Mucous Release Research Articles

Temporal dynamics of epidermal responses of guppies Poecilia reticulata to a sublethal range of waterborne zinc concentrations

This study assessed the histological changes in the epidermis of guppies Poecilia reticulata induced by waterborne zinc (Zn). Laboratory-reared P. reticulata fry were maintained individually in separate vessels containing artificial water (8 microg l(-1) Zn) to which 0, 15, 30, 60 or 120 microg l(-1) Zn was added. Their epidermal response to Zn was monitored regularly over 4 weeks. Compared with controls, mucus was rapidly released and mucous cell numbers decreased at all concentrations. Thereafter mucous release, epidermal thickness, numbers and size of mucous cells fluctuated at a rate that varied with Zn concentration, but fluctuations declined after day 18. Results clearly highlight the dynamic nature of the epidermal response to sublethal concentrations of waterborne Zn. In general, low concentrations of Zn induced a rapid response with reduced numbers and size of mucous cells and shift in mucin composition, and a subsequent thickening of the epidermis. Epidermal thickness and mucous cell area fluctuated over time but were normal after a month of exposure to low Zn concentrations. The number of mucous cells, however, remained low. Virtually all mucous cells from fish maintained in 15 and 60 microg l(-1) Zn contained acidic mucins throughout the month, whereas fish maintained at 30 microg l(-1) Zn responded by production of neutral mucins during the first 12 days followed by a mixture of neutral and acidic mucins. At 120 microg l(-1) Zn, the most dramatic effects were the gradual but sustained decrease in numbers and area of mucous cells, and the shift to acidic mucins in these cells. Thus, as concentration of Zn increased, the epidermal responses indicated a disturbed host response (dramatic decline in mucous cell numbers, with mixed composition of mucins), which may have been less effective in preventing Zn uptake across the epithelium.

In vitro evidence that cortisol directly modulates stress-related responses in the skin epidermis of the rainbow trout (Oncorhynchus mykiss Walbaum)

Exposure of fish to stressors leads to multiple changes in the skin epithelium. We investigated the role of the stress hormone cortisol in the control of these changes by exposure of pieces of skin from the rainbow trout Oncorhynchus mykiss with an in vitro tissue culture incubation procedure. The effects of 24 h exposure to 4 cortisol concentrations (0, 50, 500 and 1000 ng/ml) were determined. Numbers of mucous, mitotic and apoptotic cells were quantitatively assessed using immunohistochemical techniques, in situ DNA nick end labelling (TUNEL), as well as conventional light and scanning electron microscopy (SEM). The cortisol receptor blocker mifepristone was used to investigate whether the effects could be attributed to the direct action of the hormone via glucocorticoid receptors. Overall, cortisol had no effect on the mucous cell population at 24 h. Incubation with the receptor blocker reduced the number of mucous cells. Cell proliferation was stimulated by the addition of 50 and 500 ng/ml cortisol, but not at 1000 ng/ml. Incubation with the receptor blocker increased proliferation in the control group (0 ng/ml) only. An increase in apoptosis occurred at 500 and 1000 ng/ml cortisol. This increase was blocked by incubation with the receptor blocker, which resulted in lower numbers of apoptotic cells in all except the 0 ng/ml controls. SEM observations corroborated the quantitative data. The results indicated that the effects of stressors on the fish epidermis mentioned above are mediated by cortisol, with the exception of mucous cell release.

Identification of sugar residues in secretory glycoconjugates of the lining and glandular epithelium of the chick embryo gizzard using lectin histochemistry

Using seven horseradish peroxidase-conjugated lectins we studied the distribution of glycoconjugate sugar residues in the lining and glandular epithelium of the gizzard in chick embryos from day 7 to 21 of incubation. At the outer layer of the lining epithelium all investigated sugar residues were present from day 7 onwards, except alpha-L-fucose, which was detected later. From day 9 some cells, which contained granules characterised by D-galactose-(beta 1--->3)-N-acetyl-D-galactosamine and N-acetyl-D-glucosamine, were observed in the inner part of the lining epithelium. Afterwards, although in different time periods, all investigated oligosaccharides were detected in intercellular spaces filled with mucous material. At first, the cells of the anlage of the tubular glands appeared to be characterised by D-glucosamine and alpha-D-mannose only. Afterwards, all the investigated sugar residues were detected. At hatching, the luminal glandular secretion showed all investigated sugar residues, except alpha-L-fucose, which was typical for the early formation of the tubular glands. Our data suggest that during the development of the gizzard three time periods of mucous release can be distinguished.

