scholarly journals Usnea Acid as Multidrug Resistance (MDR) Reversing Agent against Human Chronic Myelogenous Leukemia K562/ADR Cells via an ROS Dependent Apoptosis

2019 ◽  
Vol 2019 ◽  
pp. 1-7 ◽  
Author(s):  
Wenjing Wang ◽  
Shubin Niu ◽  
Luxin Qiao ◽  
Feili Wei ◽  
Jiming Yin ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Flow Cytometry ◽  
Multidrug Resistance ◽  
Cell Viability ◽  
Leukemia Cell ◽  
Myelogenous Leukemia ◽  
Leukemia Cell Line ◽  
Phase Cell ◽  
Cell Cycle Distribution ◽  
Promising Therapeutic Approach

Purpose. Multidrug resistance (MDR) is a major obstacle in chemotherapy of leukemia treatments. In this paper, we investigated Usnea Acid (UA) as MDR reversal agent on hematologic K562/ADR cells via ROS dependent apoptosis. Methods. CCK8 assay was used to measure cell viability rate of K562/ADR. Intracellular reactive oxygen species (ROS) generation, cell cycle distribution, cell apoptosis were measured with flow cytometry, respectively. Proteins related to apoptosis were measured by Western blot. Intracellular Adriamycin accumulation was observed by confocal microscopy and measured by flow cytometry. Results. In vitro study showed intracellular Adriamycin accumulation was remarkably increased by UA. Cell viability treated with Adr (4 μM) was decreased from 89.8%  ± 4.7 to 32%  ± 8.9 by combined with UA (4 μM). Adr-induced apoptosis and G1/G0 phase cell cycle arrest were remarkably increased by UA, as well as, intracellular ROS level. However, MDR reversing activity of UA was inhibited by N-acetyl cysteine (NAC), a ROS scavenger. Conclusion. These data provide compelling evidence that UA is a promising agent against MDR in leukemia cell line and suggest a promising therapeutic approach for leukemia.

Indirubin-3-Monoxime Induces Apoptosis and Autophagy in Human Lymphocytic Leukemia Cells and Human Chronic Myelogenous Leukemia Cells.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 4106-4106
Author(s):  
Ming-Yang Lee ◽  
Jing-Jing Chuang ◽  
Yi-Wen Liu
Keyword(s):  
Cell Cycle ◽  
Cell Death ◽  
Cell Viability ◽  
Cytotoxic Effect ◽  
Leukemia Cell ◽  
K562 Cells ◽  
Lymphocytic Leukemia ◽  
Myelogenous Leukemia ◽  
Leukemia Cell Line ◽  
Normal Human

Abstract Abstract 4106 Introduction Indirubin-3-monoxime (IO) is the active ingredient of Danggui Longhui Wan, a mixture of plants that is used in traditional Chinese medicine. It is a potent inhibitor of cyclin-dependent kinases (CDKs), especially CDK2. In clinical studies demonstrated that indirubin did not cause major side effects in patients with chronic myelogenous leukemia. However, the functional action of indirubin on acute lymphoblastic leukemia (ALL) is still unclear. In the present study, we investigated that cytotoxic effect and mechanism study of IO in human acute lymphocytic leukemia cell line JM1 and human chronic myeloid leukemia cell line K562. We also analyzed the viability influence of IO in normal human granulocytes and lymphocytes. Materials & Methods After the treatment of IO, cell viability of JM1 and K562 cells was determined by WST-1 assay. The influence of IO on cell-cycle distribution was analyzed by FACScan. To identify which cell death types were induced during IO treatment we measured the caspase-3 activity for analysis the induction of apoptosis; evaluated the LDH release in the necrotic analysis and the expression level of LC3-II was determined by Western blotting analysis for autophagic cell death. Furthermore, we also analyzed the toxicity of IO in normal human granulocytes and lymphocytes by the treatment dose of IO with value of IC50. Results IO significantly affected the cell viability on JM1 but also on K562 cells in a dose dependent manner. The G2/M phase of cell cycle was arrested and sub-G1 proportion was relative increased. In addition, the finding showed that IO initiated caspase-3-dependent apoptosis in JM1 and K562 cells. The expression of autophagosome-incorporated LC3-II protein was also clearly increased once cells treated with IO. However, the necrotic phenomenon through measuring LDH release from K562 and JM1 cells could not be observed. Excitingly, by the treatment dose of IO with value of IC50, the cell viability of lymphocytes was marginally affected, whereas the cell cytotoxitity of granulocytes was not induced. Conclusions IO has the ability to induce potent cytotoxic effect on K562 and JM1 cells. The cell death after IO treatment is mainly caused by apoptosis and autophagy, not by necrosis. This effect is also associated with interrupted cell cycle. Importantly, IO has few or little cytotoxic effect on human lymphocytes and granulocytes. The results suggest that IO might be useful for clinical anti- ALL treatment. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Induction of apoptosis by Kola nut extract as a recent and promising treatment strategy for Leukemia

