concanavalin-a and Pituitary-Neoplasms

This study analyzed the structural differences of the carbohydrate chains of circulating free alpha-submit (alpha-SU) hypersecreted in various non-tumoral (primary hypothyroids, postmenopausal women, patients with chronic uremia, normal fetuses) and tumoral (gut carcinoids, TSH-, GH- and pure alpha-secreting pituitary adenomas) clinical conditions. Carbohydrate structures of free alpha-SU were investigated by means of lectin affinity chromatography using Concanavalin A (Con-A), which allows the separation of free alpha-SU in three different fractions (unbound = UB, weakly bound = WB and firmly bound = FB) depending on the nature and maturation of glycosylated chains. The concentrations of alpha-SU in serum and in Con-A fractions were measured by a sensitive and specific IRMA. Free alpha-SU hypersecreted from postmenopausal women, primary hypothyroids, and patients with chronic uremia showed similar binding patterns to Con-A, the percentage of UB fractions (UB: 44.5 +/- 1.9%, 39.5 +/- 3.8%, 48.2 +/- 5.6% respectively) being higher than both WB and FB fractions (WB: 33.2 +/- 1.4%, 30.7 +/- 4.6%, 28.5 +/- 2.1%; FB: 22.3 +/- 0.7%, 29.8 +/- 6.6%, 23.3 +/- 4.2% respectively). In normal fetuses the amount of UB fraction was very high (UB: 70.7 +/- 5.4%). Free alpha-SU from patients with TSH- and GH-secreting adenomas showed a binding pattern to Con-A significantly different from that observed in postmenopausal women taken as controls, the WB fractions being significantly higher (WB: 56.9 +/- 16.8% and 71 +/- 12.4% respectively, P < 0.001). A typical pattern of elution on Con-A, characterized by a prevalence of immature alpha-SU molecules eluted in the FB fraction, was found in patients with pure alpha-secreting adenomas. This chromatographic behavior was significantly different from that seen in the controls, as well as in other pituitary tumors and in gut carcinoids (FB: 41.8 +/- 5.0%, 22.3 +/- 0.7%, 16.8 +/- 6.6%, 10.6 +/- 2.0% respectively). Moreover, in these latter patients the pattern of free alpha-SU binding was exactly the opposite of that observed in pure alpha-secreting adenomas, with a prevalence of mature alpha-SU molecules (UB: 59.1 +/- 4.4 vs 18.3 +/- 7.2%). In conclusion, our data on Con-A affinity chromatography clearly demonstrate that carbohydrate branching of circulating free alpha-SU varies in patients with pituitary adenomas as compared with patients with gut carcinoids or other non-tumoral conditions. Moreover, the finding of a greater prop

Topics: Adenoma; Adult; Carcinoid Tumor; Chromatography, Affinity; Concanavalin A; Fetus; Glycoconjugates; Glycoprotein Hormones, alpha Subunit; Humans; Hypothyroidism; Intestinal Neoplasms; Pituitary Neoplasms; Postmenopause; Protein Binding; Uremia

Carbohydrate structures of intrapituitary and circulating TSH were studied by Concanavalin-A (Con A) and ricin lectin chromatography under different clinical conditions. Con A permits the separation of molecules differing in the extent of their carbohydrate branching, whereas ricin gives an estimation of the degree of their sialylation. Intrapituitary TSH was more retained on Con A and less sialylated than circulating hormone, suggesting that carbohydrate chains of intrapituitary molecules are less mature than those present in the circulation. A greater proportion of TSH firmly bound to Con A, compared to control values, was found in sera from fetuses and patients with uremia, TSH-secreting adenomas, and central hypothyroidism. In primary hypothyroid patients, TSH binding to Con A was similar to that found in controls, but a greater percentage of sialylated forms was seen. In central hypothyroidism patients, TSH released in response to TRH was less sialylated. Interestingly, no sialylated TSH was found in normal fetuses. In conclusion, the present data show that both TSH carbohydrate branching and sialylation may vary in different clinical conditions. As some of the above clinical conditions are known to be accompanied by variations in the bioactivity of circulating TSH, the finding of changes in TSH carbohydrate structures further supports the view that glycosylation modulates the expression of TSH biological activity.