Endothelin in Human Nasal Mucosa

Endothelin (ET), a potent vasoconstrictor and bronchoconstrictor peptide synthesized by endothelial and epithelial cells, was examined for its potential functions in human inferior turbinate nasal mucosal tissue by four techniques: (1) immunoreactive ET was localized in the mucosa by immunohistochemistry; (2) receptors for ET were identified by autoradiography employing [125I]ET; (3) ET-1 mRNA was localized by in situ hybridization; and (4) the secretory functions of ET were examined by the release of mucous and serous cell products after the addition of ET to human nasal turbinates in short-term cultures. Specific ET-1-immunoreactive material was found most extensively in small muscular arteries and in serous cells in submucosal glands. ET-1 was also found to a lower extent in the walls of venous sinusoids. [125I]ET-1 binding sites were localized by autoradiography to submucosal glands and to venous sinusoids and small muscular arterioles. mRNA for ET-1 was found most extensively in the venous sinusoids and to a lesser extent in small muscular arteries. In mucosal explant cultures, ET-1 and ET-2 stimulated lactoferrin and mucous glycoprotein release from serous and mucous cells, but ET-3 was inactive. The observations indicate that in the human nasal mucosa, ET is present in the vascular endothelium and the serous cells in submucosal glands and acts on glandular ET receptors to induce both serous and mucous cell secretion. It is also likely that ET plays a role in the regulation of vasomotor tone.

Localization of vasoactive intestinal polypeptide-receptor-immunoreactivity in human salivary glands

A monoclonal VIP-receptor antibody derived from a human adenocarcinoma cell line (HT 29) was used in combination with immunogold silver staining for the immunohistochemical demonstration of VIP-receptor (rec) immunoreactivity (IR) in paraffin-embedded human salivary glands (parotid, palatal, labial glands). VIP-rec-IR was localized to mucous endpieces of labial and—to a lesser extent—palatal glands, intercalated ducts of the parotid gland, and excretory ducts of all glands investigated. The findings correlate well with known effects of VIP on mucous release and electrolyte transport in salivary glands. Lack of VIP-rec-IR at serous acini may point to immunologically different receptor subtypes in these glands.

The effects of corticosteroids on mucous glycoprotein secretion from human airways in vitro.

In order to examine the mechanisms by which corticosteroids may benefit some patients with bronchorrhea, cultured human airways releasing [3H]glucosamine labeled mucous glycoproteins were exposed to corticosteroids, and mucus release was examined. Both dexamethasone and methylprednisolone produced dose-related suppression of the spontaneous release of radiolabeled mucous glycoproteins. The inhibitory effects of dexamethasone were maximal after 18 to 24 h and returned to control levels by 34 h. In order to study the effects of dexamethasone on stimulated mucus release, airways were exposed to dexamethasone and to the mucus secretagogues, histamine or 5-monohydroxyeicosatetraenoic acid. Both of these secretagogues stimulated radiolabeled mucous glycoprotein release from airways that had never been exposed to corticosteroids, as well as in a reduced fashion from corticosteroid-treated airways. The reduced mucus release caused by secretagogues from dexamethasone-treated airways appeared to reflect a lowered baseline secretion rate rather than a specific inhibition of either secretagogue.

Slow-reacting substances, leukotrienes C4 and D4, increase the release of mucus from human airways in vitro.

Allergic pulmonary reactions in vivo lead to airway constriction and mucous secretion, whereas in vitro lung anaphylaxis lead to mediator release and increased mucous glycoprotein secretion from cultured human airways. Using quantitation of radiolabeled mucous glycoprotein from cultured airways as a model for mucous release, the effects of two leukotrienes, LTC4 and LTD4, were assessed. BOth biosynthetic and synthetic LTC4 and LTD4 produced dose-related increases in mucous production at concentrations of 1 to 1,000 units/ml (20 to 20,000 pg [LTD4] and 24 to 24,000 pg [LTC4]/ml). These enhancing actions were seen in 11 of 11 lung cultures, and were significantly prevented by the specific SRS-A antagonist FPL 55712. Therefore, LTC4 and LTD4, two major components of the allergic mediator SRS-A, are potent mucous secretagogues and may possibly contribute to the mucous secretion observed during allergic reactions in vitro and in vivo.

Effects of arachidonic acid, monohydroxyeicosatetraenoic acid and prostaglandins on the release of mucous glycoproteins from human airways in vitro.

