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F Gremo

Publications and source records attributed to F Gremo.

At least 37 records · Page 2Linked to original sources
PubMed1992

Correlation between P19 presence and MHC class II expression in human fetal astroglial cells cocultured with HTLV-I donor cells.

The possibility of a direct infection of human brain by HTLV-I, has been studied using an in vitro model. Human fetal astroglial cells were cocultivated with irradiated HTLV-I donor cell line MT-2, and assayed for the presence of HTLV-I core protein p19 after 1 week. Fifty-six per cent of GFAP positive astrocytes showed the viral core protein p19 and increased expression of Class II MHC antigens. Electron microscopy of astroglial cells exposed to HTLV-I revealed the presence of vacuoli-like structures containing viral core protein p19. Cell intermediate filament cytoskeleton was also disorganized. Even if this study does not provide direct evidence for virus replication inside astroglial cells, all these findings suggest that HTLV-I can indeed enter the cell and exert a cytopathic effect. Therefore the results of the present study are consistent with the hypothesis that astroglial cells could be involved in demyelination processes occurring in the HTLV-I associated neurological disorders, such as human associated myelopathy and tropical spastic paraparesis.

Astrocytes↗
PubMed1991

HTLV-I neurotropism. In vitro studies.

The human T cell leukemia lymphoma virus (HTLV-I) has been recently associated to neurological disorders. In order to investigate the interaction between this virus and the central nervous system, cells of neuroectodermic origin have been exposed to HTLV-I through an experimental in vitro model. The results revealed the presence of HTLV-I virus core protein p19 inside glial cells, after a short time exposure to a chronically infected donor cell line. This suggests a possible role of a lymphotropic retrovirus in neurological diseases.

Animals↗
PubMed1990

Basic fibroblast growth factor in neuronal cultures of human fetal brain.

The presence of basic fibroblast growth factor (bFGF) was investigated in neuronal cells derived from 12 and 18 week-old human fetal brain cultures. To this purpose, the ability of bFGF to stimulate plasminogen activator (PA) production in fetal bovine aortic endothelial GM 7373 cells was used as an assay for this molecule in neuronal cell extracts. The identity of the PA-stimulating activity of neuronal cell extract with bFGF was confirmed by its high affinity for heparin and by its cross-reactivity with polyclonal antibodies to human placental bFGF. These antibodies recognized a Mr 18,000 cell-associated protein both in Western blot and in immuno-precipitation experiments. All the neurons showed bFGF immunoreactivity, as demonstrated by immunocytochemical staining, while nonneuronal cells were unstained. The data demonstrate for the first time that cultured human fetal brain neurons contain and synthesize bFGF.

Animals↗
PubMed1990

Acetylcholine-induced production of platelet-activating factor by human fetal brain cells in culture.

Platelet-activating-factor (PAF) is a potent, biologically active lipid mediator produced by several tissues, including brain. Its role in the central nervous system (CNS) is still unknown, even if its involvement in brain damage and neurotoxicity has been postulated. Its production by neural cells has been demonstrated in different species, but not in man. This paper provides evidence that PAF can be produced by human fetal neurons and/or glial cells in culture. Its synthesis dramatically increased upon stimulation with acetylcholine (ACh), and it was significantly lowered by the cholinergic receptor antagonist atropine. Almost no PAF was detected in the incubation medium, which indicated no release of PAF from cultured cells. Characterization of the cells in culture with specific monoclonal antibodies excluded the presence of endothelial cells or macrophages, which also produce PAF.

Acetylcholine↗
PubMed1990

Synthesis of urokinase-type plasminogen activator and of type-1 plasminogen activator inhibitor in neuronal cultures of human fetal brain: stimulation by phorbol ester.

Human neuronal brain cultures established from 12- and 14-week-old fetuses synthesize and secrete urokinase-type plasminogen activator (uPA) and limited amounts of tissue-type plasminogen activator (tPA). These cells also produce and secrete the endothelial cell-type PA inhibitor (PAI-1), which forms sodium dodecyl sulfate-stable tPA/PAI-1 complexes in the culture medium. Immunocytochemistry shows a predominant localization of uPA, tPA, and PAI-1 in neuronal cells, with only a very weak positivity detectable in the few glial cells present in these cultures. The protein kinase C (PKC) activator 12-O-tetradecanoylphorbol 13-acetate (TPA) stimulates the synthesis of both uPA and PAI-1, resulting in a final increase in the plasmin-generating capacity of neuronal cell cultures. No significant effect is observed, however, when cells are treated with the TPA analogue 4 alpha-phorbol 12,13-didecanoate, which is inactive as a PKC inducer, or with the neurotrophic polypeptide basic fibroblast growth factor. These data represent the first characterization of the plasmin-generating system in human fetal brain neurons and suggest a role for PKC in the modulation of uPA and PAI-1 synthesis.

