scholarly journals c-kit expression in human megakaryoblastic leukemia cell lines

Blood ◽  
1994 ◽  
Vol 83 (8) ◽  
pp. 2133-2144 ◽  
Author(s):  
ZB Hu ◽  
W Ma ◽  
CC Uphoff ◽  
H Quentmeier ◽  
HG Drexler
Keyword(s):  
Growth Factors ◽  
Cell Lines ◽  
Leukemia Cell ◽  
Northern Blotting ◽  
Lymphoma Cell ◽  
Mrna Levels ◽  
Bryostatin 1 ◽  
Rt Pcr ◽  
Megakaryoblastic Leukemia ◽  
Leukemia Cell Lines

Abstract A panel of 164 continuous human leukemia-lymphoma cell lines was analyzed for expression of c-kit using Northern blotting and reverse transcriptase-polymerase chain reaction (RT-PCR). The c-kit transcripts were detectable in cell lines assigned to the myeloid (in 7 of 29 by Northern blotting and in 4 of 8 by RT-PCR), monocytic (in 1 of 24 by Northern blotting and in 3 of 6 by RT-PCR), erythroid (in 6 of 8 by Northern blotting and in 5 of 5 by RT-PCR), and megakaryoblastic (in 10 of 10 by Northern blotting) lineages, c-kit expression was not seen by Northern blotting or RT-PCR analysis in any of the 93 lymphoid leukemia, myeloma, or lymphoma cell lines. Treatment of four megakaryoblastic cell lines with protein kinase C activators (phorbol ester 12-O-tetradecanoylphorbol 13-acetate and Bryostatin 1) led to terminal differentiation as assessed by morphologic alterations, changes in the surface marker profile, and growth arrest. These effects were associated with enhanced c-kit mRNA expression. Exposure to all- trans retinoic acid down-regulated c-kit mRNA levels, while simultaneously causing morphologic alterations in all four cell lines. Stimulation with growth factors (interleukin-3, granulocyte macrophage- colony stimulating factor, and insulin-like growth factors I and II), used to assess any role of c-kit in proliferative processes, did not lead to significant upregulation or downregulation of c-kit expression. The finding of constitutive and high expression of c-kit mRNA in all megakaryoblastic leukemia cell lines and its modulation by various reagents might further contribute to the understanding of megakaryopoietic proliferation, differentiation, and leukemogenesis.

scholarly journals c-kit expression in human megakaryoblastic leukemia cell lines

Blood ◽  
1994 ◽  
Vol 83 (8) ◽  
pp. 2133-2144 ◽  
Author(s):  
ZB Hu ◽  
W Ma ◽  
CC Uphoff ◽  
H Quentmeier ◽  
HG Drexler
Keyword(s):  
Growth Factors ◽  
Cell Lines ◽  
Leukemia Cell ◽  
Northern Blotting ◽  
Lymphoma Cell ◽  
Mrna Levels ◽  
Bryostatin 1 ◽  
Rt Pcr ◽  
Megakaryoblastic Leukemia ◽  
Leukemia Cell Lines

A panel of 164 continuous human leukemia-lymphoma cell lines was analyzed for expression of c-kit using Northern blotting and reverse transcriptase-polymerase chain reaction (RT-PCR). The c-kit transcripts were detectable in cell lines assigned to the myeloid (in 7 of 29 by Northern blotting and in 4 of 8 by RT-PCR), monocytic (in 1 of 24 by Northern blotting and in 3 of 6 by RT-PCR), erythroid (in 6 of 8 by Northern blotting and in 5 of 5 by RT-PCR), and megakaryoblastic (in 10 of 10 by Northern blotting) lineages, c-kit expression was not seen by Northern blotting or RT-PCR analysis in any of the 93 lymphoid leukemia, myeloma, or lymphoma cell lines. Treatment of four megakaryoblastic cell lines with protein kinase C activators (phorbol ester 12-O-tetradecanoylphorbol 13-acetate and Bryostatin 1) led to terminal differentiation as assessed by morphologic alterations, changes in the surface marker profile, and growth arrest. These effects were associated with enhanced c-kit mRNA expression. Exposure to all- trans retinoic acid down-regulated c-kit mRNA levels, while simultaneously causing morphologic alterations in all four cell lines. Stimulation with growth factors (interleukin-3, granulocyte macrophage- colony stimulating factor, and insulin-like growth factors I and II), used to assess any role of c-kit in proliferative processes, did not lead to significant upregulation or downregulation of c-kit expression. The finding of constitutive and high expression of c-kit mRNA in all megakaryoblastic leukemia cell lines and its modulation by various reagents might further contribute to the understanding of megakaryopoietic proliferation, differentiation, and leukemogenesis.

