BACKGROUND: The diagnosis of paroxysmal nocturnal hemoglobinuria (PNH) relies on flow cytometric demonstration of loss of glycosyl-phosphatidyl inositol (GPI)-anchored proteins from red blood cells (RBC) and white blood cells (WBC). High-sensitivity multiparameter assays have been developed to detect loss of GPI-linked structures on PNH neutrophils and monocytes. High-sensitivity assays to detect PNH phenotypes in RBCs have also been developed that rely on the loss of GPI-linked CD59 on CD235a-gated mature RBCs. The latter is used to delineate PNH Type III (total loss of CD59) and PNH Type II RBCs (partial loss of CD59) from normal (Type I) RBCs. However, it is often very difficult to delineate these subsets, especially in patients with large PNH clones who continue to receive RBC transfusions, even while on eculizumab therapy. METHODS: We have added allophycocyanin (APC)-conjugated CD71 to the existing CD235aFITC/CD59PE RBC assay allowing simultaneous delineation and quantification of PNH Type III and Type II immature RBCs (iRBCs). RESULTS: We analyzed 24 medium to large-clone PNH samples (>10% PNH WBC clone size) for PNH Neutrophil, PNH Monocyte, Type III and Type II PNH iRBCs, and where possible, Type III and Type II PNH RBCs. The ability to delineate PNH Type III, Type II, and Type I iRBCs was more objective compared to that in mature RBCs. Additionally, total PNH iRBC clone sizes were very similar to PNH WBC clone sizes. CONCLUSIONS: Addition of CD71 significantly improves the ability to analyze PNH clone sizes in the RBC lineage, regardless of patient hemolytic and/or transfusion status.

• MeSH
• antigeny CD59 metabolismus   MeSH
• buněčná diferenciace MeSH
• CD antigeny krev   fyziologie   MeSH
• diferenciální diagnóza MeSH
• erytrocyty metabolismus   patologie   MeSH
• glykoforin metabolismus   MeSH
• imunofenotypizace přístrojové vybavení   metody   normy   MeSH
• kohortové studie MeSH
• leukocyty patologie   MeSH
• lidé MeSH
• monocyty metabolismus   patologie   MeSH
• neutrofily metabolismus   patologie   MeSH
• paroxysmální hemoglobinurie krev   klasifikace   diagnóza   patologie   MeSH
• počet leukocytů přístrojové vybavení   metody   MeSH
• průtoková cytometrie přístrojové vybavení   metody   normy   MeSH
• receptory transferinu krev   fyziologie   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH

We compare the effects of Imatinib mesylate (Glivec) on chronic myeloid leukemia derived cell lines K562 and JURL-MK1. In both cell lines, the cell cycle arrests in G(1)/G(0) phase within 24 h after the addition of 1 microM Imatinib. This is followed by a decrease of Ki-67 expression and the induction of apoptosis. In JURL-MK1 cells, the apoptosis is faster in comparison with K562 cells: the caspase-3 activity reaches the peak value (20 to 30 fold of the control) after about 40 h and the apoptosis proceeds to its culmination point, the DNA fragmentation, within 48 h following 1 microM Imatinib addition. Unlike K562 cells, JURL-MK1 cells possess a probably functional p53 protein inducible by TPA (tetradecanoyl phorbol acetate) or UV-B irradiation. However, no increase in p53 expression was observed in Imatinib-treated JURL-MK1 cells indicating that the difference in the apoptosis rate between the two cell lines is not due to the lack of p53 in K562 cells. Imatinib also triggers erythroid differentiation both in JURL-MK1 and K562 cells. Glycophorin A expression occurred simultaneously with the apoptosis, even at the single cell level. In K562 cells, but not in JURL-MK1 cells, the differentiation process involved increased hemoglobin synthesis. However, during spontaneous evolution of JURL-MK1 cells in culture, the effects produced by Imatinib progressively changed from the fast apoptosis to the more complete erythroid differentiation. We suggest that the apoptosis and the erythroid differentiation are parallel effects of Imatinib and their relative contributions, kinetics and completeness are related to the differentiation stage of the treated cells.

• MeSH
• aktivace enzymů MeSH
• antigen Ki-67 metabolismus   MeSH
• antitumorózní látky farmakologie   MeSH
• apoptóza účinky záření   MeSH
• buněčná diferenciace účinky léků   MeSH
• buněčné linie MeSH
• buněčný cyklus MeSH
• erytrocyty cytologie   metabolismus   účinky léků   MeSH
• financování organizované MeSH
• glykoforin metabolismus   MeSH
• lidé MeSH
• nádorový supresorový protein p53 metabolismus   MeSH
• permeabilita buněčné membrány MeSH
• piperaziny farmakologie   MeSH
• pyrimidiny farmakologie   MeSH
• Check Tag
• lidé MeSH

• MeSH
• aktivace enzymů MeSH
• antigen Ki-67 metabolismus   MeSH
• antitumorózní látky farmakologie   MeSH
• apoptóza účinky záření   MeSH
• buněčná diferenciace účinky léků   MeSH
• buněčné linie MeSH
• buněčný cyklus MeSH
• erytrocyty cytologie   metabolismus   účinky léků   MeSH
• financování organizované MeSH
• glykoforin metabolismus   MeSH
• lidé MeSH
• nádorový supresorový protein p53 metabolismus   MeSH
• permeabilita buněčné membrány MeSH
• piperaziny farmakologie   MeSH
• pyrimidiny farmakologie   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• abstrakty MeSH