Bionatura ◽  
2021 ◽  
Vol 6 (2) ◽  
pp. 1725-1732
Author(s):  
Hamdah Alsaeedi ◽  
Rowaid Qahwaji ◽  
Talal Qadah
Keyword(s):  
Cell Cycle ◽  
Cell Line ◽  
Leukemia Cell ◽  
K562 Cells ◽  
Myelogenous Leukemia ◽  
Time Dependent ◽  
Leukemia Cell Line ◽  
Apoptotic Cells ◽  
Dependent Manner ◽  
Anti Cancer

Kola nut extracts have recently been reported to contain chemopreventive compounds providing several pharmacological benefits. This study investigated Kola nut extracts' anti-cancer activity on human immortalized myelogenous leukemia cell line K562 through apoptosis and cell cycle arrest. Fresh Kola nuts were prepared as powder and dissolved in DMSO. Different concentrations (50, 100, 150, 200, and 250 μg/ml) of working solutions were prepared. The K562 cells were treated with the different concentrations of Kola nut extract or vehicle control (10% DMSO) followed by incubation at 37°C for 24, 48, and 72 hours, respectively. Treatment activity was investigated in K562 cells; by Resazurin, and FITC/Propidium Iodide and 7-AAD stained cells to evaluate apoptotic cells and the cell cycle's progression. Inhibition of leukemia cell proliferation was observed. The extract effectively induced cell death, early and late apoptosis by approximately 30% after 24 and 48 hours incubation, and an increase in the rate of dead cells by 50% was observed after 72 hours of incubation. Also, cell growth reduction was seen at high dose concentrations (150 and 200 µg/ml), as evident by cell count once treated with Kola nut extract. The total number of apoptotic cells increased from 5.8% of the control group to 27.4% at 250 µg/ml concentration. Moreover, Kola nut extracts' effects on K562 cells increased gradually in a dose and time-dependent manner. It was observed that Kola nut extracts could arrest the cell cycle in the G2/M phase as an increase in the number of cells by 29.8% and 14.6 % were observed from 9.8% and 5.2% after 24 and 48 hours of incubation, respectively. This increase was detected in a dose and time-dependent manner. Kola nut extracts can be used as a novel anti-cancer agent in Leukemia treatment as it has shown significant therapeutic potential and therefore provides new insights in understanding the mechanisms of its action. Keywords: Kola nut extracts, Leukemia, K562 cell line, Apoptosis, Cancer.

Analysis of Histone Deacetylase Inhibitor, FK228, Effect on Chronic Myelogenous Leukemia Cell Line.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 4675-4675
Author(s):  
Seiichi Okabe ◽  
Testuzo Tauchi ◽  
Akihiro Nakajima ◽  
Goro Sashida ◽  
Masahiki Sumi ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Clinical Trial ◽  
Cell Line ◽  
Chronic Myelogenous Leukemia ◽  
Glycogen Synthase ◽  
Leukemia Cell ◽  
Myelogenous Leukemia ◽  
Leukemia Cell Line ◽  
Parental Cell Line ◽  
Chronic Myelogenous Leukemia Cell

