Search

As the transcriptional coactivator CITED2 (CBP/p300-interacting-transactivator-with-an ED-rich-tail 2) can be overexpressed in acute myeloid leukemia (AML) cells, we analyzed the consequences of high CITED2 expression in normal and AML cells. CITED2 overexpression in normal CD34(+) cells resulted in enhanced hematopoietic stem and progenitor cell (HSPC) output in vitro, as well as in better hematopoietic stem cell (HSC) engraftability in NSG (NOD.Cg-Prkdcscid Il2rgtm1Wjl/SzJ) mice. This was because of an enhanced quiescence and maintenance of CD34(+)CD38(-) HSCs, due in part to an increased expression of the cyclin-dependent kinase inhibitor CDKN1A. We demonstrated that PU.1 is a critical regulator of CITED2, as PU.1 repressed CITED2 expression in a DNA methyltransferase 3A/B (DNMT3A/B)-dependent manner in normal CD34(+) cells. CD34(+) cells from a subset of AML patients displayed higher expression levels of CITED2 as compared with normal CD34(+) HSPCs, and knockdown of CITED2 in AML CD34(+) cells led to a loss of long-term expansion, both in vitro and in vivo. The higher CITED2 expression resulted from reduced PU.1 activity and/or dysfunction of mutated DNMT3A/B. Collectively, our data demonstrate that increased CITED2 expression results in better HSC maintenance. In concert with low PU.1 levels, this could result in a perturbed myeloid differentiation program that contributes to leukemia maintenance. [ABSTRACT FROM AUTHOR]

Different mechanisms for CBFβ-MYH11 function in acute myeloid leukemia with inv(16) have been proposed such as tethering of RUNX1 outside the nucleus, interference with transcription factor complex assembly and recruitment of histone deacetylases, all resulting in transcriptional repression of RUNX1 target genes. Here, through genome-wide CBFβ-MYH11-binding site analysis and quantitative interaction proteomics, we found that CBFβ-MYH11 localizes to RUNX1 occupied promoters, where it interacts with TAL1, FLI1 and TBP-associated factors (TAFs) in the context of the hematopoietic transcription factors ERG, GATA2 and PU.1/SPI1 and the coregulators EP300 and HDAC1. Transcriptional analysis revealed that upon fusion protein knockdown, a small subset of the CBFβ-MYH11 target genes show increased expression, confirming a role in transcriptional repression. However, the majority of CBFβ-MYH11 target genes, including genes implicated in hematopoietic stem cell self-renewal such as ID1, LMO1 and JAG1, are actively transcribed and repressed upon fusion protein knockdown. Together these results suggest an essential role for CBFβ-MYH11 in regulating the expression of genes involved in maintaining a stem cell phenotype. [ABSTRACT FROM AUTHOR]

The MLL-AF9 fusion gene is associated with aggressive leukemias of both the myeloid and lymphoid lineage in infants, whereas in adults, this translocation is mainly associated with acute myeloid leukemia. These observations suggest that differences exist between fetal and adult tissues in terms of the 'cell of origin' from which the leukemia develops. Here we show that depending on extrinsic cues, human neonatal CD34+ cells are readily immortalized along either the myeloid or lymphoid lineage upon MLL-AF9 expression and give rise to mainly lymphoid leukemia in immunocompromised mice. In contrast, immortalization of adult bone marrow CD34+ cells is more difficult to achieve and is myeloid-biased, even when MLL-AF9 is expressed in purified hematopoietic stem cells (HSCs). Transcriptome analysis identified enrichment of HSC but not progenitor gene signatures in MLL-AF9-expressing cells. Although not observed in adult cells, neonatal cells expressing MLL-AF9 were enriched for gene signatures associated with poor prognosis, resistance to chemotherapeutic agents and MYC signaling. These results indicate that neonatal cells are inherently more prone to MLL-AF9-mediated immortalization than adult cells and suggest that intrinsic properties of the cell of origin, in addition to extrinsic cues, dictate lineage of the immortalized cell. [ABSTRACT FROM AUTHOR]

A letter to the editor is presented regarding the effect of MEIS1 gene's downregulation on the impairment of acute myeloid leukemia cells.

A letter to the editor is presented regarding the cytotoxicity of acute myeloid leukemia (AML) cells potentially caused by Ras-isoprenylation differences.

