Towards optimizing the protocol for untargeted profiling of urine volatiles via gas chromatography-ion mobility spectrometry. A pilot study.

The analysis of volatile organic compounds (VOCs) present in various biological samples holds immense potential for non-invasive disease diagnostics and metabolic profiling. One of the biological fluids that are suitable for use in clinical practice is urine. Given the limited quantity of VOCs in the urine headspace, it's imperative to enhance their extraction into the gaseous phase and prevent any degradation of VOCs during the thawing process. The study aimed to test several key parameters (incubation time, temperature, and thawing) that can influence urine volatilome and monitor selected VOCs for their stability. The analysis in this study was performed using a BreathSpec® (G.A.S., Dortmund, Germany) device consisting of a gas chromatograph (GC) coupled with an ion mobility spectrometer (IMS). Testing three different temperatures and incubation times yielded a low number of VOCs (9 out of 34) that exhibited statistically significant differences. However, examining three thawing conditions revealed no VOCs with statistically significant changes. Thus, we conclude that urine composition remains relatively stable despite exposure to various thermal stresses.

THE PREVALENCE OF MYOPIA IN SCHOOL-AGE CHILDREN IN SLOVAKIA AND THE COVID-19 PANDEMIC.

AIMS: The aim of the authors' study was to determine the prevalence of myopia and premyopia in children and adolescents attending primary school in a village in the north of Slovakia in the period before and after the peak of the Covid-19 pandemic. MATERIAL AND METHODS: Changes in the spherical equivalent and axial length of the eyes were measured in 2019 and 2022 in a group of 47 children and adolescents within the age range of 7-12 years at the time of first measurement. In all the individuals, refraction was measured using an automatic keratorefractometer without the use of cycloplegic eye drops, and axial length was measured with an optical biometer. Refraction was also measured using an automatic keratorefractometer after using cycloplegic eye drops. RESULTS: We recorded a significant change in the mean spherical equivalent (1.02 ±1.16 D vs. 0.72 ±1.29) and the axial length of the eyes (23.05 ±0.72mm vs. 23.30 ±0.74mm) when comparing the measurements from 2019 to 2022. The probands manifested a myopic refractive shift of -0.30 D throughout the entire cohort (-0.24 D in boys and -0.38 D in girls), and the mean axial length increased by 0.2 mm in the entire cohort (by 0.3 mm in boys and 0.2 mm in girls). The percentage of myopic and premyopic eyes increased (4.3% vs. 8.5% a 31.9% vs. 48.9%). CONCLUSION: We noted a decrease in the spherical equivalent and increase in the axial length of the eyes after the Covid-19 pandemic compared to the period before the pandemic in Slovak school-aged children. During this period, the prevalence of myopia and premyopia increased.

COVID-19 and the differences in physiological background between children and adults and their clinical consequences.

The SARS-CoV-2 pandemic has indeed been one of the most significant problems facing the world in the last decade. It has affected (directly or indirectly) the entire population and all age groups. Children have accounted for 1.7 % to 2 % of the diagnosed cases of COVID-19. COVID-19 in children is usually associated with a mild course of the disease and a better survival rate than in adults. In this review, we investigate the different mechanisms which underlie this observation. Generally, we can say that the innate immune response of children is strong because they have a trained immunity, allowing the early control of infection at the site of entry. Suppressed adaptive immunity and a dysfunctional innate immune response is seen in adult patients with severe infections but not in children. This may relate to immunosenescence in the elderly. Another proposed factor is the different receptors for SARS-CoV-2 and their differences in expression between these age groups. In infants and toddlers, effective immune response to viral particles can be modulated by the pre-existing non-specific effect of live attenuated vaccines on innate immunity and vitamin D prophylaxis. However, all the proposed mechanisms require verification in larger cohorts of patients. Our knowledge about SARS-CoV-2 is still developing.

Canakinumab as monotherapy for treatment of familial Mediterranean fever - first report in Central and Eastern Europe region.