Topics: Adenoma; Carbohydrates; Chromatography, Affinity; Concanavalin A; Female; Fetus; Humans; Hypothyroidism; Isoelectric Focusing; Male; Molecular Structure; Pituitary Gland; Pituitary Neoplasms; Pregnancy; Ricin; Thyrotropin; Uremia

Pituitary tissues were investigated for the presence of regulatory molecules that would alter the function of lymphoid cells. A novel endogenous polypeptide inhibitor of basal and mitogen-stimulated splenocyte DNA synthesis and proliferation, suppressin, was isolated from bovine pituitary glands. Suppressin is a potent inhibitor of basal and mitogen-stimulated splenocyte proliferation at picomole and nanomole concentrations with 50% inhibition occurring 2.8 x 10(-9) M. Suppressin was purified to apparent homogeneity using sequential (NH4)2SO4 precipitation, ion-exchange chromatography, and preparative native gel electrophoresis. Biochemical characterizations of suppressin showed that this inhibitory molecule was a monomeric polypeptide with (i) a Mr = 63,000 and (ii) a pI of 8.1. Finally, metabolic labeling studies using a rat pituitary tumor cell line, GH3, showed that suppressin was synthesized de novo and secreted by these cells.

Topics: Ammonium Sulfate; Animals; Antibodies; Antigens; B-Lymphocytes; Cattle; Cell Division; Chemical Precipitation; Chromatography, Ion Exchange; Concanavalin A; DNA; Electrophoresis, Polyacrylamide Gel; Lipopolysaccharides; Lymphocytes; Molecular Weight; Peptides; Pituitary Gland; Pituitary Neoplasms; Spleen; T-Lymphocytes; Tumor Cells, Cultured

Electron-immunocytochemical staining with lectin (concanavalin A: Con A) binding sites analysis was applied to study secretory granules of human pituitary adenomas and surrounding normal pituitary tissue using post-embedded serial ultrathin sections. Twelve cases of human pituitary adenoma and three specimens of normal pituitary tissue surrounding adenomas were studied: the cases were operated on between 1982 and 1984. The tumors consisted of four prolactin (PRL)-, six growth hormone (GH)-, and two adrenocorticotropic hormone (ACTH)-producing adenomas. In parallel with the detection of Con A binding sites of secretory granules, their secreting hormones were characterized electron-microscopically with the immunocytochemical horseradish peroxidase (HRP) labeling using the avidin-biotin technique. The two cases of ACTH-producing adenomas showed either weak or negative reactions with Con A on secretory granules, while normal ACTH-producing pituitary cells showed strong reactions with Con A on every secretory granule observed. Large secretory granules of PRL- or GH-producing cells showed negative reactions with Con A both in the pituitary adenoma and normal pituitary, while some small granulated or sparsely granulated adenoma cells also showed strong reactions with Con A. The complexity of human pituitary adenomas is illustrated as well as the difference in biochemical structure of normal pituitary cells and pituitary adenoma cells secreting the same specific hormone.

Topics: Adenoma; Adrenocorticotropic Hormone; Binding Sites; Concanavalin A; Cytoplasmic Granules; Growth Hormone; Histocytochemistry; Horseradish Peroxidase; Humans; Immunochemistry; Microscopy, Electron; Pituitary Gland; Pituitary Neoplasms; Prolactin; Reference Values

