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Novel biomarkers in patients with renal disease
Zakiyanov, Oskar ; Kalousová, Marta (advisor) ; Teplan, Vladimír (referee) ; Průša, Richard (referee)
Chronic kidney disease (CKD) and acute kidney injury (AKI) are major public health problems. It is important to be able to identify those at high risk of adverse outcome, CKD progression and associated cardiovascular disease. The aim of the thesis was to study novel promising biomarkers, their relationship to kidney function, chronic inflammation and/or cardiovascular risk - placental growth factor (PlGF), pregnancy associated plasma protein A (PAPP-A), matrix metalloproteinase 2 (MMP-2), matrix metalloproteinase 9 (MMP-9), soluble receptor for advanced glycation end products (sRAGE), calcium binding protein S100A12 or extracellular newly identified RAGE binding protein (EN-RAGE), and high mobility group box protein-1 (HMGB-1) in patients with renal diseases including CKD, haemodialysis (HD), AKI patients, and healthy controls for comparison. First study revealed that PlGF is elevated in patients with decreased renal function. Second study demonstrated the association of MMP-2 and PAPP-A with proteinuria in patients with CKD. Moreover, serum MMP-2, MMP-9 and PAPP-A levels significantly differed in patients with various nephropathies. EN-RAGE levels are not elevated in patients with CKD, but are related to inflammatory status. PAPP-A, EN-RAGE and HMGB-1 levels are significantly elevated, but sRAGE and PlGF...
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Laboratory diagnostics of kidney function
Šálek, Tomáš ; Palička, Vladimír (advisor) ; Jabor, Antonín (referee) ; Průša, Richard (referee)
Laboratory diagnostics of kidney function The work deal with issue of glomerular filtration rate (GFR). GFR is the most important indicator of kidney function. Its decline is associated with increased morbidity and mortality. Chronic kidney disease (CKD) is defined as decrease of GFR bellow 1.0 ml/s/1,73m2 for more than three months. Patients with CKD receive nephroprotective drugs and have reduced drugs excreted by kidneys if needed. Early detection of CKD is of clinical importance. We compare GFR estimated from serum creatinine (eGFRcreatinine) and from serum cystatin C (eGFRcystatinC) in four cohorts of patients: 101 MGUS patients, 13 methanol intoxicated patients, 1515 diabetics and 352 patients with CKD. Creatinine was measured by enzymatic method traceable to international reference material NST SRM 967 in all patients. Cystatin C was determined by standardized immunoturbidimetric method traceable to DA ERM 471. This standardized method was not available only for MGUS patients. eGFR was always calculated according to the best available validated equation. MGUS patients were calculated according to MDRD equation for eGFRcreatinine and according to Grubb equation for eGFRcystatinC. Methanol intoxicated patients were estimated according to Lund-MalmÖ equation for eGFRcreatinine and according to...
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New markers of liver fibrosis
Skřebská, Lenka ; Čepová, Jana (advisor) ; Průša, Richard (referee)
This material is talking about problems of liver fibrosis. It Is a very fundamental liver's disease which is bringing about a chronic harm of the liver's tissue. An effect of it is a process defect of fibrogenesis and fibrolysis which has to be balanced. A diagnosis of liver fibrosis on time is important for start an efficient therapy. At this time we are getting diagnosis of the liver fibrosis by liver's biopsy which is bringing many risks for patients and it is disadvantageus for them. This situation is running us to search for a new non-invasive methods. It is a lot of calculations they are using a specific and unspecific markers to determine fundamental measures of specific fibrotisation and nonspecific too. A goal of this work it is to get a contrast between a new automatic imunochemicle ELF test which is using specific markers for the liver fibrosis and using calculations APRI and the Hepascore until now. Powered by TCPDF (www.tcpdf.org)
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The Heme Catabolic Pathway in Chronic Hepatitis C
Subhanová, Iva ; Zima, Tomáš (advisor) ; Průša, Richard (referee) ; Kráslová, Ivana (referee)
This thesis focuses on the importance of the heme catabolic pathway in chronic hepatitis C (HCV). The aim is mainly to investigate, whether expresion/activity of key enzymes of the heme catabolic pathway, heme oxygenase (HMOX) and biliverdin reductase (BLVRA) in the liver and blood (study A) or promoter variations of HMOX1 and UDP- glucuronosyltransferase (UGT1A1) (study B) may be associated with the progression of fibrosis and may also predict antiviral treatment outcome in patients chronically infected with HCV. We set up a new sensitive method to quantify HMOX activity by reduction gas chromatography. We developed and extensively validated RealTime PCR assay for HMOX and BLVRA expression in the liver and peripheral blood leucocytes (PBL). The (GT)n and (TA)n dinucleotide variations in HMOX1 and UGT1A1 gene promoters, respectively, were determined by fragment analysis. No association was detected between either expression of HMOX/BLVRA or the HMOX1/ UGT1A1 promoter variants and the individual histological stages of liver disease in the HCV positive patients. A marked difference in BLVRA expression in PBL between the sustained responders (SVR) and patients with treatment failure (NVR) was detected before antiviral treatment and during the follow-up. Our data suggests, that BLVRA basal expression...