Abstract Chronic myelogenous leukemia (CML) results from transformation of hematopoietic cells by the BCR/ABL gene. Although high rates of hematologic responses to imatinib therapy, the acquired resistance to imatinib has been recognized as a major problem in the treatment of CML Histone deacetylases (HDACs) and histone acetyltransferases (HATs) regulate gene expression and cell growth. Recently, HDAC inhibitors have known as a new class of anti-cancer drugs. One of the HDAC inhibitor, FK228 (FR901228, depsipeptide) is now doing the clinical trial for the treatment of patients, such as peripheral T-cell lymphoma, but there was not known to the CML. In this study, we used the TF-1 BCR-ABL cell line, which were transfected BCR/ABL gene to the leukemia cell line, TF-1. We show here that FK228 potently induced apoptosis of TF-1 BCR-ABL cells, compare to the parental cell line, TF-1, in a dose and time depend fashion. BCR-ABL, intracellular molecular chaperone, heat shock protein 90 (HSP90), and p53 which regulate cell cycle, were acetylated after FK228 treatment, but not glycogen synthase kinase-3 β(GSK-3β) and signal-transducing activators of transcription 5 (STAT5). Histone H4 is also acetylated after FK228 treatment. In a cell cycle analysis, TF-1 BCR-ABL cells were stopped at G2-M phase after FK228 treatment. The activity of MAPK and Src kinases were blocked after FK228 treatment in a time and dose depend fashion, but p38 was activated. Inhibitor of apoptosis proteins (c-IAPs) have prevented cell death by inhibiting effectors caspases. IAPs were inhibited by FK228 and caspase3, caspase9 and poly (ADP-ribose) polymerase (PARP) were activated in a time and dose depend manner. Histone acetylation and caspase activitation were not blocked by treatment of p38 inhibitor, SB203580. Our study supports the future clinical trial of FK228 in the management of CML patients.

scholarly journals Orlistatand and Rosuvastatin Suppressed Proliferation of K562 Human Myelogenous Leukemia Cell Line through AMPK/Akt/c-Myc Signaling

2020 ◽  
Author(s):  
Behnam Mojjarad ◽  
Yaghub Pazhang
Keyword(s):  
Cell Cycle ◽  
Chronic Myeloid Leukemia ◽  
Cell Line ◽  
Myeloid Leukemia ◽  
Leukemia Cell ◽  
K562 Cells ◽  
Myelogenous Leukemia ◽  
Leukemia Cell Line ◽  
Mrna Levels ◽  
Hsp 70

Abstract Background: Chronic myeloid leukemia is a myeloproliferative cancer with worldwide incidence, has become as a clinical concern due to chemoresistance in the patients received chemotherapy. Here, we investigated the effect of Orlistat and Rosuvastatin on K562 human myelogenous leukemia cell line in vitro and attempted to illuminate their possible underlying mechanisms. Methods: Cells were exposed to Orlistat and Rosuvastatin, the inhibitors of lipogenesis, then survival and apoptosis rate of K562 cells were examined by MTT assay and flow cytometric analysis respectively. The real time-PCR analysis was used to quantify mRNA levels of Bax, Bcl-2, and Hsp-70 genes. Cell cycle analysis was performed using flow cytometry, whereas the subcellular distribution of c-Myc was measured via immunofluorescence imaging technique. Additionally, the protein level of AMPK, p-AMPK Akt-1, and p-Akt-1 were studied by western blotting. Results: The results showed Orlistat and Rosuvastatin had synergistic anticancer effects on cells and in comparison with the control group, viability and apoptosis rate decreased and increased in treated cells respectively in a dose/time-dependent manner (P<0.05). The mRNA levels of Bax increased while expression of Hsp-70 decreased (P< 0.05). K562 cells treated with Orlistat and Rosuvastatin showed a cell cycle arrest in sub-G1 phase and a decreased level of c-Myc positive cells. Upon outlining the mechanism, it was revealed that AMPK/p-AMPK and p-Akt-1/Akt-1 ratio decreased in treated cells (P< 0.05). Conclusions: Data suggest Orlistat and Rosuvastatin could synergically suppress proliferation of K562 cells through AMPK/Akt/c-Myc axis, proposing a theoretical basis for upcoming application in the treatment of chronic myeloid leukemia

scholarly journals Expression of the X-CGD gene during induced differentiation of myeloid leukemia cell line HL-60

1988 ◽  
Vol 8 (7) ◽  
pp. 2804-2810
Author(s):  
K A Barker ◽  
S H Orkin ◽  
P E Newburger
Keyword(s):  
Cell Cycle ◽  
Retinoic Acid ◽  
Leukemia Cell ◽  
Morphological Differentiation ◽  
Fold Increase ◽  
Leukemia Cell Line ◽  
Dimethyl Formamide ◽  
Cell Cycle Distribution ◽  
Mrna Levels ◽  
Induced Differentiation

