Abstract

 Background and Objectives

 Acute promyelocytic leukemia (APL) is one of the most malignant forms of acute leukemia with a fatal course of only weeks which represents 10-15% of AML in adults. Arsenic trioxide as a single agent factor (without chemotherapy) is the treatment of choice for APL patients it induces cell death through apoptosis but the mechanism by which arsenic targets apoptosis and dramatically affects gene expression remains poorly understood. Since arsenic is used as first line treatment in Iran, it is worth investigating its effect on expression of genes involved in APL.

 Materials and Methods

 In this descriptive study, to understand the underlying mechanisms of cell death induction by arsenic, we treated NB4 cell line in a dose and time dependent manner. Extracting RNA and synthesis of cDNA, gene expression of apoptotic genes in mitochondrial pathway including caspase3, Mcl-1 and Bcl-2 was analyzed through Real-Time PCR.

 Results

 Our findings showed that As₂O₃-induced cell death was paralleled by reduced expression of the antiapoptotic protein Bcl-2 but the expression of Caspase3 and Mcl-1 did not change after arsenic treatment.

 Conclusions

 These results suggest that changes in Bcl-2 gene expression may be one of the mechanisms of action of arsenic in induction of apoptosis, while Caspase3 and Mcl-1 gene expression are not affected by arsenic at the transcriptional level.

 Key words : Promyelocytic Leukemia, Acute, arsenic trioxide, Apoptosis, Caspase 3, bcl-2 Genes 

  Abstract

 Background and Objectives

 Arsenic trioxide (ATO) is an active drug in treatment of acute promyelocytic leukemia (APL), but has adverse effects on patients. In order to enhance antileukemic effectiveness and to reduce dosage of ATO, combinatorial effect of it with Azidothymidine (AZT) in apoptosis induction was evaluated on APL cell line (NB4).

 Materials and Methods

 The cells cultured and treated with 50 μM AZT and/or ATO for 48 hrs and then with apoptosis, cell cycle distribution, and P21 mRNA levels in comparison with untreated cells (control) were analyzed by flow cytometry and Real Time PCR, respectively.

 Results

  ATO led to induction of apoptosis (50.14% ± 7.12) in comparison with the control (3.9% ± 2.97) through the increment of the cell cycle arrest in G2/M. The apoptotic effect of ATO had been inhibited in cells treated with combination of AZT/ATO (24.35% ± 4.65). This inhibition was associated with the relative reduction of the cells in G2/M and relative increase of the cells in G1 phase. While ATO had suppressive effect on p21 gene expression (0.27±0.14), AZT (1.81 ± 0.21) and AZT/ATO (2.06 ± 0.32) induced it.

 Conclusions

  AZT attenuates ATO-caused apoptosis possibly through the induction of p21 expression and subsequent relative evasion from G2/M arrest and accumulation of cells in G1 phase.

  Abstract

 Background and Objectives

 About 70% of MSCs die in the early stages of transplantation into the infarcted myocardium. Several solutions have been made to address this problem. In the last decade, preconditioning of MSCs with oxidative stresses has gained a lot of attention. In this study, we have investigated the effects of preconditioning with hydrogen peroxide (H₂O₂) on the survival of MSCs and their resistance against oxidative stresses .

 Materials and Methods

 Mesenchymal stem cells from bone marrow have been cultured. Cells from the passage four were treated with 5, 10, 15, 20, 30, 40, 50, 60, 80, and 100µm concentrations of H₂O₂, and were then recovered with the fresh medium. Finally, the treated cells were exposed to 500 µM H₂O₂ as the killing condition. The percentage of survived cells was analyzed by the MTT assay kit .

 Results

 Preconditioning with 5 and 10 µ M H₂O₂ significantly increased the resistance of MSCs against the apoptosis induced by 500 µ M H₂O₂ .

 Conclusions

 Preconditioning of MSCs with oxidative stresses enhances their survival therefore, it can increase the efficacy of transplantation .

  Abstract

 Background and Objectives

 The efficacy of arsenic trioxide (ATO), as a poisonous drug, in the treatment of acute promyelocytic leukemia (APL) is widely accepted. Due to adverse effects associated with high-dose ATO, the combination therapy is a rational therapeutic strategy to enhance the effectiveness of ATO. In this regard, we used simvastatin (SV), a cholesterol lowering medication, and hypothesized that SV plus ATO would potentiate the efficacy of ATO at lower doses. 

