BACKGROUND: Aberrant expression of myeloid antigens (MyAgs) on acute lymphoblastic leukemia (ALL) cells is a well-documented phenomenon, although its regulating mechanisms are unclear. MyAgs in ALL are interpreted e.g. as hallmarks of early differentiation stage and/or lineage indecisiveness. Granulocytic marker CD66c -- Carcinoembryonic antigen-related cell adhesion molecule 6 (CEACAM6) is aberrantly expressed on ALL with strong correlation to genotype (negative in TEL/AML1 and MLL/AF4, positive in BCR/ABL and hyperdiploid cases). METHODS: In a cohort of 365 consecutively diagnosed Czech B-precursor ALL patients, we analyze distribution of MyAg+ cases and mutual relationship among CD13, CD15, CD33, CD65 and CD66c. The most frequent MyAg (CD66c) is studied further regarding its stability from diagnosis to relapse, prognostic significance and regulation of surface expression. For the latter, flow cytometry, Western blot and quantitative RT-PCR on sorted cells is used. RESULTS: We show CD66c is expressed in 43% patients, which is more frequent than other MyAgs studied. In addition, CD66c expression negatively correlates with CD13 (p < 0.0001), CD33 (p = 0.002) and/or CD65 (p = 0.029). Our data show that different myeloid antigens often differ in biological importance, which may be obscured by combining them into "MyAg positive ALL". We show that unlike other MyAgs, CD66c expression is not shifted from the onset of ALL to relapse (n = 39, time to relapse 0.3-5.3 years). Although opposite has previously been suggested, we show that CEACAM6 transcription is invariably followed by surface expression (by quantitative RT-PCR on sorted cells) and that malignant cells containing CD66c in cytoplasm without surface expression are not found by flow cytometry nor by Western blot in vivo. We report no prognostic significance of CD66c, globally or separately in genotype subsets of B-precursor ALL, nor an association with known risk factors (n = 254). CONCLUSION: In contrast to general notion we show that different MyAgs in lymphoblastic leukemia represent different biological circumstances. We chose the most frequent and tightly genotype-associated MyAg CD66c to show its stabile expression in patients from diagnosis to relapse, which differs from what is known on the other MyAgs. Surface expression of CD66c is regulated at the gene transcription level, in contrast to previous reports.
- MeSH
- akutní lymfatická leukemie metabolismus MeSH
- antigen CD33 MeSH
- antigen Lewis X biosyntéza MeSH
- antigeny CD13 biosyntéza MeSH
- antigeny diferenciační myelomonocytární biosyntéza MeSH
- buněčná membrána metabolismus MeSH
- časové faktory MeSH
- CD antigeny biosyntéza MeSH
- cytoplazma metabolismus MeSH
- dítě MeSH
- genetická transkripce MeSH
- genotyp MeSH
- glykosylace MeSH
- GPI-vázané proteiny MeSH
- imunofenotypizace MeSH
- kohortové studie MeSH
- kojenec MeSH
- lidé MeSH
- mladiství MeSH
- molekuly buněčné adheze biosyntéza MeSH
- polymerázová řetězová reakce s reverzní transkripcí MeSH
- předškolní dítě MeSH
- přežití po terapii bez příznaků nemoci MeSH
- prognóza MeSH
- průtoková cytometrie MeSH
- recidiva MeSH
- regulace genové exprese u nádorů * MeSH
- RNA metabolismus MeSH
- western blotting MeSH
- Check Tag
- dítě MeSH
- kojenec MeSH
- lidé MeSH
- mladiství MeSH
- předškolní dítě MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Geografické názvy
- Česká republika MeSH
- Názvy látek
- antigen CD33 MeSH
- antigen Lewis X MeSH
- antigeny CD13 MeSH
- antigeny diferenciační myelomonocytární MeSH
- CD antigeny MeSH
- CD33 protein, human MeSH Prohlížeč
- CD65s antigen, human MeSH Prohlížeč
- CEACAM6 protein, human MeSH Prohlížeč
- GPI-vázané proteiny MeSH
- molekuly buněčné adheze MeSH
- RNA MeSH
In this study flow cytometric and morphologic methods of apoptosis detection in human promyelocytic leukemia cell line HL-60 were compared. HL-60 cells were harvested at 4, 7, 16, 24 a 48 hours after induction of apoptosis by 3 % ethanol. Little changes were observed both by flow cytometry (decrease of forward scatter, increase of unprocessed cells staining with APO2.7 antibody) and viability determination by Trypan-blue staining until after 7 hours. However, after 4 hours morphologic changes were observed in the nuclear and cytoplasmic structures using Diff-Quik stained cytospin preparations and standard light microscopic techniques (50% apoptotic cells). The same results were obtained by flow cytometric measurement of sub-diploid DNA content (sub-G1 cells), and an increase of staining with APO2.7 antibody in cells permeabilised by digitonin prior to staining. After 7 hours almost all cells exhibited apoptotic morphology. After 16 hours the cell size (forward scatter) decreased significantly, and 54% of unprocessed cells were APO2.7 positive. After 24 hours only 6% of cells were alive (high forward scatter) and these cells were APO2.7 negative. The HL-60 cells did not proliferate during the cultivation in 3% ethanol, and after 48 hours all stained by Trypan blue. HL-60 leukemic cells were CD34-/AC133-, CD33+/CD15+, and only 2% of the cells were CD95+. Induction of apoptosis by ethanol did not enhance CD95 antigen expression.
- MeSH
- antigen AC133 MeSH
- antigen CD33 MeSH
- antigen Lewis X biosyntéza MeSH
- antigeny CD34 biosyntéza MeSH
- antigeny CD95 biosyntéza MeSH
- antigeny diferenciační myelomonocytární biosyntéza MeSH
- apoptóza účinky léků MeSH
- buněčné jádro účinky léků MeSH
- buněčný cyklus účinky léků MeSH
- časové faktory MeSH
- CD antigeny biosyntéza MeSH
- cytoplazma účinky léků MeSH
- DNA metabolismus MeSH
- ethanol farmakologie MeSH
- glykoproteiny biosyntéza MeSH
- HL-60 buňky MeSH
- kinetika MeSH
- lidé MeSH
- monoklonální protilátky metabolismus MeSH
- peptidy MeSH
- průtoková cytometrie MeSH
- viabilita buněk účinky léků MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Názvy látek
- antigen AC133 MeSH
- antigen CD33 MeSH
- antigen Lewis X MeSH
- antigeny CD34 MeSH
- antigeny CD95 MeSH
- antigeny diferenciační myelomonocytární MeSH
- CD antigeny MeSH
- CD33 protein, human MeSH Prohlížeč
- DNA MeSH
- ethanol MeSH
- glykoproteiny MeSH
- monoklonální protilátky MeSH
- peptidy MeSH
- PROM1 protein, human MeSH Prohlížeč
