The interaction of different pathogens with systemic and liver immune system is suggested an important factor in pathogenesis of various liver diseases. The variable aspects of this interaction and ...different immune system patways were subject of the present thesis. The ultimate goal of the present thesis was to investigate the reactivity of the local and systemic immune system against different disease related antigens and to demonstrate their participation in liver diseases and liver damage pathogenesis. Chronic hepatitis B infection, chronic hepatitis C infection and drug-induced liver damage were investigeted by 7 different studies using ex-vivo and in-vitro methods. In general, chronic liver diseases are associated with increased oxidative stress. Due to this fact, the role bilirubin, a potent endogenous antioxidant, was also analysed. The results of presented studies demonstrate clearly active participation of immune system to the pathogenesis of liver diseases, its direct involvement in liver damage and its effect on outcome of disease. The protective role of bilirubin in oxidative stress was confirmed. In conclusion, the results of all presented studies in this thesis illustrate the role of systemic as well as liver based immune system as a crucial player in the pathogenesis of liver...
In immune-induced liver damage the reactive metabolites may covalently bind or alter liver proteins such as cytochrome P450 enzymes, which leads to activation of the immune system. Ticlopidine is an ...inhibitor of CYP2C19 human liver cytochrome. We attempted to analyse the role of cytochrome CYP2C19 genetic polymorphism in the development of ticlopidine-induced cholestatic hepatitis and relate it with the specific immune reactivity to ticlopidine, different cytokine profiles and induction of necrosis and apoptosis within the liver tissue. Three patients with cholestatic hepatitis with ticlopidine-related liver injury, 3 patients with obstructive jaundice due to choledocholithiasis, 3 patients treated with ticlopidine without liver damage and 10 healthy individuals were studied. Genotyping for the following genotypes CYP2C19 (CYP2C19*1–3) were tested after polymerase chain reaction (PCR) by restriction fragment length polymorphism (RFLP) with Sma I and BamH I enzymes. The T cell reactivity to ticlopidine was analysed by T cell proliferation assay in PBMC against ticlopidine, tetanus toxoid antigen and phytohemagglutinin on days 0, 90, 150 and 210 after therapy withdrawal. The serum levels of INF-γ, IL-2, IL-4, IL-10, TNF-α, sFas and sFasL were measured by ELISA at the same time points. Apoptosis was analysed by TUNEL assay. All patients with cholestatic hepatitis had “slow metabolizers” genotypes in contrast to other groups. The T cell reactivity to ticlopidine was present only in all the cholestatic hepatitis patients together with substantial decrease in levels of INF-γ, IL-2 and TNF-α during all of the follow-up period. Cholestatic hepatitis patients had high apoptotic index in TUNEL assay. The genetic polymorphism of the cytochrome CYP2C19 gene is directly responsible for the susceptibility to the ticlopidine-induced liver damage. Th1 type of immune reactivity plays the key role in the pathogenesis of drug-induced hepatotoxicity.
Abstract
ARGONAUTE-2 and associated miRNAs form the RNA-induced silencing complex (RISC), which targets mRNAs for translational silencing and degradation as part of the RNA interference pathway. ...Despite the essential nature of this process for cellular function, there is little information on the role of RISC components in human development and organ function. We identify 13 heterozygous mutations in
AGO2
in 21 patients affected by disturbances in neurological development. Each of the identified single amino acid mutations result in impaired shRNA-mediated silencing. We observe either impaired RISC formation or increased binding of AGO2 to mRNA targets as mutation specific functional consequences. The latter is supported by decreased phosphorylation of a C-terminal serine cluster involved in mRNA target release, increased formation of dendritic P-bodies in neurons and global transcriptome alterations in patient-derived primary fibroblasts. Our data emphasize the importance of gene expression regulation through the dynamic AGO2-RNA association for human neuronal development.
Type I restriction-modification enzymes are multifunctional heteromeric complexes with DNA cleavage and ATP-dependent DNA translocation activities located on motor subunit HsdR. Functional coupling ...of DNA cleavage and translocation is a hallmark of the Type I restriction systems that is consistent with their proposed role in horizontal gene transfer. DNA cleavage occurs at nonspecific sites distant from the cognate recognition sequence, apparently triggered by stalled translocation. The X-ray crystal structure of the complete HsdR subunit from E. coli plasmid R124 suggested that the triggering mechanism involves interdomain contacts mediated by ATP. In the present work, in vivo and in vitro activity assays and crystal structures of three mutants of EcoR124I HsdR designed to probe this mechanism are reported. The results indicate that interdomain engagement via ATP is indeed responsible for signal transmission between the endonuclease and helicase domains of the motor subunit. A previously identified sequence motif that is shared by the RecB nucleases and some Type I endonucleases is implicated in signaling.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Epilepsy is a leading serious neurological condition worldwide and has particularly significant physical, economic and social consequences in Sub-Saharan Africa. This paper aims to contribute to the ...understanding of epilepsy prevalence in this region and how this varies by age and sex so as to inform understanding of the disease characteristics as well as the development of infrastructure, services and policies.
