Long-term health sequelae and quality of life at least 6 months after infection with SARS-CoV-2: design and rationale of the COVIDOM-study as part of the NAPKON population-based cohort platform (POP).

PURPOSE: Over the course of COVID-19 pandemic, evidence has accumulated that SARS-CoV-2 infections may affect multiple organs and have serious clinical sequelae, but on-site clinical examinations with non-hospitalized samples are rare. We, therefore, aimed to systematically assess the long-term health status of samples of hospitalized and non-hospitalized SARS-CoV-2 infected individuals from three regions in Germany. METHODS: The present paper describes the COVIDOM-study within the population-based cohort platform (POP) which has been established under the auspices of the NAPKON infrastructure (German National Pandemic Cohort Network) of the national Network University Medicine (NUM). Comprehensive health assessments among SARS-CoV-2 infected individuals are conducted at least 6 months after the acute infection at the study sites Kiel, Würzburg and Berlin. Potential participants were identified and contacted via the local public health authorities, irrespective of the severity of the initial infection. A harmonized examination protocol has been implemented, consisting of detailed assessments of medical history, physical examinations, and the collection of multiple biosamples (e.g., serum, plasma, saliva, urine) for future analyses. In addition, patient-reported perception of the impact of local pandemic-related measures and infection on quality-of-life are obtained. RESULTS: As of July 2021, in total 6813 individuals infected in 2020 have been invited into the COVIDOM-study. Of these, about 36% wished to participate and 1295 have already been examined at least once. CONCLUSION: NAPKON-POP COVIDOM-study complements other Long COVID studies assessing the long-term consequences of an infection with SARS-CoV-2 by providing detailed health data of population-based samples, including individuals with various degrees of disease severity. TRIAL REGISTRATION: Registered at the German registry for clinical studies (DRKS00023742).

Systematic review of the effects of intensive-care-unit noise on sleep of healthy subjects and the critically ill.

Intensive-care-unit (ICU) patients exhibit disturbed sleeping patterns, often attributed to environmental noise, although the relative contribution of noise compared to other potentially disrupting factors is often debated. We therefore systematically reviewed studies of the effects of ICU noise on the quality of sleep to determine to what extent noise explains the observed sleep disruption, using the Cochrane Collaboration method for non-randomized studies. Searches in Scopus, PubMed, Embase, CINAHL, Web of Science, and the Cochrane Library were conducted until May 2017. Twenty papers from 18 studies assessing sleep of adult patients and healthy volunteers in the ICU environment, whilst recording sound levels, were included and independently reviewed by two reviewers. We found that the numbers of arousals between the baseline and the ICU noise condition in healthy subjects differed significantly (mean difference 9.59; 95% confidence interval 2.48-16.70). However, there was considerable heterogeneity between studies (I2 94%, P < 0.00001), and all studies suffered from a considerable risk of bias. The meta-analysis of results was hampered by widely varying definitions of sound parameters between studies and a general lack of detailed description of methods used. It is, therefore, currently impossible to quantify the extent to which noise contributes to sleep disruption among ICU patients, and thus, the potential benefit from noise reduction remains unclear. Regardless, the majority of the observed sleep disturbances remain unexplained. Future studies should, therefore, also focus on more intrinsic sleep-disrupting factors in the ICU environment.

In vivo detection of aflatoxin-induced lipid free radicals in rat bile.

Aflatoxin B1 (AFB1), a potent hepatotoxin and hepatocarcinogen, is metabolized in the liver via cytochrome P-450 to an AFB1-8,9-epoxide intermediate. The formation of the AFB1-8,9-epoxide correlates with the pathological changes observed in numerous mammalian species. Oxidative damage has been postulated to play a major role in the mechanisms associated with AFB1-induced cytotoxicity and carcinogenecity in mammalian species. The aim of this study was to detect and identify free radical intermediates from the hepatic metabolism of AFB1 in vivo. Rat bile ducts were cannulated and rats were treated simultaneously with AFB1 (3 mg/kg i.p.) and the spin trapping agent 4-POBN (alpha-(4-pyridyl-1-oxide)-N-tert-butyl nitrone) (1 g/kg i.p.), and bile was collected over a period of 2 h at 20-min intervals. ESR spectroscopy was used to detect a carbon-centered radical adduct of 4-POBN in rat bile. The effect of metabolic inhibitors, such as deferoxamine mesylate (DFO), an iron chelator, and SKF 525A, a cytochrome P-450 inhibitor, on in vivo aflatoxin-induced free radical formation were also studied. It was found that there was a significant decrease in free radical formation by pre-treatment with both DFO and SKF 525A. This indicates that oxidation of AFB1 generates free radical species via CYP metabolism and an iron-mediated redox mechanism.

