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Immunological profile of patients with multiple sclerosis
Šubjak, David ; Fišerová, Anna (advisor) ; Růžičková, Šárka (referee)
Immunological profile of multiple sclerosis patients Abstrakt Multiple sclerosis is an autoimmune neurodegenerative disease affecting predominantly the white matter of the CNS and the spinal cord. The mechanism of disease progression is not yet fully understood. In this study we focused on a comparison of selected immunological markers between patients with multiple sclerosis who were naïve newly diagnosed, subsequently treated with Avonex (IFNβ1a) and healthy donors. The T cells (particulary cytotoxic CD8+ T cells) are the major population involved in pathogenesis of MS causing the demyelization of axons. Subpopulation of CD161+ Th cells has a potential to be very important in this process. We focused on the role of NK cells phenotype and function in autoimmune response of patients and their changes during the therapeutic intervention. Using flow cytometry we analyzed the distribution of NK, NKT, T cells and monocytes with special regard to the expression of CD161 and NKG2D molecules on their surface. We observed increase counts of CD161+ cells in subpopulations NK CD56bright , NK CD56dim , Th, Tc CD8bright , Tc CD8dim and decrease counts of NKG2D+ cells in subpopulations NK CD56bright , NK CD56dim , NKT, Th, Tc CD8bright , Tc CD8dim and monocytes. The decreased cytotoxic activity of NK cells in naïve MS...
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Cryptic Rearrangements of Human Chromosomes Associated with Autism Spectrum Disorders
Křivánková, Anna ; Šolc, Roman (advisor) ; Růžičková, Šárka (referee)
Autism spectrum disorders (ASD) are heterogeneous group of neurodevelopmental disabilities characterized by antisociality and atypical behavioral patterns. Its etiology is very complex, autism is usually formed by combining many factors. One of the causes may be genetic (gene mutation). It is known about 450 candidate genes for ASD so far. Minority of these genes occur in loci which are affected by cryptic rearrangements. These rearrangements significantly contribute to manifestation of this disorder. Patologies they cause, lead to syndromes with high penetrance for ASD such as Angelman/Prader-Willi or DiGeorge syndrome. Other loci are found on chromosome 1, 2 or 16. Due to short time of studies of cryptic rearrangements, phenotypic variability and number of patients we can expect more researches in the future. These researches are expected not to overlook the impact of the aberrations on formation of autism spectrum disorders.
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Heat shock proteins - - their role in diagnosis and prognosis of pregnancy related complications
Dvořáková, Lenka ; Hromadníková, Ilona (advisor) ; Růžičková, Šárka (referee)
Heat shock proteins increase their gene expression after exposure of cells or organisms to some forms of stress, which may be high temperature, infection, inflammation, hypoxia, lack of nutrients and water. Stressful situations for the body are also pregnancy-related complications associated with placental insufficiency - preeclampsia and IUGR, as well as other pregnancy-related complications such as fetal growth restriction and gestational hypertension. Therefore, I examined whether the occurrence of pregnancy-related complications (preeclampsia, fetal growth retardation, gestational hypertension) affect the gene expression of heat shock proteins. Five hsp systems was detected, namely Hsp27, Hsp60, Hsp70, Hsp90 and HspBP1 in placental tissue samples and whole maternal peripheral blood. Samples came from women with physiological pregnancy and from women with certain pregnancy-related complications (PE, FGR, GH). RNA was isolated from the samples. Detection of hsp expression was performed by using real-time RT-PCR and the comparative Ct method. Changes in gene expression between the test samples and reference sample were examined. To assess the difference between physiological pregnancies and pregnancies with selected pregnancy- related complications, an analysis of variance (ANOVA) was used....
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Expression analysis of new follicullar B cell populations characterized by absence of CD27 molecule and down-modulation of CD38 molecule.