In order to identify acute myeloid leukemia (AML) CD34+-specific gene expression profiles, mononuclear cells from AML patients (n=46) were sorted into CD34+ and CD34− subfractions, and genome-wide expression analysis was performed using Illumina BeadChip Arrays. AML CD34+ and CD34− gene expression was compared with a large group of normal CD34+ bone marrow (BM) cells (n=31). Unsupervised hierarchical clustering analysis showed that CD34+ AML samples belonged to a distinct cluster compared with normal BM and that in 61% of the cases the AML CD34+ transcriptome did not cluster together with the paired CD34− transcriptome. The top 50 of AML CD34+-specific genes was selected by comparing the AML CD34+ transcriptome with the AML CD34− and CD34+ normal BM transcriptomes. Interestingly, for three of these genes, that is, ankyrin repeat domain 28 (ANKRD28), guanine nucleotide binding protein, alpha 15 (GNA15) and UDP-glucose pyrophosphorylase 2 (UGP2), a high transcript level was associated with a significant poorer overall survival (OS) in two independent cohorts (n=163 and n=218) of normal karyotype AML. Importantly, the prognostic value of the continuous transcript levels of ANKRD28 (OS hazard ratio (HR): 1.32, P=0.008), GNA15 (OS HR: 1.22, P=0.033) and UGP2 (OS HR: 1.86, P=0.009) was shown to be independent from the well-known risk factors FLT3-ITD, NPM1c+ and CEBPA mutation status. [ABSTRACT FROM AUTHOR]

Recent studies in erythroid cells have shown that autophagy is an important process for the physiological clearance of mitochondria during terminal differentiation. However, autophagy also plays an important role in removing damaged and dysfunctional mitochondria. Defective mitochondria and impaired erythroid maturation are important characteristics of low-risk myelodysplasia. In this study we therefore questioned whether the autophagic clearance of mitochondria might be altered in erythroblasts from patients with refractory anemia (RA, n=3) and RA with ringed sideroblasts (RARS, n=6). Ultrastructurally, abnormal and iron-laden mitochondria were abundant, especially in RARS patients. A large proportion (52±16%) of immature and mature myelodysplastic syndrome (MDS) erythroblasts contained cytoplasmic vacuoles, partly double membraned and positive for lysosomal marker LAMP-2 and mitochondrial markers, findings compatible with autophagic removal of dysfunctional mitochondria. In healthy controls only mature erythroblasts comprised these vacuoles (12±3%). These findings were confirmed morphometrically showing an increased vacuolar surface in MDS erythroblasts compared to controls (P<0.0001). In summary, these data indicate that MDS erythroblasts show features of enhanced autophagy at an earlier stage of erythroid differentiation than in normal controls. The enhanced autophagy might be a cell protective mechanism to remove defective iron-laden mitochondria.Leukemia (2009) 23, 886–891; doi:10.1038/leu.2008.389; published online 15 January 2009 [ABSTRACT FROM AUTHOR]

Platelet production requires compartmentalized caspase activation within megakaryocytes. This eventually results in platelet release in conjunction with apoptosis of the remaining megakaryocyte. Recent studies have indicated that in low-risk myelodysplastic syndromes (MDS) and idiopathic thrombocytopenic purpura (ITP), premature cell death of megakaryocytes may contribute to thrombocytopenia. Different cell death patterns have been identified in megakaryocytes in these disorders. Growing evidence suggests that, besides apoptosis, necrosis and autophagic cell death, may also be programmed. Therefore, programmed cell death (PCD) can be classified in apoptosis, a caspase-dependent process, apoptosis-like, autophagic and necrosis-like PCD, which are predominantly caspase-independent processes. In MDS, megakaryocytes show features of necrosis-like PCD, whereas ITP megakaryocytes demonstrate predominantly characteristics of apoptosis-like PCD (para-apoptosis). Triggers for these death pathways are largely unknown. In MDS, the interaction of Fas/Fas-ligand might be of importance, whereas in ITP antiplatelet autoantibodies recognizing common antigens on megakaryocytes and platelets might be involved. These findings illustrate that cellular death pathways in megakaryocytes are recruited in both physiological and pathological settings, and that different forms of cell death can occur in the same cell depending on the stimulus and the cellular context. Elucidation of the underlying mechanisms might lead to novel therapeutic interventions.Leukemia (2006) 20, 1937–1942. doi:10.1038/sj.leu.2404385; published online 7 September 2006 [ABSTRACT FROM AUTHOR]