 Materials and Methods

 To evaluate the effects of SV and ATO treatment (in isolation or in combination) on transcriptional levels of Bax and Bcl-2, apoptosis, growth kinetic, and metabolic activity of NB-4 cells, we employed RQ-PCR, flowcytometry, trypan blue dye exclusion, and MTT assays, respectively. The data were analyzed using SPSS 21, student’s t-test and one-way ANOVA tests.

 Results

 Both SV and ATO considerably hindered the metabolic activity of NB-4 cells in a concentration-dependent manner. In addition, the co-treatment with them exerted an indicative decline in viability, and a significant augmentation in apoptotic population and in the ratio of Bax to Bcl-2 mRNA level. 

 Conclusions

 Our results have demonstrated that ATO and SV cooperate synergistically to induce cell death and to inhibit the proliferation rate of NB-4 cells. Furthermore, our results suggest that the combination treatment increased the programmed cell death rate probably through enhancing the intrinsic mitochondrial apoptotic pathway. On aggregate and in view of these data, SV showed the potency for attenuating the effective dose of ATO.

  Abstract

 Background and Objectives

 Acute lymphoblastic leukemia (ALL) is a neoplastic disorder and a common malignancy in children. Apoptosis is a normal physiological process which occurs during the maintenance of tissue hemostasis. The defect in this process leads to the development of cancer. It can be induced by a variety of treatments, such as cytotoxic chemotherapy. Hesperetin, a flavonoid isolated from the red fruits, has been examined with regard to the inhibition of proliferation and induction of apoptosis in pancreatic cells. Hesperetin also activates NOTCH-1 and tumor suppressor in carcinoid tumor.

 Materials and Methods

 In this fundamental-applied research, the cell lines of C121 (Jurkat cell) were cultured in RPMI supplemented with 10% fetal bovine serum (FBS) and Penicilin/streptomycin. The growth and survival of cells with various concentrations of hesperetin was evaluated by MTS kit. The amount of cell death by trypan blue staining and flow cytometry as well as by apoptosis kit (Partec) was examined.

 Results

 Lymphoblastic cell lines (C121) in exposure to different concentrations of hesperetin were affected and ΜM 200 drug concentration at 48 hours was reported as inhibitory concentration or IC50. The mode of cell death induced by hesperetin was found to be apoptosis, as judged by the morphologic alteration of the cells and by Anexin v conjugated with FITC and flow cytometric analysis.

 Conclusions

 Since hesperetin can induce apoptosis and reduce cell activity, it seems appropriate to be able to inhibit the growth of tumor cells.

Abstract

Background and Objectives

Prevention, control and treatment of cancer using natural materials currently have been considered as the promising strategy. In this regard, caffeine as a purine alkaloid found in various plants, including coffee, cocoa, cola and tea has recently attracted great interest because of its remarkable multifunctional inhibitory effects on cancers such as leukemia. Regarding the anti-cancer effects of caffeine, we sought to investigate the effectiveness of this methylxanthine alkaloid against NB4 acute promyelocytic leukemia cells.

Materials and Methods

In order to examine the effects of caffeine in APL, NB4 cells were cultured in the presence of different concentrations of caffeine subsequently, MTT, Caspase 3 activity assay, BrdU cell proliferation assay and quantitative real-time PCR were performed to assess the effect of caffeine on cell viability and the possible mechanisms of inducing cell death.

Results

Evaluation of DNA synthesis and cell survival using BrdU and MTT methods indicated that caffeine reduces proliferation and survival of NB4 cells in a dose and time dependent manner. The results of this study showed that elevation in the dose of caffeine induces caspase-3 activation and increases Bax and p21 gene mRNA expression levels.

Conclusions

Overall, our data illustrate that caffeine inhibits cell proliferation and induces apoptosis in acute promyelocytic leukemia NB4 cells. Thus, it can be concluded that caffeine as a substance in tea may be a useful agent for induction of cell death in malignant cells of patients with acute promyelocytic leukemia.

Key words: Acute Promyelocytic Leukemia, Apoptosis, Caffeine, Caspases

Abstract

Background and Objectives

Prevention, control and treatment of cancer using natural materials currently have been considered as the promising strategy. In this regard, caffeine as a purine alkaloid found in various plants, including coffee, cocoa, cola and tea has recently attracted great interest because of its remarkable multifunctional inhibitory effects on cancers such as leukemia. Regarding the anti-cancer effects of caffeine, we sought to investigate the effectiveness of this methylxanthine alkaloid against NB4 acute promyelocytic leukemia cells.