Kerdíková, Zuzana ; Růžičková, Šárka (advisor) ; Drda Morávková, Alena (referee)
Two novel B cell populations were characterized in peripheral blood of patients with common variable immunodeficiency and healthy controls were observed using flow cytometry in the study supported by the grant IGA MZ ČR NKT11414-3. These B cell populations were defined as CD19+ CD27- CD21+ CD38low CD24+ IgM+ FO I and CD19+ CD27- CD21+ CD38low CD24++ IgM++ cells. Since none of found populations has ever been described, the aim of this thesis was to characterize these populations with focus on analysis of variable regions of the heavy chains of immunoglobulins and genes coding proteins participating in the process of VHDHJH formation (Rag 1, Rag 2, and TdT) produced by cells of these populations. Flow cytometry, single cell sorting, single-cell RT-PCR, IgVH, Rag 1, Rag 2, and TdT specific PCR amplification and cycle sequencing were employed to perform the molecular analysis in individual B lymphocytes. Both populations in two patients with common variable immunodeficiency, two healthy controls, and in two patients with autoimmune diseases - rheumatoid arthritis and systemic lupus erythematosus (as the disease control) - were examined. Finally, the statistical analysis was used to evaluate the differences in expression of variable regions of the heavy chains of immunoglobulins and in Rag1 and 2, and...
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Characterization of biological and functional features of a new type of CD27- memory B lymphocytes.
Bajzíková, Martina ; Růžičková, Šárka (advisor) ; Šinkora, Marek (referee)
The increased frequencies of two novel B cell populations defined as IgM+ CD19+ CD27- CD21low CD38low CD24+ and IgM+ CD19+ CD27- CD21low CD38low CD24- in peripheral blood of patients with common variable immunodeficiency (CVID) compared to healthy donors were found. The aim was to search for such B cells in patients with rheumatoid arthritis (RA) and their further characterization. The production of immunoglobulin (Ig) mRNA in single B cells was analyzed using flow cytometry, single cell sorting and RT-PCR, IgVH-specific PCR, cycle sequencing and statistical analysis. The study was focused on analysis of variable regions of the heavy chains of Igs and significant differences in the usage of VH, DH and JH gene segments, mutational frequencies, distribution of silent and replacement mutations, length and composition of CDR3 regions, clonal relation and RAG gene expression in above mentioned B cell populations were found. Because of lack of the surface CD27 molecule being regarded as marker of B cells that have undergone antigen-driven germinal reactions, analyzed populations were considered as naive. However, the pattern and type of mutations suggested that these cells could represent a new type of differentiated memory/antigen- experienced B lymphocytes (in CVID less maturated) with the likely role in...
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Development of selected CD markers and their role in the phylogenesis of human immune system
Podolská, Tereza ; Růžičková, Šárka (advisor) ; Vinkler, Michal (referee)
In the first part of the thesis we investigated the origin of selected surface CD markers of human, namely CD19, CD20, CD21, CD24, CD27 and CD38 molecules. In addition, nucleotide and amino acid sequences of these molecules were compared using in silico approach. Bioinformatic databases of sequences of selected molecules at DNA, mRNA and protein level, such as GeneBank, NCBI BLAST, Homologene and OrthoDB, have been used. The intent was to identify at the domain level the first organism in which it is possible to find the searched molecule. At the N-terminal domain of the CD38 of birds, a sequence showing significant similarity to the archaebacterial flagellin domain was found. This flagellin sequence in the CD38 avian molecule is located in the region of transmembrane domain, indicating that the occurrence of this sequence might be related to the formation of the transmembrane domain. The approach used here could be implemented in comparative hybridization studies as a tool in the preparatory non-laboratory phase of the research of the presence of paralogs and orthologs in phylogenetically old species. Keywords: CD marker, immunocyte, B lymphocyte, innate and adaptive immunity, sequence database, amino acid, nucleotide
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Development of selected CD markers and their role in the phylogenesis of human immune system
Podolská, Tereza ; Růžičková, Šárka (advisor) ; Vinkler, Michal (referee)
In the first part of the thesis we investigated the origin of selected surface CD markers of human, namely CD19, CD20, CD21, CD24, CD27 and CD38 molecules. In addition, nucleotide and amino acid sequences of these molecules were compared using in silico approach. Bioinformatic databases of sequences of selected molecules at DNA, mRNA and protein level, such as GeneBank, NCBI BLAST, Homologene and OrthoDB, have been used. The intent was to identify at the domain level the first organism in which it is possible to find the searched molecule. At the N-terminal domain of the CD38 of birds, a sequence showing significant similarity to the archaebacterial flagellin domain was found. This flagellin sequence in the CD38 avian molecule is located in the region of transmembrane domain, indicating that the occurrence of this sequence might be related to the formation of the transmembrane domain. The approach used here could be implemented in comparative hybridization studies as a tool in the preparatory non-laboratory phase of the research of the presence of paralogs and orthologs in phylogenetically old species. Keywords: CD marker, immunocyte, B lymphocyte, innate and adaptive immunity, sequence database, amino acid, nucleotide
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