The pathogenesis of gastroesophageal reflux disease: a challenge in clinical physiology.
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Esophagogastric Junction↗
Biomedical subjects
S Cohen
Publications and source records attributed to S Cohen.
Epidermal growth factor stimulates tyrosine phosphorylation in the neonatal mouse: association of a M(r) 55,000 substrate with the receptor.
Administration of epidermal growth factor (EGF) to neonatal mice resulted in rapid tyrosine phosphorylation of a number of specific substrates in liver, kidney, lung, bladder, skin, and brain as detected by Western blot analysis of tissue homogenates with anti-phosphotyrosine antibodies. In the liver, three prominent EGF-dependent substrates of M(r) approximately 170,000, 120,000, and 55,000 were detected. A number of less prominent EGF-dependent substrates also were noted. Maximal tyrosine phosphorylation of pp170, pp120, and pp55 occurred within 5 min of subcutaneous injection and the levels of these phosphoproteins remained elevated for at least 45 min. Direct hepatic injection of EGF resulted in the tyrosine phosphorylation of these substrates within 60 sec of treatment. Tyrosine-phosphorylated pp170 was identified as the EGF receptor (EGFR). The tyrosine-phosphorylated pp55 substrate appeared to be associated with EGFR; both pp55 and EGFR were adsorbed to EGF-Affi-Gel, wheat germ lectin-Sepharose, and anti-EGFR antibodies bound to protein A-Sepharose. pp55 was not immunoreactive with anti-EGFR antiserum by Western blot analysis, indicating that it was not a fragment of the receptor. These results were confirmed by repeating the liver experiments using 32P-labeled neonatal mice. Increased amounts of 32P-labeled pp170 and pp55 were detected in anti-EGFR immunoprecipitates from liver extracts of EGF-treated animals as compared with controls. Phospho amino acid analysis of the 32P-labeled phosphoproteins revealed that EGF stimulated both serine and tyrosine phosphorylation in pp55 as well as in EGFR. The neonatal mouse may be a useful model for the study of signal transduction mediated by a variety of growth factors.
Animals↗
Mutagenesis of human acetylcholinesterase. Identification of residues involved in catalytic activity and in polypeptide folding.
Evidence for the involvement of Ser-203, His-447, and Glu-334 in the catalytic triad of human acetylcholinesterase was provided by substitution of these amino acids by alanine residues. Of 20 amino acid positions mutated so far in human acetylcholinesterase (AChE), these three were unique in abolishing detectable enzymatic activity (less than 0.0003 of wild type), yet allowing proper production, folding, and secretion. This is the first biochemical evidence for the involvement of a glutamate in a hydrolase triad (Schrag, J.D., Li, Y., Wu, M., and Cygler, M. (1991) Nature 351, 761-764), supporting the x-ray crystal structure data of the Torpedo californica acetylcholinesterase (Sussman, J.L., Harel, M., Frolow, F., Oefner, C., Goldman, A., Toker, L. and Silman, I. (1991) Science 253, 872-879). Attempts to convert the AChE triad into a Cys-His-Glu or Ser-His-Asp configuration by site-directed mutagenesis did not yield effective AChE activity. Another type of substitution, that of Asp-74 by Gly or Asn, generated an active enzyme with increased resistance to succinylcholine and dibucaine; thus mimicking in an AChE molecule the phenotype of the atypical butyrylcholinesterase natural variant (D70G mutation). Mutations of other carboxylic residues Glu-84, Asp-95, Asp-333, and Asp-349, all conserved among cholinesterases, did not result in detectable alteration in the recombinant AChE, although polypeptide productivity of the D95N mutant was considerably lower. In contrast, complete absence of secreted human AChE polypeptide was observed when Asp-175 or Asp-404 were substituted by Asn. These two aspartates are conserved in the entire cholinesterase/thyroglobulin family and appear to play a role in generating and/or maintaining the folded state of the polypeptide. The x-ray structure of the Torpedo acetylcholinesterase supports this assumption by revealing the participation of these residues in salt bridges between neighboring secondary structure elements.
Acetylcholinesterase↗
Exercise tolerance in panic disorder patients.
Sixteen panic patients and fifteen normal controls performed submaximal exercise testing on a bicycle ergometer. Only one patient subject panicked. Biochemical, physiological, and psychological data showed similar exercise tolerance in both patients and controls. Exercise-induced distress and lactate increment do not appear to cause panic attacks.
Adult↗
Differential long-term effect of AF64A on [3H]ACh synthesis and release in rat hippocampal synaptosomes.
