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G Fayet

Publications and source records attributed to G Fayet.

At least 55 records · Page 3Linked to original sources
PubMed1982

Reorganization of porcine thyroid cells into functional follicles in a chemically defined, serum- and thyrotropin-free medium.

In the serum-free, chemically defined medium NCTC 109, freshly isolated porcine thyroid cells aggregate and form functional follicles in culture even in the absence of thyrotropin. The follicular pattern observed under light and electron microscopy express the main morphological characteristics of in vivo thyroid cells. Follicles are large, replete with dense colloid, and the apical pole of cells is characterized by well-developed microvilli and the presence of aminopeptidase N. The index of iodide transport activity (125I-C/M ratio) decreases vs. days of culture to a resting value of about 1 or 2 at day 2. Addition of thyrotropin (200 microU/ml final concentration) at day 4 is followed by a 10-fold increase in iodide transport activity within 24 h and a 40-fold increase 4 d later. Incorporation and organification of iodide are dose dependent between 0 and 250 microU/ml thyrotropin; highest concentrations (4,000--16,000 muU/ml) are significantly inhibitory. In the absence of thyrotropin each cell synthesizes 8.2 pg thyroglobulin/d. Acute stimulation by thyrotropin at day 4 resulted in a slight decrease in the quantity of thyroglobulin present in the cell layer but in an increase in the total amount of thyroglobulin recovered in both cells and medium, reaching 34.3 pg/cell/d. The protein exported into the medium is thyroglobulin, as shown by SDS PAGE and immunological properties. Here we demonstrate that porcine thyroid cells can be maintained in culture as resting, highly differentiated, follicular-associated cells, sensitive to acute stimulation by thyrotropin.

Animals↗
PubMed1981

[Aminopeptidase N, a marker of the apical pole in thyroid epithelium of swine and in cultured cells].

An aminopeptidase N has been detected by immunofluorescence in the apical plasma membrane of porcine thyroid cells, facing the follicular lumen. Freshly isolated cells obtained by tissue trypsinization have lost their polarity and exhibited a homogeneous enzyme distribution over the whole plasma membrane. In thyrotropin-stimulated cultured cells organized into follicles, the enzyme is localized on the apical cell pole as observed in tissue. In monolayer cells, the enzyme is localized on all the surface facing the medium. In both types of cultures, fluorescence is also observed in intracytoplasmic organelles, probably lysosomes. Aminopeptidase is a marker of the apical part of the thyroid plasma membrane. Its in vitro localization depends upon cell differentiation which is in relation to culture conditions.

Aminopeptidases↗
PubMed1981

Aminopeptidase N is a marker for the apical pole of porcine thyroid epithelial cells in vivo and in culture.

An aminopeptidase N has been detected by immunofluorescence in the apical plasma membrane of porcine thyroid cells, facing the follicular lumen. Freshly isolated cells obtained by tissue trypsinization, lose their polarity and exhibit a homogeneous enzyme distribution over the whole plasma membrane. In thyrotropin-stimulated cultured cells organized into follicles, the enzyme is localized the apical cell pole. In monolayer cells, on the other hand, the enzyme is distributed over the whole surface facing the medium. In both types of cultures fluorescence is also observed in intracytoplasmic organelles. In vivo, aminopeptidase ia a marker of the apical part of the thyroid plasma membrane, but its in vitro localization depends upon cell differentiation related to the culture conditions.

Aminopeptidases↗
PubMed1981

Follicle formation and iodide metabolism in cultures of human thyroid cells.

Primary cultures were initiated using thyroid tissue obtained at operation from patients with Graves's disease. The in-vitro conditions which permitted the formation of functional follicular structures in both primary cultures and derived sub-cultures were examined. In both situations, culture without the addition of calf serum to the medium resulted in the formation of follicles in response to thyrotrophin. In primary cultures the response to stimulation by exogenous thyrotrophin was variable. However, cells derived from long-term primary monolayers responded to thyrotrophin stimulation in a more predictable manner. In sub-cultures, the ability of cells to concentrate and organify iodide was augmented in a dose-dependent fashion in response to thyrotrophin (0 to 0.2 mu./ml); maximal values of 20 to 80 times those of control cultures being obtained. While follicular structure was maintained at higher hormone concentrations iodide-trapping capacity declined. Similar effects were produced by both low and high purity thyrotrophin and by dibutyryl cyclic AMP. Thyroid cells from two patients with a genetic defect of iodide organification exhibited the same lesion in vitro.

Cells, Cultured↗
PubMed1980

In vitro conversion of porcine thyroid cells growing in monolayer into functional follicular cells.

