The work deals with the evaluation Calprotectin levels in healthy children and pediatric patients in the Czech Republic.

ABSTRACT Background Calprotectin is a protein found in the cytoplasm of neutrophile granulocytes. In the course of inflammatory bowel disease (IBD), calprotectin is released during chronic inflammation in the gut. Activation of neutrophils during the inflammation is followed by activation and secretion of pro-inflammatory molecules such as calprotectin. Calprotectin is stable in stool up to 7 days and can therefore be used as a non-invasive marker for diagnosis, treatment and measurement of the disease activity in patients with IBD. The most common method for analysis of calprotectin concentration is ELISA. This method is time-consuming and many manufactures have therefore developed rapid tests as a faster alternative for quantification of calprotectin in stool. Aim The aim of the study was to evaluate one ELISA and one rapid test from the same manufacture compare the data with the existing ELISA-method used in the laboratory for routine analysis. Methods A rapid test (CalFast) and an ELISA method (CalPrest) from Eurospital, were used for analysis of calprotectin in stool. These two methods were compared with known concentrations of calprotectin obtained by the ELISA method from Bühlmann used in the routine work.  Results The results showed poor correlation between the rapid test and the ELISA method. Furthermore, the comparison between the two ELISA-methods showed a poor correlation. Conclusion Evaluation of the two new methods showed poor correlation with the existing ELISA method from Bühlmann. Evaluation of the rapid test did not show any correlation with the two ELISA methods and the data cannot be trusted. It is difficult to conclude which of the two ELISA methods gives accurate results due to the absence of an international standard.

Background Calprotectin is a protein expressed in the cytoplasm inside the neutrophile granulocytes. During inflammatory bowel disease (IBD), the neutrophile granulocytes are involved in a complex interaction at the inflammatory area where they die and release their content into the intestinal lumen. Therefore, calprotectin in stool is a suitable marker for diagnosis and measurement of the disease-activity in patients with IBD. The most commonly used method to detect calprotectin in stool is ELISA, but the process of manual preparation of stool samples is time-consuming. Aim The objective of the study was to evaluate an extraction method that could replace manual preparation of fecal samples and to compare different methods for measuring Calprotectin in stool using two ELISA-methods from two manufacturers and one rapidtest. Methods For extraction of calprotectin from stool samples we used sample collector tubes from Epitope Diagnostics and fecal preparation kits from Roche. Two different ELISA-kits for measuring calprotectin concentration in stool were compared. Measurements of calprotectin with rapid-test from Epitope Diagnostics were also performed and were compared with the two ELISA kits. Results The results indicate a poor correlation between two extraction methods with Sample Collector Tube and Roche preparation kit. The comparison between the two ELISA-kits showed poor correlation. Evaluation of rapid test showed 33% false negative results with a cut-off value at 50 mg/kg. Conclusion Evaluation of products from Epitope Diagnostics showed poor correlation with the Bühlmann ELISA and an unreliable rapid test. Therefore, none of evaluated products from Epitope Diagnostics is accurate enough to be used for clinical diagnosis in the laboratory.

Sepsis and septic shock are conditions, with severe outcome or in many cases death. Sepsis is a systemic inflammatory response trigger by bacteraemia but systemic inflammatory response can also be triggered by major trauma, major surgery, pancreatitis, severe burns etc. The systemic inflammatory reaction initiating the evolvement of septic organ dysfunction can be modelled using endotoxin, a Gram-negative bacterial lipopolysaccharide. This thesis used a porcine experimental sepsis model to examine timing of the inflammatory response due to endotoxin infusion (Paper I) and the influence of steroid treatment on the inflammatory response in endotoxemic pigs (Paper II). Timing of steroid treatment and the role of neutrophil granulocyte activation was evaluated with pig specific NGAL assessing neutrophil activation (Paper III). A clinical observational study was performed with the aim to differentiate between sepsis and other inflammatory conditions (e.g. trauma due to major surgery) evaluated by calprotectin as a marker of neutrophil activation (Paper IV). There was a dose-dependency in endotoxin tolerance which was measured with TNF-a. Pre-exposure to endotoxin did not reduce the pulmonary response to endotoxemic challenge. In fact, both PaO2 / FiO2 and static pulmonary compliance were reduced in this group when pre-treated with endotoxin at low dose. Endotoxemic animals treated with hydrocortisone were more stable in circulatory variables than those without such treatment. This was not explained by an ability of steroids to modulate the production of NO (Nitric oxide), which has been suggested to be a mechanism of steroids in this aspect. Pre-treatment with hydrocortisone attenuated the neutrophil granulocyte response and consequently diminished the release of NGAL in plasma. Circulatory derangement was associated with high plasma NGAL levels. Urine NGAL levels did not differ among the four groups. Plasma calprotectin levels on ICU admission is a sensitive marker of systemic inflammation and are markedly increased in patients with sepsis and patients with systemic inflammatory response. Plasma Calprotectin performed better than any of the other inflammatory variables in predicting mortality at 30 days, except from the composite mortality prediction score, SAPS 3.

No description available.

