scholarly journals Effects of Apigenin and Apigenin- Loaded Nanogel on Induction of Apoptosis in Human Chronic Myeloid Leukemia Cells

2018 ◽  
Vol 7 ◽  
pp. e1008
Author(s):  
Mehrdad Hashemi ◽  
Nooshin Samadian
Keyword(s):  
Drug Delivery ◽  
Flow Cytometry ◽  
Chronic Myeloid Leukemia ◽  
Cell Line ◽  
K562 Cell ◽  
Myeloid Leukemia ◽  
Toxic Effects ◽  
Blue Staining ◽  
K562 Cell Line ◽  
Laboratory Conditions

Background: Diet plays an important role in cancer prevention. Apigenin, a flavonoid with thechemical formula C15H10O5, is abundantly present in vegetables. Vegetarian foods containing flavonoids are rich sources of bioactive compounds. Flavonoids have been utilized in herbal treatment. Nanogels are drug delivery systems based on polymers and are used in tissue engineering and for drug delivery. This study was conducted to compare the effects of apigenin and a nanodrug on the viability of the K562 cell line of chronic myeloid leukemia at different durations under laboratory conditions. Materials and Methods: Chitosan was first dissolved in 1% acetic acid, and  ethylene dichloride EDC and NHS were added to the solution. Then, the nanodrug was prepared by loading apigenin into stearate–chitosan nanogel (scs nanogel), and its physical and morphological characteristics were evaluated by TEM, DLS, and FTIR. Trypan blue staining, MTT assay, and flow cytometry were used to analyze the effects of various concentrations of apigenin and apigenin-loaded chitosan–stearate nanogel (APG–SCS) at 24, 48, and 72 h after they were applied to the K562 cell line. Results: The diameter of the nanodrug particles was measured using DLS and confirmed by TEM. The K562 cells treated with APG–SCS and with apigenin exhibited significant differences compared with the control (P < 0.05). Apoptosis was detected by flow cytometry. Conclusion: This study showed that the toxic effects of apigenin and the nanodrug improved with increasing concentrations and exposure durations compared to those in the control.The toxic effect of apigenin loaded into the stearate-chitosan nanogel was greater than apigenin, and the toxic effects of both materials were greater compared to the control under laboratory conditions.[GMJ.2018;7:e1008]

Properties of the K562 cell line, derived from a patient with chronic myeloid leukemia

1976 ◽  
Vol 18 (4) ◽  
pp. 421-431 ◽  
Author(s):  
Eva Klein ◽  
Farkas Vánky ◽  
Hannah Ben-Bassat ◽  
Hava Neumann ◽  
Peter Ralph ◽  
...  
Keyword(s):  
Chronic Myeloid Leukemia ◽  
Cell Line ◽  
K562 Cell ◽  
Myeloid Leukemia ◽  
K562 Cell Line

Imatinib-induced apoptosis could be enhanced by Triptolide in Imatinib-resistant chronic myeloid leukemia cell line through down-regulation of HIF-1αand Nrf2

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 4928-4928
Author(s):  
Bing Xu ◽  
Feili Chen ◽  
Yuejian Liu ◽  
Shiyun Wang ◽  
Rongwei Li ◽  
...  
Keyword(s):  
Chronic Myeloid Leukemia ◽  
Cell Line ◽  
K562 Cell ◽  
Myeloid Leukemia ◽  
Target Genes ◽  
Leukemia Cell Line ◽  
K562 Cell Line ◽  
Down Regulation ◽  
G Cells ◽  
Induced Apoptosis

