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Biomedical subjects

F Lamy

Publications and source records attributed to F Lamy.

At least 19 recordsLinked to original sources
PubMed2000

Phosphatidylinositol 3-kinase, protein kinase B and ribosomal S6 kinases in the stimulation of thyroid epithelial cell proliferation by cAMP and growth factors in the presence of insulin.

The proliferation of most normal cells depends on the co-operation of several growth factors and hormones, each with a specific role, but the key events involved in the action of each necessary stimulant remain largely uncharacterized. In the present study, the pathways involved in the mechanism(s) of co-operation have been investigated in primary cultures of dog thyroid epithelial cells. In this physiologically relevant system, thyroid stimulating hormone (TSH) acting through cAMP, epidermal growth factor (EGF) and phorbol esters (such as PMA) induce DNA synthesis. Their effect requires stimulation of the insulin-like growth factor-1 (IGF-1) receptor by either IGF-1 or insulin, which are not themselves mitogenic agents. In contrast, hepatocyte growth factor (HGF) is itself fully mitogenic. The results of the study demonstrate that cAMP, EGF, HGF and PMA stimulate p70 ribosomal S6 kinase (p70 S6 kinase). However, insulin/IGF-1 also stimulate p70 S6 kinase. Thus stimulation of p70 S6 kinase might be necessary, but is certainly not sufficient, for the induction of DNA synthesis and is not specific for any stimulated pathway. In contrast, phosphatidylinositol 3-kinase (PI 3-kinase) and protein kinase B (PKB) activation by insulin and HGF is strong and sustained, whereas it is weak and transient with EGF and absent in the presence of TSH or PMA. These findings suggest that: (i) stimulation of PI 3-kinases and/or PKB is not involved in the cAMP-dependent pathways leading to thyrocyte proliferation, or in the action of PMA, (ii) the stimulation of the PI 3-kinase/PKB pathway may account for the permissive action of insulin/IGF-1 in the proliferation of these cells, and (iii) the stimulation of this pathway by HGF may explain why this agent does not require insulin or IGF-1 for its mitogenic action.

Androstadienes↗
PubMed1999

Biphasic extracellular proteolytic enzyme activity in benthic water and sediment in the northwestern mediterranean Sea

In this study, we used the fact that bacteria are able to cleave a fluorogenic substrate analog (L-leucine-7-amido-4-methylcoumarin) to determine the maximal ectoproteolytic activities (Vm) and affinities (Km) of natural benthic microbial communities by the multiconcentration kinetic method. This investigation was performed during the winter and summer of 1997 with a set of 36 samples of near-bottom water and sediment collected from a coastal area and an offshore area in the western part of the Gulf of Lions. The existence of biphasic microbial ectoproteolysis was statistically confirmed for both the near-bottom water and the sediment, regardless of the spatial and seasonal conditions. Globally, 72.2% of the entire set of bacterial consortia collected at the water-sediment boundary layer showed biphasic microbial kinetics. A specific estimator of the biphasicity indicated that deep benthic bacterial consortia responded better with episodic nutrient supplies than shallower benthic bacterial consortia responded.

Journal Article↗
PubMed1997

Phosphorylation of the three Rb protein family members is a common step of the cAMP-, the growth factor, and the phorbol ester-mitogenic cascades but is not necessary for the hypertrophy induced by insulin.

Thyrotropin (TSH) through the cAMP cascade and in the presence of insulin induces the proliferation of dog thyroid cells. In this work, it is shown that TSH via cAMP causes the phosphorylation of the three members of the pRb family, pRb, p107, and p130, with the same kinetics as those observed when these cells are stimulated by mitogens acting through a tyrosine kinase receptor or through activation of kinase C. It is the first described point of convergence of cAMP-dependent and -independent mitogenic pathways in dog thyrocytes and suggests that the phosphorylation of the three proteins may be involved in the initiation of DNA synthesis in these cells. We also show that insulin, which induces hypertrophy and is permissive for the TSH mitogenic action, does not provoke the phosphorylation of any pRb family member, suggesting that none of these phosphorylations is required for this effect.