The activities of various presynaptic cholinergic parameters were determined in hippocampal synaptosomes of rats 29 weeks after intracerebroventricular injection of ethylcholine aziridinium (AF64A) (3 nmol/2 microliters/side) or vehicle (saline). Synaptosomes were preloaded with [3H]choline ([3H]Ch), treated with diisopropyl fluorophosphate to inhibit cholinesterase activity and then were assayed for their content of [3H]Ch and [3H]acetylcholine ([3H]ACh) and for their ability to synthesize and release [3H]ACh. In synaptosomes from AF64A-treated rats compared with synaptosomes from vehicle-treated rats we observed that: (i) specific uptake of [3H]Ch was reduced to 60% of control; (ii) residing [3H]ACh levels were 43% of control while residing [3H]Ch levels were 72% of control; (iii) basal and K(+)-induced [3H]ACh release were 77% and 73% of control, respectively; (iv) high K(+)-induced synthesis of [3H]ACh was only 9% of control; (v) but, choline acetyltransferase activity remained relatively high, being 80% of control. These results suggest that AF64A-induced cholinergic hypofunction is expressed by both loss of some cholinergic neurons and impairment in the functioning of the spared neurons.
Acetylcholine↗
Low density lipoprotein inhibits accumulation of nitrites in murine brain endothelial cell cultures.
Endothelial cells produce nitric oxide which is considered to serve as a major source of endothelial derived relaxing factor activity. It has been demonstrated that activation of mouse brain endothelium by TNF-alpha and IFN-gamma led to accumulation of nitrite which is presumably formed by oxidation of nitric oxide. A number of studies suggest that reactive oxygen species produced by cytokine-activated cells are involved in the conversion of nitric oxide to nitrites and nitrates. We investigated whether low density lipoprotein (LDL), acting as a radical scavenger, is able to inhibit nitrite accumulation in mouse brain endothelial cell cultures and in a cell-free system in which sodium nitroprusside was used as a source of nitric oxide. A comparison of these two models indicates the active involvement of LDL in suppressing nitrite accumulation in murine endothelial cultures.
Animals↗
Ultrastructural immunolocalization of rat oxytocin-neurophysin in transgenic mice expressing the rat oxytocin gene.
Cell-specific expression of the rat oxytocin (OT)-neurophysin transgene in mice was achieved using a construct containing both OT and vasopressin genes (Young III, W.S., Reynolds, K., Shepard, E.A., Gainer, H. and Castel, M., Cell-specific expression of the rat oxytocin gene in transgenic mice, J. Neuroendocrinol., 2 (1990) 1-9). The present study describes the distribution of the protein products of these genes in various regions of the cell, and determines whether the transgenic rat and endogenous mouse OT-neurophysins are colocalized within the same neurosecretory granules. Two monoclonal antibodies against OT-neurophysins were used: PS38 which can react with both rat and mouse OT-neurophysin (pan-specific), and PS67 which is specific for rat OT-neurophysin only. Various approaches to double immunolabeling at the ultrastructural level were employed; these included: (1) pre-embedding immunoperoxidase followed by post-embedding immunogold; (2) post-embedding immunolabeling using gold particles of different sizes; and (3) labeling of consecutive ultrathin sections with different antibodies. Results from each of these approaches showed that both in the transgenic mouse and in the rat (used as control), immunocytochemical labeling for both PS38 and PS67 occurred in the same OT-ergic neurosecretory granules. In the control mouse, only PS38 elicited labeling. Hence, it may be concluded that the protein and peptide products of the transgene and the endogenous gene for OT-neurophysin are being processed similarly in the cell and finally concentrated together in the same neurosecretory granules.
Animals↗
Inhibition of basal and tumor necrosis factor-enhanced binding of murine tumor cells to murine endothelium by transforming growth factor-beta 1.
The adherence of cells to microvascular endothelium is important in a number of processes, including inflammatory responses and metastasis. It has been demonstrated that in human models, cytokines such as TNF, IL-1, IFN-gamma increase the adhesiveness of endothelium for cells of the immune and inflammatory system by stimulating the expression of cell adhesion molecules on endothelial cell surfaces. We and others have shown similar cytokine-induced endothelial adhesiveness for tumor cells in murine and human models. In contrast to the effect of those modulators, transforming growth factor-beta (TGF-beta) has been shown to inhibit the binding of human neutrophils and T lymphocytes to human endothelium, although the mechanism of TGF-beta action remains unknown. Little is known about the effect of TGF-beta on tumor cell-endothelial interaction. In the present study, we demonstrate that TGF-beta inhibits basal and TNF-enhanced binding of murine P815 mastocytoma cells to murine microvascular endothelium (MME). The alterations in MME mediated by TGF-beta, also lead to the inhibition of adherence of murine splenocytes, thymocytes, and human lymphoblastoid cells but do not inhibit adherence of murine B16 melanoma cells. The effect of TGF-beta is transient and inhibition of the endothelial adhesive phenotype is strongest 12 to 24 h after addition of the factor to MME. The TGF-beta-mediated inhibition of P815 basal binding to endothelium is dependent on protein synthesis because cycloheximide reverses the TGF-beta effect. TGF-beta does not appear to activate classical signal transduction pathways. Inhibitors of G proteins do not abolish TGF-beta action, protein kinase C and protein kinase A activators elicit an effect opposite to that of the factor, TGF-beta does not increase intracellular cAMP levels, and finally calcium-mobilizing agents do not mimic, but rather inhibit the effect of TGF-beta. However, TGF-beta-mediated inhibition of both basal binding and TNF-enhanced P815 binding to MME is completely abolished in the presence of the protein phosphatase inhibitor okadaic acid which suggests that TGF-beta may elicit its effect by stimulating protein phosphatase activity.
