Based on simple microscopic cell morphology in blood and bone marrow smear preparations, it seems to be likely that the cell differentiation and terminal differentiation in human blood cells, and particularly in erythroid or granulocytic lineages, simultaneously reflect ageing of the lineage progenitors and terminal differentiation steps. The terminal differentiation stages of both these lineages actually appear as senescent cells. Abnormal ageing of progenitor cells may represent one of the "dysplastic" phenomena of the premature terminal differentiation state. Such state is characterized by heterochromatin condensation and nucleolar morphology similar to that in fully differentiated terminal cells of granulocytic or erythroid lineages. It should also be mentioned that in some known erythropoietic disorders, less differentiated erythroblasts may lose nuclei similarly as "normal" fully terminally differentiated cells of the erythroid cell lineage. It seems to be clear that cells in both abnormal less differentiated and terminally differentiated stages of erythroid or granulocytic lineages lose the ability to multiply similarly as senescent cells. On the other hand, the background of cell ageing and differentiation is very complicated and requires a different approach than the simple microscopic morphology at the single cell level. However, the morphology and clinical cytology at the single cell level might still contribute with complementary data to more sophisticated complex studies of that topic. In addition, the morphological approach facilitates the study of the main components of single cells in various states, including the differentiation steps or ageing.

• MeSH
• buněčná diferenciace MeSH
• buněčný rodokmen MeSH
• erytroblasty * MeSH
• erytropoéza * MeSH
• lidé MeSH
• stárnutí MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH

Reconstruction of heterogeneity through single cell transcriptional profiling has greatly advanced our understanding of the spatial liver transcriptome in recent years. However, global transcriptional differences across lobular units remain elusive in physical space. Here, we apply Spatial Transcriptomics to perform transcriptomic analysis across sectioned liver tissue. We confirm that the heterogeneity in this complex tissue is predominantly determined by lobular zonation. By introducing novel computational approaches, we enable transcriptional gradient measurements between tissue structures, including several lobules in a variety of orientations. Further, our data suggests the presence of previously transcriptionally uncharacterized structures within liver tissue, contributing to the overall spatial heterogeneity of the organ. This study demonstrates how comprehensive spatial transcriptomic technologies can be used to delineate extensive spatial gene expression patterns in the liver, indicating its future impact for studies of liver function, development and regeneration as well as its potential in pre-clinical and clinical pathology.

• MeSH
• anotace sekvence MeSH
• B-lymfocyty cytologie   metabolismus   MeSH
• dendritické buňky cytologie   metabolismus   MeSH
• endoteliální buňky cytologie   metabolismus   MeSH
• erytroblasty cytologie   metabolismus   MeSH
• genetická heterogenita * MeSH
• genová ontologie MeSH
• hepatocyty cytologie   metabolismus   MeSH
• játra cytologie   metabolismus   MeSH
• Kupfferovy buňky cytologie   metabolismus   MeSH
• makrofágy cytologie   metabolismus   MeSH
• myši inbrední C57BL MeSH
• myši MeSH
• neutrofily cytologie   metabolismus   MeSH
• stanovení celkové genové exprese MeSH
• transkriptom * MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Cyclin-dependent kinases (CDKs) play an important role in the cell-division cycle. Synthetic inhibitors of CDKs are based on 2,6,9-trisubstituted purines and are developed as potential anticancer drugs; however, they have low solubility in water. In this study, we proved that the pharmaco-chemical properties of purine-based inhibitors can be improved by appropriate substitution with the adamantane moiety. We prepared ten new purine derivatives with adamantane skeletons that were linked at position 6 using phenylene spacers of variable geometry and polarity. We demonstrated that the adamantane skeleton does not compromise the biological activity, and some of the new purines displayed even higher inhibition activity towards CDK2/cyclin E than the parental compounds. These findings were supported by a docking study, which showed an adamantane scaffold inside the binding pocket participating in the complex stabilisation with non-polar interactions. In addition, we demonstrated that β-cyclodextrin (CD) increases the drug's solubility in water, although this is at the cost of reducing the biochemical and cellular effect. Most likely, the drug concentration, which is necessary for target engagement, was decreased by competitive drug binding within the complex with β-CD.