Abstract Background The emergence of Imatinib has brought a new era for the treatment of chronic myeloid leukemia. However, resistant to Imatinib might lead to the treatment failure and death of patients. Thus, finding a drug which could enhance the Imatinib-induced apoptosis in vitro could provide the experimental base for the treatment of chronic myeloid leukemia. K562 cell line is a common cell line used in the study of chronic myeloid leukemia while K562/G cell line which is resistant to Imatinib is derived from K562 cell line. Aim This study aims to explore whether low-dose Triptolide(TPL) could enhance the Imatinib-induced apoptosis in K562/G cells and related mechanism. Methods K562/G cells were subjected to different treatments and thereafter MTT assay, flow cytometry and Western blot or RT-PCR were used to determine IC50, apoptotic status and expression of Nrf2, HIF-1α and their target genes. Results Triptolide is highly cytotoxic to K562/G cells in a concentration-dependent manner. To determine the combination effect of TPL and anticancer agents, K562/G cells were exposed to Imatinib(50μM) with or without TPL (25nM) for 24h. The apoptotic cells were determined by PI/Annexin V staining and flow cytometric analysis. Apoptotic ratio of cells treated by Imatinib together with TPL is significantly increased compared to cells treated by Imatinib alone (24.78±1.12 vs. 77.52±7.75, P<0.01). Next, the underlying mechanism was investigated. High expression of Hypoxia-inducible factor-1α (HIF-1α) has been shown to be related with bad clinical outcome in patients while Nrf2 is responsible for chemo-resistanct in cancer cells. Imatinib in combination with TPL could decrease Nrf2 and HIF-1α expression at protein and mRNA levels. Down-stream genes of Nrf2 e.g NQO1, GSR and HO-1 as well as target genes of HIF-1α e.g BNIP3, VEGF and CAIX are also down-regulated at mRNA level. Conclusion TPL could significantly increase the Imatinib-induced apoptotic ratio in K562/G cells through down-regulation of HIF-1α and Nrf2. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Modification by Heme Oxygenase Inhibitor, Tin Protoporphyrin, of Cellular Differentiation of Human Myeloid Leukemia K562 Cell Line.

1999 ◽  
Vol 22 (4) ◽  
pp. 439-440 ◽  
Author(s):  
Yukiko KOISO ◽  
Yasuyuki FUJIMOTO ◽  
Daisuke MATSUMURA ◽  
Osamu NAKAJIMA ◽  
Yuichi HASHIMOTO
Keyword(s):  
Cell Line ◽  
Heme Oxygenase ◽  
K562 Cell ◽  
Myeloid Leukemia ◽  
Cellular Differentiation ◽  
K562 Cell Line ◽  
Tin Protoporphyrin ◽  
Leukemia K562 Cell

Pharmacologic Doses of Amiloride Preferentially Induce Apoptosis and Growth Inhibition of Flt3-ITD Mutation Positive Acute Myeloid Leukemia Cell Lines

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 5004-5004
Author(s):  
Yuliya Linhares ◽  
Jade Dardine ◽  
Siavash Kurdistani
Keyword(s):  
Cell Cycle ◽  
Cell Death ◽  
Cell Cycle Arrest ◽  
Cell Line ◽  
Cell Lines ◽  
K562 Cell ◽  
Myeloid Leukemia ◽  
K562 Cells ◽  
K562 Cell Line ◽  
Cycle Arrest

