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Biomedical subjects

Y Courtois

Publications and source records attributed to Y Courtois.

At least 19 recordsLinked to original sources
PubMed1992

Cloning of two different 5' untranslated exons of bovine acidic fibroblast growth factor by the single strand ligation to single-stranded cDNA methodology.

In an attempt to characterize the 5' UTR of the aFGF mRNAs we used the new anchored PCR methodology, single strand ligation to ss-cDNAs (SLIC). In bovine brain and retina, two kinds of aFGF cDNA clones were isolated. They contained two alternative exons located 34 bp upstream to the translation initiation codon ATG. Taking into account the number of clones specific for each exon, the two mRNAs are expressed with the same ratio in both tissues. One of these bovine 5' UTR exons (136 bp) showed 81% identity to a human 5' UTR exon, the second one (323 bp) was 70% identical to the second human 5' UTR exon with a central region of 90 nucleotides showing 41% identity. The conservation of the splicing positions for these 5' UTR alternate exons in both bovine and human species, suggests that the overall structure of the aFGF gene is conserved in mammals. Furthermore, the conservation of the nucleotide sequences and of the localization of these 5' UTR exons suggests that these non-coding regions may be involved in the control of aFGF gene expression.

Amino Acid Sequence↗
PubMed1992

Lipopolysaccharide and cytokines induce a macrophage-type of nitric oxide synthase in bovine retinal pigmented epithelial cells.

The present study demonstrates that bovine retinal pigmented epithelial cells, which are neuroectodermal in origin, produce nitric oxide (NO) upon treatment with interferon-gamma in the presence of lipopolysaccharide or tumor necrosis factor-alpha. NO production was measured by the accumulation of the stable endproduct NO2-. The biosynthesis of NO requires an induction period of approximately 12 hours and continues for at least 96 hours. The synthesis was abolished by the stereoselective inhibitors of NO synthase, NG-monomethyl-L-arginine and NG-nitro-L-arginine-benzylester. Cycloheximide and dexamethasone blocked cytokine-induced NO production. The results indicate that endotoxin and cytokines are capable of inducing NO synthase of the macrophage type, in retinal pigmented epithelial cells.

Amino Acid Oxidoreductases↗
PubMed1992

Heterogeneity of 3'untranslated region of bovine acidic FGF transcripts.

A bovine aFGF genomic clone (14.2 Kb) has been isolated and characterized. This clone contains exons 2 and 3 interrupted by 6.7 Kb long intron. Exon 3 contains part of the coding region and the 3' untranslated region (3'UTR). Two overlapping cDNA clones specific for this 3'UTR have been isolated from bovine retina cDNA libraries or after amplification of RNA by the RACE technique. Analysis of these clones and RNAse protection assay demonstrate alternative termination of aFGF transcripts giving rise to differently sized 3'UTR of 2.5 Kb and at least 3.9 Kb. The sequence of these long 3'UTR is highly conserved among species (70% identity between human and rat) which suggests an important role for aFGF expression.

Amino Acid Sequence↗
PubMed1992

Biphasic effect of the mitotoxin bFGF-saporin on bovine lens epithelial cell growth: effect of cell density and extracellular matrix.

We have studied the effect of a specific FGF receptor suicide antagonist on the growth of bovine epithelial cells (BEL cells) in culture. This basic fibroblast growth factor-saporin conjugate (bFGF-SAP) has a biphasic effect on bovine lens epithelial cells (BEL cells). Whereas 0.01 nM and 0.1 nM bFGF-SAP stimulate BEL cells proliferation, 1 nM and 10 nM bFGF-SAP have the predicted toxic effects on BEL cell growth. The toxicity of bFGF-SAP is observed 2 to 3 days after the initial treatment and depends on cell density. Accordingly, the sensitivity of confluent cells to bFGF-SAP is reduced compared to sparse cells. A time course analysis reveals that bFGF-SAP is effective after a short exposure to cells and that its effects are not increased with longer treatments. Cell growth on bFGF-SAP pretreated extracellular matrix (ECM) or posterior lens capsule (PLC) is also affected. Basic FGF-SAP has been shown to bind to the extracellular material, allowing a modulation of lens cells migration and survival by a single treatment in vitro. This finding raises the possibility of its use in vivo to prevent capsules invasion by lens cells after cataract surgery.

Animals↗
PubMed1992

Immunocytochemical study of extracellular matrix components during lens and neural retina regeneration in the adult newt.

