in the formation of enlarged lateral transverse muscles. These data indicate a key role for the msh and lb genes in the specification and diversification of myoblast lineages from the lateral domain, and reveal a plasticity of cell fate within the somatic mesoderm of Drosophila.

Animals↗
PubMed1998

Complex expression pattern of the SCO-spondin gene in the bovine subcommissural organ: toward an explanation for Reissner's fiber complexity?

Bovine SCO-spondin is a glycoprotein secreted by the subcommissural organ (SCO), an ependymal derivative located in the roof of the third ventricle. It shows homology with developmental molecules involved in directional axonal growth. Using SCO-spondin cDNAs as probes, we analysed the specific expression of the corresponding gene in the bovine SCO by Northern blot and in situ hybridization (ISH). A strong expression was detected in the secretory ependymal and hypendymal cells of the SCO and the main transcripts showed a large size 14 kb. A single copy gene was revealed by Southern blot analysis of bovine genomic DNA. The presence of additional transcripts suggested a transcriptional regulation of the SCO-spondin gene. A comparative analysis of the results obtained by molecular and immunological techniques (immunoblotting and immunopurification) pointed to the presence of several SCO-spondin related proteins in the SCO encoded by the same gene. The presence in the cerebral hemispheres (CH) of a 54-kDa glycoprotein with a common epitope is discussed as a putative cleaved SCO-spondin product carried by the cerebrospinal fluid, that may act on neuronal development.

Animals↗
PubMed1998

Effect of synthetic peptides derived from SCO-spondin conserved domains on chick cortical and spinal-cord neurons in cell cultures.

SCO-spondin is a newly identified protein, strongly expressed in the subcommissural organ (SCO), an ependymal differentiation of the brain. When secreted into the cerebrospinal fluid at the entrance to the Sylvian aqueduct, it condenses and forms Reissner's fiber. Several conserved domains have previously been characterizedin SCO-spondin, e.g., thrombospondin type 1 repeats (TSRs), low-density lipoprotein receptor (LDLr) type A repeats, and epidermal-growth-factor-like domains, which are potent sites of protein-protein interaction. To clarify the role of this protein on neuronal development, we have tested the effect of oligopeptides, the sequences of which include highly conserved amino acids of TSRs, LDLr type A repeats and a potent site of attachment to proteoglycan, on cortical and spinal-cord neurons in primary cell cultures. One of these peptides (WSGWSSCSRSCG), corresponding to a SCO-spondin TSR sequence, markedly increases adhesivity and neuritic outgrowth of cortical neurons and induces an opposite effect on cortical and spinal-cord neuronal aggregation. These effects are specific, as no response is observed with the scrambled sequence of this peptide. Another peptide (WGPCSVSCG) is only slightly active on adhesivity and neuritic outgrowth of cortical neurons and has no effect on spinal-cord neurons. Peptides derived from other conserved domains of SCO-spondin are not effective under our experimental conditions. Thus, SCO-spondin may be responsible for at least a part of the effects previously observed on neuronal cells cultured in the presence of Reissner's fiber. In addition, SCO-spondin seems to interfere with neuronal development and/or axonal guidance during ontogenesis of the central nervous system in modulating side-to-side interactions and neuritic outgrowth.

Amino Acid Sequence↗
PubMed1997

Invertebrate retroviruses: ZAM a new candidate in D.melanogaster.

ZAM, a new retroelement of Drosophila melanogaster, was identified as a mutational insertion at the white locus. It displays all the structural features of a vertebrate retrovirus. Its three open reading frames encode predicted products resembling the products of the gag, pol and env genes of retroviruses. Its transcription gives rise to an 8.6 kb full-length RNA and a 1.7 kb spliced message for the env gene. The latter encodes an envelope protein that is typical of elements having an extracellular phase of the life cycle. The identification of a ZAM envelope retrogene provides evidence that ZAM is mobilized through a reverse trancriptional process in the germ line of flies. We report that ZAM is distributed differently among D.melanogaster strains. Two stocks out of >15 tested display a ZAM high copy number, with numerous copies distributed on chromosomal arms. This high copy number is associated with a high transcriptional rate of ZAM. The existence of these two categories of strains offers a new genetic system in which the properties of a potential invertebrate retrovirus can be tested.

Amino Acid Sequence↗
PubMed1997

In vitro differentiation of chick spinal cord neurons in the presence of Reissner's fibre, an ependymal brain secretion.

The subcommissural organ (SCO), which belongs to the circumventricular organs, is a specialized ependymal structure of the brain that secretes glycoproteins into the cerebrospinal fluid (CSF) which condense to form a thread-like structure, the Reissner's fibre (RF). Regarding the presence of this ependymal brain secretion all along the central canal of the developing spinal cord, we analysed a putative developmental activity of RF on neuronal spinal cord cells. The effects of RF proper and soluble RF-material were examined in primary cultures of dissociated spinal cord cells from day 6 chicken embryos. In serum-containing mixed glial/neuronal cell cultures, both RF and soluble RF-material promoted neuronal survival. This effect was blocked by addition of specific antibodies raised against bovine RF into the culture medium. In serum-free neuron-enriched cultures, no neuronal survival activity was observed; however, under these conditions RF proper induced neuronal aggregation and neuritic outgrowth of spinal cord cells. Interestingly, neurites extending from the aggregates appeared mainly unfasciculated. Our results suggest a direct modulation of cell-cell interactions by SCO/RF glycoproteins and an indirect survival effect on neurons. These data strengthen the hypothesis of the involvement of SCO/RF complex in the development of the central nervous system (CNS) and are discussed regarding molecular features of SCO-spondin, a novel glycoprotein recently identified in this complex.