Abstract Abstract 5004 Introduction: Amiloride is an FDA approved potassium-sparing diuretic which targets Na+/H+ exchanger isoform 1 (NHE1). NHE1 is responsible for the regulation of the intracellular pH, as well as cell-cycle and apoptosis. In supra-pharmacologic concentrations, amiloride non-specifically inhibits protein kinases. Recent study demonstrated that proapoptotic effect of amiloride in CML cell lines is linked to the modulation of the alternative splicing of Bcl-x, HIPK3, and BCR/ABL genes and is independent of pHi. Here, we demonstrate that pharmacologic doses of amiloride preferentially induce growth inhibition, cell cycle arrest and apoptosis in Flt3-ITD positive acute myeloid leukemia cell lines as compared to Bcr-Abl positive leukemia cell line. Our data suggests that amiloride may have an effect on Flt3 signaling and that its treatment potential for Flt3-ITD positive acute myeloid leukemia needs to be explored. Methods: MV4-11, MOLM13 and K562 cells lines in log-phase growth were used for the experiments. Analysis of the baseline Flt3 expression and phosphorylation status was assessed via Flt3 immunoprecipitation and Western blotting for Flt3 and phosphotyrosine. Cells were incubated with various amiloride concentrations; equal volume dilutions of DMSO were used for control. Cell counting and trypan blue exclusion viability was performed on TC10 Bio-Rad automated cell counter. The cell cycle analysis was performed applying propidium iodide staining. To assess for apoptosis and cell death, we used annexin V/PI staining kit and flow cytometry. Results: MOLM13 and MV4-11 cell lines carry activating Flt3-ITD mutation. We confirmed the expression and constituative activation of Flt3 in MOLM13 and MV4-11 cells with Western blotting. Flt3 protein was not detectable in K562 cell line. Amiloride at 0.025 mM and 0.05 mM completely inhibited the growth of MV4-11 cells after 24 hrs of treatment with no significant increase in total or live cell numbers at 72 hrs, but only mildly affected K562 cell proliferation. While the above amiloride concentrations caused cell death in MV4-11 and MOLM13 cell lines, there was no increased cell death in K562 cells. Incubation of MOLM13 and MV4-11 cell lines with 0.05 mM amiloride for 20 hrs induced cell cycle arrest. In MV4-11 cell line, the proportion of S phase cells after amiloride treatment was 15.4% (SD=5.4%) as compared to 31.3% (SD=1.4%) in control. MOLM13 cell line demonstrated 15.3% (SD=4.7%) of cells in S after amiloride treatment as compared to 35.3% (SD=2.4%) cells in S phase in control treatment. In K562 cell line, there was less effect with 52% (SD=4.2%) of cells in S phase in control as compared to 37% (SD=8.9%) in amiloride treatment. MV4-11 and MOLM 13 cell lines were more sensitive than K562 cells to amiloride induced apoptosis with 28.8% (control 12.7%) of MV4-11 cells, 11.4% (control 7.4%) of MOLM13 cells, and 11.4% (control 8.6%) of K562 cells being apoptotic after 20 hr treatment with 0.05mM amiloride. At 72 hrs of amiloride treatment 34% (control 1.5%) of MV4-11 cells, 17% (control 5%) of MOLM13 cells and 11% of K562 cells (control 8.9%) were apoptotic. Amiloride had similar effect on the number of dead cells with no increase in total cell death in K562 cell line. Upon treatment with increasing amiloride concentrations, there was dose-dependent increase in cell death and apoptosis in all three cell lines with K562 line showing relative resistance to amiloride. Discussion: Our results demonstrate that amiloride induces cell cycle arrest and inhibits proliferation of Flt3-ITD positive cell lines MV4-11 and MOLM13 as well as K562 cell line at a pharmacologic concentration of 0.05 mM. Both, cell cycle arrest and antiproliferative effect are more pronounced in Flt3-ITD positive cells lines while it is mild in Bcr-Abl positive K562 cell line. Pharmacologic doses of amiloride induce cell death and apoptosis in Flt3-ITD positive cell lines but not in K562 cell line. Both, Bcr-Abl and Flt3 signaling stimulates proliferation and inhibits apoptosis in myeloid leukemia cells. Our study suggests that amiloride may induce cell cycle arrest and apoptosis via modulating Flt3 signaling cascade. We are currently investigating the effects of amiloride on Flt3 phosphorylation. In conclusion, our data suggests that amiloride presents a potential treatment option for Flt3-ITD positive acute myeloid leukemia. Disclosures: No relevant conflicts of interest to declare.