We have conducted an immunocytochemical study of fibronectin, laminin, heparan sulfate proteoglycans and nidogen-entactin during lens and neural retina regeneration in the adult newt from 0 to 60 days. In the normal eye, fibronectin was detected in the corneal stroma and Descemet's membrane, in dorsal and ventral irises and lens capsule but not in Bowman's membrane of the cornea. In normal neural retina, fibronectin was found in Bruch's and inner limiting membranes. Heparan sulfate proteoglycans gave a slight signal in both irises and the lens capsule. Nidogen-entactin distribution in the cornea was similar to that of fibronectin; it was absent from the stroma of both irises, and the signal was weak in the pigmented iris epithelium. Nidogen-entactin was not detected in the lens capsule and inner limiting membrane of the neural retina but was present in Bruch's membrane. During the first 15 days of lens regeneration, fibronectin and nidogen-entactin decreased but did not disappear from the pupillary margin of both irises, and no signal was obtained for laminin and heparan sulfate proteoglycans. From day 15 to day 60 fibronectin and nidogen-entactin increased in both irises and lens capsule. The signal for laminin was restricted to the lens capsule. Heparan sulfate proteoglycans gave a slight signal in both irises and in the lens capsule. During the first 25 days of neural retina regeneration, fibronectin was the first to appear in Bruch's membrane and the cell border of the new neuroepithelium and remained during the entire process. Laminin appeared after 41 days in the inner limiting and Bruch's membranes, but by day 50 it appeared as a weak signal only in the inner limiting membrane. Heparan sulfate proteoglycans were not detected at any of the regeneration stages studied. Nidogen-entactin was only detected in Bruch's membrane and around the cells and blood vessels of the new neural retina. Later it was detected in the inner limiting membrane but not in Bruch's membrane. Thus, the results obtained showed that extracellular matrix components do change during both lens and neural retina regeneration. These changes may play an important role during both regenerating processes.

Animals↗
PubMed1992

Recombinant human basic fibroblast growth factor (Rh-bFGF) in three different wound models in rabbits: corneal wound healing effect and pharmacology.

Prior to a clinical trial in humans, we studied the effect and pharmacological distribution of recombinant human basic fibroblast growth-factor (Rh-bFGF) in vivo. Healing experiments on de-epithelialized rabbits corneas (n = 24 animals) compared the efficacy of three bFGF doses to controls and revealed a significantly increased healing rate for both 200 ng and 500 ng per application Rh-bFGF treatment groups compared to the control groups. To assess possible side effects of Rh-bFGF (500 ng topically applied for up to 7 days, twice daily), ten rabbits were involved in a model of an anterior keratectomy wound (performed with Draeger's roto-keratome to a depth of 0.15 mm). Light microscopy of thin sections of treated corneas showed an increased fibrogenesis in the anterior stroma with a more pronounced activation of keratocytes. No evidence for abnormal neovascularization or inflammation was observed when compared to control corneas. Ocular penetration and systemic distribution of topically applied labelled 125I FGF was assessed in three models (iodine vapour epithelial burn, anterior keratectomy and penetrating autokeratoplasty) in 24 rabbits. No intraocular penetration of bFGF occurred as shown by direct gamma counting. Macroautoradiography showed a selective labelling of epithelial basement membrane when denuded and intact, as previously described. Evidence for systemic absorption of breakdown products was confirmed by heparin-sepharose chromatography of blood and urine samples. Under these conditions, we suggest that topical Rh-bFGF promotes corneal wound healing without morphological adverse reaction or intraocular and systemic penetration.

Animals↗
PubMed1992

Basic fibroblast growth factor high and low affinity binding sites in developing mouse brain, hippocampus and cerebellum.

Fibroblast growth factors (FGFs), first extracted from brain and retina, are potent neurotrophic factors. They stimulate neuroblast proliferation and neuron differentiation and survival. In order to study the spatial and temporal distribution of the target cells in the mouse brain we studied by autoradiography and quantified by image analysis 125I-bFGF binding sites as a function of development. We have revealed the presence of two types of specific bFGF receptors. One is heparitinase sensitive and is co-localized with heparan sulfate proteoglycans of the basement membranes (meninges, choroid plexus and blood vessels). It is not developmentally regulated and corresponds to the low affinity receptors. It may be a storage form. The second type is heparitinase resistant and is modified during development, matching, in the adult, layering of the hippocampus and cerebellum. At 13 days of embryonic development there is a preferential distribution of silver grains on the ecto- and neuroectodermal tissues. In the adult, the labeling is localized on the neural process layers. It likely corresponds to the specific binding to cell high affinity receptors. Binding patterns according to the developmental stages of the brain can be correlated with mitotic, migration and differentiation phases of the neuronal cells.

