Modulation of adenylate cyclase activity by sulfated glycosaminoglycans I. Inhibition by heparin of gonadotropin- stimulated ovarian adenylate cyclase

Heparin inhibits (I₅₀ = 2 μg/ml) the activity of luteinizing hormone and human chorionic gonadotropin-stimulated adenylate cyclase in purified rat ovarian plasma membranes. Unstimulated enzyme activity and activity stimulated by NaF, GTP or guanosine 5′-(β,γ-imido)triphosphate were inhibited to a lesser extent. Human chorionic gonadotropin binding to this membrane preparation was inhibited by hepatin (I₅₀ = 6 μg/ml). The inhibition with respect to hormone concentration was of a mixed type for hormone binding and adenylate cyclase stimulation. Inhibition by heparin was not eliminated at saturating hormone concentration. The degree of inhibition was unaffected by the order in which enzyme, hormone and heparin were introduced into the assay system. Herapin (3 μg/ml) did not affect the pH activity relationship of basal and hormone-stimulated adenylate cyclase activity and did not change the dependence of enzyme activity on magnesium ion concentration. The inhibitory action of heparin cannot be solely attributed to interference with either catalysis or hormone binding. The possibility is considered that the highly charged herapin molecule interferes with enzyme receptor coupling, by restricting the mobility of these components or by effecting their conformation.     

Ghrelin potentiates TSH-induced expression of the thyroid tissue-specific genes thyroglobulin, thyroperoxidase and sodium-iodine symporter, in rat PC-Cl3 Cells

Ghrelin is a 28-amino-acid peptide that stimulates pituitary growth-hormone secretion and modulates food-intake and energy metabolism in mammals. It is mainly secreted by the stomach, but it is also expressed in many other tissues such as cartilage or the thyroid gland. In the present study we have analyzed by RT-PCR and using immunohistochemistry and immunofluorescence the expression and tissue distribution of ghrelin and its functional receptor (GHS-R type 1α) in thyroid cell-lines and in normal and pathological rat thyroid tissue. Additionally, by measuring the incorporation of BrdU, we have investigated if, as previously noted for FRTL-5 cells, ghrelin enhances the proliferation rate in the PC-Cl3 rat-thyrocyte cell-line. Finally, we have determined the stimulatory effect of ghrelin on TSH-induced expression of the tissue-specific key genes involved in the synthesis of thyroid hormone: thyroglobulin, thyroperoxidase and sodium-iodine symporter. Our data provide direct evidence that C-cell secreted ghrelin may be involved in the paracrine regulation of the thyroid follicular cell function.     

Cat eye syndrome and growth hormone deficiency with pituitary anomalies: A case report and review of the literature

Cat eye syndrome is a rare congenital disease characterized by the existence of a supernumerary chromosome derived from chromosome 22, with a variable phenotype comprising anal atresia, coloboma of the iris and preauricular tags or pits. We report a girl with cat eye syndrome, presenting short stature, with growth hormone deficiency due to posterior pituitary ectopia. Short stature is a common feature of this syndrome, and the association with a structural pituitary anomaly has been described, however growth hormone deficiency and the underlying mechanisms are rarely reported. A review on short stature and growth hormone deficiency in cat eye syndrome is conducted.     

Differential distribution of orexin-A-like and orexin receptor 1 (OX1R)-like immunoreactivities in the Xenopus pituitary

Immunohistochemical techniques were employed to investigate orexin-A-like and orexin receptor 1 (OX1R)-like immunoreactivities in the Xenopus pituitary gland. Orexin-A-immunoreactive cells were mainly scattered in the posterior half of the pars distalis. They corresponded to thyroid-stimulating hormone (TSH)-containing cells and so far have not corresponded to other types of pituitary adenocytes. On the other hand, OX1R-immunoreactive cells were mainly distributed in the anterior half of the pars distalis and corresponded to prolactin (PRL)-containing cells; however, we found that OX1R-immunoreactive cells did not correspond to other types of adenocytes in the Xenopus pituitary. These results suggest that an orexin-A-like substance secretes with and/or without TSH from TSH-containing cells and that the peptide modulates the functions of PRL-containing cells via OX1R in a paracrine fashion.     

In vitro translation of distinct mRNAs coding for the precursors of porcine LH subunits

Conditions are described to characterize and estimate the precursors of porcine LH alpha and beta subunits and indirectly their specific mRNAs. Poly(A) RNAs extracted from castrated male pig anterior pituitaries were translated in a wheat-germ system in the presence of [35S] cysteine and [35S] methionine. The translation products were precipitated by antisera directed against reduced and carboxymethylated LH alpha and beta subunits and analyzed by high resolution electrophoresis. It is shown that the precursors of pLH alpha and beta subunits are located in two distinct congruent to 15 K proteins and represent--on the basis of the incorporation of the [35S] labeled aminoacids into proteins--congruent to 0.12% and 0.05% respectively of the total translation products. It is suggested that in the pig, as in other species, the LH alpha and beta subunits are encoded by two distinct mRNAs, and at variance with other species the leader sequence of LH alpha mRNA is longer than that of LH beta mRNA.     