Animals↗
PubMed1996

Thyrotropin via cyclic AMP induces insulin receptor expression and insulin Co-stimulation of growth and amplifies insulin and insulin-like growth factor signaling pathways in dog thyroid epithelial cells.

Despite the similarity of their receptors and signal transduction pathways, insulin is regarded as a regulator of glucose, protein, and lipid metabolism, whereas insulin-like growth factors (IGF-I and IGF-II) mainly act as mitogenic hormones. In the dog thyroid primary culture model, the triggering of DNA synthesis by thyrotropin (TSH) through cAMP, or by cAMP-independent factors including epidermal growth factor, hepatocyte growth factor and phorbol esters, requires insulin or IGFs as comitogenic factors. In the present study, in TSH-treated cells, IGF-I receptors and insulin receptors were paradoxically equivalent in their capacity to elicit the comitogenic pathway, which, however, was mediated only by IGF-I receptors in dog thyroid cells stimulated by cAMP-independent mitogens. Moreover, prior cell exposure to TSH or forskolin increased their responsiveness to insulin, IGF-I, and IGF-II, as seen on DNA synthesis and activation of a common insulin/IGF signaling pathway. To understand these observations, binding characteristics and expression of insulin and IGF-I receptors were examined. To analyze IGF-I receptor characteristics, the unexpected interference of a huge presence of IGF-binding proteins at the cell membrane was avoided using labeled Long R3 IGF-I instead of IGF-I. Strikingly, TSH, through cAMP, time-dependently induced insulin binding and insulin receptor mRNA and protein accumulation without any effect on IGF-I receptors. These findings constitute a first example of an induction of insulin receptor gene expression by a cAMP-mediated hormone. In dog thyroid cells, this allows low physiological insulin concentrations to act as a comitogenic factor and might explain in part the enhanced responsiveness to IGFs in response to TSH. This raises the possibility that TSH-insulin interactions may play a role in the regulation of thyroid growth and function in vivo.

Animals↗
PubMed1996

Expression and subcellular localization of CDK2 and cdc2 kinases and their common partner cyclin A in thyroid epithelial cells: comparison of cyclic AMP-dependent and -independent cell cycles.

Dog thyroid epithelial cells in primary culture constitute a model of positive control of DNA synthesis initiation and G0-S prereplicative phase progression by cyclic AMP as a second messenger for TSH. In tis early steps, this mitogenic control is quite distinct from cyclic AMP-independent mitogenic cascades elicited by growth factors. We demonstrate here that TSH (cyclic AMP) and EGF+serum (cyclic AMP-independent) stimulations cooperate and finally converge on proteins that control the cell cycle machinery. This convergence included a common induction of the expression of cyclin A and p34cdc2, and to a lesser extent of p33/38cdk2, which was already expressed in quiescent thyroid cells, and common changes of cdc2 and CDK2 phosphorylations as evidenced by electrophoretic mobility shifts. Kinetic differences in these processes after stimulation by TSH or EGF+serum or by these factors in combination correlated with differences in cell cycle kinetics. Moreover, an immunofluorescence analysis of these proteins using the double labeling of PCNA as a marker of each cell cycle phase shows: (1) a previously undescribed nuclear translocation of CDK2 before S phase initiation; (2) a sudden increase of cdc2 nuclear immunoreactivity at G2/mitosis transition. These data support the roles of CDK2 and cdc2 at G1/S and G2/mitosis transitions, respectively. (3) We were unable to demonstrate in individual cells a strict association between the nuclear appearance of cyclin A and G1/S transition, and an association of cyclin A and CDK2 with PCNA-stained DNA replication sites. On the other hand, the lengthening of G2 phase in the TSH/cyclic AMP-dependent thyroid cell cycle was associated with a stabilization of Tyr15 inhibitory phosphorylation of cdc2 and an especially high nuclear concentration of cyclin A and CDK2. We hypothesize that high nuclear accumulation of cyclin A and CDK2 during G2 phase could be causative in the cyclic AMP-dependent delay of mitosis onset.

Animals↗
PubMed1996

c-Myc expression is controlled by the mitogenic cAMP-cascade in thyrocytes.