Animals↗
PubMed1992

Glial cell localization of acidic fibroblast growth factor-like immunoreactivity in the optic nerve of young adult and aged mammals.

The number of axons in the optic nerve decreases with age and this degeneration is greater in patients suffering from Alzheimer's disease. Alterations in the role of neurotrophic factors could lead to this degeneration. Acidic fibroblast growth factor (aFGF)-like immunoreactivity was examined by indirect immunofluorescence on cryostat sections incubated with a rabbit polyclonal antiserum specific for aFGF. Staining was observed by photonic microscopy on optic nerves of Wistar rats (1- to 25-month-old), bovine animals (0.5- to 7-year-old) and normal human adults (24-, 34-, 54- and 84-year-old). In the three species studied, the results show that (1) glial cells were stained in the nuclear region and (2) aFGF-like immuno-reactivity was present over a large age span in adult subjects. Endogenous aFGF may have trophic effects on retinal ganglion cells and their axons throughout the adult life span.

Adult↗
PubMed1992

Human recombinant bFGF stimulates corneal endothelial wound healing in rabbits.

We have previously shown that bovine, human placenta extracted and recombinant human basic Fibroblast Growth Factor (bFGF) are effective in enhancing corneal epithelial wound healing in vivo. In the present study, we investigated the effect of rh-bFGF on the regeneration of injured rabbit endothelium. A standardized wound was created by scraping of endothelial cells with a special device within the boundaries of a central epithelial trephine mark of 7 mm in diameter. A single dose of 1.5 micrograms rh-bFGF was injected into the anterior chamber immediately after wounding, while control eyes received the vehicle only (n = 27). Functional recovery and wound closure rates were assessed by means of ultrasonic pachymetry, corneal button wet weight, endothelial vital staining as well as direct computer assisted surface analysis of Janus green stained corneal buttons. Measurements were carried out 1, 2, 4, and 7 days after injury. Morphological evaluation and cell counts at D4 and D7 were also performed. Significant stimulation of endothelial regeneration in rh-bFGF treated eyes, was observed with all methodological approaches. These results demonstrate the effectiveness of rh-bFGF in enhancing experimental corneal endothelial wound healing and advocate for a possible clinical application of this growth factor in order to preserve endothelial cell function or to promote healing of this important monolayer in case of disease or injury.

Animals↗
PubMed1992

[Health effects of chronic exposure to tobacco smoke on a non-smoker population].

A study devised to evaluate the effects of chronic exposure to tobacco smoke on the health of a population of non-smokers was conducted in a prison. Fourteen volunteers among male, non-smoking prisoners in good health and without history of lung disease were put for thirty days in a cell that was already occupied by three smokers. These subjects were examined on arrival and on the 30th day of their imprisonment. The examination included filling a questionnaire concerning daily habits and food, respiratory function tests, measurement of CO in the expired air, measurement of nicotine, cotinine, thiocyanates and cadmium concentrations in blood and in urine, and measurement of mutagenic substances excreted. The purpose of the study was to detect possible changes in the variables tested between the first and last days of confinement. No significant difference could be demonstrated within this lapse of time. These results are in agreement with the data available in the literature. In the present state of our knowledge, it is difficult to find evidence of a biological impact, notably on the concentrations of the specific indicators nicotine and cotinine, in subjects passively exposed to tobacco smoke.

Adult↗
PubMed1992

Fibroblast growth factor stimulates photoreceptor differentiation in vitro.