In the present study, we aimed to elucidate the mechanism responsible for constitutive NF-kappaB DNA-binding activity in AML cells. Intervening in aberrant signaling pathway provides a rational approach for in vivo targeting of AML cells. Constitutive NF-kappaB DNA-binding activity was observed in 16 of 22 (73%) investigated AML cases and was, in general, associated with resistance to spontaneous apoptosis. Indeed, inhibition of NF-kappaB activity by the NF-kappaB inhibitor SN-50 peptide resulted in enhanced chemotherapy-induced apoptosis. In the majority of cases, constitutive NF-kappaB activity was mediated by a Ras/PI3 kinase (PI3-K)/protein kinase B (PKB)-mediated pathway. The PI3-K inhibitor Ly294002 and the Ras inhibitor L-744832 both inhibited PKB phosphorylation and NF-kappaB DNA-binding activity. The constitutive activation of Ras GTP-ase was caused by mutations in the gene encoding for N-Ras in 29% of the cases. The constitutive NF-kappaB activity could so far not be ascribed to the autocrine production of growth factors or to mutations in the Flt3 receptor, since anti-GM-CSF, -IL-1, -IL6, -TNFalpha or the tyrosine kinase inhibitor AG1296 did not affect the NF-kappaB DNA-binding activity. The present study demonstrates that Ras activation is an important pathway for triggering the NF-kappaB pathway in AML cells. [ABSTRACT FROM AUTHOR]

In the present study, we aimed to elucidate the mechanism responsible for constitutive NF-?B DNA-binding activity in AML cells. Intervening in aberrant signaling pathway provides a rational approach for in vivo targeting of AML cells. Constitutive NF-?B DNA-binding activity was observed in 16 of 22 (73%) investigated AML cases and was, in general, associated with resistance to spontaneous apoptosis. Indeed, inhibition of NF-?B activity by the NF-?B inhibitor SN-50 peptide resulted in enhanced chemotherapy-induced apoptosis. In the majority of cases, constitutive NF-?B activity was mediated by a Ras/PI3 kinase (PI3-K)/protein kinase B (PKB)-mediated pathway. The PI3-K inhibitor Ly294002 and the Ras inhibitor L-744832 both inhibited PKB phosphorylation and NF-?B DNA-binding activity. The constitutive activation of Ras GTP-ase was caused by mutations in the gene encoding for N-Ras in 29% of the cases. The constitutive NF-?B activity could so far not be ascribed to the autocrine production of growth factors or to mutations in the Flt3 receptor, since anti-GM-CSF, -IL-1, -IL6, -TNFa or the tyrosine kinase inhibitor AG1296 did not affect the NF-?B DNA-binding activity. The present study demonstrates that Ras activation is an important pathway for triggering the NF-?B pathway in AML cells.Leukemia (2004) 18, 103-112. doi:10.1038/sj.leu.2403145 [ABSTRACT FROM AUTHOR]

In the present study, we analysed the expression and localization of p21Waf1/Cip1 in normal and malignant haematopoietic cells. We demonstrate that in normal monocytic cells, protein kinase C (PKC)-induced p21 gene activation, which is nuclear factor-?B (NF-?B) independent, results in predominantly cytoplasmic localized p21 protein. In acute monocytic leukaemia (M4, M5), monocytic blasts (N=12) show constitutive cytoplasmic p21 expression in 75% of the cases, while in myeloid leukaemic blasts (N=10), low nuclear and cytoplasmic localization of p21 could be detected, which is also PKC dependent. Constitutive p21 expression in monocytic leukaemia might have important antiapoptotic functions. This is supported by the finding that in U937 cells overexpressing p21, VP16-induced apoptosis is significantly reduced (20.0±0.9 vs 55.8±3.8%, P<0.01, N=5), reflected by a reduced phosphorylation of p38 and JNK. Similarly, AML blasts with high cytoplasmic p21 were less sensitive to VP16-induced apoptosis as compared to AML cases with low or undetectable p21 expression (42.25 vs 12.3%, P<0.01). Moreover, complex formation between p21 and ASK1 could be demonstrated in AML cells, by means of coimmunoprecipitation. In summary, these results indicate that p21 has an antiapoptotic role in monocytic leukaemia, and that p21 expression is regulated in a PKC-dependent and NF-?B-independent manner.Leukemia (2003) 17, 2113-2121. doi:10.1038/sj.leu.2403106 Published online 14 August 2003 [ABSTRACT FROM AUTHOR]