Induction of Differentiation of B-Cell Leukemia Cell Lines JVM-2 and EHEB bj Bryostatin 1

1993 ◽  
Vol 10 (1-2) ◽  
pp. 135-142 ◽  
Author(s):  
Zhen-Bo Hu ◽  
Suzanne M. Gignac ◽  
Cord C. Uphoff ◽  
Hilmar Quentmeier ◽  
Klaus G. Steube ◽  
...  
Keyword(s):  
B Cell ◽  
Cell Lines ◽  
Leukemia Cell ◽  
Cell Leukemia ◽  
Bryostatin 1 ◽  
Leukemia Cell Lines ◽  
B Cell Leukemia ◽  
Induction Of Differentiation

Alternation of Cellular Morphology and Proliferation Induced by microRNA in Human Leukemia Cell Line, UT-7/EPO.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 4203-4203
Author(s):  
Nobuyoshi Kosaka ◽  
Yusuke Yamamoto ◽  
Nami Nogawa ◽  
Keiichi Sugiura ◽  
Hiroshi Miyazaki ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Cell Line ◽  
Cell Lines ◽  
Macrocytic Anemia ◽  
Leukemia Cell ◽  
Leukemia Cell Line ◽  
Human Leukemia ◽  
Mrna Levels ◽  
Rt Pcr ◽  
Morphological Studies

Abstract Mature microRNA (miRNA) originated from primary miRNA (pri-miRNA) is a new group of potential regulator for cell differentiation, apoptosis, proliferation and oncogenesis. Some miRNAs were recently identified in hematopoietic cells, while the roles of miRNAs in erythrocytic and megakaryocytic cells had not been well examined. As a first step to explore for miRNAs specific for hematopoietic lineage, the expressions of several known primary microRNAs in erythrocytic and megakaryocytic cell lines, such as TF-1, HL-60, HEK293 and UT-7 leukemia cells, were examined by RT-PCR. We consequently focused on the pri-miR-10a, a primary transcript of miR-10a located within Hox gene clusters, and found the significant expression in TF-1 cells and UT-7/EPO cells. The UT-7/EPO cells were a subline established from the original UT-7 cells, as well as UT-7/GM and UT-7/TPO cells; therefore it was suitable for the further comparative analysis. Interestingly, in UT-7/EPO cells, the expression of pri-miR-10a increased under stimulation of erythropoietin (EPO; 1U/mL and 10U/mL). Based on these observations, it was postulated that pri-miR-10a might involve in modulating erythrocyte differentiation or proliferation. To clarify the role of pri-miR-10a in UT-7/EPO, we have established clonal cell lines by transfecting UT-7/EPO cells with either the control vector or the pri-miR-10a expression vector pCMV-pri-miR10a. Overexpression of pri-miR-10a in the UT-7/EPO cell line (miR10a-UT-7/EPO) was confirmed by RT-PCR. MiR10a-UT-7/EPO showed higher proliferation rate even at low concentration of EPO (0.1 mU/mL). Overexpression of pri-miR-10a did not appear to affect HOXB4 and HOXA1 expression, as similar mRNA levels were seen in both cell lines. It was notable that the cellular size of miR10a-UT-7/EPO became larger than its parental cells. Morphological studies of miR10a-UT-7/EPO were performed in detail. It is possible that miR-10a was capable to modulate morphological features particularly in cellular size relating to cell cycle regulation. For instance, loss of the E2F family members result in marked macrocytic anemia with megaloblastic features in adult mice (Mol Cell. 2000 Aug;6(2):281–91., Mol Cell Biol. 2003 May;23(10):3607–22., Blood. 2006 Aug 1;108(3):886–95.). Data presented here hypothesized that the roles of miR-10a in erythroid cells are tightly associated with cell cycle.

