Coexistence of gland mucous cell-type mucin and lysozyme in gastric gland mucous cells

Class III mucin, identified by paradoxical concanavalin A staining, is confined to gastric gland mucous cells and is an essential component of the gastric surface mucous gel layer. The pretreatment required has hampered the application of this method to electron microscopic studies. Antibody HIK1083 reacts selectively with class III mucins. The present study was undertaken to explore, electron microscopically, the immunoreactivity of the human stomach to HIK1083. We examined normal mucosa from resected human stomachs (five cases; formalin-fixed, paraffin-embedded) and gastric biopsy specimens from patients with early gastric cancer [nine cases; glutaraldehyde- and osmium-fixed, epoxy-embedded (seven cases) and half-strength Karnovsky’s solution-fixed, Lowicryl K4M-embedded (two cases)]. Immunostaining with HIK1083 and anti-lysozyme antibody was examined under light and electron microscopes. Gland mucous cells were labeled with HIK1083, and lysozyme was detected in some gland mucous cells and surface mucous cells. Electron microscopically, the secretory granules of gland mucous cells contained a single electron-dense core. HIK1083-positive mucins and lysozyme coexisted in the secretory granules of gastric gland mucous cells. HIK1083-reactive mucins and lysozyme were distributed in the matrix and in the dense core of these secretory granules, respectively. HIK1083 can be used for electron immunohistochemistry. Accepted: 1 December 1999     

Cystic fibrosis and the relationship between mucin and chloride secretion by cultures of human airway gland mucous cells

We investigated how cystic fibrosis (CF) alters the relationship between Cl(-) and mucin secretion in cultures of non-CF and CF human tracheobronchial gland mucous (HTGM and CFTGM, respectively) cells. Biochemical studies showed that HTMG cells secreted typical airway mucins, and immunohistochemical studies showed that these cells expressed MUC1, MUC4, MUC5B, MUC8, MUC13, MUC16, and MUC20. Effects of cumulative doses of methacholine (MCh), phenylephrine (Phe), isoproterenol (Iso), and ATP on mucin and Cl(-) secretion were studied on HTGM and CFTGM cultures. Baseline mucin secretion was not significantly altered in CFTGM cells, and the increases in mucin secretion induced by mediators were unaltered (Iso, Phe) or slightly decreased (MCh, ATP). Across mediators, there was no correlation between the maximal increases in Cl(-) secretion and mucin secretion. In HTGM cells, the Cl(-) channel blocker, diphenylamine-2-carboxylic acid, greatly inhibited Cl(-) secretion but did not alter mucin release. In HTGM cells, mediators (10(-5) M) increased mucin secretion in the rank order ATP > Phe = Iso > MCh. They increased Cl(-) secretion in the sequence ATP > MCh ≈ Iso > Phe. The responses in Cl(-) secretion to MCh, ATP, and Phe were unaltered by CF, but the response to Iso was greatly reduced. We conclude that mucin secretion by cultures of human tracheobronchial gland cells is independent of Cl(-) secretion, at baseline, and is unaltered in CF; that the ratio of Cl(-) secretion to mucus secretion varies markedly depending on mediator; and that secretions induced by stimulation of β-adrenergic receptors will be abnormally concentrated in CF.     

Signal transduction in eclosion hormone-induced secretion of ecdysis-triggering hormone