Autoinflammatory disorders (AID) are characterized by spontaneous attacks of acute inflammation with a broad spectrum of clinical symptoms. Ongoing inflammation and reoccurrence of acute flares can lead to the development of amyloidosis. One group of AID is represented by monogenic periodic fever syndromes while familial Mediterranean fever (FMF) is the most common form of AID from this group. Its prevalence in Central and Eastern Europe was reported to be very low. We report a case of FMF patient with a very severe clinical course of FMF and intolerance to colchicine, which is a gold standard for FMF treatment. The clinical effect of the application of anakinra was insufficient and accompanied with side effects and low tolerability. Switching to canakinumab (human monoclonal antibody against IL-1ß) at dose of 150 mg every 4 weeks induced a rapid remission of the disease activity and inflammatory markers. However, due to relapse of acute flares after three weeks from application, the escalation of dose to 300 mg every 4 weeks induced a complete remission of symptoms and significantly improved the quality of life. This is the first report of successful canakinumab administration in FMF patient in Central and Eastern Europe, a region with very low incidence of FMF (Tab. 1, Ref. 16).

The Role of Ion Channels to Regulate Airway Ciliary Beat Frequency During Allergic Inflammation.

Overproduction of mucus is a hallmark of asthma. The aim of this study was to identify potentially effective therapies for removing excess mucus. The role of voltage-gated (Kir 6.1, KCa 1.1) and store-operated ion channels (SOC, CRAC) in respiratory cilia, relating to the tracheal ciliary beat frequency (CBF), was compared under the physiological and allergic airway conditions. Ex vivo experiments were designed to test the local effects of Kir 6.1, KCa 1.1 and CRAC ion channel modulators in a concentration-dependent manner on the CBF. Cilia, obtained with the brushing method, were monitored by a high-speed video camera and analyzed with ciliary analysis software. In natural conditions, a Kir 6.1 opener accelerated CBF, while CRAC blocker slowed it in a concentration-dependent manner. In allergic inflammation, the effect of Kir 6.1 opener was insignificant, with a tendency to decrease CBF. A cilio-inhibitory effect of a CRAC blocker, while gently reduced by allergic inflammation, remained significant. A KCa 1.1 opener turned out to significantly enhance the CBF under the allergic OVA-sensitized conditions. We conclude that optimally attuned concentration of KCa 1.1 openers or special types of bimodal SOC channel blockers, potentially given by inhalation, might benefit asthma.

Airway Defense Control Mediated via Voltage-Gated Sodium Channels.

Expression of voltage-gated sodium channels (Nav) takes place in the airways and the role of Nav1.7 and Nav1.8 in the control of airway's defense reflexes has been confirmed. The activation of Nav channels is crucial for cough initiation and airway smooth muscle reactivity, but it is unknown whether these channels regulate ciliary beating. This study evaluated the involvement of Nav1.7 and Nav1.8 channels in the airway defense mechanisms using their pharmacological blockers in healthy guinea pigs and in the experimental allergic asthma model. Asthma was modeled by ovalbumin sensitization over a period of 21 days. Blockade of Nav1.7 channels significantly decreased airway smooth muscle reactivity in vivo, the number of cough efforts, and the cilia beat frequency in healthy animals. In the allergic asthma model, blockade of Nav1.8 efficiently relieved symptoms of asthma, without adversely affecting cilia beat frequency. The study demonstrates that Nav1.8 channel antagonism has a potential to alleviate cough and bronchial hyperreactivity in asthma.

[Factors modifying cord blood IgE levels - a pilot study]. / Faktory modifikujúce koncentráciu IgE v pupocníkovej krvi - pilotná stúdia.