The obligate human pathogen Neisseria gonorrhoeae successfully overcomes host strategies to limit essential nutrients, termed “nutritional immunity” by expression of TonB-dependent transporters (TdTs): outer membrane receptors that facilitate nutrient transport in an energy-dependent manner. N. gonorrhoeae encodes eight TdTs, five of which facilitate utilization of iron or iron-chelates from host derived proteins including transferrin, lactoferrin and hemoglobin, in addition to siderophores from neighboring bacteria. The transferrin utilization system was previously shown to be critical for establishing infection in human males; demonstrating the possible contributions of TdTs to gonococcal pathogenesis. As such, studies describing the biological function and contribution to pathogenesis of the remaining three uncharacterized TdTs (TdfG, TdfH and TdfJ) are needed. In this study we report that neither TdfG, TdfH nor TdfJ are heme receptors as gonococcal heme utilization occurs passively, independent of energy derived from the TonB system. We also report that TdfH and TdfJ are zinc (Zn) regulated and identify virulence associated regulators that modulate expression of these TdTs, which is in some cases strain-specific. We report that both TdfH and TdfJ contribute to Zn acquisition in N. gonorrhoeae and we characterize TdfH as a calprotectin receptor. Calprotectin, an immune effector protein highly expressed in neutrophils, has antimicrobial activity due to its ability to sequester Zn and Mn. We present evidence that TdfH confers resistance to calprotectin and that TdfH facilitates gonococcal calprotectin binding and Zn accumulation in the presence or absence of calprotectin. Finally, we demonstrate that TdfH expression enhances N. gonorrhoeae NET survival. These studies identify for the first time a novel gonococcal defense strategy to host-mediated nutritional immunity, in which N. gonorrhoeae, via the TdT TdfH, utilizes calprotectin as a Zn source neutralizing its antimicrobial activity. These studies have yielded novel insights into the function and regulation of TdfG, TdfH and TdfJ in N. gonorrhoeae and have laid the framework for future investigation of TdT-mediated Zn acquisition and its role in bacterial pathogenesis.

Apie 70% vaikų, sergančių lėtiniais artritais, suaugusiųjų amžių pasiekia su tam tikru negalios lygiu ir kasdienės veiklos apribojimu. Todėl ypač aktualu nagrinėti vaikams lėtinių artritų priežastis bei prognostinius veiksnius ankstyvose sąnarių ligos stadijose. Mūsų darbo tikslas buvo nustatyti ar ūminėje ligos fazėje nustatomų laboratorinių, imunologinių ir interleukinų (MRP8/14 (kalprotektinas), IL-6, IL-33) rodiklių pokyčiai, funkcinės būklės vertinimas bei ligos aktyvumas gali prognozuoti vaikams lėtinę sąnarių ligą. Atliktas tyrimas parodė, kad artritu susirgusiems vaikams MRP8/14 (kalprotektino) ir Il-6 koncentracijos buvo reikšmingai didesnės nei kontrolinės grupės vaikams tiek serume, tiek sinoviniame skystyje (p<0,01). Kalprotektinas ir IL-6 ūminėje ligos stadijoje stipriai koreliavo su ligos aktyvumo rodikliais (ENG, CRB, sąnarių pobūdžiu (poliartritu), rytiniu sąnarių sustingimu). Taip pat nustatėme, kad didelės kalprotektino (> 5785 ng/ml) ir IL-6 (> 5,5 pg/ml) koncentracijos ūminėje artrito stadijoje buvo nustatytos tiems pacientams, kuriems sąnarių liga įgavo lėtinę eigą. Vertinome tai, kaip reikšmingus lėtinės sąnarių ligos kriterijus. Šiame darbe sergančių artritų vaikų funkcinė būklė ir vaiko gyvenimo kokybė pirmą kartą Lietuvoje buvo įvertinta naujai įdiegiamu ne tik Lietuvoje, bet ir pasaulyje, įvairiapusiu jaunatvinių artritų vertinimo klausimynu JAMAR, bei duomenys palyginti su seniau Lietuvoje naudojamu vaiko sveikatos būklės vertinimo klausimynu... [toliau žr. visą tekstą] / The earlier studies proved that nearly 70% of children with chronic arthritis reach the adulthood with a certain level of disability and restriction of daily activities. Therefore, it is very important to analyse the causes of arthritis and prognostic factors in the early stages of disease. The aim of our study was to determine whether the changes of laboratory, immunological and interleukins (MRP8/14 (calprotectin), IL-6, IL-33) indexes, assessment of functional condition, and disease activity may predict a chronic arthritis in children. The study showed that MRP8/14 (calprotectin) and Il-6 concentrations in the serum and synovial fluid of children with arthritis were significantly higher than in the control group of healthychildren (p<0.01). During the acute stage of disease, calprotectin and IL-6 concentrations strongly correlated with variables of disease activity (ESR, CRP, polyarthritis, morning stiffness of joints). Also, the analysis showed that high concentrations of calprotectin (> 5785ng/ml) and IL-6 (> 5.5 pg/ml) at the acute stage of arthritis were determined for those patients, whose were developed chronic arthritis. It was evaluated as significant criterion of the chronic joint disease. For the first time in Lithuania, the functional ability and the quality of life in children with arthritis were assessed using a newly introduced multidimensional assessment questionnaire for juvenile arthritis (JAMAR), and data was compared... [to full text]