Free radical generation in the cochlea during combined exposure to noise and carbon monoxide: an electrophysiological and an EPR study.

Ototoxicity following combined exposure to noise and carbon monoxide (CO) is known to result in more severe permanent threshold shifts than exposure to noise alone. We have previously demonstrated that such potentiation of noise-induced auditory impairment by CO can be prevented by the administration of a nitrone spin-trapping agent. Although such protection implicates injury via free radical pathways, drug-induced protection does not provide direct evidence for the presence of free radicals in the cochlea. The objective of this study was to demonstrate the actual presence of nitrone spin adducts in the cochlea following simultaneous exposure to noise and CO. Using electrophysiological end-points, the protective effects of the nitrone spin-trapping agent alpha-(4-pyridyl-1-oxide)-N-tert-butylnitrone (POBN) were assessed following combined exposure of adult male Long Evans hooded rats to noise and CO. In addition, an ex-vivo evaluation of POBN spin adducts was done by electron paramagnetic resonance spectroscopy (EPR). The noise used was octave band noise with center frequency 13.6 kHz at 100 dB(Lin) for a duration of 2 h. The level of CO used was 1200 ppm. Electrophysiological results demonstrate that POBN protects against combined exposure to noise plus CO. The EPR study demonstrates POBN spin adducts in the cochleae of animals exposed to noise plus CO. Therefore, this study provides evidence to the hypothesis that ototoxicity due to noise plus CO exposure is mediated via free radicals.

Nf-kappab activation is associated with free radical generation and endotoxemia and precedes pathological liver injury in experimental alcoholic liver disease.

Endotoxemia and oxidative stress activate nuclear factor kappa B (NF-kappaB) in alcoholic liver injury. In alcohol-fed rats, activation of NF-kappaB is associated with the development of necro-inflammatory changes in the liver. Whether activation of NF-kappaB occurs prior to development of liver injury is unknown. We determined whether activation of NF-kappaB preceded histopathological liver changes. Male Wistar rats were fed a liquid diet containing ethanol by continuous infusion through permanently implanted gastric tubes. Radical intermediates detected by spin trapping were measured in bile prior to killing. After 2 weeks of treatment, samples of liver tissue were obtained for histopathological examination, for evaluation of NF-kappaB, and determination of messenger RNA levels of cytokines, chemokines and cyclo-oxygenase-2. No pathological changes in liver were seen after 2 weeks of intragastric feeding. However, activation of NF-kappaB was seen in the livers from ethanol-fed rats. In addition, elevated mRNA levels of hepatic pro-inflammatory cytokines (TNF-alpha and IL12), chemokines MIPIalpha and MIP-2) and cyclo-oxygenase-2 were seen in association with activation of NF-kappaB and increased levels of free radicals and endotoxin. Thus, activation of NF-kappaB, associated with elevated mRNA levels of pro-inflammatory stimuli, precedes the histopathological liver changes in experimental alcoholic liver disease in rats.

Antioxidants and gadolinium chloride attenuate hepatic parenchymal and endothelial cell injury induced by low flow ischemia and reperfusion in perfused rat livers.