Materials and Methods

In order to examine the effects of caffeine in APL, NB4 cells were cultured in the presence of different concentrations of caffeine subsequently, MTT, Caspase 3 activity assay, BrdU cell proliferation assay and quantitative real-time PCR were performed to assess the effect of caffeine on cell viability and the possible mechanisms of inducing cell death.

Results

Evaluation of DNA synthesis and cell survival using BrdU and MTT methods indicated that caffeine reduces proliferation and survival of NB4 cells in a dose and time dependent manner. The results of this study showed that elevation in the dose of caffeine induces caspase-3 activation and increases Bax and p21 gene mRNA expression levels.

Conclusions

Overall, our data illustrate that caffeine inhibits cell proliferation and induces apoptosis in acute promyelocytic leukemia NB4 cells. Thus, it can be concluded that caffeine as a substance in tea may be a useful agent for induction of cell death in malignant cells of patients with acute promyelocytic leukemia.

Key words: Acute Promyelocytic Leukemia, Apoptosis, Caffeine, Caspases

Abstract

Background and Objectives

Prevention, control and treatment of cancer using natural materials currently have been considered as the promising strategy. In this regard, caffeine as a purine alkaloid found in various plants, including coffee, cocoa, cola and tea has recently attracted great interest because of its remarkable multifunctional inhibitory effects on cancers such as leukemia. Regarding the anti-cancer effects of caffeine, we sought to investigate the effectiveness of this methylxanthine alkaloid against NB4 acute promyelocytic leukemia cells.

Materials and Methods

In order to examine the effects of caffeine in APL, NB4 cells were cultured in the presence of different concentrations of caffeine subsequently, MTT, Caspase 3 activity assay, BrdU cell proliferation assay and quantitative real-time PCR were performed to assess the effect of caffeine on cell viability and the possible mechanisms of inducing cell death.

Results

Evaluation of DNA synthesis and cell survival using BrdU and MTT methods indicated that caffeine reduces proliferation and survival of NB4 cells in a dose and time dependent manner. The results of this study showed that elevation in the dose of caffeine induces caspase-3 activation and increases Bax and p21 gene mRNA expression levels.

Conclusions

Overall, our data illustrate that caffeine inhibits cell proliferation and induces apoptosis in acute promyelocytic leukemia NB4 cells. Thus, it can be concluded that caffeine as a substance in tea may be a useful agent for induction of cell death in malignant cells of patients with acute promyelocytic leukemia.

Abstract

Background and Objectives

Acute promyelocytic leukemia (APL) is characterized by a specific t(15;17), distinct morphologic picture, and a clinical coagulopathy that contributes to the morbidity and mortality of the disease. Telomerase is consistently activated in nearly all APL patients and telomerase-mediated telomere stabilization is responsible for unlimited replicative potential of the disease. This study aimed to investigate the effects of designed oligonucleotide as a telomerase inhibitor on NB4 cells.

Materials and Methods

NB4 leukemic cells were treated with various concentrations of oligonucleotide by transfection method using lipofectamin 2000. The inhibitory effect of oligonucleotide on cell metabolic activity and cell viability was assessed by MTT assay. Quantitative real-time PCR were applied to investigate alteration of Bax and Bcl-2 mRNA levels.

Results

Oligonucleotide decreased cell viability index and exerted cytotoxic effect against NB4 leukemic cells; we found that exposing cells with Oligonucleotide at 40 pMol for 48 h induced cell death and apoptotic effects on NB4 cells. Furthermore, transcriptional suppression of Bcl-2 and upregulation of Bax were found upon NB4 treatment by oligonucleotide.

Conclusions

Based on the short telomere length and high terlomerase activity in APL as well as inhibitory effect of oligonucleotide against NB4 cells, anti-telomerase-based therapy might be regarded as a successful strategy in APL therapy.

Abstract

Background and Objectives

Acute lymphoblastic leukemia (ALL) is the most common childhood malignancy. The side effects of chemotherapeutic drugs have primarily lead to the increased use of natural products for cancer treatment. Juniperus excelsa, a medicinal herb, was reported to show antiproliferative effects on various cancer cells. In this study, cytotoxic and apoptotic effects of extract from aerial parts of Juniperus excelsa plant were investigated on acute lymphoblastic leukemia cell lines, Nalm-6 and Reh.  

