Mechanisms of the adenosine A2A receptor-induced sensitization of esophageal C fibers.

Clinical studies indicate that adenosine contributes to esophageal mechanical hypersensitivity in some patients with pain originating in the esophagus. We have previously reported that the esophageal vagal nodose C fibers express the adenosine A2A receptor. Here we addressed the hypothesis that stimulation of the adenosine A2A receptor induces mechanical sensitization of esophageal C fibers by a mechanism involving transient receptor potential A1 (TRPA1). Extracellular single fiber recordings of activity originating in C-fiber terminals were made in the ex vivo vagally innervated guinea pig esophagus. The adenosine A2A receptor-selective agonist CGS21680 induced robust, reversible sensitization of the response to esophageal distention (10-60 mmHg) in a concentration-dependent fashion (1-100 nM). At the half-maximally effective concentration (EC50: ≈3 nM), CGS21680 induced an approximately twofold increase in the mechanical response without causing an overt activation. This sensitization was abolished by the selective A2A antagonist SCH58261. The adenylyl cyclase activator forskolin mimicked while the nonselective protein kinase inhibitor H89 inhibited mechanical sensitization by CGS21680. CGS21680 did not enhance the response to the purinergic P2X receptor agonist α,ß-methylene-ATP, indicating that CGS21680 does not nonspecifically sensitize to all stimuli. Mechanical sensitization by CGS21680 was abolished by pretreatment with two structurally different TRPA1 antagonists AP18 and HC030031. Single cell RT-PCR and whole cell patch-clamp studies in isolated esophagus-specific nodose neurons revealed the expression of TRPA1 in A2A-positive C-fiber neurons and demonstrated that CGS21682 potentiated TRPA1 currents evoked by allylisothiocyanate. We conclude that stimulation of the adenosine A2A receptor induces mechanical sensitization of nodose C fibers by a mechanism sensitive to TRPA1 antagonists indicating the involvement of TRPA1.

Studies on the regulation of transient lower esophageal sphincter relaxations (TLESRs) by acid in the esophagus and stomach.

Transient lower esophageal sphincter relaxation (TLESR) is the major mechanism of gastroesophageal reflux, but the regulation of TLESR by stimuli in the esophagus is incompletely understood. We have recently reported that acid infusion in the esophagus substantially (by 75%) increased the number of meal-induced TLESR in healthy subjects. We concluded that the TLESR reflex triggered by gastric distention with meal was enhanced by the stimulation of esophageal nerves by acid. However, the possibilities that the acid infused into the esophagus acts after passing though lower esophageal sphincter in stomach to enhance TLESR, or that the acid directly initiates TLESR from the esophagus were not addressed. Here, we evaluated the effect of acid infusion into the proximal stomach on meal-induced TLESR (study 1) and the ability of acid infusion into the esophagus to initiate TLESR without prior meal (study 2). We analyzed TLESRs by using high-resolution manometry in healthy subjects in paired randomized studies. In study 1, we found that acid infusion into the proximal stomach did not affect TLESRs induced by standard meal. The number of meal-induced TLESRs following the acid infusion into the proximal stomach was similar to the number of meal-induced TLESRs following the control infusion. In study 2, we found that acid infusion into the esophagus without prior meal did not initiate TLESRs. We conclude that the increase in the meal-induced TLESRs by acid in the esophagus demonstrated in our previous study is not attributable to the action of acid in the stomach or to direct initiation of TLESR from the esophagus by acid. Our studies are consistent with the concept that the stimuli in the esophagus can influence TLESRs. The enhancement of TLESR by acid in the esophagus may contribute to pathogenesis of gastroesophageal reflux in some patients.

Deep nasal inspirations increase the cough threshold in children with mild asthma.

Asthma is a chronic inflammatory disease characterized by bronchospasms accompanied with frequent coughing, the pathogenesis of which is not clear. In healthy adults deep inspirations (DIs) provide a protective effect against bronchoconstriction triggered by methacholine inhalation, which correlates with the number of accompanying cough efforts. The aim was to study the effect of deep nasal inspirations representing the voluntary equivalent of the sniff-like aspiration reflex on the capsaicin-induced cough in children with mild asthma. The cough reflex sensitivity to capsaicin was determined using a compressed air-driven nebulizer in 21 children (8 girls and 13 boys of median age 13.3 year) suffering from mild asthma (FEV(1)∼80%). The effect of five previous DIs through the nose was examined on the elicitability of two and five or more cough efforts (C2, C5). Under control conditions, the concentration of 20.86 (14.58-29.8) µmol/l of capsaicin provoked two cough efforts (C2). After five DIs similar reaction required significantly higher concentrations of capsaicin: 29.02 (18.88-44.6) µmol/l; P=0.016. Five or more cough efforts (C5) were not significantly changed after previous DIs 161.49 (77.31-337.33) µmol/l and without DIs 141.52 (68.77-291); P=0.54. A series of five deep inspirations decreases the cough reflex sensitivity to evoke two efforts (C2) in children with mild asthma. The inhibitory effect of similar DIs disappeared after repeated applications of increasing doses of capsaicin, aiming to evoke five or more cough efforts, suggesting a reflex character of protective effect of DIs.