The thyroid "microsomal" antigen is an epitope on the thyrotropin receptor

Antimicrosomal antibodies are present in the sera of most patients with autoimmune thyroiditis, and Graves' disease. It has, in general, been difficult to separate antimicrosomal activity from that directed against the thyrotropin (TSH) receptor in Graves' IgG preparations. The "microsomal" antigen has been localized to the endoplasmic reticulum and microfollicular aspect of thyrocytes; its structure is however unknown. In an attempt to identify the thyroid microsomal antigen, we studied the interaction of Hashimoto's IgG with high microsomal antibody titre and negative for thyroglobulin with purified thyroid plasma and light microsomal membranes. We allowed Hashimoto's, Graves', and control IgGs to bind to protein blots of thyroid plasma membranes resolved on SDS-PAGE under non-reducing conditions. All seven Hashimoto's IgG at a concentration of 2 mg/ml interacted with an M approximately 197,000 polypeptide corresponding to the TSH holoreceptor. By contrast to Graves' IgG (which were positive at 1 mg/ml), however, this binding was not blocked by pretreatment of the protein blots with TSH. Normal IgGs showed no binding at concentrations of up to 2 mg/ml. Both Hashimoto's and Graves' IgG interacted with TSH-affinity column-purified receptor preparations. Two of the Hashimoto's IgGs induced adenylate cyclase activation in thyroid plasma membranes, three inhibited TSH-stimulated enzyme activation, and two were without effect. Two classes of autoantibodies, other than TSH receptor directed, were encountered; one class raised to antigens common to all seven patients and another class unique to individual patients, eg, Mr 210,000 and Mr 20,000 polypeptides. We propose that the TSH receptor has multiple epitopes (functional domains), and the one to which antimicrosomal antibody bind is likely to be spatially separated from that with which Graves' IgG and TSH interact. Differences in affinity or number of sites allows for the demonstration of Graves' IgG against a background of antimicrosomal antibody.     

Effect of chronic desipramine treatment on neurotransmitter-thyrotropin releasing hormone—thyrotropin interactions in the rat

Long-term administration of the antidepressant drug, desipramine (20 mg/kg/day, orally for 28 days), decreased the stimulatory effect of the ₂-adrenoceptor agonist, clonidine (250 g/kg, i.p.) on thyrotropin (TSH) secretion in the rat, but did not alter basal TSH secretion. -Adrenoceptor-mediated inhibition of TSH secretion by isoproterenol (1 mg/ kg, i.p.) was unaffected by chronic desipramine treatment, as were the stimulatory effect of TSH-releasing hormone (TRH, 5 g/kg, i.v.) on TSH release and its inhibition by the -adrenoceptor antagonist, phentolamine (2 mg/kg, i.p.). These findings suggest that chronic desipramine treatment induces subsensitivity of ₂-adrenoceptors which modulate TSH secretion in the rat while not affecting -adrenoceptor-mediated inhibition of TSH release. These findings suggest that pituitary TRH receptors are unchanged but that changes occurred at the hypothalamic level in ₂-adrenoceptor-mediated stimulation of TRH release. Although cerebral -adrenoceptors have been shown convincingly to be down-regulated after chronic desipramine treatment, their function in the hypothalamic TRH system after 28 days of treatment with desipramine appears to be unimpaired.     

Chronobiological Aspects of Weight Loss in Obesity: Effects of Different Meal Timing Regimens

A series of short- and long-lasting experimental protocols of different meal timing regimes were performed in obese subjects to assess the possible occurrence of (1) a different metabolic fate of nutrients; (2) a phase shift of circadian rhythms of metabolic and hormonal parameters strictly related to nutrition; (3) a different weight loss. (A) In a short-lasting protocol (3 days) IS obese subjects were fed a hypocaloric diet (684 kcal/day) (a) at 10 hr only, (b) at 1800 hr only; (c) at 1000 hr, 1400 hr and 1800 hr, or (d) studied during a 36-hr fasting. Measures of calorimetry (R.Q., CHO and lipid oxidations, energy expenditure), hormones (plasma Cortisol, insulin, HGH, urinary catecholamines), urinary electrolytes (Na, K) and vital parameters (body temperature, heart rate, blood pressure) were carried out at 4-hr intervals for three days. A significantly higher lipid oxidation and a lower CHO oxidation were documented with the meal at 1800 hr, in comparison with the meal at 1000 hr. CHO and lipid oxidation circadian rhythms appeared the most affected by meal timing. (B) In a long-lasting protocol (18 days) 10 obese subjects were fed the same hypocaloric diet (a) at 1000 hr only and (b) at 1800 hr only. Calorimetric measures were performed every other day for 2 hr preceding each meal. Before and after the 18-days single meal period, body temperature, plasma Cortisol, PRL and TSH were recorded (δt = 4 hr). A higher lipid oxidation and a lower CHO oxidation were again demonstrated with the meal at 18 hr. Minimal changes of hormonal circadian rhythms were documented suggesting that the hypothalamus-hypophysis network is scarcely affected by meal timing. Weight loss did not vary in both short- and long-term protocol.