The objective of this study was to determine whether Kupffer cells contribute to parenchymal and endothelial cell damage induced by ischemia-reperfusion in perfused rat livers. Parenchymal and endothelial cell injury were determined by measuring activities of lactate dehydrogenase (LDH) and purine nucleoside phosphorylase (PNP), respectively, in the effluent perfusate of livers subjected to 60 min of low flow ischemia followed by 30 min of reperfusion. Upon reperfusion, LDH and PNP activities increased significantly within the first 10 min of reperfusion and remained elevated over control values throughout the duration of reperfusion. Pretreatment with gadolinium chloride, an inhibitor of Kupffer cell function, significantly decreased LDH and PNP efflux during reperfusion by approximately 60% and 50%, respectively. When Kupffer cells were stimulated by vitamin A pretreatment, PNP efflux was doubled during reperfusion. Vitamin E pretreatment attenuated LDH and PNP release by approximately 70% during reperfusion compared to enzyme release in untreated livers. Moreover, the water-soluble antioxidants superoxide dismutase and desferrioxamine reduced reperfusion injury, whereas catalase had no effect on enzyme release. These results demonstrate that superoxide anions released from Kupffer cells are involved in oxidative damage to endothelial cells as well as hepatocytes during the early stages of hepatic reperfusion.

Asthma education: creating a partnership.

This article advances the theory that the key to creating an effective partnership is teaching asthma patients what to self-treat, how to self-treat, and when to consult a clinician. The five comanaging rules that the health educator is encouraged to emphasize with the adult asthma patient are: know your own unique asthma symptoms and triggers; keep written records; see appropriate specialists; know your medicines and follow your action plan; and accept no treatment you do not understand. Current research shows asthma to be a chronic inflammatory disorder of the airways. In susceptible individuals, this inflammation causes recurrent episodes of wheezing, breathlessness, chest tightness, and cough, particularly at night and in the early morning. The stepwise approach to asthma therapy divides asthma into several levels of severity. However, patients at any level of severity can have mild, moderate, or severe exacerbations. Asthma triggers; how to use a metered dose inhaler (MDI), a dry powder inhaler (DPI), and a peak flow meter; and how to follow an asthma action plan are thoroughly covered. The last section of the article deals at length with the indications for and actions of long-term-control medications, used to achieve and maintain control of persistent asthma, and quick-relief medications, used to treat symptoms and exacerbations.

Pronounced hepatic free radical formation precedes pathological liver injury in ethanol-fed rats.

BACKGROUND: The role of free radicals in alcoholic liver injury remains uncertain. These experiments were conducted to measure radical formation in rats that were fed alcohol along with either fish oil or saturated fats, which cause different types of liver pathology. METHODS: Liquid diets containing alcohol or isocaloric dextrose were administered to rats by intragastric infusion for 2 weeks. Radical intermediates detected by spin trapping were measured in bile. RESULTS: In rats that were fed alcohol plus fish oil, biliary concentrations of trapped radicals, which most likely originated from lipids, were 6-fold higher than in controls that were fed fish oil plus dextrose. High rates of radical formation persisted 24 hr after alcohol withdrawal, when all alcohol had been metabolized. In contrast, diets containing alcohol and medium chain triglycerides did not stimulate lipid radical formation. CONCLUSIONS: High rates of lipid radical formation were observed only in rats that were fed alcohol in combination with a fish oil diet, and a persistent flux of radical formation continued after alcohol withdrawal. These radical phenomena precede serious liver pathology, which develops after longer periods of fish oil plus alcohol diets.

Effect of low flow ischemia-reperfusion injury on liver function.

The release of liver enzymes is typically used to assess tissue damage following ischemia-reperfusion. The present study was designed to determine the impact of ischemia-reperfusion on liver function and compare these findings with enzyme release. Isolated, perfused rat livers were subjected to low flow ischemia followed by reperfusion. Alterations in liver function were determined by comparing rates of oxygen consumption, gluconeogenesis, ureagenesis, and ketogenesis before and after ischemia. Lactate dehydrogenase (LDH) and purine nucleoside phosphorylase (PNP) activities in effluent perfusate were used as markers of parenchymal and endothelial cell injury, respectively. Trypan blue staining was used to localize necrosis. Total glutathione (GSH + GSSG) and oxidized glutathione (GSSG) were measured in the perfusate as indicators of intracellular oxidative stress. LDH activity was increased 2-fold during reperfusion compared to livers kept normoxic for the same time period whereas PNP activity was elevated 5-fold under comparable conditions. Rates of oxygen consumption, gluconeogenesis, and ureagenesis were unchanged after ischemia, but ketogenesis was decreased 40% following 90 min ischemia. During reperfusion, the efflux rates of total glutathione and GSSG were unchanged from pre-ischemic values. Significant midzonal staining of hepatocyte nuclei was observed following ischemia-reperfusion, whereas normoxic livers had only scattered staining of individual cells. Reperfusion of ischemic liver caused release of hepatic enzymes and midzonal cell death, however, several major liver functions were unaffected under these experimental conditions. These data indicate that there were negligible changes in liver function in this model of ischemia and reperfusion despite substantial enzyme release from the liver and midzonal cell death.

