Alterations in N-methyl-D-aspartate receptor subunits in primary sensory neurons following acid-induced esophagitis in cats

The excitatory amino acid glutamate plays an important role in the development of neuronal sensitization and the ionotropic N-methyl-d-aspartate receptor (NMDAR) is one of the major receptors involved. The objective of this study was to use a cat model of gastroesophageal reflux disease (GERD) to investigate the expression of the NR1 and NR2A subunits of NMDAR in the vagal and spinal afferent fibers innervating the esophagus. Two groups of cats (Acid-7D and PBS-7D) received 0.1 N HCl (pH 1.2) or 0.1 M PBS (pH 7.4) infusion in the esophagus (1 ml/min for 30 min/day for 7 days), respectively. NR1 splice variants (both NH(2) and COOH terminals) and NR2A in the thoracic dorsal root ganglia (DRGs), nodose ganglia (NGs), and esophagus were evaluated by RT-PCR, Western blot, and immunohistochemistry. Acid produced marked inflammation and a significant increase in eosinophil peroxidase and myeloperoxidase contents compared with PBS-infused esophagus. The NR1-4 splice variant gene exhibited a significant upregulation in DRGs and esophagus after acid infusion. In DRGs, NGs, and esophagus, acid infusion resulted in significant upregulation of NR1 and downregulation of NR2A subunit gene expression. A significant increase in NR1 polypeptide expression was observed in DRGs and NGs from Acid-7D compared with control. In conclusion, long-term acid infusion in the cat esophagus resulted in ulcerative esophagitis and differential expressions of NR1 and NR2A subunits. It is possible that these changes may in part contribute to esophageal hypersensitivity observed in reflux esophagitis.     

The fibroblast growth factor-binding protein FGF-BP

Fibroblast growth factors (FGFs) are important regulators of cell migration, proliferation and differentiation, e.g., during embryogenesis and wound healing, and under several pathological conditions including tumor growth and tumor angiogenesis. Since heparin-binding FGFs are tightly bound to heparansulfate proteoglycans, and therefore, trapped in the extracellular matrix, their release through the action of an FGF-binding protein (FGF-BP) is one of the critical steps in FGF bioactivation. FGF-BP expression is highly tissue specific and strictly regulated through different promoter elements. Besides its role in embryogenesis and wound healing, FGF-BP is upregulated in several tumors and it is associated especially with early stages of tumor formation, where angiogenesis plays a critical role. Concomitantly, in several mouse tumor models, targeting of FGF-BP by ribozymes or RNA interference (RNAi) abolishes or reduces tumor growth and tumor angiogenesis. This indicates that FGF-BP can be rate-limiting for tumor growth and serves as an angiogenic switch molecule, and that it represents an increasingly promising target molecule in anti-tumor therapy.     

Relative contribution of various airway protective mechanisms to prevention of aspiration during swallowing

Deglutitive airway protective mechanisms include glottal closure, epiglottal descent, and anterosuperior displacement of the larynx. Aspiration of swallowed material may occur during the pre-, intra-, or postpharyngeal phase of swallowing. Our objectives were to determine the relative contribution of the airway protective mechanisms during each phase of swallow in 14 decerebrated cats before and after suprahyoid myotomy, epiglottectomy, and unilateral cordectomy. After myotomy, superior excursions of the hyoid, thyroid, and cricoid cartilages and anteroposterior diameter of maximum upper esophageal spincter (UES) opening were significantly diminished, but the incidence of pharyngeal residue significantly increased (P < 0.05). No aspiration was observed in the predeglutitive period. After myotomy, the incidence of aspiration significantly increased in both intra- and postdeglutitive periods. Epiglottectomy did not alter aspiration incidence, but unilateral cordectomy resulted in a 100% incidence of intra- and postdeglutitive aspiration. In conclusion, glottal closure constitutes the primary mechanism for prevention of intra- and postdeglutitive aspiration, but laryngeal elevation may assist this function. Bolus pulsion without laryngeal distraction can open the UES, but at risk of aspiration due to decreased pharyngeal clearance. The epiglottis provides no apparent airway protection during any phase of swallowing.     