• Klíčová slova
• 2,6,9-trisubstituted purine, adamantane, cyclin-dependent kinase, cytotoxicity, molecular docking, β-cyclodextrin,
• MeSH
• adamantan chemie   MeSH
• antitumorózní látky chemie   farmakologie   MeSH
• beta-cyklodextriny chemie   MeSH
• buňky K562 MeSH
• cyklin-dependentní kinasa 2 antagonisté a inhibitory   MeSH
• inhibitory proteinkinas farmakologie   MeSH
• lidé MeSH
• MFC-7 buňky MeSH
• puriny chemie   MeSH
• vztahy mezi strukturou a aktivitou MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• adamantan MeSH
• antitumorózní látky MeSH
• beta-cyklodextriny MeSH
• CDK2 protein, human MeSH Prohlížeč
• cyklin-dependentní kinasa 2 MeSH
• inhibitory proteinkinas MeSH
• puriny MeSH

Current multiagent chemotherapy regimens have improved the cure rate in acute leukemia patients, but they are highly toxic and poorly efficient in relapsed patients. To improve the treatment approaches, new specific molecules are needed. The G-quadruplexes (G4s), which are noncanonical nucleic acid structures found in specific guanine-rich DNA or RNA, are involved in many cellular events, including control of gene expression. G4s are considered as targets for the development of anticancer agents. Heterocyclic molecules are well known to target and stabilize G4 structures. Thus, a new series of 2,9-bis[(substituted-aminomethyl)phenyl]-1,10-phenanthroline derivatives (1a-i) was designed, synthesized, and evaluated against five human myeloid leukemia cell lines (K562, KU812, MV4-11, HL60, and U937). Their ability to stabilize various oncogene promoter G4 structures (c-MYC, BCL-2, and K-RAS) as well as the telomeric G4 was also determined through the fluorescence resonance energy transfer melting assay and native mass spectrometry. In addition, the more bioactive ligands 1g-i were tested for telomerase activity in HuT78 and MV4-11 protein extracts.

• Klíčová slova
• 1, 10-phenanthroline, FRET melting, G-quadruplex, G4 ligands, antiproliferative activity, leukemia,
• MeSH
• akutní myeloidní leukemie farmakoterapie   patologie   MeSH
• antitumorózní látky chemická syntéza   chemie   farmakologie   MeSH
• buňky K562 MeSH
• fenantroliny chemická syntéza   chemie   farmakologie   MeSH
• G-kvadruplexy účinky léků   MeSH
• HL-60 buňky MeSH
• lidé MeSH
• ligandy MeSH
• nádorové buněčné linie MeSH
• racionální návrh léčiv MeSH
• rezonanční přenos fluorescenční energie MeSH
• telomerasa metabolismus   MeSH
• U937 buňky MeSH
• vztahy mezi strukturou a aktivitou MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• 1,10-phenanthroline MeSH Prohlížeč
• antitumorózní látky MeSH
• fenantroliny MeSH
• ligandy MeSH
• telomerasa MeSH

Erythropoietin (EPO) downregulates hepcidin expression to increase the availability of iron; the downregulation of hepcidin is mediated by erythroferrone (ERFE) secreted by erythroblasts. Erythroblasts also express transferrin receptor 2 (TFR2); however, the possible role of TFR2 in hepcidin downregulation is unclear. The purpose of the study was to correlate liver expression of hepcidin with the expression of ERFE and TFR2 in murine bone marrow and spleen at 4, 16, 24, 48, 72 and 96 h following administration of a single dose of EPO. Splenic Fam132b expression increased 4 h after EPO injection; liver hepcidin mRNA was decreased at 16 h. In the spleen, expression of TFR2 and transferrin receptor (TFR1) proteins increased by an order of magnitude at 48 and 72 h after EPO treatment. The EPO-induced increase in splenic TFR2 and TFR1 was associated with an increase in the number of Tfr2- and Tfr1-expressing erythroblasts. Plasma exosomes prepared from EPO-treated mice displayed increased amount of TFR1 protein; however, no exosomal TFR2 was detected. Overall, the results confirm the importance of ERFE in stress erythropoiesis, support the role of TFR2 in erythroid cell development, and highlight possible differences in the removal of TFR2 and TFR1 from erythroid cell membranes.