Aromatic hydroxylation in PBN spin trapping by hydroxyl radicals and cytochrome P-450.

Phenyl N-tert-butylnitrone (PBN) is widely used as a spin trapping agent, but is not useful detecting hydroxyl radicals because the resulting spin adduct is unstable. However, hydroxyl radicals could attack the phenyl ring to form stable phenolic products with no electron paramagnetic resonance signal, and this possibility was investigated in the present studies. When PBN was added to a Fenton reaction system composed of 25 mM H(2)O(2) and 0.1 mM FeSO(4), 4-hydroxyPBN was the primary product detected, and benzoic acid was a minor product. When the Fe(2+) concentration was increased to 1.0 mM, 4-hydroxyPBN concentrations increased dramatically, and smaller amounts of benzoic acid and 2-hydroxyPBN were also formed. Although PBN is extensively metabolized after administration to animals, its metabolites have not been identified. When PBN was incubated with rat liver microsomes and a reduced nicotinamide adenine dinculeotide phosphate (NADPH)-generating system, 4-hydroxyPBN was the only metabolite detected. When PBN was given to rats, both free and conjugated 4-hydroxyPBN were readily detected in liver extracts, bile, urine, and plasma. Because 4-hydroxyPBN is the major metabolite of PBN and circulates in body fluids, it may contribute to the pharmacological properties of PBN. But 4-hydroxyPBN formation cannot be used to demonstrate hydroxyl radical formation in vivo because of its enzymatic formation.

Modeling implicit reorganization in continuum descriptions of protein-protein interactions.

The determination of free energies that govern protein-protein recognition is essential for a detailed molecular understanding of biological specificity. Continuum models of macromolecular interactions, in which the solvent is treated by an implicit representation and the proteins are treated semi-microscopically, are computationally tractable for estimating free energies, yet many questions remain concerning their accuracy. This article reports a continuum analysis of the free-energy changes underlying the binding of 31 interfacial alanine substitutions of two complexes of the antihen egg white lysozyme (HEL) antibody D1.3 bound with HEL or the antibody E5.2. Two implicit schemes for modeling the effects of protein and solvent relaxation were examined, in which the protein environment was treated as either homogeneous with a "protein dielectric constant" of epsilon(p) = 4 or inhomogeneous, with epsilon(p) = 4 for neutral residues and epsilon(p) = 25 for ionized residues. The results showed that the nonuniform dielectric model reproduced the experimental differences better, with an average absolute error of +/-1.1 kcal/mol, compared with +/-1.4 kcal/mol for the uniform model. More importantly, the error for charged residues in the nonuniform model is +/-0.8 kcal/mol and is nearly half of that corresponding to the uniform model. Several substitutions were clearly problematic in determining qualitative trends and probably required explicit structural reorganization at the protein-protein interface.

Nursing strategies for smoking cessation.

The purpose of this article is to provide the staff nurse with an overview of the Agency for Health Care Policy and Research Smoking Cessation guidelines. The authors outline the practical components of the guidelines to enable nurses in any clinical setting to implement various smoking cessation strategies according to individual patient needs. Treatment options, including behavioral modification and a comprehensive review of pharmacological therapy, are discussed.

Continuous monitoring of in vivo nitric oxide formation using EPR analysis in biliary flow.