Materials and Methods

In this basic research, Nalm-6 and Reh were cultured and then they were treated with various concentrations of J. excelsa extract for 48 and 72 h and then the cell viability was evaluated by using MTT assay. Apoptosis also was assessed by caspase 3 activity assay and flow cytometry following Annexin V and Propidium iodide staining. Statistical analysis was assessed by one-way ANOVA test and SPSS 23.

Results

MTT assay results show that J. excelsa extract concentrations of 3, 4 and 5 µg/ml significantly reduce percentage of alive cells (p < 0.001). Flow cytometry results also show that J. excelsa extract significantly increase percentage of apoptotic cells compared with control groups (p < 0.001). Caspase 3 activity assay results show that caspase 3 activity was significantly increased in treated cells (p < 0.05).

Conclusions 

Our study shows that extract from aerial parts of J. excelsa has cytotoxic and apoptotic effects on Nalm-6 and Reh cells.

Abstract

Background and Objectives

Acute lymphoblastic leukemia (ALL) is the most common malignancy among children. The use of natural compounds has invoked a lot of interest in treatment of malignancies due to the adverse effects of chemotherapy treatment. Artemisia annua was reported to show cytotoxic effects on various cancer cell lines. In this study, cytotoxic effects of Artemisia annua extract were investigated on acute lymphoblastic leukemia cell lines.  

Materials and Methods

In this basic research, Nalm-6 and Reh cells were cultured and then treated with various concentrations of Artemisia annua extract; afterwards, the cell viability was evaluated using MTT assay for 48 and 72 h. The Caspase 3 activity assay and flow cytometry following Annexin V and Propidium iodide staining were used to assess apoptosis. Statistical analysis was evaluated by one-way ANOVA test.

Results

A. annua extract showed IC50 of 90 µg/ml on Nalm-6 and IC50 of 70 µg/ml on Reh cells after 48 h; the cytotoxic effect after 72 h was noticeable (p < 0.001). Flow cytometry results also show that A. annua extract concentration of 40 µg/ml increases the percentage of apoptotic cells compared with control groups (p < 0.05). Significant increase in Caspase 3 activity was observed after treatment with Artemisia annua compared with control groups (p < 0.01).

Conclusions 

Our results show that methanolic extract of aerial parts of A. annua exerting cytotoxic and inhibitory effects on Nalm-6 and Reh cells.

Key words: Acute Lymphoid Leukemia, Artemisia annua, Apoptosis

Abstract

Background and Objectives

The frequency of deregulated PI3K in acute lymphoblastic leukemia (ALL) coupled with the critical role of this signaling pathway in the acquisition of chemo-resistant phenotype lend compelling weight to the application of PI3K inhibitors for the treatment of ALL. In this study we aimed to evaluate the effect of selective p110δ inhibitor, GS-1101 on acute lymphoblastic leukemia Nalm-6 cells.

Materials and Methods

Nalm-6 cells were treated with different concentrations of GS-1101. In order to determine the cytotoxic and anti-proliferative effects of the drug, MTT and trypan blue exclusion assays were performed, respectively. Afterwards the effect of GS-1101 on cell cycle progression was evaluated using flowcytometry. Finally, Rq-PCR was applied to evaluate the mRNA expression level of cell cycle- and apoptotic-related genes in GS-1101-treated cells.

Results

Our results showed that GS-1101 not only reduced the number of viable cells but also hampered the metabolic activity of inhibitor-treated Nalm-6 cells both in dose- and time-dependent manner. Moreover, we found that the anti-proliferative effect of GS-1101 is mediated, at least partially, through the induction of G1 arrest as a result of up-regulated p21 expression level and accumulation of cells in sub-G1 phase of cell cycle. The results of Rq-PCR also demonstrated that GS-1101 exerts an inductive effect on the mRNA expression level of Bax, as the most important pro-apoptotic gene.

Conclusions

p110δ isoform inhibitor, GS-1101 shows both apoptotic and anti-proliferative effect on Nalm-6 cells through inducing cell-cycle arrest via up-regulation of cycline-dependent kinase inhibitor, p21 and upregulation of pro-apoptotic-related gene.