RASSF2 Functions As a Tumor Suppressor in t(8;21) AML Via Promotion of Apoptosis

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 3590-3590
Author(s):  
Samuel A Stoner ◽  
Russell Dekelver ◽  
Miao-Chia Lo ◽  
Dong-Er Zhang
Keyword(s):  
Gene Expression ◽  
Tumor Suppressor ◽  
Cell Lines ◽  
Hematopoietic Cells ◽  
Leukemia Cell ◽  
Leukemia Cells ◽  
Mrna Levels ◽  
Human Cd34 ◽  
Leukemia Cell Lines ◽  
Cd34 Cells

Abstract The t(8;21) chromosomal translocation is one of the most common chromosomal translocations associated with acute myeloid leukemia (AML), found in approximately 12% of de novo AML cases. The majority of these cases are classified as FAB-subtype M2 AML. The t(8;21) results in the stable fusion of the AML1 (RUNX1) and ETO (RUNX1T1) genes. The AML1-ETO fusion protein is composed of the N-terminal portion of AML1, which includes the DNA-binding Runt-homology domain, and nearly the full-length ETO protein. The primary accepted mechanism by which AML1-ETO promotes leukemia development is through the aberrant recruitment of transcriptional repression/activation complexes to normal AML1 target genes. Therefore, the identification of individual genes or biological pathways that are specifically disrupted in the presence of AML1-ETO will provide further molecular insight into the pathogenesis of t(8;21) AML and lead to the possibility for improved treatment for these patients. We identified RASSF2 as a gene that is specifically downregulated in (2-4 fold) in total bone marrow of t(8;21) patients compared to non-t(8;21) FAB-subtype M2 AML patients by analyzing publicly available gene expression datasets. Similarly, using a mouse model of t(8;21) AML we found Rassf2 mRNA levels to be nearly 30-fold lower in t(8;21) leukemia cells compared to wild-type Lin-Sca-cKit+ (LK) myeloid progenitors. Gene expression analysis by RT-qPCR in leukemia cell lines confirmed that RASSF2 mRNA levels are significantly downregulated (8-10-fold) in both Kasumi-1 and SKNO-1 t(8;21) cell lines as compared to a similar non-t(8;21) HL-60 cell line and to primary human CD34+ control cells. In addition, expression of AML1-ETO in HL-60 or CD34+ cells results in a decrease in RASSF2 mRNA expression, which further suggests that RASSF2 is a target for regulation by AML1-ETO. Assessment of published ChIP-seq data shows that AML1-ETO binds the RASSF2 gene locus at two distinct regions in both primary t(8;21) AML patient samples and in the Kasumi-1 and SKNO-1 cell lines. These regions are similarly bound by several important hematopoietic transcription factors in primary human CD34+ cells, including AML1, ERG, FLI1, and TCF7L2, implicating these two regions as important for the regulation of RASSF2 expression during blood cell differentiation. Overexpression of RASSF2 in human leukemia cell lines using an MSCV-IRES-GFP (MIG) construct revealed that RASSF2 has a strong negative effect on leukemia cell proliferation and viability. The overall percentage of GFP-positive cells in MIG-RASSF2 transduced cells markedly decreased compared to MIG-control transduced cells over a period of 14 days. This effect was primarily due to significantly increased apoptosis in the RASSF2 expressing cell populations. Similarly, we found that expression of RASSF2 significantly inhibits the long-term self-renewal capability of hematopoietic cells transduced with AML1-ETO in a serial replating/colony formation assay. AML1-ETO transduced hematopoietic cells were normally capable of serial replating for more than 6 weeks. However, AML1-ETO transduced cells co-expressing RASSF2 consistently had reduced colony number and lost their ability to replate after 3-4 weeks. This was due to a dramatically increased rate of apoptosis in RASSF2 expressing cells. RASSF2 is reported to be a tumor suppressor that is frequently downregulated at the transcriptional level by hypermethylation in primary tumor samples, but not healthy controls. Here we have identified RASSF2 as a target for repression, and demonstrated its tumor suppressive function in t(8;21) leukemia cells. Further insights into the molecular mechanisms of RASSF2 function in AML will continue to be explored. Disclosures No relevant conflicts of interest to declare.