Production of monoclonal antibodies against target schistosomal antigen secreted in the urine of Schistosoma mansoni-infected patients

This study was undertaken to develop an immunodiagnostic test of active human schistosomiasis mansoni using a monoclonal antibody which targets urinary schistosomal antigen. Polyclonal antisera raised in rabbits against the processed urine of Schistosoma mansoni-infected patients showed very high and significant reactivity with ES product of ova compared with other different S. mansoni antigens. The monoclonal antibody (4.23) was reactive with repetitive epitopes of S. mansoni soluble egg antigen and ES product of ova with molecular mass range of 65–23 kDa and 80–23 kDa, respectively. It recognised different stages of the parasite life-cycle, with no cross reaction with Fasciola or hydatid antigen. MAbs were characterised by isotyping, immunoelectrophoresis, SDS–PAGE and the enzyme-linked immunoelectrotransfer blot technique, ELISA, and their recognition of carbohydrate or protein antigenic epitopes by periodate oxidation and trichloroacetic acid treatment of the antigen. It was used for detection of circulating schistosomal antigen in an antigen capture antibody sandwich ELISA on sera and urines of 58 S. mansoni-infected patients, 17 S. haematobium-infected patients, 15 parasite-free negative healthy controls and sera from 13 schistosomiasis-free patients harbouring Fasciola or hydatid infections. The percentage sensitivity of the assay in the serum of S. mansoni-infected patients was 98.4% and in urine 94.8%. A positive correlation was found between the number of faecal S. mansoni eggs and the circulating antigen, both in serum and in urine. Antigen circulating in urine correlated with that in the sera of S. mansoni patients. These data provide a sensitive and non-invasive method almost comparable with the use of sera for immunodiagnosis of schistosomiasis and an indirect way to reflect the intensity of infection. Australian Society for Parasitology.     

Synthesis and antiplatelet effects of an isoxazole series of glycoprotein IIb/IIIa antagonists

Despite the excellent in vitro potency of a series of benzamide glycoprotein IIb/IIIa antagonists, which have been reported previously, poor in vivo potency in the inhibition of platelet aggregation was observed when the most potent inhibitor XU057 was dosed intravenously to dogs. In this communication, we report that replacement of the benzamide in XU057 with an isoxazolecarboxamide resulted in significant improvement in in vivo potency. More importantly, the analogue XU065 showed an excellent oral antiplatelet effect in dogs.     

Antiangiogenesis efficacy of nitric oxide donors

Angiogenesis is a complex process involving endothelial cell migration, proliferation, invasion, and tube formation. Inhibition of these processes might have implications in various angiogenesis‐mediated disorders. Because nitric oxide (NO) is known to play a key role in various vascular diseases, the present study was undertaken to determine the role of NO in angiogenesis‐mediated processes using the NO donor, S‐nitroso N‐acetyl penicillamine (SNAP) and S‐nitroso N‐acetyl glutathione (SNAG). The antiangiogenic efficacy of these NO donors was examined using in vivo and in vitro model systems. The in vitro studies demonstrated the ability of SNAP to inhibit cytokine fibroblast growth factor (FGF2)‐stimulated tube formation and serum‐induced cell proliferation. The inhibitory effect on cell proliferation by SNAP concentrations above the millimolar range was associated with significant shifts in the concentration of NO metabolites. Furthermore, using the mouse Matrigel implant model and the chick chorioallantoic membrane (CAM) models, SNAP demonstrated maximal inhibitory efficacy (85–95% inhibition) of cytokine (FGF2)‐induced neovascularization in both in vivo models. SNAP and SNAG resulted in 85% inhibition of FGF2‐induced neovascularization in the mouse Matrigel model when given at 5 mg/kg/day infusion in minipumps during 14 days and 87% inhibition of angiogenesis induced by FGF2 in the CAM when administered a single dose of 50 μg. Thus, NO donors might be a useful tool for the inhibition of angiogenesis associated with human tumor growth, or neovascular, ocular, and inflammatory diseases. J. Cell. Biochem. 80:104–114, 2000. © 2000 Wiley‐Liss, Inc.     