To explore the possible cross-talk between the IL-6 and TGF-beta1 pathways in AML blast cells, the effect of TGF-beta1 pretreatment on IL-6-induced STAT3 tyrosine phosphorylation was studied. A reduction of STAT3 tyrosine phosphorylation after TGF-beta1 pretreatment was observed in four out of 40 AML cases (10%), although all of the AML cases responded to TGF-beta1 by means of SMAD3 translocation. The reduced IL-6-mediated STAT3 tyrosine phosphorylation after pre-treatment with TGF-beta1 was associated with apoptosis and coincided with the degradation of certain cellular proteins, including JAK1 and -2 and Tyk2, without affecting the ERK expression and phosphorylation. Furthermore, treatment of AML blasts with the cytostatic agent VP16, as an alternative way to induce apoptosis, resulted in a similar degree of degradation of JAK kinases and concomitant reduction of IL-6-mediated STAT3 tyrosine phosphorylation. Although degradation of JAK kinases could be rescued by incubating the cells with the pan-caspase inhibitor Z-VAD-fmk, the attenuating effect of TGF-beta1 treatment on the STAT3 tyrosine phosphorylation was still partly present. It was shown that in AML cells cultured in the presence of Z-VAD-fmk, TGF-beta1 pretreatment resulted in a reduction of JAK1 phosphorylation upon IL-6 stimulation. Expression of SOCS1 and -3 could be ruled out as a possible cause of reduced JAK1 phosphorylation levels in the investigated AML case. [ABSTRACT FROM AUTHOR]

In the present study, we examined the underlying mechanism, which causes the constitutive tyrosine phosphorylation of signal transducer and activator of transcription 5 (STAT5) in acute myeloid leukemia (AML) blasts. Constitutive STAT5 phosphorylation was observed in 18 of 26 (69%) patients with AML. The constitutive STAT5 phosphorylation was caused by different mechanisms. In the majority of the investigated cases (71% (12 of 17)) constitutive STAT5 phosphorylation was associated with autophosphorylation of the type III receptor tyrosine kinase Flt3. In 47% (eight of 17) of these cases autophosphorylation of Flt3 coincided with tandem duplications of the Flt3 gene, resulting in constitutive phosphorylation of the receptor, while 24% (four of 17) of the cases demonstrated STAT5 phosphorylation and Flt3 autophosphorylation without mutations. In addition, a subset of AML cases (29% (five of 17)) had no autophosphorylation of the Flt3 receptor, but demonstrated constitutive STAT5 phosphorylation, which was partly due to autocrine growth factor production. All AML cases with high STAT5 and Flt3 phosphorylation demonstrated, in general, a lower percentage of spontaneous apoptosis, compared to AML blasts with no spontaneous STAT5 phosphorylation. Addition of the receptor tyrosine III kinase inhibitor AG1296 strongly inhibited STAT5 phosphorylation and enhanced the percentage of apoptotic cells without modulating the Bcl-xl protein levels. These data indicate that in the majority of AML cases the constitutive STAT5 phosphorylation is caused by Flt3 phosphorylation mostly due to mutations in the receptors and associated with a low degree of spontaneous apoptosis. [ABSTRACT FROM AUTHOR]