Identification of AHI-1 Involvement in Mediating Cellular Resistance to Imatinib Mesylate in Chronic Myeloid Leukemia Cells

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 3201-3201
Author(s):  
Donna DeGeer ◽  
Leon Zhou ◽  
Min Chen ◽  
Yun Zhao ◽  
Ali G Turhan ◽  
...  
Keyword(s):  
Chronic Myeloid Leukemia ◽  
Protein Expression ◽  
Cell Line ◽  
Myeloid Leukemia ◽  
K562 Cells ◽  
K562 Cell Line ◽  
Cellular Resistance ◽  
Expression Levels ◽  
Total Protein Expression

Abstract Chronic myeloid leukemia (CML) is a clonal multilineage myeloproliferative disorder arising from the neoplastic transformation of a pluripotent hematopoietic stem cell that acquires a unique BCR-ABL fusion gene. The BCR-ABL oncoprotein displays constitutively elevated tyrosine kinase activity that deregulates cellular proliferation and apoptosis control through effects on several common signal transduction cascades, including the PI3K/AKT, JAK2/STAT5, and NF-kB pathways. The current first line treatment for CML involves administration of the tyrosine kinase inhibitor imatinib mesylate (IM) that has shown promise in treating chronic phase CML patients. However, early relapses and IM-resistant disease have emerged and are frequently associated with mutations in the BCR-ABL kinase domain that affect inhibitor binding. AHI-1 (Abelson helper integration site 1) is a recently discovered oncogene that has been demonstrated to be highly deregulated in a CML cell line (K562) and in primary leukemic stem/progenitor cells from CML patients. AHI-1 contains several unique domains that are indicative of signalling functions, including both an SH3 and a WD40-repeat domain. We have recently demonstrated that overexpression of murine Ahi-1 is able to transform IL-3 dependent Baf3, resulting in cells able to grow in the absence of growth factors. When these transduced cells were injected into sublethally irradiated NOD/SCID immunodeficient mice, the mice developed leukemia, demonstrating the oncogenic properties of Ahi-1. Interestingly, these in vitro and in vivo effects can be enhanced by co-transduction of BCR-ABL in these cells. In addition, a direct interaction between AHI-1 and BCR-ABL at endogenous levels was identified in K562 cells and this protein interaction complex further mediated IM response/resistance in CML stem/progenitor cells. To further investigate AHI-1’s involvement in mediating this cellular resistance to IM, AHI-1 was either stably overexpressed in K562 cells by transduction of EF1a-AHI-1-IRES-YFP lentivirus or was suppressed in K562 cells using a lentiviral-mediated RNA interference approach. Interestingly, overexpression of AHI-1 in K562 cells significantly increased cellular survival in the presence of 1, 5 and 10 uM of IM as measured by a viability assay; survival of these cells was similar to that observed in an IM resistant K562 cell line reported to be highly resistant to IM in vitro. Furthermore, suppression of AHI-1 had the opposite effect, with cells displaying heightened sensitivity to IM at concentrations as low as 1 uM. Phosphorylation and protein expression levels of several proteins known to be involved in BCR-ABL signalling, including JAK2, STAT5, AKT and NF-kB (P105, P50, and P65 subunits), were then quantified by Western blot analysis. Interestingly, elevated phosphorylation and protein expression levels of JAK2, and STAT5 and total protein expression levels NF-kB p105/p55 subunits were observed in both the AHI-1 overexpressing K562 cells and IM resistant K562 cells, while reduced phosphorylation and protein expression of these same proteins was observed in AHI-1 suppressed K562 cells. Differential expression of phosphorylated NF-kB p65 subunit at serine 536 was observed, while total protein expression levels did not significantly differ. Phosphorylated AKT expression levels were only affected in AHI-1 suppressed K562 cells, and total AKT protein expression was not affected in AHI-1 overexpressed or suppressed cells. Interestingly, AHI-1 protein expression was highly elevated at endogenous levels in the IM resistant K562 cells relative to a parental K562 cell line. These findings suggest that AHI-1 may play an important role in mediating cellular resistance to IM through activation of several signalling proteins involved in BCR-ABL signalling pathway, including JAK2 and STAT5.

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