Analysis of FoxM1 Expression and Regulation of Cell Cycle, and Anti-Leukemic Effectss by FoxM1 Knockdown on Leukemia Cells Transfected with siRNA FoxM1.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 4320-4320
Author(s):  
Satoki Nakamura ◽  
Takaaki Ono ◽  
Yuya Sugimoto ◽  
Miki Kobayashi ◽  
Naohi Sahara ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Cyclin D1 ◽  
Cell Lines ◽  
Leukemia Cell ◽  
Aurora Kinase ◽  
Leukemia Cells ◽  
Rt Pcr ◽  
Aurora Kinase B ◽  
Leukemia Cell Lines ◽  
Foxm1 Expression

Abstract [Background] FoxM1, a member of the Fox transcription factor family, plays an important cell cycle regulator of both the transition from G1 to S phase and progression to mitosis. FoxM1 expression was also found to be up-regulated in some solid tumors (basal cell carcinomas, hepatocellular carcinoma, and primary breast cancer). These results suggested that FoxM1 plays a role in the oncogenesis of malignancies. However, it is unknown whether FoxM1 expression contributes to the development or progression of leukemia cells. Therefore, we investigated whether and how FoxM1 regulated the cell cycle of leukemia cells. [Methods] The cells used in this study were human leukemia cell lines, K562, HL60, U937 cells. For analysis of FoxM1 mRNA, RT-PCR was performed in all cell lines. For analysis of proliferation and mitotic regulatory proteins (p27, p21, Skp2, Cdc25B, Cyclin D1, Survivin, and Aurora kinase B) in leukemia cells, MTT assays and western blot were performed in all cell lines untransfected or transfected with siRNA FoxM1, respectively. For cell cycle analysis, flow cytometory analysis was performed in leukemia cells untransfected or transfected with siRNAFoxM1 by PI staining. [Results] In all leukemia cell lines, the expression of FoxM1B mRNA were significantly higher than normal MNCs. In K562, HL60, and U937 cells transfected with the siRNA FoxM1, suppression of FoxM1 caused a mean 71% (range 62 to 80%) reduction in S phase cells and a mean 4.4-fold (range 3.2 to 5.6-fold) increase in G2/M phase cells compared to untransfected cells. MTT assay demonstrated that the proliferation of the siRNA FoxM1 transfected cells was inhibited compared to the untransfected cells at 2, 3, 4, or 5 days after siRNA FoxM1 transfection. FoxM1 has been reported to regulate transcription of essential mitotic regulatory genes. We showed that FoxM1 knockdown by siRNA in leukemia cells reduced protein and mRNA expression of Aurora kinase B, Survivin, Cyclin D1, Skp2 and Cdc25B, while increased protein expression of p21and p27 in RT-PCR and western blot analysis. [Conclusions] In this study, we report in the first time that FoxM1 is overexpressed in myeloid leukemia cells. These results demonstrated that expression of FoxM1 is an essential transcription factor for growth of leukemia cells, and regulate expression of the mitotic regulators, Cdc25B, Cyclin D1, Survivin, Aurora kinase B, and p21. Moreover, we showed that FoxM1 regulated the expression of Skp2 protein, which is known to promote degradation of the cell cycle regulator p27. Our study found that inhibition of FoxM1 expression in leukemia cells suppressed their growth in vitro. Therefore, FoxM1 might be a new potential target of therapy for leukemias. We will have further study whether the level of FoxM1 expression in leukemia cells is correlated with patient survival or sensitivity for chemotherapy.