In dog thyroid epithelial cells in primary culture, thyrotropin (TSH), acting through cAMP, induces proliferation and differentiation expression, whereas epidermal growth factor (EGF) and phorbol esters induce proliferation and dedifferentiation. In these cells, we have detailed the regulation by cAMP of the c-myc protooncogene mRNA and protein. The cAMP signaling pathway induces a biphasic increase of c-myc mRNA and protein. c-Myc protein accumulation follows the abundance and kinetics of its mRNA expression. Using in vitro elongation of nascent transcripts to measure transcription and actinomycin D (AcD) chase experiments to study mRNA stability, we have shown that in the first phase cAMP releases a transcriptional elongation block. No modification of transcriptional initiation was observed. After 30 min of treatment with TSH, c-myc mRNA was also stabilized. During the second phase, cAMP stabilization of the mRNA disappears and transcription is again shut off. Thus, in a tissue in which it stimulates proliferation and specific gene expression, cAMP regulates biphasically c-myc expression by mechanisms operating at the transcriptional and posttranscriptional levels.

Amino Acid Sequence↗
PubMed1995

Herpesvirus-like DNA sequences in patients with Mediterranean Kaposi's sarcoma.

DNA sequences closely related to herpesvirus-like sequences have been found in AIDS-associated Kaposi's sarcoma. Using PCR, we found herpesvirus-like DNA sequences in Kaposi's lesions and normal adjacent skin in five patients with Mediterranean Kaposi's sarcoma. We did not find these sequences in tissues from patients without Kaposi's sarcoma. Semi-quantitative PCR revealed many more herpesvirus-like sequences in Kaposi's lesions than in unaffected skin. Our results reinforce the hypothesis that an infectious agent closely related to gamma-herpesvirus is implicated in the pathogenesis of Mediterranean and AIDS-associated Kaposi's sarcoma.

Acquired Immunodeficiency Syndrome↗
PubMed1995

Rapid purification and identification of calcyphosine, a Ca(2+)-binding protein phosphorylated by protein kinase A.

A method is presented for the rapid purification of dog thyroid calcyphosine, a protein previously identified as a major substrate for cyclic AMP-dependent protein kinase in dog thyroid slices stimulated by thyrotropin [Lecocq, Lamy and Dumont (1979) Eur. J. Biochem. 102, 147-152]. The protein was previously identified as a spot on two-dimensional gels and is now purified in its native form by a procedure involving three chromatographic steps. Homogeneous calcyphosine identified by SDS/PAGE, immunoblotting and peptide sequencing can be obtained within 7 h. As for calmodulin, Ca(2+)-dependent conformational changes can be shown by Ca(2+)-dependent hydrophobic interaction chromatography using phenyl-Sepharose. Unlike calmodulin, calcyphosine is a substrate for protein kinase A.

Amino Acid Sequence↗
PubMed1994

[Gastroduodenal tolerance of methylprednisolone. Study of oral versus intravenous administration in healthy volunteers].

OBJECTIVES: To endoscopically evaluate the tolerance of gastroduodenal mucosa to methylprednisolone given orally and intravenously. METHODS: Thirty two healthy volunteers (age range 18-39 years) were divided randomly into two groups of 16 each (8 males and 8 females). All were Caucasians, gave their informed consent and were considered normal after a complete clinical and laboratory work-up including gastroduodenal fibroscopy. Methylprednisolone (500 mg) was administered for three consecutive days at 9 a.m., orally in one group and intravenously in the second group. No other drugs were being taken and alcohol and smoking were prohibited from day 0 to day 11. Tolerance was evaluated on days 4 and 11 based on clinical examination, blood pressure, heart rate, oral temperature, body weight, blood and urine chemistry and by video-recorded gastroduodenal endoscopy. Two independent endoscopists, uninformed of the patient's regimen, scored lesions from 0 (normal) to 5 (more than 25 lesions including at least 2 erosions). In case of abnormal findings, follow-up was continued to normalization. RESULTS: Endoscopically detectable lesions (stage I) attributed to corticosteroid therapy were observed in 4 subjects in the oral group and in 5 in the intravenous group. All regressed spontaneously. Duodenal lesions were observed only after oral administration while lesions of gastric mucosa were mostly found after intravenous administration. Systemic effects included abdominal pain after oral intake, 1 case of insomnia and bitter taste in the mouth after intravenous administration. CONCLUSIONS: These findings suggest that the effect of corticosteroid therapy, on the gastric mucosa, is basically systemic, and on the duodenal mucosa, basically local. No severe manifestations were observed after high-dose methylprednisolone given orally or by intravenous injection.