The role of Aß in Alzheimer's Disease as an Evolutionary Outcome of Optimized Innate Immune Defense.

Alzheimer's Disease is a progressive manifestation of aging associated with accumulated Amyloid ß. It remains frustratingly unclear why this protein accumulates and how it contributes to Alzheimer's Disease pathology. In one recent hypothesis, Amyloid ß is suggested to function as an antimicrobial peptide in innate immune defense within the brain, where Amyloid ß gains toxicity when it becomes abundant. This essay proposes an evolutionary explanation for why Amyloid ß expression is regulated at an optimum based on its function as a defense and how this leads to disease. Among its potential physiological functions, Amyloid ß confers benefits to reduce direct pathogen damage while this simultaneously entails cellular cost of defense. Optimal Amyloid ß expression occurs when the gain in fitness from an incremental increase is balanced by the marginal cost of this increase. It proposes that natural selection acting upon the young favored systems to maintain Amyloid ß at an optimal level through mechanisms that induce the defense and repress its expression. With age, the force of natural selection declines and permits mechanisms of negative feedback repression to degenerate. Consequently, Amyloid ß is expressed beyond its optimum. Age also elevates cumulative pathogen exposure, reduces pathogen barriers and reactivates latent pathogens. The net effect is elevated, chronic induction of Amyloid ß in the brain. The model recommends attention to innate immune negative regulation in the brain to discover ways to restore these functions toward a youthful state in the elderly.

Workshop: tuning the 'cough center'.

The Workshop considered the mechanisms whereby the 'cough center' could be tuned by various afferent inputs. There were particular presentations on the effects of inputs from the nose, mouth, respiratory tract and lungs, cerebral cortex, somatic tissues and the pharynx. From all these sites cough induced from the lungs could be increased or decreased in its strength or modified in its pattern. Thus 'tuning' of cough could be due to the interaction of afferent inputs, or to the sensitization or desensitization of brainstem neural pathways. The pattern of response depended on the 'type' of cough being studied and, in some instances, on the timing of the sensory input into the brainstem. Cough inputs could also affect various 'non-cough' motor outputs from the brain, although this was not the main theme of the Workshop. The main conclusion was that cough is not a stereotyped output from the medullary 'cough center', but that its pattern and strength depend on many afferent inputs acting on the 'cough center'.

Distinction of cough from other sounds produced by daily activities in the upper airways.

OBJECTIVES: The aim of this study was to validate the successfulness of our developed system for distinction between cough and other sounds which are present in daily human activities from the upper airways. BACKGROUND: To date, methods used for monitoring of cough sound were primarily subjective. A reliable measure of cough is needed so that the severity of cough in various patients and the effectiveness of treatment can be assessed. METHODS: Sounds of induced cough and sneezing, voluntary throat and nasopharynx clearing, forced ventilation and laughing, snoring, eructation, loud swallowing, and nasal blowing were studied. Characteristics of the sound events in 20 volunteers were calculated using the time-domain, spectral and non-linear analysis. The classification tree was constructed for classification between cough and non-cough sounds. We have validated the usefulness of our developed algorithm against subjective cough counts, which were performed by two trained observers. RESULTS: The value of sensitivity for distinction between cough and other sounds was 86% and the value of specificity was 91%. The value of sensitivity for distinction between voluntary and induced cough sounds was 96% and specificity was 43%. The value of sensitivity between cough sounds and voluntary throat clearing was 96% and specificity was 85%. The value of sensitivity between cough sounds and induced sneezing was 95% and specificity was 93%. CONCLUSION: We have developed an algorithm for distinction between cough and other sounds with a relatively high degree of accuracy (Tab. 1, Fig. 5, Ref. 15).

Droplet digital PCR as a novel dia-gnostic tool.