Cerebral cortical representation of external anal sphincter contraction: effect of effort

The external anal sphincter (EAS) plays a critical role in maintaining fecal continence; however, cerebral cortical control of voluntary EAS contraction is not completely understood. Our aims were to determine the cortical areas associated with voluntary EAS contraction and to determine the effect of two levels of sphincter contraction effort on brain activity. Seventeen asymptomatic adults (ages 21-48, 9 male) were studied using functional magnetic resonance imaging (fMRI) to detect brain activity. Studies were done in two stages. In stage 1 (10 subjects, 5 male), anal sphincter pressure was monitored from a catheter-affixed bag. Subjects performed maximal and submaximal EAS contractions during two fMRI scanning sessions consisting of alternating 10-s intervals of sustained contraction and rest. In stage 2 studies, seven subjects (4 male) performed only maximum effort sphincter contractions without a catheter. EAS contraction was associated with multifocal fMRI activity in sensory/motor, anterior cingulate, prefrontal, parietal, occipital, and insular regions. Total cortical activity volume was significantly larger (P < 0.05) for maximal (5,175 +/- 720 microl) compared with submaximal effort contractions (2,558 +/- 306 microl). Similarly, percent fMRI signal change was significantly higher (P < 0.05) for maximal (4.8 +/- 0.1%) compared with submaximal effort contractions (2.2 +/- 0.1%). Cortical region-of-interest analysis showed the incidence of insular activation to be more common in women compared with men. Other cortical regions showed no such gender differences. fMRI activity detected in stage 2 showed similar regions of cortical activation to those of the stage 1 study. Willful contraction of the EAS is associated with multifocal cerebral cortical activity. The volume and intensity of cerebral cortical activation is commensurate with the level of contractile effort.     

Characterization of the cerebral cortical representation of heartburn in GERD patients

Although symptoms arising from the esophagus such as heartburn and pain can at times become challenging clinical problems, esophageal viscerosensation, especially with regard to chemical stimulation in humans, is incompletely understood. Our aims were 1) to characterize and ascertain the reproducibility of cerebral cortical registration of heartburn and 2) to elucidate the differences between these findings and those of esophageal subliminal acid stimulation in asymptomatic controls. We studied 11 gastroesophageal reflux disease (GERD) patients (9 males, 30-55 yr) and 15 healthy controls (8 males, 21-49 yr). Cerebral cortical functional magnetic resonance imaging (fMRI) activity was recorded twice in each subject, during two 5-min intervals of 0.1 N HCl, separated by 5 min of NaCl perfusion. Time from onset of acid perfusion to instant of fMRI signal increase and first report of heartburn averaged 1.60 +/- 0.80 and 1.85 +/- 0.60 min, respectively. Average maximum percent signal increase in the GERD patients (16.3 +/- 3.5%) was significantly greater than that of healthy controls (3.8 +/- 0.9%; P < 0.01). Temporal fMRI signal characteristics during heartburn were significantly different from those of subliminal acid stimulation in controls (P < 0.01). Activated cortical regions included sensory/motor, parieto-occipital, cingulate and prefrontal regions, and the insula. There was 92% concordance between the activated Brodmann areas in repeated studies of GERD patients. Cortical activity associated with perceived and unperceived esophageal acid exposure in GERD patients and healthy controls, respectively, involves multiple brain regions but occurs more rapidly and with greater intensity in GERD patients than the activity in response to subliminal acid exposure in healthy controls. The cortical pain-processing pathway seems to be involved in perception of esophageal acid exposure and could explain the variations encountered in clinical practice defining this sensation.     

Upper esophageal sphincter function during gastroesophageal reflux events revisited

Upper esophageal sphincter (UES) function during gastroesophageal reflux events is not completely elucidated because previous studies addressing this issue yielded conflicting results. We reexamined the UES pressure response to intraluminal esophageal pressure and pH changes induced by reflux events. We studied 14 healthy, asymptomatic volunteers (age 49 +/- 6 yr) and 7 gastroesophageal reflux disease patients (age 48 +/- 5 yr). UES pressure, intraesophageal pressure, and pH were monitored at the distal, middle, and proximal esophagus concurrently in the supine position 1 h before and 2 h after a 1,000-calorie meal. A total of 321 reflux events were identified by the development of abrupt reflux-induced intraesophageal pressure increase (IPI); 285 events occurred in patients and 36 in control subjects. In control subjects 33 of 36 and in patients 252 of 285 IPI events were associated with a pH drop. Among patients and control subjects, 99% and 100%, respectively, of all IPI events irrespective of pH drop were associated with abrupt increase in UES pressure (34 +/- 2 and 27 +/- 6 mmHg, respectively). The average percentage of maximum UES pressure increase over prereflux values ranged between 66% and 96% (control subjects) and 34% and 122% (patients). IPIs induced by both acidic and nonacidic reflux events evoke strong UES contractile responses.