Endothelin expression in human megakaryoblastic leukemia cell lines and normal platelet precursors

1997 ◽  
Vol 68 (2) ◽  
pp. 91-97 ◽  
Author(s):  
Marie-Noelle Mathieu ◽  
Daniel Vittet ◽  
Marie-France Laliberté ◽  
François Laliberté ◽  
Isabelle Nonotte ◽  
...  
Keyword(s):  
Cell Lines ◽  
Leukemia Cell ◽  
Megakaryoblastic Leukemia ◽  
Normal Platelet ◽  
Leukemia Cell Lines

The Activity of Combined Topoisomerase Inhibitors Is Augmented by Octreotide in the Treatment of Lymphoid Leukemias.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 2475-2475
Author(s):  
Dimitrios T.P. Trafalis ◽  
Konstantinos Lilakos ◽  
Daphne Koumbi ◽  
Panayiotis Dalezis ◽  
Maria K. Angelopoulou ◽  
...  
Keyword(s):  
Cell Lines ◽  
Antitumour Activity ◽  
Leukemia Cell ◽  
Mrna Levels ◽  
In Vitro Growth ◽  
Antagonistic Effects ◽  
Leukemia Cell Lines ◽  
Topo Ii

Abstract Clinical protocols combining a topoisomerase I (topo I) and a topoisomerase IIalpha (topo II inhibitor) have shown major responses against several tumours including acute leukemia and myelodysplastic syndromes. However, combinations with topoisomerase targeting drugs should be considered with caution because antagonistic effects have been observed when administering camptothecin or topotecan (topo I inhibitors) with doxorubicin (topo II inhibitor). Octreotide (OCT) is an eight amino-acid peptide, which attains its biological effects on target cells by binding preferentially to sst2 and, to a lesser extent, to sst3 and sst5 somatostatin receptors (SS-Rs). The presence of SS-Rs in human lymphoid leukemia cell lines, in malignant lymphomas and in lymphoproliferative diseases is clearly detected. We studied the in vitro effect of combinations of OCT with doxorubicin (DOX) and topotecan (TP) on cell growth and viability in four human lymphoblastic leukemia cell lines (CCRF-CEM, RPMI-8226, JURKAT, MOLT-4) and on the Topo I, IIalpha and sst2 expression in JURKAT, MOLT-4 leukemia cell lines, as well as the in vivo effect on rodent P388 lymphocytic and L1210 lymphoid leukemias. In vitro growth inhibition and cytotoxicity were evaluated with the MTT colormetric metabolic assay. Topo I, IIalpha and sst2 mRNAs were detected with RT-PCR and the quantification of the electrophoresed specific PCR products was accomplished with Molecular Imager FX. In vivo antitumour activity was estimated by the % survival ratio of treated to untreated (control) mice and the ratio of 70-day tumour free survivors (cures). The in vitro growth inhibition and cytotoxicity that were induced by the DOX and TP combinations were neither synergistic nor additive and were similar to the activity of DOX alone. However, with the addition of OCT to the DOX and TP combinations a significant (p<0.001) synergistic effect was resulted in all tested cell lines. Treatment of cell lines with DOX produced almost a total consumption of Topo I and IIalpha mRNAs and with TP induced increase of Topo IIalpha mRNA levels (0.5–1.5 folds). OCT is clearly upregulates the Topo IIalpha expression (>2.5 folds) and restores Topo I mRNA production in cells treated with DOX. The sst2 mRNA levels were not affected in any case. In vivo antitumour activity of DOX and TP combinations was neither synergistic nor additive and it was similar to the activity of DOX alone. The addition of OCT to DOX and TP combinations produced an important synergistic antitumour effect increasing significantly survival time and cures (p<0.01) in both P388 and L1210 leukemias. Our data indicate that the antagonistic effects of Topo I and Topo II inhibitors may be due to effects on the regulation of topoisomerases expression. OCT significantly enhances the antileukemic activity of combinations with such important anticancer drugs, upregulating Topo I and IIalpha expression.