Animals↗
Substrate inhibition of acetylcholinesterase: residues affecting signal transduction from the surface to the catalytic center.
Amino acids located within and around the 'active site gorge' of human acetylcholinesterase (AChE) were substituted. Replacement of W86 yielded inactive enzyme molecules, consistent with its proposed involvement in binding of the choline moiety in the active center. A decrease in affinity to propidium and a concomitant loss of substrate inhibition was observed in D74G, D74N, D74K and W286A mutants, supporting the idea that the site for substrate inhibition and the peripheral anionic site overlap. Mutations of amino acids neighboring the active center (E202, Y337 and F338) resulted in a decrease in the catalytic and the apparent bimolecular rate constants. A decrease in affinity to edrophonium was observed in D74, E202, Y337 and to a lesser extent in F338 and Y341 mutants. E202, Y337 and Y341 mutants were not inhibited efficiently by high substrate concentrations. We propose that binding of acetylcholine, on the surface of AChE, may trigger sequence of conformational changes extending from the peripheral anionic site through W286 to D74, at the entrance of the 'gorge', and down to the catalytic center (through Y341 to F338 and Y337). These changes, especially in Y337, could block the entrance/exit of the catalytic center and reduce the catalytic efficiency of AChE.
Acetylcholinesterase↗
Localization of p35 (annexin I, lipocortin I) in normal adult rat kidney and during recovery from ischemia.
The 35-kDa protein (p35, lipocortin I, annexin I), originally discovered as a Ca(++)-dependent substrate for the EGF receptor tyrosine kinase, binds Ca++ and phospholipids, is developmentally regulated in embryos and has restricted expression in adults. Immunohistochemistry of normal rat kidney shows that p35 is enriched in epithelia of Bowman's capsule, the macula densa, and medullary/papillary collecting ducts, suggesting that p35 is related to specialized renal functions. Light staining is observed in the thick ascending limb; elsewhere, immunoreactivity is nil. Since renal recovery from ischemia involves both hyperplasia and hypertrophy and reportedly is accelerated by EGF, we examined p35 distribution during this process. After 48 hours of recovery, both the distribution and amount of renal p35 are altered. Immunoblots show p35 levels increased at least threefold in whole-kidney homogenates. The expression of p35 is still highly restricted in recovering kidneys; however, the thick ascending limb now stains heavily. This is the first documentation of alterations in annexin levels during a pathophysiologic response. However, our attempts to discern effects of exogenous EGF on the recovery from ischemia were negative for both mitotic index and renal function assays.
Animals↗
Kinetics of transdermal penetration of an organic ion pair: physostigmine salicylate.
Physostigmine salicylate was delivered from a series of solvents consisting of isopropyl myristate, isopropyl alcohol (IPA), and their mixtures across dermatomed human skin. The apparent steady-state fluxes over the time range of the test, obtained separately for physostigmine and its corresponding salicylate, indicate a consistent trend toward higher values for the salicylate in the series tested. The ratios of salicylate fluxes to physostigmine fluxes ranged from 1.1 to 3.34, the higher ratios being obtained at a volume fraction of IPA exceeding 0.7. Ionization of the ion pair at the pH of the hydrated stratum corneum immediately after its partitioning into the membrane, followed by differential diffusion of the species across the membrane, is consistent with the kinetics of penetration. It is proposed that the apparent volume of distribution of physostigmine is larger than that of salicylate and, hence, a smaller concentration difference across the diffusion barrier exists for physostigmine. This hypothesis can explain the lower flux of physostigmine to conform to Fick's first law of diffusion and the assumption of equal molar transfer to the skin of both species. The hypothesis implies that if steady state appears to have been reached for the faster migrating salicylate over the time range tested then the apparent steady state of physostigmine is not a true one. Increasing the salicylate content in one of the donor solutions by eight times over that of physostigmine decreased the saturation concentration of physostigmine but not in its flux. Increasing the physostigmine content by 6.5 times over that of salicylate in the same donor solution did not change either the flux or the salicylate concentration but decreased the permeability coefficient of physostigmine.(ABSTRACT TRUNCATED AT 250 WORDS)
Administration, Cutaneous↗
Effect of verapamil on human intestinal transit.