BACKGROUND: Nowadays, modern treatment methods for cancer patients are based on targeting specific molecules involved in cellular signaling system associated with tumor initiation and progression. The success of such approach depends on a correctly chosen dia-gnostic test with high sensitivity that identifies the occurrence and level of bio-markers in patients to select those who will respond and benefit from the treatment. The development of new technologies and the upgrades of the known ones contribute to the innovations in molecular characterization of cancer, which allows the detection of patients mutational status with high sensitivity and specificity. PURPOSE: Here, we discuss the utilization of the third-generation type of polymerase chain reaction (PCR), droplet digital PCR (ddPCR), in the molecular dia-gnostics of oncology diseases. According to the studies reported in our review, ddPCR represents a promising tool in genetic profiling of cancer patients. Therefore, the optimization and precise validation may enable gradual implementation of ddPCR into clinical practice in the field of oncology.

Cough reflex sensitivity and fractional exhaled nitric oxide in children with asthma.

Individual studies have suggested the utility of fractional exhaled nitric oxide (FeNO) measurement in detecting cough-variant asthma and eosinophilic bronchitis in patients with chronic cough. The aim of this study was to clarify a correlation of cough reflex sensitivity and fractional exhaled nitric oxide in asthmatic children. 25 children with asthma and 15 controls were submitted to cough reflex sensitivity measurement - capsaicin aerosol in doubling concentrations (from 0.61 to 1250 micromol/l) was inhaled by a single breath method. Concentrations of capsaicin causing two (C2) and five coughs (C5) were reported. Fractional exhaled nitric oxide (FeNO) measurement was included. Asthmatic children' (11 boys and 14 girls, mean age 9+/-1 years) and control group (unconfirmed diagnosis of asthma) (6 boys and 9 girls, mean age 8+/-1 years) were included into the study. FeNO vs. C2 in asthma (Spearman´s rank correlation: -0.146, p=0.49); FENO vs. C5 in asthma (Spearman´s rank correlation: -0.777, p=0.71). We found that there is no correlation between cough reflex sensitivity and fractional exhaled nitric oxide either in children with asthma or in the control group.

Relationship between cough reflex sensitivity and body mass index in children with chronic cough - a pilot study.

Obesity is characterized by chronic, low-grade systemic inflammation. Obesity may also be associated with chronic cough. The aim of this pilot study was to clarify relation of cough reflex sensitivity and body mass index (BMI) in children with chronic cough. Altogether 41 children having symptoms of chronic cough were submitted to cough reflex sensitivity measurement. We assessed the relation of cough reflex sensitivity (CKR) due to BMI. Cough reflex sensitivity was defined as the lowest capsaicin concentration which evoked two (C2) or five (C5) coughs. Capsaicin aerosol in doubling concentrations (from 0.61 to 1250 micromol/l) was inhaled by a single breath method (KoKo DigiDoser; nSpire heath Inc, Louisville, CO, USA), modified by the addition of an inspiratory flow regulator valve (RIFR; nSpire heath Inc, Louisville, CO, USA). BMI was calculated. Pulmonary function was within normal range. Concentrations of capsaicin causing two (C2) and five coughs (C5) were reported. Children (22 boys and 19 girls, mean age 6.8 years) cough reflex sensitivity (median, with the Inter-Quartile Range) for C2 was 19.5 (73.4) micromol/l; for C5 it was 78.1 (605.5) micromol/l. We have noticed statistically significant relation of the cough reflex sensitivity (C5) and body mass index (P<0.0001); however, the effect size was small, R2=0.03. Increase of body mass index in one unit is associated with -34.959 micromol/l decrease of C5. We did not find a statistically significant relation between C2 and BMI (P=0.41). The median value of CKR (C2) in boys is not statistically significantly different than the median value of CKR (C2) in girls (P-value 0.5). The median value of CKR (C5) in boys is not statistically significantly different than the median value of CKR (C5) in girls (P-value 0.5). Increase of body mass index in children suffering from chronic cough relates to decrease of cough reflex sensitivity (C5 value).

A mutant Drosophila insulin receptor homolog that extends life-span and impairs neuroendocrine function.

The Drosophila melanogaster gene insulin-like receptor (InR) is homologous to mammalian insulin receptors as well as to Caenorhabditis elegans daf-2, a signal transducer regulating worm dauer formation and adult longevity. We describe a heteroallelic, hypomorphic genotype of mutant InR, which yields dwarf females with up to an 85% extension of adult longevity and dwarf males with reduced late age-specific mortality. Treatment of the long-lived InR dwarfs with a juvenile hormone analog restores life expectancy toward that of wild-type controls. We conclude that juvenile hormone deficiency, which results from InR signal pathway mutation, is sufficient to extend life-span, and that in flies, insulin-like ligands nonautonomously mediate aging through retardation of growth or activation of specific endocrine tissue.