The expression of the X-CGD gene, which encodes the heavy-chain subunit of the phagocyte cytochrome b, was studied during induced myeloid differentiation of HL-60 cells. Incubation of the cells with a combined regimen of retinoic acid and dimethyl formamide resulted in granulocytic morphological differentiation and acquisition of nitroblue tetrazolium reduction, a measure of superoxide generation. During the 5-day course of induced differentiation, the levels of X-CGD mRNA transcripts rose 13-fold, with a 2-fold increase detectable within 3 h of exposure to retinoic acid. Relative transcription rates for the X-CGD gene, determined by nuclear runoff, increased two- to eightfold after 24 to 72 h of induced differentiation. However, the greater change in X-CGD mRNA levels than that in transcription rates implies the involvement of posttranscriptional regulation as well. Fractionation by centrifugal elutriation into phases of the cell cycle showed expression of X-CGD transcripts predominantly in G1 cells before induction and in all phases of the cell cycle 24 h after induction. Thus the rapid increase in X-CGD expression in induced cells reflects the acquisition of functional competence and not the concomitant cessation of proliferation or shift in cell cycle distribution.

Silencing of TEL/AML1 In Definitive Leukemic Cells Does Not Impair Cell Survival

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 3229-3229
Author(s):  
Marketa Zaliova ◽  
Jozef Madzo ◽  
Gunnar Cario ◽  
Jan Trka
Keyword(s):  
Gene Expression ◽  
Cell Cycle ◽  
Flow Cytometry ◽  
Cell Viability ◽  
Fusion Gene ◽  
Global Gene Expression ◽  
Leukemic Cells ◽  
Cell Cycle Distribution ◽  
Wild Type ◽  
Egfp Reporter

Abstract Abstract 3229 The most frequent structural chromosomal aberration in childhood acute lymphoblastic leukemia t(12;21) generates TEL/AML1 fusion gene. Resulting TEL/AML1 protein probably acts as an aberrant transcription factor that deregulates AML1-dependent transcription but its target genes and thus also the exact role in leukemic cells remain unknown. In vivo studies showed that TEL/AML1 itself is not sufficient to cause leukemia but may induce a preleukemic state characterized by the increased numbers of multipotent or B-cell progenitors with an incomplete block of differentiation. Despite its role for leukemia establishment the relevance of TEL/AML1 fusion gene for leukemia persistance has not been studied enough.To address this question and to explore the possibility of TEL/AML1-targeted therapy, we studied the effects of RNAi-mediated TEL/AML1 silencing on leukemic cells. As the only siRNA used for TEL/AML1 silencing published so far (Diakos et al, Blood, 2007) targets also the wild type AML1 (46% transcript reduction, our data), our first goal was to identify efficient and TEL/AML1-specific siRNA. We designed eleven different siRNAs spanning the fusion point of TEL/AML1 lacking the total sequence homology to wild type TEL and AML1 alleles to avoid their silencing. These 11 siRNAs were tested in HeLa cells transgenic for TEL/AML1-ires2-EGFP reporter. After lipofection into HeLa cells the efficiency of individual siRNAs was measured as a decrease of EGFP reporter fluorescence by flow cytometry. The best five siRNAs, that induced 50–58% silencing of the EGFP reporter, were tested at the mRNA level in TEL/AML-positive leukemic cell line. 24h after electroporation of siRNAs, when the silencing reached its maximum, two most efficient siRNAs induced 58% and 57% TEL/AML1 transcript reduction, respectively. We achieved 61% TEL/AML1 transcript reduction with the pool of both siRNAs while there was only slight reduction (14%) of wild type AML1 transcript. We used this efficient and specific siRNA pool to silence TEL/AML1 in REH and UOC-B6 TEL-AML1 positive cell lines and studied its effect on cell viability, proliferation and global gene expression. Applying two rounds of siRNA electroporation within 48 hours interval we achieved 74% and 86% TEL/AML1 protein knockdown in REH and UOC-B6 cells, respectively. We used trypan blue staining followed by optical microscopy to monitor cell viability and staining with annexin V and propidium idode to assess apoptosis rate by flow cytometry. Analysis of DNA content using staining with propidium iodide was performed to assess cell-cycle distribution. Incorporation of nucleoside analog was measured by flow cytometry to analyse de novo DNA synthesis as an indicator of proliferation rate. Despite the common expectation derived from studies on other fusion oncogenes (BCR/ABL, AML1/ETO, E2A/PBX), TEL/AML1 silencing neither decreased cell viability, nor induced apoptosis. On the contrary, TEL/AML1 depletion was accompanied by the slight but significant increase in the fraction of S-phase cells and corresponding rise in proliferation rate. Opposite effects on cell cycle distribution and proliferation were induced when we silenced wild type AML1. These findings support our hypothesis that TEL/AML1 may block previously established AML1 function in G1/S progression through the cell cycle. In line with the lack of effect on cell viability and discreet effect on cell-cycle distribution and proliferation we found no significant changes in global gene expression pattern upon TEL/AML1 depletion. Our data indicate, that TEL/AML1 is dispensable for the survival of definitive leukemic cells. This work was supported by grants MSM0021620813 and MZOFNM2005. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Orlistatand and Rosuvastatin Suppressed Proliferation of K562 Human Myelogenous Leukemia Cell Line through AMPK/Akt/c-Myc Signaling