Abstract

Background and Objectives

PI3K/Akt pathway plays a key role in cell growth and proliferation. Constitutive activation of this pathway is detectable in 50-70% of patients with acute promyelocytic leukemia (APL). Previous studies showed that PI3K activity in APL cells is mainly due to overexpression of p110δ  isoform. In an effort to investigate the effect of PI3K inhibition in acute promyelocytic leukemia, APL-derived NB4 cells were subjected to different concentrations of Idelalisib, as a potent p110δ-specific inhibitor.

Materials and Methods

In this experimental study, we examined the effect of Idelalisib on the viability and metabolic activity of NB4 cells. Moreover, flowcytometery and quantitative RQ-PCR were applied to investigate both induction of apoptosis and transcriptional activity of apoptosis-related genes, respectively.

Results

The results revealed that Idelalisib not only decreased cell viability and metabolic activity in a dose- and time-dependent manner, but also induced cell death through the apoptotic pathway in NB4 cells. Our data also delineated that Idelalisib-induced apoptosis was mediated through induction of Bax and PUMA coupled with decreased expression of Mcl-1.

Conclusions

Based on P110δ activity in APL patients and the potent efficacy of Idelalisib in NB4 cells, it is tempting to suggest this inhibitor, as either single agent or in combination with common medications, for the treatment of patients with APL.

Abstract
Background and Objectives
The expression of Numb as a cell-fate determinant gene leads to inhibition of leukemic cell growth. The high levels of Musashi2 (Msi2) results in down-regulation of Numb in chronic myeloid leukemia cells. In this study we knocked down Msi2 using RNA interference (RNAi) strategy and investigated the effects of this knock down on the expression of Musashi2-Numb axis and genes involved in proliferation and apoptosis.
 
Materials and Methods
In this experimental study, synthetic double stranded siRNAs designed to target human Msi2 were transfected to K562 cells using Lipofectamine in duplicate. Changes in the expression levels of Msi-2, Numb, P21, and Bcl-2 genes 48h after transfection were evaluated by real-time PCR. Induction of apoptosis in transfected leukemic cells was determined using Annexin-PI staining and flowcytometry analysis. Data were analyzed by t-test.
 
Results
We found that upon Msi2 suppression, the expression levels of the cell-fate determinant Numb showed two-fold increase in K562 cells (p < 0/05). Msi2 down-regulation and subsequent increase in Numb expression levels caused the elevated expression of p21, as a cell cycle regulator (p < 0/05). In addition, Msi2 down-regulation promoted cell apoptosis via the down-regulation of Bcl-2 expression (p < 0/05). We observed that the rates of apoptosis in leukemic increased to 52% and 60%, 24h and 48h after transfection.
 
Conclusions 
It seems that Msi2 could be an option for targeting leukemic cells and its down-regulation through RNAi strategy may lead to induction of apoptosis in leukemic cells. This approach may open up new opportunities for leukemia therapy.
 

 

Abstract
Background and Objectives
Telomerase-targeted therapy for cancer has received great attention because telomerase is expressed in almost all cancer cells but is inactive in most normal somatic cells. This study aimed to investigate the effects of telomerase inhibitor MST-312, a chemically modified derivative of epigallocatechin gallate (EGCG), on apoptosis of myeloma cell line U266.
 
Materials and Methods
In an Experimental study, U266 cells were treated with different concentrations of MST-312 at different times; then, cell viability by trypan blue exclusion assay, cell metabolic activity by MTT assay, and cell apoptosis by Annexin V Apoptosis Detection Kit were measured. To further investigate apoptosis, BAX and BCL2 gene expression of the treated cells was investigated by the quantitative Real-Time PCR.
 
Results
MST-312 exerted a dose-dependent short-term cytotoxic effect on myeloma cells. Over 50% decrease in the viability of treated cells was seen in 8 μM concentration of MST-312 within 48 h. The cytotoxic effect of MST-312 was concentration-dependent, with approximately 25, 46, and 62% reduction in metabolic activity after 48 h exposure to 2, 4, and 8 μM of MST-312, respectively. Gene expression analysis showed downregulation of antiapoptotic gene Bcl-2 and upregulation of apoptopic gene BAX.
 
Conclusions 
Inhibition of telomerase activity by MST-312 represents a novel treatment strategy for Multiple Myeloma cancer.