• Klíčová slova
• erythroferrone, exosomes, hepcidin, transferrin receptor,
• MeSH
• cytokiny genetika   metabolismus   MeSH
• erythropoetin farmakologie   MeSH
• erytroblasty účinky léků   metabolismus   MeSH
• exozómy metabolismus   MeSH
• hepcidiny genetika   metabolismus   MeSH
• játra metabolismus   MeSH
• kultivované buňky MeSH
• myši inbrední C57BL MeSH
• myši MeSH
• receptory transferinu genetika   metabolismus   MeSH
• slezina metabolismus   MeSH
• svalové proteiny genetika   metabolismus   MeSH
• zvířata MeSH
• Check Tag
• mužské pohlaví MeSH
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• cytokiny MeSH
• Erfe protein, mouse MeSH Prohlížeč
• erythropoetin MeSH
• hepcidiny MeSH
• receptory transferinu MeSH
• svalové proteiny MeSH
• TFR2 protein, mouse MeSH Prohlížeč

Measurement of BCR activator of RhoGEF and GTPase -ABL proto-oncogene 1, non-receptor tyrosine kinase (BCR-ABL1) mRNA levels by reverse transcription quantitative polymerase chain reaction (RTqPCR) has been critical to treatment protocols and clinical trials in chronic myeloid leukaemia; however, interlaboratory variation remains a significant issue. Reverse transcriptase droplet digital PCR (RTddPCR) has shown potential to improve testing but a large-scale interlaboratory study is required to definitively establish this. In the present study, 10 BCR-ABL1-positive samples with levels ranging from molecular response (MR)1·0 -MR5·0 were tested by 23 laboratories using RTddPCR with the QXDX BCR-ABL %IS kit. A subset of participants tested the samples using RTqPCR. All 23 participants using RTddPCR detected BCR-ABL1 in all samples to MR4·0 . Detection rates for deep-response samples were 95·7% at MR4·5 , 78·3% at MR4·7 and 87·0% at MR5·0 . Interlaboratory coefficient of variation was indirectly proportional to BCR-ABL1 level ranging from 29·3% to 69·0%. Linearity ranged from 0·9330 to 1·000 (average 0·9936). When results were compared for the 11 participants who performed both RTddPCR and RTqPCR, RTddPCR showed a similar limit of detection to RTqPCR with reduced interlaboratory variation and better assay linearity. The ability to detect deep responses with RTddPCR, matched with an improved linearity and reduced interlaboratory variation will allow improved patient management, and is of particular importance for future clinical trials focussed on achieving and maintaining treatment-free remission.

• Klíčová slova
• BCR-ABL1, CML, Quality, RTddPCR, external quality assessment (EQA),
• MeSH
• bcr-abl fúzové proteiny krev   MeSH
• buňky K562 chemie   MeSH
• chronická myeloidní leukemie krev   MeSH
• HL-60 buňky chemie   MeSH
• klinické laboratoře MeSH
• lidé MeSH
• lineární modely MeSH
• nádorové biomarkery krev   MeSH
• polymerázová řetězová reakce s reverzní transkripcí metody   MeSH
• reagenční diagnostické soupravy MeSH
• reprodukovatelnost výsledků MeSH
• testování odbornosti laboratoří * MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• multicentrická studie MeSH
• práce podpořená grantem MeSH
• srovnávací studie MeSH
• Geografické názvy
• Asie MeSH
• Evropa MeSH
• Severní Amerika MeSH
• Názvy látek
• bcr-abl fúzové proteiny MeSH
• BCR-ABL1 fusion protein, human MeSH Prohlížeč
• nádorové biomarkery MeSH
• reagenční diagnostické soupravy MeSH