Previous studies have demonstrated that SH2-containing inositol phosphatase (SHIP) is involved in the control of B cell, myeloid cell and macrophage activation and proliferation. The goal of the present study was to examine the role of SHIP during proliferation and apoptosis in cells of the erythroid lineage. Wild-type and catalytically inactive SHIP proteins were overexpressed in the erythropoietin (EPO)-dependent cell line AS-E2. Stable overexpression of catalytically inactive SHIP decreased proliferation and resulted in prolonged activation of the extracellular signal-regulated protein kinases ERK1/2 and protein kinase B (PKB), while wild-type SHIP did not affect EPO-mediated proliferation or phosphorylation of ERK and PKB. When AS-E2 cells were EPO deprived a significant increase in apoptosis was observed in clones overexpressing wild type. Mutational analysis showed that this increase in apoptosis was independent of the enzymatic activity of SHIP. The enhanced apoptosis due to overexpression of SHIP was associated with an increase in caspase-3 and -9 activity, without a distinct effect on caspase-8 activity or mitochondrial depolarization. Moreover, in cells overexpressing SHIP apoptosis could be reduced by a caspase-3 inhibitor. These data demonstrate that in the erythroid cell line AS-E2 overexpression of catalytically inactive SHIP reduced proliferation, while overexpression of wild-type SHIP had no effect. Furthermore, overexpression of SHIP enhanced apoptosis during growth factor deprivation by inducing specific caspase cascades, which are regulated independently of the 5-phosphatase activity of SHIP. [ABSTRACT FROM AUTHOR]

The multidrug resistance proteins (MRPs) MRP1, MRP2, MRP3, MRP5 and P-glycoprotein (P-gp) act in concert with each other to give a net resultant pump function in acute myeloid leukemia (AML). The aim of the present study was to analyze the activity of these proteins, which might be upregulated at relapse as compared with de novo AML due to clonal selection. The mRNA expression and activity of P-gp and the MRPs were determined with RT-PCR and flow cytometry, in conjunction with phenotype, as measured with the monoclonal antibodies CD34, CD38 and CD33, in 30 paired samples of de novo and relapsed AML. P-gp and MRP activity varied strongly between the cases (rhodamine 123 efflux-blocking by PSC833: 5.4+/-7.7, and carboxyfluorescein efflux-blocking by MK-571: 4.3+/-6.7, n = 60). P-gp and MRP activity were increased in 23% and 40% of the relapse samples, and decreased in 30% and 20% of the relapse samples, respectively (as defined by a difference of >2 x standard deviation of the assays). Up- or downregulation of mRNA expression was observed for MDR1 (40%), MRP1 (20%), MRP2 (15%), MRP3 (30%), and MRP5 (5%). Phenotyping demonstrated a more mature phenotype in 23% of the relapsed AML cases, and a more immature phenotype in 23% of the relapses, which was independent of the karyotypic changes that were observed in 50% of the studied cases. P-gp and MRP activity correlated with the phenotypic changes, with higher P-gp and MRP activities in less mature cells (r = -0.66, P < 0.001 and r = -0.31, P = 0.02, n = 58). In conclusion, this study shows that P-gp and MRP activity are not consistently upregulated in relapsed AML. However, P-gp and MRP activities were correlated with the maturation stage as defined by immune phenotype, which was observed to be different in 46% of the relapses. [ABSTRACT FROM AUTHOR]

In the present study we examined in more detail the dual role of the c-JUN N-terminal kinase (JNK) and p38 stress-activated protein kinase pathways in mediating apoptosis or cellular activation in hematopoietic cells. Growth factor deprivation of the erythroleukemic cell line TF-1 led to apoptosis which was associated with an enhanced activity of JNK and p38 and immediate dephosphorylation of the extracellular signal-regulated kinases (ERKs). Enhanced activity of p38 and JNK was not only observed during apoptosis but also in TF-1 cells stimulated with IL-1. IL-1 rescued TF-1 cells from apoptosis. In this case, the upregulation of p38 and JNK was associated with an enhanced activity of ERK. By using SB203580, a specific inhibitor of the p38 signaling pathway, it was demonstrated that p38 plays a pivotal role in the apoptotic process. SB203580 repressed the apoptotic process to a large extent. In contrast, PD98059, a specific inhibitor of the ERK pathway, counteracted the suppressive effects of SB203580 and IL-1 on the apoptotic process indicating that the protective effect of SB203580 and IL-1 might be the result of a shift in the balance between the ERK1/2 and p38/JNK route. This was also supported by experiments with TF-1 cells overexpressing the Shc protein that demonstrated a significantly lower percentage of apoptotic cells, which coincided with higher ERK activity. Finally, the IL-1 and SB203580-mediated effects were associated with an enhanced nuclear factor-kappaB (NF-kappaB) and activator protein-1 (AP-1) binding activity, which could also be blocked by PD98059. These data demonstrate a dual function of the p38 pathway whereby other factors, such as ERK kinases, AP-1 and NF-kappaB, might determine the final cellular response. [ABSTRACT FROM AUTHOR]