Inka cells of insect epitracheal glands (EGs) secrete preecdysis and ecdysis-triggering hormones (PETH and ETH) at the end of each developmental stage. Both peptides act in the central nervous system to evoke the ecdysis behavioral sequence, a stereotype behavior during which old cuticle is shed. Secretion of ETH is stimulated by a brain neuropeptide, eclosion hormone (EH). EH evokes accumulation of cGMP followed by release of ETH from Inka cells, and exogenous cGMP evokes secretion of ETH. The secretory responses to EH and cGMP are inhibited by the broad-spectrum kinase inhibitor staurosporine, and the response to EH is potentiated by the phosphatase inhibitor calyculin A. Staurosporine did not inhibit EH-evoked accumulation of cGMP. Changes in cytoplasmic Ca2+ in Inka cells during EH signaling were monitored via fluorescence ratioing with fura-2-loaded EGs. Cytoplasmic Ca2+ increases within 30-120 s after addition of EH to EGs, and it remains elevated for at least 10 min, corresponding with the time course of secretion. Secretion is increased in dose-dependent manner by the Ca2+-ATPase inhibitor thapsigargin, a treatment that does not elevate glandular cGMP above basal levels. The secretory response to EH is partially inhibited in glands loaded with EGTA, while cGMP levels are unaffected. These findings suggest that EH activates second messenger cascades leading to cGMP accumulation and Ca2+ mobilization and/or influx and that both pathways are required for a full secretory response. cGMP activates a staurosporine-inhibitable protein kinase. We propose that Ca2+ acts via a parallel cascade with a time course that is similar to that for cGMP activation of a cGMP-dependent protein kinase.     

Signal transduction in plant cells.

In plants, unlike animals, signal transduction studies are in their infancy. While intracellular Ca2+ appears to have second messenger functions, attempts to show that protein kinases, inositol phosphates and cyclic AMP are involved in signal transduction in plants have run into considerable difficulty.     

Chloride secretion by cultures of pig tracheal gland cells

Malfunction of airway submucosal glands contributes to the pathology of cystic fibrosis (CF), and cell cultures of CF human airway glands show defects in Cl(-) and water transport. Recently, a transgenic pig model of CF (the CF pig) has been developed. Accordingly, we have developed cell cultures of pig airway gland epithelium for use in investigating alterations in gland function in CF. Our cultures form tight junctions (as evidenced by high transepithelial electrical resistance) and show high levels of active anion secretion (measured as amiloride-insensitive short-circuit current). In agreement with recent results on human airway glands, neurohumoral agents that elevate intracellular Ca(2+) potently stimulated anion secretion, while elevation of cAMP was comparatively ineffective. Our cultures express lactoferrin and lysozyme (serous gland cell markers) and MUC5B (the main mucin of airway glands). They are, therefore, potentially useful in determining if CF-related alterations in anion transport result in altered secretion of serous cell antimicrobial agents or mucus.     

基因转录爆发的信号转导机制

随着单分子检测技术水平的提高,学术界发现了与传统认知大相径庭的转录现象——转录爆发。基因转录不是连续平稳且波动较小的过程,而是表现出发生时间极不规律、信使RNA产量波动极大的特征。转录爆发本质上是编码和传递调控信号的方式。本文综述了转录爆发现象、转录爆发的两态模型、转录爆发信号转导机制以及其重要的生物学意义。最后,指出了该领域亟待解决的问题。     

Signal transduction by mitochondrial oxidants

The production of mitochondrial reactive oxygen species occurs as a consequence of aerobic metabolism. Mitochondrial oxidants are increasingly viewed less as byproducts of metabolism and more as important signaling molecules. Here, I review several notable examples, including the cellular response to hypoxia, aspects of innate immunity, the regulation of autophagy, and stem cell self-renewal capacity, where evidence suggests an important regulatory role for mitochondrial oxidants.     

G regulatory proteins and muscarinic receptor signal transduction in mucous acini of rat submandibular gland

The involvement of G regulatory proteins in muscarinic receptor signal transduction was examined in electrically permeabilized rat submandibular acinar cells. The guanine nucleotide analog, GTP gamma S, caused the dose dependent hydrolysis of membrane phosphatidylinositol 4,5-bisphosphate to release IP3. This response was insensitive to pertussis toxin treatment and was duplicated by NaF but not by GDP beta S. Enhanced IP3 synthesis was observed with a combination of GTP gamma S and carbachol. Exogenous IP3, as well as carbachol and GTP gamma S, provoked the release of sequestered 45Ca2+ from non-mitochondrial stores. In intact cells, carbachol significantly reduced the level of cyclic AMP induced by the beta-adrenergic agonist, isoproterenol, to 69% of its normal value. Pertussis toxin abolished this inhibitory action of carbachol on cyclic nucleotide levels. These results suggest that muscarinic receptors are coupled to two separate G regulatory proteins in submandibular mucous acini-the pertussis toxin-insensitive Gp of the phosphoinositide transduction pathway associated with elevated cytosolic calcium levels, and the pertussis toxin-sensitive Gi inhibitory protein of the adenylate cyclase complex.     