Dissociated newborn rat retinal cells were maintained in monolayer culture for periods of up to 11 d. When grown in the absence of exogenous growth factors, 1-2% of the total neuronal population expressed opsin (the photopigment that is specific for maturing photoreceptors). Addition of a single dose of 10 ng/ml basic fibroblast growth factor (bFGF) to the culture medium induced an average increase of sixfold in the numbers of neurons expressing opsin. This supplementation had little effect on the total number of differentiated neurons or of glial cells when measured at the same time points. Furthermore, another specific class of retinal neurons, the amacrine cells, showed no changes following exposure to this growth factor. Two other growth factors known to exert neurotrophic effects, epidermal and nerve growth factor, were without effect. The effect of bFGF was dose dependent, with highly significant differences being observed with as little as 100 pg/ml, and with 700 pg/ml eliciting half-maximal stimulation; maximal effects were observed at 10 ng/ml. Induction of opsin expression by low concentrations of bFGF was blocked completely by an antiserum directed specifically against bFGF, but not by preimmune serum immunoglobulins. This increase in the number of photoreceptors expressing opsin following exposure to bFGF could have been due to either increased cell survival, increased proliferation of progenitor cells, or increased differentiation of immature photoreceptors. There was no increase in overall cell survival under the experimental conditions used, and double labeling immunocytochemistry combined with autoradiographic analysis of 3H-thymidine uptake showed that proliferation of neuronal precursors was not enhanced by the addition of bFGF. In contrast to these observations, cultures established from older (postnatal day 3) retina revealed large numbers of opsin-expressing photoreceptors in all culture plates, with or without added growth factors. This reduction in the stimulatory effects of bFGF with increasing postnatal age is consistent with the period of sensitivity being limited to the cycling of neuronal precursors. It is possible that a bFGF-like molecule is secreted by neighboring cells such as the retinal pigmented epithelium, to participate in retinal development and differentiation. To our understanding, this molecule is the first protein identified to influence specifically the differentiation of photoreceptor cells.

Aging↗
PubMed1991

Acidic fibroblast growth factor (aFGF)-like immunoreactivity in the optic nerve.

Acidic fibroblast growth factor (aFGF)-like immunoreactivity was examined in the optic nerves of 1- to 25-month-old Wistar rats, 0.5- to 7-year-old bovine animals and normal human adults (24 and 35 years old), using cryostat sections incubated with a rabbit polyclonal antibody specific for aFGF. The immunoreactivity was associated with glial cells, and was localized predominantly in the nucleus. The presence of endogenous aFGF in the optic nerve of adult subjects and 'old' rats suggests that aFGF could play a role in the survival of retinal ganglion cells and their axons during aging.

Adult↗
PubMed1991

Molecular cloning of RI-HB, a heparin binding protein regulated by retinoic acid.

RI-HB is an extracellular heparin binding protein regulated by retinoic acid and essentially expressed during embryogenesis. This study reports the cloning and sequencing of the cDNA that encodes RI-HB. The sequence of RI-HB contains 121 amino acid residues and is very rich in basic amino acids and cysteines. This sequence was compared to those of HBGAM and MK protein, two other heparin binding proteins exhibiting growth and/or neurotrophic activities. Northern blot analysis indicates that RI-HB mRNA is strongly expressed during early chicken embryogenesis and that it is induced by retinoic acid treatment of chicken fibroblasts and myotubes in culture.

Animals↗
PubMed1991

A new heparin binding protein regulated by retinoic acid from chick embryo.

A 19 KDa heparin binding protein was previously purified from chicken embryos. Essentially localized within basement membranes in early embryonic tissues, this protein is very rich in basic and cystein residues. Its N-terminal fragment is similar to corresponding fragment of two other proteins expressed during embryogenesis and postnatal period. Its synthesis and secretion are induced by retinoic acid in chicken myoblasts and fibroblasts. This new retinoic acid induced heparin binding protein (RI-HB) does stimulate neurite outgrowth and proliferation on PC12 cells. These results suggest that retinoic acid could regulate some aspect of differentiation and development by inducing the synthesis of a new family of growth and neurotrophic factors.

Adrenal Gland Neoplasms↗
PubMed1991

In vitro kinetics of basic fibroblast growth factor diffusion across a reconstituted corneal endothelium.

We have studied the ability of bFGF to traverse and be trapped within basement membranes. An extract of EHS tumor (matrigel) coated on culture chamber filters was used as an in vitro model of basement membranes. Our results showed a slow diffusion of bFGF dependent on the amount of low affinity binding sites present within the matrigel. High amounts of bFGF and heparin increased the initial rate of diffusion by displacement of bFGF bound to matrigel. An in vitro corneal endothelium model (endothelial cells overlying matrigel) was also developed. This monolayer, with a weak permeability, decreased the kinetic rate of bFGF diffusion compared with matrigel alone. These results indicate that modulation of bFGF distribution in a tissue by a basement membrane is dependent on bFGF concentration, basement membrane composition, permeability of associated cells, and local presence of heparin. This selective control may be a regulating step in bFGF action.

Animals↗