Molecular forms of immunoreactive adrenocorticotropin (ACTH), beta-lipotropin (beta-LPH) (beta-endorphin (beta-END), human NH-2-terminal (hNT) of pro-opiomelanocortin (POMC), and gamma-3-melanotropin (gamma-3-MSCH) were studied in plasma, CSF and urine of a patient with Nelson's syndrome by molecular sieving and concanavalin A (Con A)-sepharose chromatography. In the culture tumor medium of the tumor cells, and in the plasma and CSF, these compounds were found mainly in forms corresponding in molecular weight to the authentic peptides, with the exception of gamma-3-MSH. Stimulation of the pituitary tumor by synthetic ovine corticotropin-releasing factor (CRF 1-41) caused a 171-468% increase in vivo (60 min) and 453-953% increase in vitro (3h incubation) in the levels of POMC derived peptides; it increased the relative amount of beta-END in vivo, and that of beta-LPH in vitro. Molecular sieving chromatography of urine samples revealed that beta-LPH and hNT are extensively degraded by the kidney. By contrast, ACTH showed no significant renal degradation before the removal of the pituitary adenoma. However, following pituitary surgery, only smaller fragments of immunoreactive (IR) ACTH were detected in the urine. These results suggest no major abnormal metabolic pathway for POMC in Nelson's syndrome, although the proportions of various peptides derived from the precursor could be different in vivo from those after in vitro incubation under basal conditions and during CRF stimulation. The results also indicate differences in the renal handling of ACTH in POMC hypersecretory states.

Topics: Adrenocorticotropic Hormone; beta-Endorphin; beta-Lipotropin; Body Fluids; Chromatography; Concanavalin A; Endorphins; Female; Humans; Middle Aged; Nelson Syndrome; Peptide Fragments; Peptides; Pituitary Neoplasms; Pro-Opiomelanocortin; Radioimmunoassay

The lectin-binding properties of serum alpha subunit were studied by lectin affinity chromatography. Normal individuals and most patients with pituitary tumours produced alpha subunit which bound specifically to Concanavalin A-Sepharose (Con A). Some patients with pituitary tumours produced both Con A-reactive alpha subunit and alpha subunit which did not bind to Con A. Concanavalin A-Sepharose-binding alpha subunit from all sources bound strongly to Ricinus communis agglutinin-Sepharose after treatment with neuraminidase. Serum alpha subunit from those patients with pituitary tumours, which did not bind to Con A, bound to wheat germ agglutinin-Sepharose, exhibiting both weakly binding and strongly binding forms. Serum alpha subunit from both patients and controls, which did bind to Con A, showed only weak affinity for wheat germ agglutinin-Sepharose. Neither the low affinity nor the high affinity of serum alpha subunit from any source for wheat germ agglutinin-Sepharose was affected by neuraminidase. These findings show that (a) the predominant pattern of glycosylation of serum alpha subunit from normal controls is a Con A-reactive, biantennate complex oligosaccharide and (b) that the structural alteration which results in serum alpha subunit which does not bind to Con A in some patients with pituitary tumours is not an absence of carbohydrate, rather the alpha subunit contains highly branched, either complex or hybrid oligosaccharides.

Topics: Adult; Chromatography, Affinity; Chromatography, Gel; Chromatography, Ion Exchange; Concanavalin A; Female; Glycoprotein Hormones, alpha Subunit; Humans; Lectins; Male; Middle Aged; Oligosaccharides; Peptide Fragments; Pituitary Hormones, Anterior; Pituitary Neoplasms; Plant Lectins; Protein Binding; Sepharose; Wheat Germ Agglutinins

Most mammalian cells are capable of growth in culture only when they are supplied with an appropriate substrate to which they can adhere and spread. To prepare suitable substrates different lectins were attached onto polystyrene tissue-culture dishes after coating with polylysine. GH3-cells (a pituitary-tumor-cell line) were seeded into the culture dishes containing serum-free, hormone-supplemented medium. When succinylated Concanavalin A (s-Con A), which binds specifically to mannose residues, is attached to the surface an extraordinary spreading of GH3-cells is induced within 15 to 20 min after seeding. Other lectins with a different sugar-binding specificity are less effective in inducing cell spreading. However, the cell spreading depends not only on the substrate-attached lectins but also on the hormones used in the proliferation-culture of GH3-cells. Both types of molecules found in the microenvironment of a cell, the matrix-fixed sugar-binding proteins and the diffusive hormones, are responsible for the regulation of the behaviour of the mammalian cell. It is suggested that the interaction of the cell-surface carbohydrates with the plasma-membrane-bound lectins of contiguous cells plays a central role in such processes, especially in in-vivo.

Topics: Animals; Cell Adhesion; Cell Line; Cell Movement; Concanavalin A; Culture Media; Hormones; Kinetics; Lectins; Mannose; Pituitary Neoplasms; Structure-Activity Relationship