Signal transduction mechanisms mediating secretion in goldfish gonadotropes and somatotropes.

The intracellular signal transduction mechanisms mediating maturational gonadotropin and somatotropin secretion in goldfish are reviewed. Several major signaling mechanisms, including changes in intracellular [Ca2+], arachidonic acid cascades, protein kinase C, cyclic AMP/protein kinase A, calmodulin, nitric oxide, and Na+/H+ antiport, are functional in both cell types. However, their relative importance in mediating basal secretion and neuroendocrine-factor-regulated hormone release differs according to cell type. Similarly, agonist- and cell-type-specificity are also present in the transduction pathways leading to neuroendocrine factor-modulated maturational gonadotropin and somatotropin release. Specificity is present not only in the actions of different regulators within the same cell type and with the same ligand in the two cell types, but this also exists between isoforms of the same neuroendocrine factor within a single cell type. Other evidence suggests that function-selectivity of signaling may also result from differential modulation of Ca2+ fluxes from different sources. The interaction of different second messenger systems provide the basis by which regulation of maturational gonadotropin and somatotropin release by multiple neuroendocrine factors can be integrated at the target cell level.     

Cultures of human tracheal gland cells of mucous or serous phenotype

There are two main epithelial cell types in the secretory tubules of mammalian glands: serous and mucous. The former is believed to secrete predominantly water and antimicrobials, the latter mucins. Primary cultures of human airway gland epithelium have been available for almost 20 yr, but they are poorly differentiated and lack clear features of either serous or mucous cells. In this study, by varying growth supports and media, we have produced cultures from human airway glands that in terms of their ultrastructure and secretory products resemble either mucous or serous cells. Of four types of porous-bottomed insert tested, polycarbonate filters (Transwells) most strongly promoted the mucous phenotype. Coupled with the addition of epidermal growth factor (EGF), this growth support produced “mucous” cells that contained the large electron-lucent granules characteristic of native mucous cells, but lacked the small electron-dense granules characteristic of serous cells. Furthermore, they showed high levels of mucin secretion and low levels of release of lactoferrin and lysozyme (markers of native serous cells). By contrast, growth on polyethylene terephthalate filters (Cyclopore) in medium lacking EGF produced “serous” cells in which small electron-dense granules replaced the electron-lucent ones, and the cells had high levels of lactoferrin and lysozyme but low levels of mucins. Measurements of transepithelial resistance and short-circuit current showed that both “serous” and “mucous” cell cultures possessed tight junctions, had become polarized, and were actively secreting Cl.     

Signal transduction in T cells: an overview

The mitogenic in vitro activation of human peripheral blood lymphocytes is severely depressed in space as well as in models of low gravity conditions on ground. The mechanism of T-cell activation is very complex; 3 signals are required for full activation. A series of experiments performed in space and in modeled low gravity on ground have shown that a failure in the delivery of the second signal--interleukin-2--is one of the reasons for the impaired activation. The cytoskeleton plays a key role in several steps of the mitogenic activation; (1) in the binding of the mitogen to the cell membrane and the subsequent patching and capping of the receptors, and (2) in the transduction of the signals from the membrane to the nucleus. Changes in the cytoskeletal structures of vimentin and tubulin observed in cells exposed to low gravity conditions may have influenced the correct signal transduction.     

Signal transduction by reactive oxygen species

Although historically viewed as purely harmful, recent evidence suggests that reactive oxygen species (ROS) function as important physiological regulators of intracellular signaling pathways. The specific effects of ROS are modulated in large part through the covalent modification of specific cysteine residues found within redox-sensitive target proteins. Oxidation of these specific and reactive cysteine residues in turn can lead to the reversible modification of enzymatic activity. Emerging evidence suggests that ROS regulate diverse physiological parameters ranging from the response to growth factor stimulation to the generation of the inflammatory response, and that dysregulated ROS signaling may contribute to a host of human diseases.