Animals↗
PubMed1997

Reissner's fibre promotes neuronal aggregation and influences neuritic outgrowth in vitro.

Reissner's fibre is the condensed form of glycoproteins secreted by the subcommissural organ; it extends through the central canal to the caudal end of the spinal cord. The effect of Reissner's fibre was assessed on dissociated embryonic chick cortical neuronal cells grown in chemically defined medium, by using two cell culture systems: (1) low-density cultures, in which neuronal cells remained evenly distributed; (2) high-density cultures, in which neuronal cells aggregated, displaying prominent neuritic outgrowth. Reissner's fibre, when added to low-density cultures, induced neuronal aggregation and neuritic outgrowth but this effect was restricted to an area centred around Reissner's fibre. Reissner's fibre, when added to high-density cultures, potentiated cell aggregation. Antibodies inhibiting the neural cell adhesion molecule or N-cadherin, and soluble Reissner's fibre material (reported previously to have anti-aggregative activity), did not prevent the aggregative activity induced by Reissner's fibre. Neuronal cells showed a similar reaction pattern when heparin or Reissner's fibre was added to the culture. These results suggest that the subcommissural organ/Reissner's fibre complex has multifunctional activities and may modulate cell-cell interactions during the development of the central nervous system.

Animals↗
PubMed1997

An in vitro cell culture system for the aggregation of embryonic chick central nervous system neurons.

The use of dissociated neuronal cell cultures is a very widespread technique. It is useful to study specific interactions between cells and resolve molecular mechanisms underlying neural development and function. For instance, the extended family of neurotrophic factors was identified and further studied especially using such techniques. Several growth factors have also been studied in this way as well as other developmental molecules. In this paper we describe a method of culturing chick cortical cells, in the complete absence of serum, which results in an enhanced aggregation of neurons by few days in culture. This cell culture system is particularly convenient to perform functional analyses of various molecules involved in neuronal cell adhesion mechanisms, such as extracellular matrix proteins or cell adhesion molecules, that require the establishment of in vitro paradigms in order to analyze their influence on cell-substratum and cell-cell interactions, as previously reported. We have successfully studied the effect of specific glycoproteins from the subcommissural organ on neuronal cell adhesion using this cell culture system.

Animals↗
PubMed1997

The Drosophila toucan (toc) gene is required in germline cells for the somatic cell patterning during oogenesis.

We have characterized a new gene, called toucan, that is expressed and required in germline cells to promote proper differentiation of the somatic follicle cells. toucan mutant ovaries are defective in (i) the enclosure of newly formed germline cysts by the follicle cells, (ii) the formation of interfollicular stalks, (iii) the migration of the follicle cells over the oocyte and (iv) the formation of the eggshell. Overexpression of a toucan cDNA in the germline leads to the production of longer interfollicular stalks than wild-type ovaries, a phenotype that is the exact opposite of the toucan mutant phenotype. This observation shows that the formation of the interfollicular stalks depends not only on interactions among the somatic cells but also requires a germline signal. Moreover, dominant interactions have been observed between toucan and certain alleles of the daughterless, Notch and Delta genes, each of which is required in the somatic cells for the formation of egg chambers. toucan encodes for a large protein with a coiled-coil domain but has no other homology with known proteins. We propose that toucan participates in the production or localization of a germline-specific signal(s) that is required for the patterning of the follicular epithelium.

Animals↗
PubMed1996

Soluble material from Reissner's fiber displays anti-aggregative activity in primary cultures of chick cortical neurons.

The subcommissural organ (SCO), which belongs to the circumventricular organs, is a specialized ependymal structure of the brain that secretes glycoproteins into the cerebrospinal fluid (CSF) which condense to form a thread-like structure, Reissner's fiber (RF). The effects of soluble material released by RF were examined in primary cultures of dissociated cortical cells from embryonic (day 8) chick brain. Under serum-free conditions, the presence in the cultures of soluble RF material markedly impaired neuronal cell aggregation. This effect was completely blocked by addition into the culture medium of specific antibodies raised against bovine RF. The anti-aggregative effect of soluble RF material is observed on poly-L-lysine as well as on different extracellular matrix proteins including collagen and laminin, but was less effective on fibronectin. The continuous exposure of the cultures to soluble RF material for 7 days significantly decreased choline acetyltransferase activity. On the other hand, soluble RF material did not appear to have mitogenic activity on neuronal cultures. Modulation of cell-cell interactions by SCO/RF glycoproteins strengthens the hypothesis of the involvement of RF in developmental events of the central nervous system.

Analysis of Variance↗