[Genetic and molecular background in autoimmune diabetes mellitus]. / Genetický a molekulový podklad vývoja autoimunitného diabetes mellitus.

Type 1 diabetes mellitus (T1 DM) is caused by autoimmune-mediated and idiopathic beta-cell destruction of the pancreatic islets of Langerhans resulting in absolute insulin deficiency. Susceptibility to T1 DM is influenced by both genetic and environmental factors. It is generally believed that in genetically susceptible individuals, the disease is triggered by environmental agents, such as viral infections, dietary factors in early infancy, or climatic influences. Many candidate genes for diabetes have been reported; those within the Major Histocompatibility Complex being among the most important. The most common autoantigens are insulin, glutamic acid decarboxylase 65, insuloma-associated antigen 2, and zinc transporter ZnT8. The destruction of beta-cells is mediated mainly by cellular mechanisms; antibodies only seem to reflect the ongoing autoimmune processes and are not directly involved in the tissue damage. They, however, appear prior to the onset of insulin deficiency which makes them suitable for use in the prevention of the disease.

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.

Comparison of selected skin decontaminant products and regimens against VX in domestic swine.

An anesthetized domestic swine model was used to compare the efficacy and cross-contamination potential of selected skin decontaminant products and regimens against the chemical warfare agent, VX. Animals topically exposed to 2x, 3x or 5x LD(50) VX showed typical signs of organophosphate nerve agent poisoning, including miosis, salivation, mastication, dysrhythmias, and respiratory distress prior to death. Animals were exposed to 5x LD(50) VX and then decontaminated 45 min later with the reactive skin decontamination lotion (RSDL), Fuller's earth (FE), 0.5% hypochlorite, or soapy water. Survival was 100% when the reactive skin decontamination lotion or FE was utilized, although 50% of Fuller's earth-decontaminated animals exhibited serious signs of VX poisoning. Decontamination of VX-treated animals with 0.5% hypochlorite was less effective but also increased survival. Soapy water was ineffective in preventing lethality. Blood cholinesterase levels were not predictive of clinical outcome in decontaminated animals. The potential of "decontaminated" VX in open wounds to cause poisoning was assessed by vigorously mixing 5x LD(50) VX with the test decontaminants for 5 min and then placing the mixture onto a full-thickness skin wound. Soapy water was ineffective in preventing lethality. Although treatment with dry Fuller's earth prevented death and all signs of organophosphate poisoning, a significant proportion of treated animals decontaminated with Fuller's earth in aqueous suspension exhibited serious signs of organophosphate poisoning, suggesting that live agent may be desorbed from Fuller's earth when it is exposed to a liquid environment. Animals treated with reactive skin decontamination lotion or 0.5% hypochlorite-VX mixtures showed no signs of organophosphate poisoning during the 6- h test period.

Swine models in the design of more effective medical countermeasures against organophosphorus poisoning.

Although the three most commonly used large mammal species in the safety assessment of drugs remain the dog, the macaque and the marmoset, swine, especially minipigs, have also been widely used over the years in many toxicological studies. Swine present a number of interesting biological and physiological characteristics. Similarities in skin properties with humans have led to extensive in vitro and in vivo studies. There is a specific interest in cardiovascular research, as well as in anaesthesiology and critical care medicine due to common features of swine and human physiology. Although knowledge of swine brain structure and functions remains incomplete, data does exist. The multiple blood sampling that is necessary in pharmacokinetic and toxicokinetic studies are possible, as well as multiparametric monitoring and interventions with equipment used in human clinical settings. Practicality (handling), scientific (stress reduction) and ethical (invasive monitoring) reasons have led research teams to incorporate anaesthesia into their paradigms which makes the analysis of data increasingly difficult. Although not substantiated by scientific data, the swine appears to have an intermediate position in the scale of public perception between non-human primates and animals commonly referred to as pets (i.e. dogs and cats) and rodents. The benefits of the swine model justify the use of these animals in the design of more effective medical countermeasures against known chemical warfare agents (nerve agents, vesicants and lung damaging agents). Exposure to organophosphorus (OP) pesticides represents a severe health issue in developing countries, while OP intoxication with the more lethal military nerve agents is not only of military concern but also a terrorist threat. Tailoring therapeutic regimens to the reality of OP poisoning is of the utmost importance when little experimental data and sparse human clinical data are available in the decision making process. We will present some of the advantages and disadvantages of the swine model in OP countermeasures elaborating on two examples. First, we will present the issues related to the use of anaesthesia during experimental OP poisoning and second we will show how results from experiments with swine can be integrated into a kinetic-based dynamic model to evaluate oxime efficacy. A better knowledge of OP poisoning in swine (comparative toxicokinetics, pharmacokinetics and biochemistry) is definitely necessary before accepting it as a first choice non-rodent model. However, there exists a large amount of data in the model on anaesthesia and different types of shock favouring their use for evaluation of complex situations such as the anaesthesia of OP poisoned patients and combined injuries.