AIM: The complex influence of internal and external environmental factors on the individual and his/her immune system and the lack of suitable markers to assess and reduce the risk of the development of allergies during the lifetime can explain the continuous increase in the number of people affected by some form of allergy. According to the results of some studies, cord blood IgE level could be a useful early marker for assessing the risk of atopic diseases, but the studies showed controversial results. In addition, several authors discuss the origin of these antibodies (synthesis in utero, peripartum contamination from maternal blood or placental transfer). The aim of our pilot study was to investigate the possible influence of modifying factors on cord blood IgE level. MATERIAL AND METHODS: Our group of patients consisted of 184 retrospectively selected neonates (98 boys, 53.3% and 86 girls, 46.7%) from whom cord blood was collected and cord blood IgE level was measured 25 years ago (PRIST method). The impact of selected modifying factors (sex, type of delivery or month of birth) on cord blood IgE level was assessed retrospectively. RESULTS: Higher cord blood IgE levels were found in boys than in girls, in neonates born by Caesarean section than in those born by natural delivery, and in those born in the winter months than in other seasons of the year. Our findings are in agreement with those of other authors. CONCLUSION: Based on our results and those of others, we assume that the selected factors affect the cord blood IgE levels to varying degrees. These facts should be taken into consideration while interpreting the cord blood IgE levels.

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.

Acid sensitivity of the spinal dorsal root ganglia C-fiber nociceptors innervating the guinea pig esophagus.

BACKGROUND: Gastroesophageal reflux can cause high acidity in the esophagus and trigger heartburn and pain. However, because of the esophageal mucosal barrier, the acidity at the nerve terminals of pain-mediating C-fibers in esophageal mucosa is predicted to be substantially lower. We hypothesized that the esophageal dorsal root ganglia (DRG) C-fibers are activated by mild acid (compared to acidic reflux), and express receptors and ion channels highly sensitive to acid. METHODS: Extracellular single unit recordings of activity originating in esophageal DRG C-fiber nerve terminals were performed in the innervated esophagus preparation ex vivo. Acid was delivered in a manner that bypassed the esophageal mucosal barrier. The expression of mRNA for selected receptors in esophagus-specific DRG neurons was evaluated using single cell RT-PCR. KEY RESULTS: Mild acid (pH = 6.5-5.5) activated esophageal DRG C-fibers in a pH-dependent manner. The response to mild acid at pH = 6 was not affected by the TRPV1 selective antagonist iodo-resiniferatoxin. The majority (70-95%) of esophageal DRG C-fiber neurons (TRPV1-positive) expressed mRNA for acid sensing ion channels (ASIC1a, ASIC1b, ASIC2b, and/or ASIC3), two-pore-domain (K2P) potassium channel TASK1, and the proton-sensing G-protein coupled receptor OGR1. Other evaluated targets (PKD2L1, TRPV4, TASK3, TALK1, G2A, GPR4, and TDAG8) were expressed rarely. CONCLUSIONS & INFERENCES: Guinea pig esophageal DRG C-fibers are activated by mild acid via a TRPV1-independent mechanism, and express mRNA for several receptors and ion channels highly sensitive to acid. The high acid sensitivity of esophageal C-fibers may contribute to heartburn and pain in conditions of reduced mucosal barrier function.

Acid infusion into the esophagus increases the number of meal-induced transient lower esophageal sphincter relaxations (TLESRs) in healthy volunteers.

BACKGROUND: Transient lower esophageal sphincter relaxation (TLESR) is the major mechanism of gastroesophageal reflux (GER) but the regulation of TLESR by stimuli in the esophagus is incompletely understood. If stimuli in the esophagus can influence TLESR, then such regulation may perpetuate or limit GER. We addressed the hypothesis that acid in the esophagus enhances TLESRs. METHODS: We evaluated the effect of acid infusion into the distal esophagus on TLESRs evoked by a standard meal in a paired randomized study in healthy subjects. TLESRs were evaluated by using high resolution manometry (HRM). KEY RESULTS: We found that acid in the esophagus enhanced meal-induced TLESRs. Compared to control infusion the number of TLESRs (median [interquartile range]) was increased during 2 h following the acid infusion (11 [9-14] vs 17 [12.5-20], p < 0.01). The average duration of individual TLESRs was not affected. The time-course analysis revealed that a robust increase in TLESRs occurred already in the first hour when the number of TLESRs nearly doubled (6 [5.5-7.5] vs 11 [7.5-12.5], p < 0.05). In contrast to the enhancement of TLESRs, the number of swallows was not changed. CONCLUSIONS & INFERENCES: The acid infusion into the esophagus increases the number of meal-induced TLESRs in healthy subjects. Our results provide evidence for the concept that the stimuli in the esophagus can influence TLESRs. The regulation of TLESR by stimuli in the esophagus may contribute to pathogenesis of GER in some patients.