A method to continuously monitor the nitric oxide (NO) level in anesthetized rats, using an in vivo trapping reaction of NO by iron-dithiocarbamate complex, is reported. Previously, we developed a method of monitoring NO in bile samples containing an NO complex excreted from the liver (Anal. Biochem. 243, 8-14, 1996). In the present study, we modified the method so that the bile flows directly through the EPR sample cell. Rats were injected with low doses of lipopolysaccharide (LPS) to induce NO formation and were later anesthetized. After cannulation, the bile duct was connected to the inlet of the EPR sample cell and the trapping agent iron complex of D-N-methylglucamine dithiocarbamate (MGD-Fe) was administered. The EPR signal level from NO complex of MGD-Fe in the flowing bile was continuously monitored. Using this method, immediate changes in in vivo NO level in rats were observed following administration of drugs that can affect NO formation. In addition, a continuous intravenous saline containing MGD-Fe made the EPR signal level stable and improved animal condition as well as survival time. Therefore, this method has two merits; (1) one can continuously monitor NO formation until it reaches the maximum level; (2) a rapid change in NO level after intervention can be followed. Using this method, we tested the effect of the substrate L-arginine and inhibitors for NO synthase activity and NO synthase induction. The sensitivity of the present method was tested by monitoring NO formation in rats after exposure to ionizing radiation.

Free radical formation during ketamine anesthesia in rats: a cautionary note.

Ketamine is a useful anesthetic agent with good analgesic properties; however, when ketamine was used to anesthetize rats for spin trapping studies of alcohol-induced free radicals, liver extracts contained a strong electron paramagnetic resonance (EPR) signal of a novel radical. The same EPR signal was observed in liver extracts when rats which had not received alcohol were anesthetized with ketamine. When ketamine was added to liver microsomes and NADPH, a nitroxide radical derived from ketamine could be detected in organic extracts. When the spin trapping agent POBN was also added, microsomes produced both a ketamine nitroxide radical and a spin adduct. Similar results were obtained during ketamine oxidation by hydrogen peroxide in a tungstate-catalyzed reaction, or in a Fenton reaction system. The data suggest that the secondary amine group of ketamine can be oxidized to a stable nitroxide which produces an EPR spectrum in the absence of a spin trapping agent. The POBN spin adduct detected may be from a carbon-centered radical in the cyclohexanone ring of ketamine. Because several types of radicals can be formed from ketamine, this agent may not be appropriate as an anesthetic for many types of in vivo spin trapping experiments.

Mechanisms for metabolism of ethanol to 1-hydroxyethyl radicals in rat liver microsomes.

Experiments have been designed to reevaluate mechanisms for metabolism of ethanol to 1-hydroxyethyl radicals (HER) in rat liver microsomes. The variables tested include addition of azide, catalase, superoxide dismutase, and deferoxamine, or use of phosphate or Tris buffers. The results indicate that several mechanisms of HER formation are possible, depending on the experimental conditions used to study this process. In the presence of phosphate buffer, which has been used extensively in spite of its ability to chelate iron, HER formation is quite sensitive to changes in hydrogen peroxide availability. These results suggest that Fenton-type reactions produced the oxidizing intermediate responsible for conversion of ethanol to a free radical in phosphate buffer. However, in Tris buffer, HER formation was inhibited markedly by addition of superoxide dismutase, whereas catalase or azide had little effect. These data indicate that the apparent mechanism of radical formation may be influenced by the choice of buffer used. HER formation was almost abolished by the combination of superoxide dismutase and deferoxamine in both buffers, suggesting little enzymatic HER formation by the cytochrome P450 enzymes. When changes in HER formation were compared with rates of ethanol oxidation, it was inferred that 25 to 50% of the acetaldehyde formed during microsomal ethanol oxidation under different experimental conditions could arise via the HER intermediate.

Free radical formation in livers of rats treated acutely and chronically with alcohol.

The spin trapping method was used to assess formation of free radical intermediates in vivo before and after acute alcohol administration to rats. Ascorbyl radicals and spin adducts of dietary alcohol or endogenous compounds, such as lipids, were detected with higher frequency in bile from alcohol-fed rats than in corresponding samples from rats fed control diets. When alcohol was given acutely to these animals, the 1-hydroxyethyl radical metabolite of ethanol was also formed at higher rates in livers of rats that had been fed ethanol chronically. Furthermore, formation of lipid radicals was enhanced after acute alcohol administration. These data support the hypothesis that chronic alcohol administration causes development of oxidative conditions in the liver, which subsequently lead to formation of differing types of radicals. Liver microsomes from alcohol-fed rats also metabolized ethanol to the 1-hydroxyethyl radical at higher rates than controls.