Attenuation of Apoptosis Stimulating Protein of p53-1 (ASPP1) Associates with Therapy Failure in Acute Myeloid Leukemia (AML)

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 1556-1556
Author(s):  
Marcus M. Schittenhelm ◽  
Max Kaiser ◽  
Gunnar Blumenstock ◽  
Kerstin Maria Kampa-Schittenhelm
Keyword(s):  
Bone Marrow ◽  
Protein Expression ◽  
Cell Lines ◽  
Western Blotting ◽  
Promoter Region ◽  
Leukemia Cell ◽  
Mrna Levels ◽  
Primary Therapy ◽  
Qrt Pcr ◽  
Leukemia Cell Lines

Abstract ASPP1 belongs to a family of p53-binding proteins and enhances apoptosis by stimulation of p53-transactivation of selected proapoptotic target genes. It is preferentially expressed in hematopoietic stem cells (HSC) and together with p53 preserves the genomic integrity of the HSC pool. Consequently, attenuated expression of ASPP1 which is linked to methylation of the promoter region has been associated with malignant transformation and development of acute lymphoblastic leukemia and lymphomas. We now provide evidence that ASPP1 is highly altered in AML suggesting a role in leukemogenesis as well as therapy response. ASPP1 mRNA and protein expression levels of freshly isolated native patient samples (68) and healthy bone marrow donors (29) were determined by qRT-PCR and western immunoblotting. Statistical analyseswere performed. To explore implications of attenuated ASPP1 levels with regard to apoptosis induction and proliferation, ASPP1-expressing leukemia cell lines (MOLM14, Jurkat), native patient blasts or native bone marrow donor samples were stably silenced using a retroviral shRNA approach. Vice versa, ASPP1 was stably overexpressed in AML cell lines expressing per se low ASPP1 levels. Expression was thereby confirmed by qRT-PCR and western blotting. XTT viability and annexin V-based apoptosis assays were performed using standard chemotherapeutics in comparison to empty vector controls. Decitabine was used as an epigenetic sensitizer via hypomethylation of the promoter region. ASPP1 mRNA expression was found to be frequently and highly statistically significantly (p=0.001) attenuated in AML. Low ASPP1 mRNA levels thereby translated into attenuated protein expression. Retroviral ASPP1-interference lead to perturbed proliferation capacities (up to 3-fold increase) and attenuated apoptosis upon standard chemotherapeutics in leukemia cell lines as well as native leukemia blasts. As expected, overexpression of ASPP1 resulted in significantly attenuated proliferation and higher induction of apoptosis in all tested cell lines and patient blasts. Intriguingly, epigenetic therapy using the hypomethylating agent decitabine resulted in upregulation of ASPP1 expression in leukemia cells with originally low basal ASPP1 levels as confirmed by qRT-PCR and western blotting. Consequently, decitabine pretreatment sensitized these patient samples towards chemotherapy with a favorable proapoptotic overall efficacy compared to chemotherapy alone. Our results demonstrate that dysfunctional regulation of ASPP1 expression likely contributes to the biology of leukemogenesis and to primary therapy resistance in a subgroup of patients with acute leukemia and seems to be linked to hypermethylation. Prospective clinical studies are warranted to evaluate the roleas a biomarker for risk stratification in leukemia patients and for monitoring therapy responses. Disclosures No relevant conflicts of interest to declare.

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