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Optimizing of the regime of marker's examination of clinically important infections in blood donors
Dušková, Daniela ; Tesař, Vladimír (advisor) ; Borovanský, Jan (referee) ; Průša, Richard (referee)
Project title: Optimalization of the regime of marker's examination of clinically important infections in blood donors Project author: Daniela Dušková, M.D. Project supervisor: prof. Vladimír Tesař, M.D., Dr.Sc., MBA, FASN The aim of this project is to contribute to the discussion about introducing the methods of molecular biology into the routine blood donor testing in the transfusions departments in the Czech Republic. The theoretical part includes a brief history and some turning points in transfusion medicine. The next part within the theoretical section is dedicated to the problems of infectious diseases concerning transfusion and the general examination processes used during the selection of blood donors. The end of the theoretical part concentrates on existing possibilities of markers' examination of clinically important infections in blood donors, including the list of processes performed in the Czech Republic, the European Union and other countries. The practical part describes this study, ie. the routine screening test of blood donors using the CMIA method (a routine method) and using RT-Real Time PCR method (a molecular biology method) for detecting infectious markers (HCV, HBV, HIV). Within this part, the principle of both methods and the process of actual examinations are described in...
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Determination of the concentration of alpha-1-antitrypsin in the stool by immunoassay method
Plačková, Marie ; Kocna, Petr (advisor) ; Průša, Richard (referee)
Determination of the concentration of α1 - antitrypsin in the stool is a diagnostic indicator of inflammatory diseases of the small and the large intestine, especially malabsorption syndrome. α1 - antitrypsin belongs to the family of plasma proteins with antiproteinase effect. α1 - antitrypsin is synthesized in liver, in small amount in macrophage and is a protease inhibitor of serine proteases sercected from neutrophils. α1 - antitrypsin is acute phase protein. Higher α1 - antitrypsin values are in early phase of inflammation associated with raised CRP and other pozitive acute phase proteins. Fecal α1 - antitrypsin clearance is a sensitive and specific marker of protein loss. For α1 - antitrypsin determination in stool samples ELISA method can be used. ELISA is noncompetetive immunoassay used to detect presence of antibody or an antigen in a sample. The aim of this work was to compare two ELISA sets (Immundiagnostik and Ridascreen) used for determination α1 - antitrypsin in the stool. Then examine stability of α1 - antitrypsin in the stool and in extract prepared from stool in various storing conditions temperature and time. After this establish this method as routine in laboratory. 20 patient stool samples were examined to compare ELISA sets. Samples were suggested to be α1 - antitrypsin...
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Study of protein changes in patients with nephrotic syndrome and Anderson-Fabry disease
Vojtová, Lucie ; Zima, Tomáš (advisor) ; Teplan, Vladimír (referee) ; Průša, Richard (referee)
Background: Heavy proteinuria may be caused by either increased glomerulal basement membrane permeability or membrane or podocyte structural damage, and also by impairment of secretion-reabsorption tubular processes. In this study, 60 patients with nephrotic proteinuria and other diagnoses (lupus nephritis, membranous nephropathy, IgA nephropathy, Wegener's granulomatosis) and 20 patients with Anderson-Fabry disease (AFD),which is an X-linked genetic disorder with deficient a-galactosidase A activity, were analysed by the 2D electrophoresis method. The main aim of this work was to investigate possible differences in urine proteins in nephropaties, between healthy controls and AFD patients and to identify abnormal proteins as potential biomarkers of disease. Methods: The urine proteins were devided by isoelectric focusing method using polyacrylamide strips (pH 3-10 linear). The second dimensional SDS electrophoresis was performed in 12 % polyacrylamide gel. The proteins were visualized by silver method and selected proteins were identified by MALDI-TOF MS. The gels were evaluated by Phoretix 2D expression software 2005. Results: We found out that without adding protease inhibitors we can detect proteolysis, with increased quantity of proteins manifested in the area about 10 kDa and decreased quantity...