A parallel systematic analysis of Medline, Embase and Global Health returned 32 studies that satisfied pre-defined quality criteria. Relevant data was extracted, tabulated and analyzed. We modelled the available information and used the UN population figures for Africa to determine the age-specific and overall burden of epilepsy.
Active epilepsy was estimated to affect 4.4 million people in Sub-Saharan Africa, whilst lifetime epilepsy was estimated to affect 5.4 million. The prevalence of active epilepsy peaks in the 20-29 age group at 11.5/1000 and again in the 40-49 age group at 8.2/1000. The lowest prevalence value of 3.1/1000 is seen in the 60+ age group. This binomial pattern is also seen in both men and women, with the second peak more pronounced in women at 14.6/1000.
The high prevalence of epilepsy, especially in young adults, has important consequences for both the workforce and community structures. An estimation of disease burden would be a beneficial outcome of further research, as would research into appropriate methods of improving health care for and tackling discrimination against people with epilepsy.
Raman microscopy permits structural analysis of protein crystals in situ in hanging drops, allowing for comparison with Raman measurements in solution. Nevertheless, the two methods sometimes reveal ...subtle differences in structure that are often ascribed to the water layer surrounding the protein. The novel method of drop-coating deposition Raman spectropscopy (DCDR) exploits an intermediate phase that, although nominally "dry," has been shown to preserve protein structural features present in solution. The potential of this new approach to bridge the structural gap between proteins in solution and in crystals is explored here with extrinsic protein PsbP of photosystem II from Spinacia oleracea. In the high-resolution (1.98 Å) x-ray crystal structure of PsbP reported here, several segments of the protein chain are present but unresolved. Analysis of the three kinds of Raman spectra of PsbP suggests that most of the subtle differences can indeed be attributed to the water envelope, which is shown here to have a similar Raman intensity in glassy and crystal states. Using molecular dynamics simulations cross-validated by Raman solution data, two unresolved segments of the PsbP crystal structure were modeled as loops, and the amino terminus was inferred to contain an additional beta segment. The complete PsbP structure was compared with that of the PsbP-like protein CyanoP, which plays a more peripheral role in photosystem II function. The comparison suggests possible interaction surfaces of PsbP with higher-plant photosystem II. This work provides the first complete structural picture of this key protein, and it represents the first systematic comparison of Raman data from solution, glassy, and crystalline states of a protein.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Quantum mechanical calculations using the Marcus equation are applied to compare the electron-transfer probability for two distinct crystal structures of the Escherichia coli protein WrbA, an ...FMN-dependent NAD(P)H:quinone oxidoreductase, with the bound substrate benzoquinone. The calculations indicate that the position of benzoquinone in a new structure reported here and solved at 1.33 Å resolution is more likely to be relevant for the physiological reaction of WrbA than a previously reported crystal structure in which benzoquinone is shifted by ∼5 Å. Because the true electron-acceptor substrate for WrbA is not yet known, the present results can serve to constrain computational docking attempts with potential substrates that may aid in identifying the natural substrate(s) and physiological role(s) of this enzyme. The approach used here highlights a role for quantum mechanical calculations in the interpretation of protein crystal structures.
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IJS, KILJ, NUK, PNG, UL, UM
The burden of non-communicable diseases is growing, particularly in developing countries. The greatest economic burden is due to dementia, the prevalence of which is rising with increasing longevity. ...In Africa, where the rate of increase of elderly persons is the fastest in the world, dementia is normally dismissed as a part of normal ageing. The lack of awareness means that many patients are suffering undiagnosed. This review aims to assess the information on the prevalence of dementia in Africa in order to estimate the current burden.
A parallel search of Medline, EMBASE and Global Health limited to post-1980 found only 10 relevant studies. Data on prevalence and risk factors were extracted and analysed. We modelled the available information and used the UN population figures for Africa to determine the age-specific and overall burden of dementia.
The overall prevalence of dementia in adults older than 50 years in Africa was estimated to be about 2.4%, which translates to 2.76 million people living with a disease in 2010. About 2.10 millions of them live in Sub-Saharan Africa. Prevalence was the highest among females aged 80 and over (19.7%) and there was little variation between regions. Alzheimer disease was the most prevalent cause of dementia (57.1%) followed by vascular dementia (26.9%). The main risk factors were increasing age, female sex and cardiovascular disease.
Information on dementia prevalence in Africa is very limited. Further research will not only provide a more reliable estimate of prevalence, and consequently the burden of disease, but will also raise awareness of the problem. This is critical in promoting help-seeking behaviour and generating the political commitment to make dementia a public health priority in Africa.
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