Spin trapping studies of alcohol-initiated radicals in rat liver: influence of dietary fat.

We conducted spin trapping experiments to test the effects of acute and chronic alcohol consumption in livers from rats that had been fed either high fat (35% of energy) or low fat (12% of energy) liquid diets. Rats were anesthetized with isoflurane, the spin trapping agent POBN [alpha-(4-pyridyl-1-oxide)-N-t-butylnitrone] was administered by intravenous injection, and bile samples were collected for electron paramagnetic resonance (EPR) analyses. Two different types of EPR spectra were observed in bile from the animals in these studies. One set of spectral lines was from the 1-hydroxyethyl radical adduct of POBN, which was conclusively identified by injecting the rats with [1-13C]ethanol. The EPR signals of a second type of radical adduct in bile could be observed both before and after acute administration of ethanol. Although the radical(s) responsible for this second series of signals could not be conclusively identified, it is likely that lipid radicals were formed under these conditions and trapped by POBN. For both types of radical adducts, the most intense EPR signals were observed in rats that had been fed alcohol in combination with a high fat diet for 2 wk before the experiments. These results confirm and extend previous data indicating that high levels of dietary fat enhance alcohol-associated free radical formation in the liver.

Comparison of salicylate and D-phenylalanine for detection of hydroxyl radicals in chemical and biological reactions.

Hydroxylation of salicylate and D-phenylalanine was measured to test the usefulness of these compounds for hydroxyl radical (HO(•)) detection in chemical and biological systems. When HO(•) were produced by the photolytic decomposition of hydrogen peroxide, nearly equal amounts of 2,5- and 2,3-dihydroxybenzoic acid (DHBA) were produced from salicylate, with catechol as a minor product. In the photolytic reaction, nearly equal concentrations of p-,m-, and o-tyrosine were formed from D-phenylalanine. When salicylate or D-phenylalanine was present with Fenton reagents or in iron(II) autoxidation systems, the relative proportions of hydroxylated products were similar to those observed after photolysis, although less total products were usually detected. In contrast, when similar experiments were conducted with isolated hepatic microsomes and perfused livers, 2,5-DHBA was the primary product from salicylate, and p-tyrosine was the major product from D-phenylalanine. Cytochrome P-450 enzymes can hydroxylate salicylate to produce 2,5-DHBA, and it is likely that phenylalanine hydroxylase produces most of the p-tyrosine detected in hepatic tissues. Thus, although both salicylate and D-phenylalanine are useful probes for hydroxyl radical formation in chemical systems, hydroxylated products formed from enzymatic reactions complicate interpretation of data from both compounds in vivo.

Hepatic nitric oxide formation: spin trapping detection in biliary efflux.

A new spin trapping method has been developed to continuously monitor nitric oxide (NO) formation in rat liver in vivo. The method is based on the reaction of NO with iron chelates of N-methylglucamine dithio-carbamate (MGD-Fe), resulting in the formation of room-temperature stable EPR-active nitrosyl complexes of MGD-Fe. Rats were injected with various doses of lipopolysaccharide (LPS) to induce NO synthase activity and were later anesthetized with isoflurane. After cannulation of the bile duct, MGD-Fe was administered by iv injection, and samples of bile were collected for EPR analyses. The EPR spectra of bile from LPS-pretreated rats contained characteristic three-line signals of NO trapped by the MGD-Fe complex, while bile from control rats that were not treated with LPS did not contain similar EPR signals. The detection limit of this method was estimated to be 5 microM. Only weak signals from NO could be detected in plasma or urine under these conditions, suggesting that the biliary NO adducts did not originate in extra-hepatic tissues. The reliability of this method was verified by administering an inhibitor for NO synthase induction, alpha-phenyl-N-t-butylnitrone (PBN), or the NO synthase inhibitor N-nitro-L-arginine methyl ester (L-NAME) to LPS-treated rats. NO detected in bile was significantly decreased by both PBN and L-NAME, which is consistent with results obtained from studies using previously established methods for NO formation.