2020 ◽  
Author(s):  
Behnam Mojjarad ◽  
Yaghub Pazhang
Keyword(s):  
Cell Cycle ◽  
Chronic Myeloid Leukemia ◽  
Cell Line ◽  
Myeloid Leukemia ◽  
Leukemia Cell ◽  
K562 Cells ◽  
Myelogenous Leukemia ◽  
Leukemia Cell Line ◽  
Mrna Levels ◽  
Hsp 70

Abstract Background: Chronic myeloid leukemia is a myeloproliferative cancer with worldwide incidence, has become as a clinical concern due to chemoresistance in the patients received chemotherapy. Here, we investigated the effect of Orlistat and Rosuvastatin on K562 human myelogenous leukemia cell line in vitro and attempted to illuminate their possible underlying mechanisms. Methods: Cells were exposed to Orlistat and Rosuvastatin, the inhibitors of lipogenesis, then survival and apoptosis rate of K562 cells were examined by MTT assay and flow cytometric analysis respectively. The real time-PCR analysis was used to quantify mRNA levels of Bax, Bcl-2, and Hsp-70 genes. Cell cycle analysis was performed using flow cytometry, whereas the subcellular distribution of c-Myc was measured via immunofluorescence imaging technique. Additionally, the protein level of AMPK, p-AMPK Akt-1, and p-Akt-1 were studied by western blotting.Results: The results showed Orlistat and Rosuvastatin had synergistic anticancer effects on cells and in comparison with the control group, viability and apoptosis rate decreased and increased in treated cells respectively in a dose/time-dependent manner (P<0.05). The mRNA levels of Bax increased while expression of Hsp-70 decreased (P< 0.05). K562 cells treated with Orlistat and Rosuvastatin showed a cell cycle arrest in sub-G1 phase and a decreased level of c-Myc positive cells. Upon outlining the mechanism, it was revealed that AMPK/p-AMPK and p-Akt-1/Akt-1 ratio decreased in treated cells (P< 0.05).Conclusions: Data suggest Orlistat and Rosuvastatin could synergically suppress proliferation of K562 cells through AMPK/Akt/c-Myc axis, proposing a theoretical basis for upcoming application in the treatment of chronic myeloid leukemia.

scholarly journals Expression of the X-CGD gene during induced differentiation of myeloid leukemia cell line HL-60.

1988 ◽  
Vol 8 (7) ◽  
pp. 2804-2810 ◽  
Author(s):  
K A Barker ◽  
S H Orkin ◽  
P E Newburger
Keyword(s):  
Cell Cycle ◽  
Retinoic Acid ◽  
Leukemia Cell ◽  
Morphological Differentiation ◽  
Fold Increase ◽  
Leukemia Cell Line ◽  
Dimethyl Formamide ◽  
Cell Cycle Distribution ◽  
Mrna Levels ◽  
Induced Differentiation

The expression of the X-CGD gene, which encodes the heavy-chain subunit of the phagocyte cytochrome b, was studied during induced myeloid differentiation of HL-60 cells. Incubation of the cells with a combined regimen of retinoic acid and dimethyl formamide resulted in granulocytic morphological differentiation and acquisition of nitroblue tetrazolium reduction, a measure of superoxide generation. During the 5-day course of induced differentiation, the levels of X-CGD mRNA transcripts rose 13-fold, with a 2-fold increase detectable within 3 h of exposure to retinoic acid. Relative transcription rates for the X-CGD gene, determined by nuclear runoff, increased two- to eightfold after 24 to 72 h of induced differentiation. However, the greater change in X-CGD mRNA levels than that in transcription rates implies the involvement of posttranscriptional regulation as well. Fractionation by centrifugal elutriation into phases of the cell cycle showed expression of X-CGD transcripts predominantly in G1 cells before induction and in all phases of the cell cycle 24 h after induction. Thus the rapid increase in X-CGD expression in induced cells reflects the acquisition of functional competence and not the concomitant cessation of proliferation or shift in cell cycle distribution.

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