Evidence for a TSH-controlled ectophosphotyrosine phosphatase in pig thyroid cultured cells.

In porcine thyroid cells in primary culture, thyrotropin (TSH) provokes important morphological changes associated with specific thyroidal function recovery. TSH influences cell-cell interactions and follicular morphogenesis. In the present report, we identify an ectotyrosine phosphatase activity controlled by chronic treatment with TSH. Specific substrates and various inhibitors allowed us to characterize this activity. This tyrosine phosphatase is located at the outer surface of thyroid cells as demonstrated by phosphotyrosylhistone hydrolysis. Control of cell viability and trypsin treatment confirm this extracellular localization. These data show that TSH activates ectotyrosine phosphatase activity, suggesting a role for surface protein phosphorylation in regulating specific functions of porcine thyroid cells in suspension.

Flavonoids extracted from fonio millet (Digitaria exilis) reveal potent antithyroid properties.

Digitaria exilis (fonio) is a tiny variety of millet commonly eaten by inhabitants of semiarid regions. A sample of fonio collected right in the middle of a severely iodine-depleted goitrous endemic was submitted to phytochemical investigations in order to assess the potential contributory roles played by vegetable molecules to the goitrogenic processes. The total content of flavonoids amounts to 500 mg/kg of the edible whole cereal grains. Their extraction and identification fail to detect the C-glycosylflavones described in other millet varieties but point out the presence of apigenin (A = 150 mg/kg) and of luteolin (L1 = 350 mg/kg). Ten percent of A and 80% of L1 are present in free form, whereas the remaining 90% of A and 20% of L1 are bound as O-glycosylflavones. Both A and L1 aglycones manifest strong anti-thyroid peroxidase (TPO) activities, resulting in a significant reduction of the hormonogenic capacity of this enzyme. In addition, L1 significantly depresses the cyclic AMP phosphodiesterase, implying a concomitant overproduction of the thyrotropin-dependent nucleotide. These last unreported data are regarded as counteracting to some extent the TPO-mediated goitrogenic properties of L1. Since fonio is devoid of other molecules likely to interfere with the thyroid function, our results are directly and casually attributed to A and L1 found in the customary diet.

Effects of sodium saccharin diet on fat-cell lipolysis: evidence for increased function of the adenylyl cyclase catalyst.

OBJECTIVE: To evaluate the effect of a 14 days' sodium saccharin (NaS) diet on lipolysis and cyclic-AMP accumulation in isolated rat white epididymal adipocytes. ANIMALS: Male Wistar rats (3 weeks old) were fed, for 14-days, ad libitum with a regular diet supplemented with or without 1-5% NaS dissolved in drinking water. MEASUREMENTS: Lipolysis and cAMP accumulation were assessed on isolated adipocytes. Adenylyl cyclase activities were measured on membrane fractions prepared from isolated adipocytes. The levels of Gs and Gi proteins were determined by Western blot analysis using specific antisera. RESULTS: Only high dietary NaS (5%) affected significantly the body growth and food consumption. Feeding rats with 2.5% of NaS increased both basal and isoproterenol-stimulated lipolysis and cAMP production. A 14-days diet of rats with 2.5% aspartame did not reproduce the effects of NaS on lipolysis and cAMP production. In fat-cell membranes of 2.5% NaS-treated rats, basal and stimulated-adenylyl cyclase activities were increased by 200% whatever the agonists used: GTP, GTP[S], [AIF4]-, isoproterenol or forskolin in the presence of Mg2+ or Mn2+ with or without GDP[S]. These effects cannot be explained by modifications of the expression of Gs and Gi proteins. The level of Gs alpha subunits was not affected by NaS treatment while the level of Gi alpha 1/2 was slightly increased. The stimulatory effect of NaS on adenylyl cyclase activity appears to be specific to adipocyte when compared with thyroid, brain or heart membrane fractions. CONCLUSION: Based on these data, and on the fact that cAMP regulates the lipolytic rate, we conclude that NaS diet increases lipolysis and cAMP formation in fat-cells by modifying the activity of the adenylyl cyclase catalyst(s).

Partial characterization of a glycosylphosphatidyl-inositol (GPI) in porcine thyroid cultured cells.

The aim of this study was to provide information on the structure of glycosylphosphatidylinositol (GPI) and to characterize this novel phospholipid isolated from pig thyroid. We investigated the incorporation of different radioactive precursors: [3H]glucosamine, [3H]inositol, [3H]oleic acid, [32P]orthophosphate and demonstrated the presence of these moieties in the structure of GPI. After labelling and hydrochloric acid hydrolysis, the major part of radioactivity comigrated with glucosamine used as marker. Cleavage of GPI by nitrous acid deamination indicated the presence of a glycosidic bond between the lipid moiety and glucosamine. The fatty acid composition of diacylglycerol was also studied by gas chromatography. Oleic acid was found preferentially to other fatty acids. In a previous paper we reported that a chronic treatment with 0.1 mU/ml thyrotropin (TSH) of thyroid cultured cells for 3 days produced a large increase in the turnover rate of GPI and concomitantly the release of a water-soluble product of GPI hydrolysis: an inositolphosphateglycan (IPG). These findings confirm that GPI may play a role in membrane signalling systems and thyroid cell regulation.

[Lipoprotein (a). An additional marker of atherosclerosis]. / La lipoprotéine (a). Un marqueur additionnel de l'athérosclérose.

Lipoprotein Lp(a) is a plasma lipoprotein which possesses many similarities to low density lipoprotein (LDL) in its physical and chemical properties. The major protein constituent of both lipoproteins is apolipoprotein B100 (apo B100); however, Lp(a) is unique in that it contains an additional distinct antigen, the (a)-antigen, attached to apo B100 by one or more disulphide bridges. The (a)-glycoprotein has recently been shown to have a striking amino-acid sequence homology with plasminogen; so, Lp(a) seems to be a potential bridge between the fields of atherosclerosis and thrombosis. Metabolic studies have made it clear that Lp(a) is not a product derived from other apo B-containing lipoproteins, but is secreted by the liver as a distinct mature lipoprotein. Although a relationship between elevated serum Lp(a) levels and the occurrence of atherosclerotic diseases had been postulated by several investigators, little is known today about the role of this lipoprotein and/or the mechanism whereby it might predispose to atheroma. However, the new knowledge on the structure of Lp(a) being more and more rapidly acquired, should facilitate the understanding of the mechanism of its atherogenicity and its physiopathological role.

Purification and characterization of phospholipase A2 inhibitory proteins from pig thyroid gland.

A 32 kDa phospholipase A2 inhibitory protein was isolated from pig thyroid gland after calcium precipitation and fast protein liquid anion-exchange chromatography. SDS-polyacrylamide gel electrophoresis revealed the purity of the protein. The protein activity was assessed by the inhibition of pancreatic phospholipase A2 on [3H]oleic acid-labelled Escherichia coli membranes as substrate and on the prostaglandin E2 production of cultured thyroid cells. The amino acid composition and the isoelectric point were quite similar to those of endonexin previously described in other tissues or cells. The cross-reactivity of a polyclonal antibody against a 32 kDa lipocortin from human peripheral blood mononuclear cells with our thyroidal 32 kDa protein confirmed its lipocortin nature. Before the purification by fast protein liquid chromatography, the Ca2+ pellet contained lipocortin I (35 kDa and its core protein 33 kDa) identified by its cross-reactivity with a polyclonal antibody.