Myelodysplasia (MDS) is mostly characterized by a normal or increased number of normoblasts in the bone marrow and an impaired in vitro colony formation. In the present study we analyzed whether this might be due to a disconnection between proliferation and differentiation. CD34+/CD36- sorted bone marrow cells of 18 MDS patients were cultured in a clonogenic and suspension culture assay in the presence of erythropoietin (Epo) and mast cell growth factor (MGF). Burst-forming units erythroid (BFU-E, 75 +/- 88/10(4) CD34+ cells, X +/- s.d.) and colony-forming units E (CFU-E) were observed in eight of the 13 cases (62%) with refractory anemia with or without ring sideroblasts (RA and RARS) and one of the five cases with RA with excess of blasts or in transformation (RAEB and RAEB-T). Suspension cultures with CD34+/CD36- sorted cells with Epo plus MGF demonstrated an 8.9 +/- 6.5-fold expansion after 7 days in cases with >10 BFU-E/10(4) CD34+/CD36- cells while cases with <10 BFU-E/10(4) CD34+/CD36- cells demonstrated 1.0 +/- 0.8-fold expansion especially in cases with RAEB/RAEB-T. FACS and morphology analysis after 7 days of suspension culture demonstrated partial differentiation along the erythroid lineage in cases with RA/RARS (75%) and RAEB/RAEB-T (66%) reflected by the presence of erythroblasts and normoblasts with variable expression of CD34, CD36 and Glycophorin A. In cases with erythroid colony formation 69 +/- 24% of the cells were CD34-/CD36+ and in cases with <10 BFU-E/10(4) CD34+ cells 18 +/- 16% of cells were CD34-/CD36+. Iron staining showed the presence of ring sideroblasts in two cases with RARS indicating that the cells originate from the abnormal erythroid clone. Finally, it was shown that cases with an impaired proliferative response demonstrate an enhanced binding of Annexin-V on CD34+ cells during the first days of the cell suspension culture phase. These results suggest that a defect in the proliferative response is most pronouncedly expressed in MDS whereas a subpopulation of cells retain the capacity to differentiate between transition to a terminated stage. [ABSTRACT FROM AUTHOR]

The transcription factor NF-kappaB plays an important role in the regulated expression of cytokines in human monocytes. A p100 subunit of NF-kappaB has IkappaB-like properties by sequestering the p65 transactivating subunit in the cytosol of cells. In transient transfection assays we demonstrated that p100 has an inhibitory effect on the NF-kappaB-dependent IL-6 promoter activity. In view of this finding, we studied the regulation of the p100 subunit in human monocytes in response to LPS, the inflammatory cytokines IL-1beta and TNF-alpha and lymphokines. The results demonstrate that LPS, IL-1beta, and TNF-alpha induce p100 expression at mRNA and protein level while IFN-gamma, IL-3 and IL-4/IL-10 have no effect. The induction of p100 expression was shown to be mediated by a two-fold increase in the p100 transcription rate and a two-fold increase in p100 mRNA stability. Furthermore the p100 mediated upregulation was dependent on a tyrosine kinase dependent pathway rather than the protein kinase C pathway. NF-kappaB is a complex of either p50 homodimers or a p50/p65 heterodimer. The latter is known to strongly autoregulate p100 transcription. We therefore examined the composition of NF-kappaB induced by LPS vs the different lymphokines. LPS-induced NF-kappaB showed a distinct p65 supershift whereas the composition of NF-kappaB induced by different lymphokines did not show a change in p65. We conclude that the p100 subunit of the transcription factor NF-kappaB is induced by different inflammatory mediators while lymphokines fail to induce p100 expression which may be caused by the induction of NF-kappaB predominantly consisting of p50 homodimers. [ABSTRACT FROM AUTHOR]