Human lung explants maintained in culture for 7 d incorporate [(3)H]glucosamine into mucous glycoproteins. Ethanol-precipitable, glucosamine-labeled mucous secretion was measured, and the effects of different pharmacologic agents upon this secretion were investigated. Anaphylaxed human lung generates prostaglandin (PG) synthesis and increased mucous release. Arachidonic acid (AA), PGA(2), PGD(2), and PGF(2alpha) significantly increased mucous glycoprotein release, whereas PGE(2) significantly reduced release. Evidence which suggests that lipoxygenase products of AA augment mucous release includes the following: (a) Nonsteroidal anti-inflammatory drugs (NSAID: acetylsalicylic acid and indomethacin) increase mucous release while preventing prostaglandin formation. (b) The increase in mucous release induced by AA or NSAID is additive once the agents are combined. (c) Several nonspecific lipoxygenase inhibitors (eicosa-5,8,11,14-tetraynoic acid; vitamin E; nordihydroguaiaretic acid; and alpha-naphthol) inhibit mucous release. Three additional lines of evidence directly indicate that monohydroxyeicosatetraenoic acid (HETE) causes increased mucous release: (a) the addition of a mixture of synthetic HETE (24-600 nM) increases mucous release; (b) pure 12-HETE (1-100 nM) also increases mucous release; (c) mucous release is increased synergistically by the combination of HETE and NSIAD. These data taken together demonstrate that HETE are capable of increasing mucous release and that conditions which may influence HETE production alter mucous release. Thus, although not directly demonstrating HETE production by human airways, the data strongly suggest that lipoxygenase products of AA in airways may profoundly influence mucous release; and it seems possible that lipoxygenase inhibitors may have a role in treating bronchorrhea.

Immunologic and neuropharmacologic stimulation of mucous glycoprotein release from human airways in vitro.

Human bronchial airways obtained after surgical resection were maintained in tissue culture for 24-48 h. Incorporation of [(3)H]- or [(14)C]-glucosamine, [(14)C]threonine, or Na(2)[(35)S]O(4) to the culture media resulted in biosynthesis of two radiolabeled glycoproteins-one filtering in the exclusion volume of Sepharose 2B, and the other filtering with an approximate molecular weight of 400,000. Both fractions had similar elution patterns from DEAE-cellulose anion exchange chromatography. [(3)H]Glucosamine was incorporated equally into the two fractions. The effects of anaphylaxis, histamine, and several neurohormones upon the release of [(3)H]glucosamine-labeled glycoproteins were analyzed, making no attempt to separate the two glycoprotein fractions. Three lines of evidence were found suggesting that mast-cell degranulation increases mucous release from cultured airways. (a) Supernatant fluids from anaphylaxed peripheral human lung that contained 200-400 ng/ml histamine and 400-1,000 U/ml slow-reacting substance of anaphylaxis (SRS-A) increased release by 40+/-18%. (b) The addition of antigen to IgE-sensitized airways led to the release of 26+/-7% of the total histamine and a 36+/-14% increase in mucous release. (c) Reversed anaphylaxis with anti-IgE antibodies induced a 36+/-6% release of histamine from the airways and an increase in the release of mucous glycoproteins of 25+/-9%. Exogenous histamine added to airways increased mucous glycoprotein release, an effect prevented by cimetidine, an H-2 antagonist. Selective histamine H-2, but not H-1 agonists increased mucous glycoprotein release, suggesting the possibility that anaphylaxis of airways results in increased mucous glycoprotein release partly through histamine H-2 stimulation.A cholinomimetic agonist, methacholine, increased mucous release; this response was prevented by atropine which alone had no effect. No response to beta-adrenergic stimulation with either isoproterenol or epinephrine was noted. However, alpha-adrenergic stimulation with either norepinephrine combined with propranolol or phenylephrine alone resulted in dose-related increases in glycoprotein release. Both alpha-adrenergic and cholinergic stimulation of human tissues induce the formation of guanosine 3',5'-phosphoric acid (cyclic GMP), and 8-bromo cyclic GMP added to the airways led to increased mucous secretion. Thus, it seems likely that neurohormones capable of stimulating cyclic GMP formation in human airways may lead to increased mucous glycoprotein release.

Efect of adrenergic stimilation on ion transport across skin of living frogs

1. 1. The influence of the intravenous administration of catecholamines and a β adrenergic blocking agent on short-circuit current ( I s ) skin resitance and sodium flux was studied with living anesthetized frog. 2. 2. Epinephrine (α plus β adrenergic stimulation) adn isoproterenol (β stimulation) produced an increase of I s , which was not explained by sodium transport changes, and mucous release. 3. 3. In contrast to epinephrine, isoproterenol decreased skin resistance and increased sodium outflux. 4. 4. Pronethalol (a β adrenergic blocking agent) decreased I 3, net sodium flux, and increased skin resistance. 5. 5. The results indicate that ion transport across the skin, in vivo, is influenced by α and β adrenergic stimulation. They support the concept that the two stimuli produce opposing effects on sodium transport, i.e. α stimulation decreases and β stimulation increases sodium permeability of the skin.