Successful treatment of extreme acute lead intoxication.

Severe acute lead intoxications are rare and are associated with accidental or purposeful ingestion. There were only few cases of severe to fatal poisonings reported in literature in children. We report a case of acute lead intoxication in a child with extremely high lead blood level of 20.4 micromol/L (422.7 microg/dL), who was treated with chelation and in whom significant organ dysfunction did not develop. Documented significant high level above 3.37 micromol/L (corresponding to 70 microg/dL) in this patient persisted for approximately 24 h. Adequate, single or combined chelatation therapy in early phase of acute lead poisoning is essential for the further patient's outcome.

Association of oxidative stress and GST-T1 gene with childhood bronchial asthma.

There is increasing evidence that bronchial asthma is associated with increased oxidative stress. Reactive oxygen species are produced in each organism as by-products of several essential reactions and can damage biomolecules. Asthma is a complex chronic inflammatory disorder of the airways, with many candidate genes suspected as being important in its development. Glutathione S-transferase (GST) is such a gene due to its role in protection against oxidative stress. In the present study, we examined the hypothesis that increased oxidative stress and polymorphism in the GST-T1 gene are associated with childhood asthma. We found that the amount of sulfhydryl groups significantly decreased and the content of thiobarbituric acid-reactive substances increased in the group of asthmatic children, compared with healthy controls. The GST-T1 null genotype was more frequent among the asthma patients. These results suggest that the GST-T1 null genotype and increased oxidative stress may play a role in the asthma pathogenesis in children.

Cough reflex sensitivity in various phenotypes of childhood asthma.

Cough is a major symptom in some children with asthma, but the relationship between cough and the severity of asthma is defined insufficiently. As cough represents common problem of pediatrics, several objective methods for its assessment were developed. Cough reflex sensitivity (CRS) test with capsaicin is one of the most important tools for studying cough. In the present study, we aimed to study the CRS in various phenotypes of childhood asthma. We found that, in general, CRS was increased in asthmatic children compared with controls. The most evident increase of CRS was observed during acute asthma exacerbation, in children suffering from asthma with concomitant allergic rhinitis, and in atopic asthmatics. Interestingly, we noted a significant decline in lung function after capsaicin CRS. Various laboratory and clinical characteristics of asthmatic children influence cough sensitivity to a different extent. Cough reflex sensitivity measurement can add valuable information beside the commonly used spirometric and inflammometric methods in the management of asthmatic children.

Factors attributable to the level of exhaled nitric oxide in asthmatic children.

BACKGROUND: Asthma is a heterogeneous disease with variable symptoms especially in children. Exhaled nitric oxide (FeNO) has proved to be a marker of inflammation in the airways and has become a substantial part of clinical management of asthmatic children due to its potential to predict possible exacerbation and adjust the dose of inhalant corticosteroids. OBJECTIVES: We analyzed potential factors that contribute to the variability of nitric oxide in various clinical and laboratory conditions. MATERIAL AND METHODS: Study population consisted of 222 asthmatic children and 27 healthy control subjects. All children underwent a panel of tests: fractioned exhaled nitric oxide, exhaled carbon monoxide, asthma control test scoring, blood sampling, skin prick tests, and basic spirometry. RESULTS: FeNO and other investigated parameters widely changed according to clinical or laboratory characteristics of the tested children. Asthmatics showed increased levels of FeNO, exhaled carbon monoxide, total serum IgE, and higher eosinophilia. Boys had higher FeNO levels than girls. We found a significant positive correlation between FeNO levels and the percentage of blood eosinophils, %predicted of forced vital capacity, total serum IgE levels, and increasing age. CONCLUSIONS: Various phenotypes of children's asthma are characterized by specific pattern of the results of clinical and laboratory tests. FeNO correlates with total serum IgE, blood eosinophilia, age, and some spirometric parameters with different strength. Therefore, the coexistence of atopy, concomitant allergic rhinitis/rhinoconjunctivitis, and some other parameters should be considered in critical evaluation of FeNO in the management of asthmatic children.