In refractory anemia (RA) and refractory anemia with ringed sideroblasts (RARS) a discrepancy is observed between the decreased in vitro erythroid colony formation and the normal or increased number of normoblasts in the bone marrow. To study the in vivo and in vitro erythropoiesis in more detail erythron transferrin uptake (ETU), soluble transferrin receptor (sTfR) and erythroid in vitro colony formation were performed in 24 patients with RA and five patients with RARS. These results were correlated with bone marrow morphology and transfusion dependency. Increased (mean, 124.9; range, 74-225 micromol/l blood/day) and normal (mean, 60.6; range, 50-71) ETU values were observed in 51% and 28% of the cases, whereas 21% of the cases demonstrated a diminished ETU value (mean, 35.8; range, 28-46), which correlated significantly with sTfR in cases with RA (P < 0.05, r = 0.64). A significant difference in ETU values was observed between RA (mean, 77.6; range, 28-189) and RARS (mean, 144.0; range, 59-225, P < 0.05). Most of the cases (73%) with increased ETU values showed an augmented percentage of erythroblasts in the bone marrow, which was inversely related with the serum Epo levels (P < 0.05, r = 0.51). However no correlation was found between the ETU values and the in vitro erythroid colony formation. Transfusion dependency was associated with normal to increased ETU levels (P < 0.05) and cytogenetic abnormalities (P < 0.05). These observations demonstrate that different patterns of defects can be observed in the erythropoiesis of RA and RARS patients whereby normal to increased ETU levels and the presence of cytogenetic abnormalities differentiate between cases of RA with ineffective erythropoiesis associated with regular transfusions and cases who are relatively transfusion independent. [ABSTRACT FROM AUTHOR]

In the present study it was investigated whether and by which mechanisms the co-administration of interleukin-3 (IL-3) and the P-glycoprotein blocker PSC 833 can augment mitoxantrone (MX) and daunorubicin (DAU) cytotoxicity in two human growth factor dependent P-glycoprotein (P-gp) positive myeloid leukemic cell lines, Mo-7 and GF-D8. Cytotoxicity was determined in MTT assay. Increased cytotoxicity occurred in Mo-7 cells preincubated with 24h IL-3 followed by 1 h MX (cell survival: 85% +/- 6 vs 68% +/- 2, at 0.05 microM MX, P < 0.005) or DAU (79% +/- 8 vs 62% +/- 9 at 0.8 microg/ml DAU, P < 0.05). Similar results were obtained for the GF-D8 cell line. In this cell line, at 0.5 microM MX the cell survival decreased from 84% +/- 13 to 61% +/- 19 (P < 0.05) and at 5.0 microg/ml DAU from 102% +/- 8 to 69% +/- 5, (P < 0.002). The IL-3 administration did not affect the P-gp and bcl-2 protein expression, cellular MX concentration or MX efflux but coincided with an increased percentage of cells in S-phase and topoisomerase II (topo II)-alpha mRNA and topo II activity especially in the Mo-7 cell line. PSC 833 enhanced DAU cytotoxicity in both cell lines. The administration of IL-3 plus PSC 833 in the Mo-7 cell line resulted in an additive effect on DAU cytotoxicity. At 0.8 microg/ml DAU and 2 microg/ml PSC 833, the percentage surviving cells decreased from 62% +/- 9 in the absence of IL-3 to 37% +/- 3 in the presence of IL-3 (P < 0.01). The additive effect of combined treatment was most pronounced in GF-D8 cells which also had the highest P-gp expression. In contrast, PSC 833 did not modulate the MX effects, irrespective of the presence of IL-3. In summary, the results demonstrate that the combined administration of IL-3 and PSC 833 can enhance the cytotoxic effects of DAU but not MX in these P-gp positive cell lines whereas the effects of MX could be modulated by factors which influence topo II activity. [ABSTRACT FROM AUTHOR]

Discusses whether deletion of the multidrug resistance protein 1 (MRP1) gene has any prognostic impact on acute myeloid leukemia (AML) patients with an inversion on chromosome 16. Function of MRP family of proteins; Investigation on AML patients to find out correlation between MRP1 deletion and remission duration.

Presents a breakpoint analysis by fluorescence in situ hybridization in myelodysplastic syndromes t(3;12)(q26;p13) and expression of human gene ET11. Analysis of metaphase spreads from short cultures of bone marrow cells by hybridization method; Information on various cases reported with myelodysplastic syndromes.

Comments on the analysis of in vitro development of haemopoietic progenitors in myelodysplastic syndromes. Complications associated with myelodysplastic syndromes; Questions related to the separation of candidate progenitor cells; Other problems related to the analysis made on myelodysplastic syndromes.