Cough reflex sensitivity in children with suspected and confirmed gastroesophageal reflux disease.

Experimental and clinical studies performed in adults revealed that gastresophageal reflux disease (GORD) is associated with an appreciable increase in cough reflex sensitivity (CRS). The association between respiratory diseases and GORD is also present in children, but there is little evidence that GORD without aspiration of refluxate (proximal reflux) is a frequent cause of cough in children. The aim of this study was to find out whether CRS in children with GORD will be changed compared with healthy children, and if so, to determine the role of proximal vs. distal reflux in these changes. CRS and 24-h esophageal pH monitoring were performed in 20 children of whom 13 had confirmed GORD and 7 were suspected to have GORD. The control group consisted of 27 healthy children. For assessing the CRS, each subject inhaled 12 capsaicin aerosol concentrations (0.61-1250 micromol/l) at 1 min intervals. CRS was defined as the lowest capsaicin concentration that evoked minimally 2 coughs (C2). CRS in the group of children with suspected GORD [C2: 17.0 micromol/l (6.4-45.6 micromol/l)] and with confirmed GORD [C2: 13.4 micromol/l (3.6-50.9 micromol/l)] were significantly elevated (P<0.05) compared with healthy children [C2: 72.1 micromol/l (25.5-203.9 micromol/l)]. According to the parameters of 24-h pH monitoring, a significantly higher exposure to acid was present in the distal compared with proximal oesophagus. CRS changes correlated negatively with the distal, but not proximal, esophageal acid exposure. In conclusion, CRS changes in children suffering from GORD are similar to those described in adult patients with GORD. It is plausible that the main role in increased CRS in children with GORD play episodes of distal acid refluxes.

Enhanced forebrain nitric oxide synthase activity in epileptic fowl.

Nitric oxide synthase (NOS) activity was examined in forebrain, cerebellum and optic lobes of adult domestic fowl, having a hereditary primary generalized convulsive disorder. NOS was approximately 2-fold higher in only the forebrain of adult epileptic fowl compared to non-epileptic (carrier) hatchmates. A significant increase in NOS was also evident in forebrains of 1-day-old epileptic chicks. Ca(2+)-dependency experiments confirmed that these increments were principally due to type I NOS (NOS-I). Induction of convulsions by intermittent photic stimulation did not affect pre-existing forebrain NOS-I activity. The present data suggest that an enhanced NO signaling may ensue in selected regions of the brain as an adaptive response to hereditary epileptogenesis.

Immobilization of Russian VX skin depots by localized cooling: implications for decontamination and medical countermeasures.