Porcine thyroid cells in primary cultures form either monolayers or, when they are stimulated by thyrotropin (TSH), follicles. This system, monolayer-follicle associated cell culture, determines two morphofunctional states of the thyroid cell in vitro. These cells divide, when grown in monolayer. In this article we describe the precise conditions which allow the conversion of monolayer cells into follicles. The rate of cell growth was lowered using a serum-free medium. Cells were concentrated, stimulated by TSH and cultured on poly-L-lysine pretreated plastic flasks. Light and electron microscope studies show that cells reorganize into follicles containing thyroglobulin. Active iodide transport by the cells, as well as detection of thyroid hormones in the cell culture media, demonstrate that these follicles are functional. Formation of monolayers from follicles is in vitro a spontaneous phenomenon. It is linked to the loss of cell polarity, iodide transport, synthesis of hormones and to the decrease of the number of TSH receptor-sites. These main characteristics of differentiation may be regained in vitro after conversion of monolayer thyroid cells into active follicles up to at least generation five.

Animals↗
PubMed1979

Biosynthesis of the basal lamina as a result of interaction between thyroid and mesenchymal cells in culture.

Porcine thyroid cells were cultured alone or in mixed cultures with mesenchymal cells. The formation of a basal lamina in vitro was investigated ultrastructurally. Follicular reassociation of thyroid cells occurred in both types of culture; however, it was followed by formation of the basal lamina only when mesenchymal cells were present. The present findings suggest an epithelial origin of the basal lamina resulting from an interaction with mesenchymal cells.

Animals↗
PubMed1979

Demonstration of growth in porcine thyroid cell culture.

Eagle's minimum essential medium supplemented with 20 per cent newborn calf serum (N.C.S.) allows porcine thyroid cell survival but not cell growth in vitro. In NCTC 109 medium supplemented with 20 per cent N.C.S. these cells actively grow and may be serially propagated. Cell population doubling time expressed as DNA doubling value is 3.5 days at 37 degrees C in 95 per cent air-5 per cent CO2. Thyrotropin does not affect porcine thyroid cell multiplication in vitro but stimulates the plating efficiency in primary cultures to about 130 per cent of controls. Cell selection was obtained by replacing media with Earle's balanced salt solution. This operation provoked death of nearly all cells by day 18 but subsequent addition of growth medium resulted in proliferation of epithelial cell clones. From generation 2 to generation 8, cells produce thyroglobulin but they do not actively trap iodide nor form follicles when thyrotropin is added to the media. Cell selection, demonstration of growth, as well as freeze-storage techniques described in this paper permit selection and storage of porcine thyroid cells and the potential constitution of cell collections.

Animals↗
PubMed1977

Active transport of iodide in isolated porcine thyroid cells. Application to an in vitro bioassay of thyrotropin.

Porcine thyroid cells were cultured for 2 days with or without dibutyryl cyclic AMP or thyrotropin (TSH). Then they were isolated post-culture by a gentle treatment with a calcium chelating agent. Some characteristics of the iodide transport system were studied in these thyroid cell suspensions. Iodide influx is a saturable, temperature- and energy-dependent phenomenon. It is blocked by ouabaïn, N-ethylmaleimide, dinitrophenol, cardiotoxin, low Na+ concentration and harmaline. Only 3% of the intracellular iodide is trichloracetic-acid-insoluble at equilibrium. The apparent Michaelis constant (Km) of the transport system is 30 micronM. For monolayer cells, the decrease of C/M ratio, increase of apparent Km, and decrease of Vmax between day 0 (freshly isolated cells) and day 6, indicate a loss of iodide-trapping ability up to passive diffusion. To the contrary, high values of C/M and normal Km (30 micronM) are observed in TSH follicles from reassociated cells. At iodide equilibrium, thryotropin, prostaglandins E1 and E2 and long-acting thyroid stimulator (LATS), induce a fast release of iodide. This release is dose-dependent in the first 5 min. It has been used to develop a bioassay of TSH and a fast detection of LATS.

Animals↗
PubMed1975

Thyrotropin-receptor interaction and cyclic AMP-mediated effects in thyroid cells.

Thyroid cells in culture constitute a suitable system for the study of thyroid gland function and its regulation. The natural thyroid stimulator (TSH) induces the in vitro reorganization of thyroid cells into three-dimensional follicles morphologically and metabolically similar to gland follicles. In contrast, nonstimulated cells develop as a monolayer and in a concerted manner rapidly lose the enzymes involved in iodine metabolosm and the aptitude to bind TSH to specific receptor sites. The morphogenetic action of TSH and its ability to maintain the specific metabolic properties of thyroid cells in culture are mediated by cyclic AMP via new RNA and new protein synthesis. Therefore comparison of the properties of a given cell type in morphologically and metabolically different status should provide a valuable tool for studying the TSH mechanism of action. Using 125-I-labeled TSH of high purity, high specific radioactivity, and preserved biologic potency, TSH interaction with intact cells and their derived plasma membranes was studied. At both the cellular and the sub-cellular level a very good agreement was found for the values of the rate and equilibrium constants of labeled TSH binding. A single type of high-affinity low-capacity site was revealed. In contrast, in both systems dissociation of bound labeled TSH was not of single-order kinetics and showed two kinetic components with half-lives of 3 and 30 min. An excellent correlation between half-stimulation of adenylate cyclase and iodide transport mechanism activation, and the dissociation constant of TSH binding, was found, indicating that the in vitro system studied was relevant to physiologic regulation.

Animals↗