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Analysis of bone marrow plasma in order to identify new biomarkers in childhood acute lymphoblastic leukemia
Kováč, Martin ; Hrušák, Ondřej (advisor) ; Schwarz, Jiří (referee) ; Petrák, Jiří (referee)
Leukemia, a malignant disorder of lymphoid or myeloid progenitor cells, is the most frequent malignant disease of childhood. The most common subtype is acute lymphoblastic leukemia (ALL). In acute leukemia, several clinical symptoms may be caused by soluble proteins secreted by AL cells into the bone marrow (BM) microenvironment. We aimed at identifying proteins in BM plasma of children with ALL, which may be responsible for ALL aggressiveness or for microenvironment-mediated survival of ALL cells. LBMp (leukemic bone marrow plasma) at the diagnosis of ALL was compared to CBMp (control bone marrow plasma) and CPBp (control peripheral blood plasma) using cytokine antibody array and two-dimensional electrophoresis (2-D PAGE). Cytokine antibody array for 79 proteins identified 23 proteins expressed differentially with a statistical significance of p<0.05; of those, 2 proteins (TIMP-1 and LIF) withstood Bonferroni correction for multiple comparisons (p<0.00064). On the other hand little difference was observed between CBMp and CPBp. Using 2-D PAGE, we were able to detect 397 protein spots per gel, among which 16 showed statistically significant differences in protein levels, when comparing LBMp and CBMp by MALDI-TOF analysis. The highest difference has been found for haptoglobin in LBMp and retinol binding...
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Rare B lymphocyte populations in the context of peripheral B lymphopoiesis in humans
Grígelová, Andrea ; Růžičková, Šárka (advisor) ; Hrušák, Ondřej (referee)
B-cells of peripheral blood in humans represent a heterogeneous cellular environment displaying many important functions in the immune system. Recently, there is an increasing amount of evidence that B cell subpopulations are involved in the pathogenesis of many different diseases. However, there is little or no knowledge on how the individual differentiation stages of B lymphocytes are involved in pathological processes, and how they are distributed and represented under the physiological state and under pathological conditions. There is a reasonable assumption that, as with dendritic cells, NK / NKT cells and T lymphocytes, also B cell populations will contain minor and/or rare subpopulations reaching relative frequencies in the range of 0.01% to 0, 1 ‰. The primary aim of this thesis was to investigate the extent of phylogenetic and ontogenetic heterogeneity of the peripheral B cell population and lymphopoietic tissues on the basis of a comparative study across different vertebrate species. Another goal of the work was to use polychromatic flow cytometry with 183 individuals, out of them 50 controls and 133 patients with different (immune) pathologies or tumors in order to identify an optimal combination of surface features, and to use it to detect and demonstrate the existence of minor/rare...
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Analysis of bone marrow plasma in order to identify new biomarkers in childhood acute lymphoblastic leukemia
Kováč, Martin ; Hrušák, Ondřej (advisor) ; Schwarz, Jiří (referee) ; Petrák, Jiří (referee)
Leukemia, a malignant disorder of lymphoid or myeloid progenitor cells, is the most frequent malignant disease of childhood. The most common subtype is acute lymphoblastic leukemia (ALL). In acute leukemia, several clinical symptoms may be caused by soluble proteins secreted by AL cells into the bone marrow (BM) microenvironment. We aimed at identifying proteins in BM plasma of children with ALL, which may be responsible for ALL aggressiveness or for microenvironment-mediated survival of ALL cells. LBMp (leukemic bone marrow plasma) at the diagnosis of ALL was compared to CBMp (control bone marrow plasma) and CPBp (control peripheral blood plasma) using cytokine antibody array and two-dimensional electrophoresis (2-D PAGE). Cytokine antibody array for 79 proteins identified 23 proteins expressed differentially with a statistical significance of p<0.05; of those, 2 proteins (TIMP-1 and LIF) withstood Bonferroni correction for multiple comparisons (p<0.00064). On the other hand little difference was observed between CBMp and CPBp. Using 2-D PAGE, we were able to detect 397 protein spots per gel, among which 16 showed statistically significant differences in protein levels, when comparing LBMp and CBMp by MALDI-TOF analysis. The highest difference has been found for haptoglobin in LBMp and retinol binding...
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