Apie 70% vaikų, sergančių lėtiniais artritais, suaugusiųjų amžių pasiekia su tam tikru negalios lygiu ir kasdienės veiklos apribojimu. Todėl ypač aktualu nagrinėti vaikams lėtinių artritų priežastis bei prognostinius veiksnius ankstyvose sąnarių ligos stadijose. Mūsų darbo tikslas buvo nustatyti ar ūminėje ligos fazėje nustatomų laboratorinių, imunologinių ir interleukinų (MRP8/14 (kalprotektinas), IL-6, IL-33) rodiklių pokyčiai, funkcinės būklės vertinimas bei ligos aktyvumas gali prognozuoti vaikams lėtinę sąnarių ligą. Atliktas tyrimas parodė, kad artritu susirgusiems vaikams MRP8/14 (kalprotektino) ir Il-6 koncentracijos buvo reikšmingai didesnės nei kontrolinės grupės vaikams tiek serume, tiek sinoviniame skystyje (p<0,01). Kalprotektinas ir IL-6 ūminėje ligos stadijoje stipriai koreliavo su ligos aktyvumo rodikliais (ENG, CRB, sąnarių pobūdžiu (poliartritu), rytiniu sąnarių sustingimu). Taip pat nustatėme, kad didelės kalprotektino (> 5785 ng/ml) ir IL-6 (> 5,5 pg/ml) koncentracijos ūminėje artrito stadijoje buvo nustatytos tiems pacientams, kuriems sąnarių liga įgavo lėtinę eigą. Vertinome tai, kaip reikšmingus lėtinės sąnarių ligos kriterijus. Šiame darbe sergančių artritų vaikų funkcinė būklė ir vaiko gyvenimo kokybė pirmą kartą Lietuvoje buvo įvertinta naujai įdiegiamu ne tik Lietuvoje, bet ir pasaulyje, įvairiapusiu jaunatvinių artritų vertinimo klausimynu JAMAR, bei duomenys palyginti su seniau Lietuvoje naudojamu vaiko sveikatos būklės vertinimo klausimynu... [toliau žr. visą tekstą] / The earlier studies proved that nearly 70% of children with chronic arthritis reach the adulthood with a certain level of disability and restriction of daily activities. Therefore, it is very important to analyse the causes of arthritis and prognostic factors in the early stages of disease. The aim of our study was to determine whether the changes of laboratory, immunological and interleukins (MRP8/14 (calprotectin), IL-6, IL-33) indexes, assessment of functional condition, and disease activity may predict a chronic arthritis in children. The study showed that MRP8/14 (calprotectin) and Il-6 concentrations in the serum and synovial fluid of children with arthritis were significantly higher than in the control group of healthychildren (p<0.01). During the acute stage of disease, calprotectin and IL-6 concentrations strongly correlated with variables of disease activity (ESR, CRP, polyarthritis, morning stiffness of joints). Also, the analysis showed that high concentrations of calprotectin (> 5785ng/ml) and IL-6 (> 5.5 pg/ml) at the acute stage of arthritis were determined for those patients, whose were developed chronic arthritis. It was evaluated as significant criterion of the chronic joint disease. For the first time in Lithuania, the functional ability and the quality of life in children with arthritis were assessed using a newly introduced multidimensional assessment questionnaire for juvenile arthritis (JAMAR), and data was compared... [to full text]

The overall aim of this thesis was to study epidemiological and clinical changes in the natural history of Crohn’s disease, its phenotype, the need for surgery and pharmacological therapy over time, as well as the role of faecal calprotectin as a biomarker of pathophysiology and disease course. An increased incidence and prevalence of Crohn’s disease was seen in the period 1963-2010. The proportion of patients with non-stricturing, non-penetrating disease behaviour at diagnosis increased, suggesting that either patients with Crohn’s disease are diagnosed earlier in their disease course today or that the Crohn’s disease phenotype is changing. A decrease in complicated disease behaviour, an increased use of immunomodulators, and a reduced frequency of surgical procedures five years after Crohn’s diagnosis was observed. The decrease in surgery at five years seemed to be explained mainly by a decrease in early surgery within three months from diagnosis, likely reflecting an increased proportion of patients with non-stricturing, non-penetrating disease. This suggests that the introduction of new treatment alternatives alone does not explain the reduction in surgery rates, and an increasing proportion of patients with uncomplicated disease at diagnosis may also play an important role. Subclinical mucosal inflammation, mirrored by increased NFkB activity and increased neutrophil activity (i.e. FC and MPO expression), was observed in healthy twin siblings in both discordant monozygotic and discordant dizygotic twin pairs with IBD. These findings strongly support the hypothesis of an ongoing subclinical mucosal inflammation at the molecular level in healthy first-degree relatives of IBD patients. Baseline FC as well as consecutive FC measurements predict relapse in IBD. The doubling of FC value increased the risk of relapse by 101% in the following three months. This increased risk attenuates with time by 20% for every three month period since the sample was obtained.

No description available.