Brain↗
PubMed1989

Muscarinic receptor subclasses in retinal cultures: effect of corticosterone.

We have previously shown that exogenously administered corticosterone affects muscarinic receptor binding in the chick embryo retina. Analysis with the selective antagonist pirenzepine has shown that both muscarinic receptor subclasses M1 and M2 are present in treated retinas. On the contrary, only M2 is detectable in controls. Moreover, receptor affinity for agonists is modified by hormone treatment. Since these studies did not show whether or not the hormone directly influences retinal cells, a similar study was performed on retinal tissue cultures. Cells were treated at day 5 in vitro for 24 hr with 1.10(-8) M corticosterone. Scatchard analysis of results obtained with 3H-quinuclidinyl benzilate (3H-QNB) binding showed no difference between treated and control cultures. However, displacement experiments demonstrated that both M1 and M2 were present in treated cultures, whereas controls had only M2. Also, receptor affinity for the agonist carbachol was modified, as already observed with in vivo studies. In addition, a new phenomenon was observed: treated cultures had a significantly higher number of cells. The possibility that the hormone can prevent cell death or stimulate cell mitosis is considered.

Animals↗
PubMed1989

Developmental expression of intermediate filament proteins in the chick embryo retina: in vivo and in vitro comparison.

An immunocytochemical study was performed on the chick embryo retina to detect the changes in intermediate filament (IF) composition in correlation with development. Results showed that all three major neurofilament subunits (NF L, M and H) were expressed from very immature neurons at the same time. No delayed expression of NF-H was observed. Vimentin (VT) was also present from very early stages in both neuronal and glial cells, but it was not detectable at hatching. Its distribution changed in retinal layers throughout development. It co-existed with NF in neurons, and conversely, it preceded the appearance of the typical glial fibrillar acidic protein (GFA-P), which was detected in radial fibres of Müller cells from day 16 of incubation. These in vivo data were compared with immunocytochemical staining of retinal cultures, grown either on collagen or on polylysine. NF-L and GFA-P were not detectable in vitro. Moreover, the early expression of NF subunits was shown to be influenced by the type of substratum. The contemporary presence of vimentin and neurofilaments was clearly shown both in neuronal cell bodies and processes. The results also show that in the chick retina the pattern of composition of intermediate filaments changes with maturation. Moreover, in vitro conditions influence the expression of IF components. These data are of interest when performing in vitro analysis of the effect of exogenous substances affecting neuronal maturation and differentiation.

Animals↗
PubMed1988

Differential expression of neurofilament subunits in the developing corpus callosum.

In the course of development, corticocortical axons seem to first appear in a labile state from which they either mature into a stable state or are eliminated. These state transitions may be related to cytoskeletal modifications. By immunohistochemistry and immunobiochemistry we found that, in the corpus callosum of the cat, the heavy (200 kDa) subunit of neurofilaments (NF) becomes progressively more visible during the first postnatal month. This aspect of cytoskeletal maturation parallels the developmental loss of callosal axons, i.e. probably the stabilization of the axons which are not eliminated. A similar maturation of the heavy subunit was observed in the visual cortical areas 17 and 18. The medium (150 kDa) and to a lesser extent the light (70 kDa) NF subunits are already present a few days after birth.

Aging↗
PubMed1988

Acetylcholine and dopamine promote the production of platelet activating factor in immature cells of chick embryonic retina.

We have previously shown that platelet-activating factor (PAF), a naturally occurring lipid mediator of cell-to-cell communication, was produced by 3-day-old chick retina stimulated with acetylcholine (ACh) and dopamine (DA), but not with other neurotransmitters. ACh and DA stimulated PAF synthesis via a dithiothreitol (DTT)-insensitive cholinephosphotransferase, without affecting the acetyltransferase pathway, which was stimulated only by the calcium ionophore A23187. Therefore, we attempted to study the effects of neurotransmitters on PAF production and on the activities of the DTT-insensitive cholinephosphotransferase and acetyltransferase in the developing chick embryo retina up to hatching. Our results show that PAF was produced already at 8 days of development, when retinal cells are still rather immature and ganglion and Mueller cells are the only differentiated cells. The stimulation of PAF production occurred with ACh and not with other neurotransmitters. In older stages, DA also stimulated PAF production, as already described in the chick after hatching. DTT-insensitive cholinephosphotransferase and acetyltransferase activities were present in 8-day-old embryos, the earliest stage analyzed. Both enzymatic activities increased with age; DTT-insensitive cholinephosphotransferase increased rapidly from day 12 up to day 18, whereas acetyltransferase activity increased linearly up to the time of hatching. To promote PAF production, ACh and DA activate DTT-insensitive cholinephosphotransferase, but not acetyltransferase.(ABSTRACT TRUNCATED AT 250 WORDS)

Acetylcholine↗
PubMed1987

Heterogeneity of muscarinic cholinergic receptors in the developing human fetal brain: regional distribution and characterization.