Chronic myeloid leukemia (CML) is a malignant hematopoietic disorder distinguished by the presence of a BCR‑ABL1 fused oncogene with constitutive kinase activity. Targeted CML therapy by specific tyrosine kinase inhibitors (TKIs) leads to a marked improvement in the survival of the patients and their quality of life. However, the development of resistance to TKIs remains a critical issue for a subset of patients. The most common cause of resistance are numerous point mutations in the BCR‑ABL1 gene, followed by less common mutations and multiple mutation-independent mechanisms. Recently, exosomes, which are extracellular vesicles excreted from normal and tumor cells, have been associated with drug resistance and cancer progression. The aim of the present study was to characterize the exosomes released by imatinib‑resistant K562 (K562IR) cells. The K562IR‑derived exosomes were internalized by imatinib‑sensitive K562 cells, which thereby increased their survival in the presence of 2 µM imatinib. The exosomal cargo was subsequently analyzed to identify resistance‑associated markers using a deep label‑free quantification proteomic analysis. There were >3,000 exosomal proteins identified of which, 35 were found to be differentially expressed. From this, a total of 3, namely the membrane proteins, interferon‑induced transmembrane protein 3, CD146 and CD36, were markedly upregulated in the exosomes derived from the K562IR cells, and exhibited surface localization. The upregulation of these proteins was verified in the K562IR exosomes, and also in the K562IR cells. Using flow cytometric analysis, it was possible to further demonstrate the potential of CD146 as a cell surface marker associated with imatinib resistance in K562 cells. Taken together, these results suggested that exosomes and their respective candidate surface proteins could be potential diagnostic markers of TKI drug resistance in CML therapy.

• Klíčová slova
• chronic myeloid leukemia, imatinib mesylate, drug resistance, proteomics, exosome, tyrosine kinase inhibitor, surface marker,
• MeSH
• antigen CD146 metabolismus   MeSH
• antigeny CD36 metabolismus   MeSH
• apoptóza účinky léků   MeSH
• bcr-abl fúzové proteiny antagonisté a inhibitory   genetika   MeSH
• buňky K562 MeSH
• chemorezistence MeSH
• chronická myeloidní leukemie farmakoterapie   genetika   patologie   MeSH
• exozómy účinky léků   metabolismus   MeSH
• imatinib mesylát farmakologie   terapeutické užití   MeSH
• inhibitory proteinkinas farmakologie   terapeutické užití   MeSH
• lidé MeSH
• membránové proteiny metabolismus   MeSH
• nádorové buněčné linie MeSH
• proteiny vázající RNA metabolismus   MeSH
• viabilita buněk účinky léků   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• antigen CD146 MeSH
• antigeny CD36 MeSH
• bcr-abl fúzové proteiny MeSH
• BCR-ABL1 fusion protein, human MeSH Prohlížeč
• IFITM3 protein, human MeSH Prohlížeč
• imatinib mesylát MeSH
• inhibitory proteinkinas MeSH
• membránové proteiny MeSH
• proteiny vázající RNA MeSH

Progenitor cells of the human erythroid and granulocytic cell lineages are characterized by the presence of several nucleoli. One of these nucleoli is larger and possesses more fibrillar centres than others. Such nucleolus is apparently dominant in respect of both size and main nucleolar function such as nucleolar-ribosomal RNA transcription. Such nucleolus is also visible in specimens using conventional visualization procedures, in contrast to smaller nucleoli. In the terminal differentiation nucleated stages of the erythroid and granulocytic development, dominant nucleoli apparently disappeared, since these cells mostly contained very small nucleoli of a similar size with one fibrillar centre. Thus, the easily visible dominant nucleoli appear to be useful markers of the progenitor cell state, such as proliferation, and differentiation potential.