The chemical weapon nerve agent known as Russian VX (VR) is a potent organophosphorus (OP) compound that is much less studied than its VX analogue with respect to toxicity, as well as to the effectiveness of several known countermeasures against it. An anaesthetized domestic swine model was utilized to assess several approaches in mitigating its toxicity, including the utility of cooling VR treated skin to increase the therapeutic window for treatment. The 6h LD50 for VR topically applied on the ear was 100 µg/kg. Treatment of VR exposed animals (5 × LD50) with pralidoxime (2PAM) very poorly regenerated inhibited blood cholinesterase activity, but was partially effective in preventing signs of OP poisoning and increasing survival. In contrast, treatment with the Hagedorn oxime HI-6 reactivated cholinesterase, eliminated all signs of poisoning and prevented death. Decontamination with the Reactive Skin Decontaminant Lotion (RSDL) 15 min after VR exposure was completely effective in preventing death. Cooling of the VR exposure sites for 2 or 6h prevented signs of OP poisoning and death during the cooling period. However, these animals died very quickly after the cessation of cooling, unless they were treated with oxime or decontaminated with RSDL. Blood analyses showed that cooling of agent exposure sites delayed the entry of VR into the bloodstream. Medical treatment with HI-6 and to a lesser extent 2PAM, or decontamination with RSDL are effective in protecting against the toxic effects of cutaneous exposure to VR. Immobilizing this agent (and related compounds) within the dermal reservoir by cooling the exposure sites, dramatically increases the therapeutic window in which these medical countermeasures are effective.

The therapeutic use of localized cooling in the treatment of VX poisoning.

The organophosphate (OP) nerve agent VX is a weaponized chemical warfare agent that has also been used by terrorists against civilians. This contact poison produces characteristic signs of OP poisoning, including miosis, salivation, mastication, dysrhythmias and respiratory distress prior to death. Although successful treatment of OP poisoning can be obtained through decontamination and/or oxime reactivation of agent-inhibited cholinesterase, medical countermeasures that increase the therapeutic window for these measures would be of benefit. An anaesthetized swine model was utilized to examine the effects of lethal VX exposure to the skin, followed by cooling the exposure site prior to decontamination or treatment. The cooling was simply accomplished by using crushed ice in grip-seal plastic bags applied to the exposure sites. Cooling of skin exposed to lethal doses of VX significantly increased the window of opportunity for successful decontamination using the Reactive Skin Decontaminant Lotion(®) (RSDL(®)) or treatment with the oxime antidotes HI-6 and 2PAM. Analyses of blood VX levels showed that cooling acted to slow or prevent the entry of VX into the bloodstream from the skin. If the exposure site is known, the simple and non-invasive application of cooling provides a safe means with which to dramatically increase the therapeutic window in which decontamination and/or antidote treatment against VX are life-saving.

Toxicokinetics of tabun enantiomers in anaesthetized swine after intravenous tabun administration.

In the present study, we report the first in vivo toxicokinetic study of tabun (O-ethyl-N,N-dimethylphosphoramidocyanidate). The toxicokinetics of the enantiomers of tabun were investigated in anesthetized swine after intravenous administration of 3xLD(50) (161.4mug/kg) tabun. Blood samples were taken for gas chromatographic-mass spectrometric determination of the tabun enantiomers and for measurement of the activity of red blood cell acetylcholinesterase (AChE) and plasma butyrylcholinesterase (BChE). The tabun enantiomers could be quantified in swine blood to a minimum concentration of 3.0pg/ml (18.5pM) and could be detected to a minimum concentration of 1.0pg/ml (6.2pM). The concentration-time profiles of both tabun enantiomers were best described by a bi-exponential equation. The elimination of (+)-tabun and (-)-tabun were comparable in the initial phase. In the terminal phase a remarkable difference was found, with terminal half lives of 11.5min for (+)-tabun and 23.1min for (-)-tabun. (+)-Tabun showed a markedly longer persistence in vivo than (+)-enantiomers of other G-type nerve agents and could be detected in all swine at least up to 30min post-injection, (-)-tabun at least up to 90min post-injection. These results demonstrate a rather rapid elimination of tabun enantiomers in vivo and may provide a toxicokinetic basis for the further development and optimization of medical countermeasures against this nerve agent.

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.