The ontogeny of muscarinic cholinergic receptors has been studied in different regions of the human fetal brain. For a comparison, the same study has been carried out on newborn and premature brain. Regarding on the areas examined (frontal cortex, cerebellum, hippocampus, thalamus and basal ganglia) either an increase or a decrease of receptor density during gestation was observed. Thus, the ontogeny of the receptors follows a different pattern in areas which differ in function, cholinergic innervation and embryological origin. However, in all the regions the affinity of the binding site for the ligand [3H]quinuclidinyl benzilate [3H]QNB was very similar to that reported for muscarinic receptors from adult mammalian brain. Data obtained from agonist binding (acetylcholine and carbachol) revealed the presence of a high (H)- and a low-affinity binding site (L) from 10 weeks of gestation. The selective antagonist pirenzepine (PZ) also distinguished two different muscarinic receptor subtypes, which however had higher affinity than that seen in adult brain. In conclusion, during ontogeny, the muscarinic acetylcholine receptor shares some but not all of the pharmacological properties shown in the adult brain.

Basal Ganglia↗
PubMed1986

Production of platelet-activating factor by chick retina.

In the present study it is demonstrated that platelet-activating factor (PAF) was produced by chick retinas, upon stimulation with neurotransmitters such as acetylcholine (ACh), dopamine, or with calcium ionophore A23187, but not upon stimulation with gamma-amino-n-butyric acid, L-glycine, L-glutamate, epinephrine, or histamine. PAF produced in response to ACh, dopamine, or A23187 was not released into supernatants but was extractable from retinas. The amounts of extractable PAF increased after sonication of stimulated retinas. While no PAF activity could be recovered from unstimulated retinas, small amounts of this lipid can be detected following sonication of the tissue. The amount of extractable PAF from ACh-, dopamine-, or A23187-stimulated retinas was dependent upon the incubation time and concentration of the agonists. PAF was identified on the basis of chemical and lipase treatments, biological activity with washed rabbit platelets, behavior on thin layer chromatography, and high pressure liquid chromatography. Control cell preparations (leukocytes, erythrocytes, and embryogenic fibroblasts) did not produce PAF upon neurotransmitter stimulation. ACh and dopamine promoted PAF production by increasing dithiothreitol-insensitive cholinephosphotransferase activity, without affecting the acetyltransferase activity. In contrast, the A23187 ionophore stimulated the acetyltransferase activity but did not affect the dithiothreitol-insensitive cholinephosphotransferase.

Acetylcholine↗
PubMed1985

Heterogeneity of muscarinic cholinergic receptors in the developing chick embryo retina.

Heterogeneity of muscarinic cholinergic receptors was investigated in chick embryo retina throughout development and in chicks immediately after hatching. The presence of a homogeneous receptor population was evidenced by antagonist binding. The affinity of antagonists increased up to day 14 of incubation, when synaptogenesis occurs. After this stage, it remained substantially unchanged. The number of receptors increased in embryos until hatching. On the contrary, agonists, such as acetylcholine and carbachol, bound to two (high- and low-affinity) binding sites. Through development, the affinity of both significantly increased until day 14, further substantiating the hypothesis of a maturation of the receptor pattern which precedes synapse formation. Muscarinic cholinergic binding seems to identify 3 critical steps in retinal neuronal development. The first is between 7 and 9 days of incubation, the second when synaptogenesis occurs and the third after initiation of function.

Acetylcholine↗
PubMed1985

Mature and immature synaptosomal membranes have a different lipid composition.