Administration, Oral↗
PubMed1994

Characterization and identification as cofilin and destrin of two thyrotropin- and phorbol ester-regulated phosphoproteins in thyroid cells.

Using separation of total cellular proteins by two dimensional (2-D) gel electrophoresis (isoelectric focusing/SDS-PAGE) we have characterized two regulated proteins, p21 and p19, in dog thyroid cells. We have used the same 2-D gel technique to purify these proteins before their trypsin cleavage and partial sequencing. Three peptides were sequenced in the case of p19 and two peptides in the case of p21. The Swiss-Prot protein sequence database revealed that p19 was identical to destrin/ADF (actin depolymerizing factor) and p21 to cofilin, two closely related and widely distributed actin-binding proteins. This was further verified by cross-reactivity with specific antibodies against brain cofilin and chicken ADF. We have demonstrated, using 2-D gel electrophoresis with a nonequilibrium pH gradient in the first dimension (nonequilibrium pH gradient in the first dimension (nonequilibrium pH gradient electrophoresis/SDS-PAGE) that, in the thyroid cell, cofilin and destrin/ADF were present, under control conditions, in two forms: a phosphorylated and an unphosphorylated one. Thyrotropin (TSH), through cyclic AMP, provoked a very rapid dephosphorylation of these two proteins, which was already maximal after 20 min of action, whereas their dephosphorylation in response to 12-O-tetradecanoylphorbol-13-acetate (TPA) was slower. This suggests that dephosphorylation of cofilin and destrin/ADF by TSH could be implicated in the disruption of actin-containing stress fibers and in the reorganization of microfilaments induced by this hormone. Epidermal growth factor, which does not induce acute morphological changes in thyroid cells, did not affect the state of phosphorylation of cofilin and destrin/ADF except for a delayed decrease (after 24 h) of destrin/ADF phosphorylation. A 10% dimethyl sulfoxide treatment of thyroid cells also induced rapid dephosphorylation of destrin and cofilin. This was accompanied by a reorganization of actin microfilaments that clearly resembles the one induced by TSH and by the appearance of intranuclear cofilin-containing rods. However, these rod structures were not observed in response to TSH, forskolin, or TPA, suggesting that dephosphorylation of cofilin correlates with the reorganization of actin microfilaments but not with the nuclear transport of cofilin. We propose that the dephosphorylation of destrin and cofilin could be involved in the TSH-stimulated macropinocytic activity, a key process in thyroid hormone secretion.

Actin Depolymerizing Factors↗
PubMed1994

Immunocyte enumeration in duodenal biopsies of migraine without aura patients with or without food-induced migraine.

The possibility of an IgE-mediated allergic mechanism in food-induced migraine remains controversial. Twenty consecutive migraine patients, 11 with food-induced migraine, 9 without, were investigated for determination and counting of immunocyte populations (IgA, IgM, IgE, IgG containing cells) by biopsies at duodenum 2 level. Conventional histology and plasmocyte populations were not significantly different between the two groups of migraine patients. This study does not support the existence of an IgE-mediated allergic mechanism in food-induced migraine.

Adult↗
PubMed1994

Cloning of cDNA specifically involved in the thyroid cAMP mitogenic pathway.

The activation of the cyclic AMP cascade in dog and human thyroid cells in primary culture induces the expression of differentiated gene expression, hyperfunction and proliferation. These programs are developed simultaneously in quiescent dedifferentiated cells. In this paper the strategy followed by our group to define the genes involved in the cAMP mitogenic cascade is outlined.

Animals↗
PubMed1993

Phosphorylation of mitogen-activated protein kinases is involved in the epidermal growth factor and phorbol ester, but not in the thyrotropin/cAMP, thyroid mitogenic pathway.