Although constipation is a well-known side effect of calcium channel blockers such as verapamil, this side effect has not been evaluated in a quantitative manner. In a double-blind, randomized, crossover trial, the effect of verapamil (240 mg/day) was compared to placebo in 15 normal male volunteers. Subjects recorded their bowel movements and any side effects. Scintigraphy was used to quantitate gastric emptying, small intestinal transit, and colonic transit. In the study period of four days, verapamil did not change the frequency, consistency, or passage of bowel movements. A significantly increased number of side effects was noted during verapamil treatment--notably abdominal pain and dry mouth. The slope of gastric emptying was not significantly different for verapamil (0.012 +/- 0.02) than for placebo (0.013 +/- 0.001). Distal ileum filling was also not different for verapamil (0.41 +/- 0.13%/min) than placebo (0.33 +/- 0.05%/min). Progression of the colonic geometric center was significantly delayed at 48 hr by verapamil (5.2 +/- 0.4 vs 6.2 +/- 0.23; P less than 0.01). This study suggests that the constipating effect of verapamil is due to a delay of colonic transit and not due to an effect on upper gastrointestinal transit.
Adult↗
The future of gastrointestinal practice and education: self-regulation or governmental control?
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Gastroenterology↗
AGA governing board policy statement on training and education.
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Certification↗
The evolution of public policy in the American Gastroenterological Association.
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Gastroenterology↗
Inhibition of choline efflux results in enhanced acetylcholine synthesis and release in the guinea-pig corticocerebral synaptosomes.
Synthesis and release of [3H]acetylcholine ([3H]ACh) were measured in synaptosomes from the guinea pig cerebral cortex after preloading with [3H]choline ([3H]Ch). We demonstrate here that inhibition of choline (Ch) efflux results in an increase in acetylcholine (ACh) synthesis and release. Our findings are as follows: (1) inhibition of [3H]Ch efflux by hemicholinium-3 (HC-3) (100 microM), increased the levels of both the released (116% of control) and the residing (115% of control) [3H]ACh. (2) The muscarinic agonist, McN-A-343 (100 microM), which was previously shown to inhibit Ch efflux, also increased the released (121% of control) and the residing (109% of control) [3H]ACh. (3) Omission of Na+ ions (which are required for Ch transport) from the incubation medium had similar effects to those observed with McN-A-343 and HC-3. These results suggest inverse relationships between Ch efflux on one hand, and ACh synthesis and release on the other hand. (4) Depolarization with 50 mM K+, or with the K+ channel blocker, 4-aminopyridine (100 microM), also increased the total level of [3H]ACh (113 and 107% of nondepolarized synaptosomes, respectively). However, whereas conditions that inhibit Ch transport such as HC-3, McN-A-343 and "no sodium" increased both the residing and the released [3H]ACh depolarization with high K+ or 4-aminopyridine reduced the residing (79 and 87% of control, respectively) and increased only the released [3H]ACh (182 and 148% of control, respectively).(ABSTRACT TRUNCATED AT 250 WORDS)
(4-(m-Chlorophenylcarbamoyloxy)-2-butynyl)trimethy↗
Effects of occupational stress on the physical and psychological health of women in a microelectronics plant.
Physical and psychological effects of occupational stress were examined in a sample of 552 female blue collar employees of a microelectronics facility. After controlling for demographic and biological risk factors, non-work life events, and solvent exposure, job-related conflict was associated with depressive symptomatology, severe headaches, lightheadedness weakness/fatigue, rashes, and presence of multiple symptoms. Job demands were only associated with multiple symptoms. Solvent exposure did not interact with either of the job stress measures synergistically to increase symptom reports. Although the main effects of social support, when present, were in the direction of reducing symptomatology, the interactive effects of social support and stress on health were inconsistent and dependent on the source of support.
Adult↗
Controlled protein release from polyethyleneimine-coated poly(L-lactic acid)/pluronic blend matrices.
Protein release from degradable polymer matrices, composed of poly(L-lactic acid) and its blends with Pluronic surfactant, was investigated with and without the aqueous coating of an adsorptive water-soluble polymer, polyethyleneimine (PEI). PEI is a highly branched cationic polymer containing primary, secondary, and tertiary amino groups in its backbone. The treatment of PEI for PLA/Pluronic blend films exhibited a remarkable decrease in the "burst" release of protein at an initial stage and a significant extension in the protein release period. Protein release profiles could be controlled by varying PEI treatment time and its concentration. Our results suggest that PEI diffuses into the polymer matrices and crosslinks protein molecules by ionic interactions. Such a PEI-protein network near the surface region of matrix may act as a diffusional barrier for further release of protein molecules.
Delayed-Action Preparations↗