Cough reflex sensitivity in adolescents with diabetic autonomic neuropathy.

OBJECTIVE: Diabetic autonomic neuropathy (DAN) is one of the chronic complications of diabetes mellitus which can involve one or more organ systems. DAN without apparent symptoms is more often in childhood and adolescence. While heart rate variability (HRV) and Ewing's battery of cardiovascular tests are regarded as a gold standard for the diagnosis of DAN, the examination of cough reflex sensitivity (CRS) is another possibility. The aim of this study was to compare HRV and CRS in children with diabetes mellitus. MATERIAL AND METHODS: Sixty one patients (37 girls, 24 boys) aged 15-19 suffering from diabetes mellitus type 1 completed the study. Based on HRV, patients were divided into 2 groups - with DAN (n=25) and without DAN (n=32), 4 patients were excluded because of ambiguous results. CRS was studied in each patient by inhalation of gradually increasing concentration of capsaicin. RESULTS: Subjects with DAN required a significantly higher concentration of capsaicin needed to evoke 2 coughs (median 625 micromol/l, IQR 68.4-625.0 micromol/l vs. median 29.3 micromol/l, IQR 9.8-156.3 micromol/l, P<0.001) and 5 coughs (median 2500.0 micromol/l, IQR 1250.0-2500.0 micromol/l vs. median 312.5 micromol/l, IQR 117.2-625.0 micromol/l, P<0.001) compared with those without DAN. Moreover, a strong negative correlation between HRV and CRS was established. CONCLUSION: Diabetes mellitus lowers the cough response. Cough reflex sensitivity appears to be another sensitive method for the evaluation of DAN in diabetes.

Cardiac autonomic control in adolescents with primary hypertension.

BACKGROUND: Impairment in cardiovascular autonomic regulation participates in the onset and maintenance of primary hypertension. OBJECTIVE: The aim of the present study was to evaluate cardiac autonomic control using long-term heart rate variability (HRV) analysis in adolescents with primary hypertension. SUBJECTS AND METHODS: Twenty two adolescent patients with primary hypertension (5 girls/17 boys) aged 14-19 years and 22 healthy subjects matched for age and gender were enrolled. Two periods from 24-hour ECG recording were evaluated by HRV analysis: awake state and sleep. HRV analysis included spectral power in low frequency band (LF), in high frequency band (HF), and LF/HF ratio. - RESULTS: In awake state, adolescents with primary hypertension had lower HF and higher LF and LF/HF ratio. During sleep, HF was lower and LF/HF ratio was higher in patients with primary hypertension. CONCLUSIONS: A combination of sympathetic predominance and reduced vagal activity might represent a potential link between psychosocial factors and primary hypertension, associated with increased cardiovascular morbidity.

Exhaled carbon monoxide as a new marker of respiratory diseases in children.

Among modern methods included in diagnostic algorithms for various diseases, analyses of expired breath and its condensate acquire increasing importance. Various markers can be determined in the exhaled air, especially volatile gaseous compounds: nitrogen oxide (NO), carbon monoxide (CO), hydrocarbons and 8-isoprostanes. In contrast to NO, CO can serve as a marker of inflammation and oxidation stress. The representation of CO in the exhaled breath (eCO) changes in various diseases of the respiratory and other systems. Among the respiratory diseases in which the use of eCO measurement seems to be perspective and beneficial are bronchial asthma, airways infections, cystic fibrosis, and primary ciliary dyskinesia. The observation of eCO concentrations represents a modern, simple, available, and well reproducible method for the diagnosis of many diseases of respiratory system in children and for the observation of progression, severity of the disease, and response to therapy.