In dog thyroid epithelial cells (thyrocytes) in primary culture, thyrotropin (TSH) acting through cAMP induces proliferation and differentiation expression, whereas epidermal growth factor (EGF) and tumor-promoting phorbol esters induce proliferation and dedifferentiation. In these cells we have demonstrated mitogen-activated protein (MAP) kinase phosphorylation by 32P labeling and two-dimensional gel electrophoresis and by immunodetection with anti-MAP kinase and anti-phosphotyrosine antibodies after one- or two-dimensional gel electrophoresis. MAP kinase localization was demonstrated by immunochemical staining. We show the following results. (i) As in other systems, EGF and phorbol esters induced p42 and p44 MAP kinases phosphorylation on tyrosine, serine, and threonine. This effect was rapid, peaking after 5 and 15 min, respectively, followed by a slow decline thereafter. It preceded a translocation of MAP kinase immunoreactivity from cytoplasm to nucleus. (ii) Carbamylcholine, a potent stimulator of the Ca(2+)-phosphatidylinositol cascade which is unable to induce DNA synthesis, stimulated MAP kinases phosphorylation and nuclear staining with kinetics similar to those observed after EGF action, indicating that MAP kinase phosphorylation was not sufficient for mitogenesis. (iii) The cAMP-dependent mitogenic cascade elicited by TSH and forskolin did not involve the phosphorylation and nuclear translocation of p42 and p44 MAP kinases at any time during the entire prereplicative phase. Activation of MAP kinases by phosphorylation is therefore not a necessary step in the G0-G1 transition in this mitogenic cascade.

Animals↗
PubMed1993

General inhibition by transforming growth factor beta 1 of thyrotropin and cAMP responses in human thyroid cells in primary culture.

Transforming growth factor beta 1 (TGF beta 1) mRNA has previously been identified in human thyroid cells and this agent has been shown to inhibit DNA synthesis in thyroid cells of some other species. In normal human thyroid cells in primary culture, TGF beta 1 inhibited inconstantly the low basal DNA synthesis and strongly the stimulation of DNA synthesis by epidermal growth factor (EGF) and serum, and by thyroid-stimulating hormone (TSH) acting through cAMP. This inhibition, by TGF beta 1, of the TSH and cAMP-dependent DNA synthesis was associated with an inhibition of PCNA (proliferating cell nuclear antigen) synthesis. TGF beta 1 almost completely abolished the cAMP induced stimulation of iodide uptake and thyroperoxidase synthesis. It thus, like EGF, also acts as a dedifferentiating agent. Investigation of the pattern of protein synthesis by two-dimensional gel electrophoresis revealed that while TGF beta 1, by itself, increased the synthesis of only one protein, a tropomyosin isoform, it inhibited most of the effects of cAMP on protein synthesis (35 out of 45 cAMP-regulated proteins were affected). It also reversed the effect of cAMP on the morphology of the thyrocytes. The fact that TGF beta 1 did not affect the increase in cAMP provoked by TSH in human thyroid cells while inhibiting most of the effects of dibutyryl cAMP in these cells suggests an action at a step distal to cAMP generation.(ABSTRACT TRUNCATED AT 250 WORDS)

Blood Physiological Phenomena↗
PubMed1993

Selenium and the thyroid: how the relationship was established.

Several hypotheses concerning consequences of selenium deficiency on iodine metabolism can be proposed on the basis of experimental studies in rats and from epidemiological and experimental studies in humans. By decreasing intracellular GSH peroxidase activity, selenium deficiency may increase hydrogen peroxide (H2O2) supply and lead over several weeks to the thyroid atrophy observed in myxoedematous cretins. By improving thyroid hormone synthesis and by decreasing peripheral thyroxin (T4) deiodination, selenium deficiency could protect fetal brain T4 supply and thus prevent neurologic cretinism. Selenium deficiency may protect against iodine deficiency by decreasing T4 metabolism--and thus iodide leakage and--perhaps also by increasing H2O2 supply and thyroid hormone synthesis and thus thyroid efficiency.

Animals↗