Subfractionation of the optic tectum in chick embryos results in the isolation of two fractions enriched in synaptosomes (fraction A and fraction B). In chicks after hatching, this fractionation results in the isolation of a single synaptosomal fraction (fraction B) and of a fraction enriched in myelin membranes devoid of synaptosomes (fraction A). The lipid composition of synaptosomal fractions (A and B) and corresponding synaptosomal plasma membranes has been analyzed and compared to the lipid composition of similar fractions isolated from 2-3 day-old chicks. The phospholipid composition of fraction A in embryos was mainly represented by phosphatidylcholine (PC) and phosphatidylethanolamine (PE). The PE content was significantly lower than that of PC, which accounted for by approximately 50%. Sphingomyelin (SP) and phosphatidylinositol (PI) accounted for by only 6% of the total membrane phospholipids. Fraction A isolated from the young chicks showed many significant changes. PC accounted for by approximately 40% and PE made up 35%. The amount of phosphatidylserine (PS) and SP increased. These data parallel our previous morphological observations, which showed that fraction A contains immature synaptosomes in embryos but myelin membranes and no synaptosomes in the young chicks. Fraction B has been shown to contain synaptosomes at all stages considered. It possessed in embryos a lipid composition similar to fraction A, except that PC content was higher in young embryos. The analyses on membrane fractions confirmed these results. On the contrary, this fraction showed many significant changes after hatching. The content of PC was significantly reduced, PE/PC ratio was significantly increased as well as ethanolamine plasmalogen (PLE) content. The percentage of PS, PI and SP were increased. The composition of fatty acids of the total fraction of phospholipids was also examined. The results parallel the observations on phospholipid classes.

Age Factors↗
PubMed1985

Muscarinic receptor subclasses in the chick embryo retina: influence of corticosterone treatment.

The present study was performed on retinas of chick embryos receiving at day 8 of incubation an intracerebral injection of 0.02 microgram of corticosterone. We had previously shown with the use of [3H]quinuclidinylbenzilate [( 3H]QNB) that such treatment induced the appearance of two muscarinic binding sites in the treated retinas, whereas only one was detectable in the controls. In the present study we investigated muscarinic cholinergic receptor subclasses with agonist and antagonist binding. Agonist binding was studied by varying the concentrations of carbachol and acetylcholine (10(-9) M-10(-5) M) in the presence of a constant concentration (0.2 nM) of [3H]QNB. Two subpopulations of receptors were revealed, a high- and a low-affinity receptor, in both treated and control retinas. However, in the hormone-treated retinas, the two subpopulations significantly differed from the controls in their affinity and in their relative percentage among the total receptor population. Moreover, using pirenzepine, an antagonist known to have the capacity to distinguish between muscarinic cholinergic subclasses, two receptor subpopulations were found to be present in the hormone-treated retinas but a single one in the controls. It is suggested that hormone treatment can either induce the appearance of a new subclass of muscarinic cholinergic receptors or favor the maturation of a population of retinal cells having these receptors. Pirenzepine binding in retinas from intact embryos of 7, 9, and 11 days of incubation revealed one receptor subpopulation. Thus, these findings are more consistent with the hypothesis that corticosterone effects the target cells, either inducing changes in muscarinic receptor and/or modifying the receptor environment.

Acetylcholine↗
PubMed1984

Effects of corticosterone on chick embryonic retinal cells in culture.

Corticosterone has been shown to affect several patterns of glial cell and neuronal development. We have previously reported that exogenously administered corticosterone preferentially accumulated into the retinas of 8-day-old chick embryos. Moreover, we observed that it affects muscarinic cholinergic binding. Thus, we investigated the effect of different concentrations of corticosterone on retinal cells in culture. Retinas were dissected from 8-day-old embryos, dissociated and cells plated on salt-precipitated collagen. At day 5, cultures were treated with corticosterone (from 10(-9) M to 10(-7) M) for 24 h. Controls received either Dulbecco's Modified Eagle Medium (DMEM) plus 10% fetal calf serum (FCS) or DMEM only. Results show that the main effect of the hormone was inhibition of neuronal process outgrowth. Also cell aggregation, flat cell proliferation and confluency are altered in hormone-treated cultures. All these effects are reversible and can be attributed to hormone effect and not to serum deprivation.

Animals↗
PubMed1983

Muscarinic cholinergic binding in chick embryo retino-tectal system: effects of corticosterone.

Muscarinic binding sites were measured using the radioligand [3H]quinuclidinyl benzilate (QNB) in the retina and tectum of 11-day-old chick embryos, after intracerebral administration of 0.02 microgram of corticosterone at 8 days of incubation. This age was chosen because the hormone preferentially accumulates in retinas at 8 days of development. Hormone treatment significantly affected the affinity of 3H-QNB-binding sites in retinas and slightly affected the affinity in treated tecta, whereas the number of binding sites remained unchanged. The specific binding was determined with either atropine or unlabeled QNB. Scatchard plot analysis of specific 3H-QNB binding revealed the presence of nonsaturable binding at high 3H-QNB concentrations (6-11 nM) in the treated retinas, but not in controls. It can be concluded from these data that the hormone has a primary effect on retinal cells during early growth in the chick embryo. The possibility that the hormone delays maturation of specific populations of retinal cells is considered in the discussion.

Animals↗