• MeSH
• buněčná diferenciace MeSH
• buněčné dělení MeSH
• buněčné jadérko fyziologie   ultrastruktura   MeSH
• buněčné jádro ultrastruktura   MeSH
• buněčný rodokmen MeSH
• erytroidní prekurzorové buňky ultrastruktura   MeSH
• granulocyty ultrastruktura   MeSH
• lidé MeSH
• prekurzorové buňky granulocytů ultrastruktura   MeSH
• RNA ribozomální metabolismus   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• RNA ribozomální MeSH

• MeSH
• buněčné klony MeSH
• buňky K562 MeSH
• CRISPR-Cas systémy MeSH
• genový knockout MeSH
• homozygot MeSH
• hyperhomocysteinemie farmakoterapie   genetika   MeSH
• kultivované buňky MeSH
• kyselina listová terapeutické užití   MeSH
• lidé MeSH
• megaloblastová anemie farmakoterapie   genetika   MeSH
• mladiství MeSH
• posunová mutace MeSH
• recidiva MeSH
• sekvenční delece MeSH
• sekvenování exomu MeSH
• sodíko-vodíkový výměnný transportér 1 nedostatek   genetika   MeSH
• vitamin B 12 terapeutické užití   MeSH
• Check Tag
• lidé MeSH
• mladiství MeSH
• mužské pohlaví MeSH
• Publikační typ
• dopisy MeSH
• kazuistiky MeSH
• práce podpořená grantem MeSH
• Názvy látek
• kyselina listová MeSH
• SLC9A1 protein, human MeSH Prohlížeč
• sodíko-vodíkový výměnný transportér 1 MeSH
• vitamin B 12 MeSH

ISWI chromatin remodeling ATPase SMARCA5 (SNF2H) is a well-known factor for its role in regulation of DNA access via nucleosome sliding and assembly. SMARCA5 transcriptionally inhibits the myeloid master regulator PU.1. Upregulation of SMARCA5 was previously observed in CD34+ hematopoietic progenitors of acute myeloid leukemia (AML) patients. Since high levels of SMARCA5 are necessary for intensive cell proliferation and cell cycle progression of developing hematopoietic stem and progenitor cells in mice, we reasoned that removal of SMARCA5 enzymatic activity could affect the cycling or undifferentiated state of leukemic progenitor-like clones. Indeed, we observed that CRISPR/cas9-mediated SMARCA5 knockout in AML cell lines (S5KO) inhibited the cell cycle progression. We also observed that the SMARCA5 deletion induced karyorrhexis and nuclear budding as well as increased the ploidy, indicating its role in mitotic division of AML cells. The cytogenetic analysis of S5KO cells revealed the premature chromatid separation. We conclude that deleting SMARCA5 in AML blocks leukemic proliferation and chromatid cohesion.

• Klíčová slova
• AML, CRISPR, SMARCA5, SNF2H, leukemia, therapeutic target,
• MeSH
• adenosintrifosfatasy nedostatek   metabolismus   MeSH
• akutní myeloidní leukemie * enzymologie   genetika   patologie   MeSH
• buňky K562 MeSH
• chromatidy * genetika   metabolismus   MeSH
• chromozomální proteiny, nehistonové nedostatek   metabolismus   MeSH
• genový knockout * MeSH
• lidé MeSH
• nádorové buněčné linie MeSH
• nádorové proteiny * nedostatek   metabolismus   MeSH
• proliferace buněk * MeSH
• Check Tag
• lidé MeSH
• mužské pohlaví MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• adenosintrifosfatasy MeSH
• chromozomální proteiny, nehistonové MeSH
• nádorové proteiny * MeSH
• SMARCA5 protein, human MeSH Prohlížeč