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Pregnancy proteins - molecular biological and biochemical analysis
Muravská, Alexandra ; Kalousová, Marta (advisor) ; Černá, Marie (referee) ; Průša, Richard (referee)
The aim of this thesis was to establish methods for selected PAPP-A (Pregnancy- Associated Plasma Protein A) gene polymorphisms analysis and to study genetic background of PAPP-A and biochemical background of PAPP-A and PlGF (Placental Growth Factor) in relation to risk pregnancy. Secondly, the aim was to establish method for two-dimensional (2D) electrophoresis of amniotic fluid. Methods for analysis of ten PAPP-A gene polymorphisms were established. These polymorphisms, PAPP-A and PlGF levels were studied in together 165 women in third trimester pregnancies complicated with threatening preterm labor (n=98), preeclampsia (n=35), IUGR (Intrauterine Growth Restriction) (n=34) and ICP (Intrahepatic Cholestasis of Pregnancy) (n=15). 114 healthy pregnant women served as controls. The method for 2D electrophoresis of amniotic fluid was established. Preeclamptic patients had significantly higher frequency of TT genotype of Cys327Cys (C/T) PAPP-A gene polymorphism compared to controls. Patients with ICP had increased serum levels of PAPP-A compared to controls, in patients with threatening preterm labor PAPP-A levels were rather decreased. PlGF levels did not differ from control group in patients with ICP and threatening preterm labor. Positive correlation was found between PAPP-A and PlGF in group of...
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Significance of determination SAA of comparison with CRP and procalcitonin
Daňková, Michaela ; Zadina, Jiří (advisor) ; Průša, Richard (referee)
The purpose of the bachelor thesis is to evaluate serum amyloid (SAA) as a diagnostic marker in selected patient groups and to compare the results of SAA with those of C-reactive protein (CRP) and procalcitonin (PCT). SAA, CRP and PCT are acute-phase proteins, whose blood concentration significantly increases within a few hours after an inflammatory stimulus. The synthesis, caused by proinflammatory cytokines, takes place in the liver. CRP is the most and longest used acute-phase reactant in routine practice. As indicators of inflammation, SAA and especially procalcitonin have been used more lately. Although SAA is an acute-phase reactant as sensitive as CRP, it is not employed so often in the diagnosis since a method suitable for routine diagnosis was not developed until recently. Briefly describing the acute-phase proteins, the theoretical part elaborates on the characterization and comparison of SAA, CRP and PCT. It further outlines methods for the determination of individual analytes. The experimental part focuses on defining selected patient groups and the immunonephelometric method for measuring SAA by the Immage 800 biochemical analyzer. The data obtained in three selected patient groups have been statistically evaluated. The aim of the thesis has been to evaluate the correlation of inflammatory...
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Determination......
Mrňák, David ; Kocna, Petr (advisor) ; Průša, Richard (referee)
The main interest of this work is in determination of pancreatic elastase 1 in the stool. The determination of the value of the pancreatic elastase 1 in the stool is important for specify of the function of the exocrine pancreas and it have high specificity and sensitivity. It is used for both long observation of patients with chronically insufficiency of the pancreas, also as a screening test for pancreatic illness and in differential diagnoses of malabsorbent syndrome. We can determine values of the pancreatic elastase 1 in the stool with using the ELISA method. ELISA means enzyme linked immunoassay and it principle is based on sandwich reaction between two monoclonal antigens with high specifity against human pancreatic elastase 1. The main objective is to determine the degree of variability in the values of the pancreatic elastase 1 in group of healthy subjects. The study will take place on four individuals and their samples will be investigated with the ELISA method in monthly intervals for six months. ELISA method was processed by a set from firm ScheBo®. After the completition of the test it is clear, that there is a high fluctuation of the values of the pancreatic elastase 1 in this group. Values of the difference between each individual measurement exceeded the limit of 70 percent....
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