The impact of exosomes derived from B-cell acute lymphoblastic leukemia as a growth factor on bone marrow mesenchymal stromal cells.

Background The incidence of various types of cancers, including leukemia, is on the rise and many challenges in both drug resistance and complications related to chemotherapy appeared. Recently, the development and application of extracellular vesicles (EV) such as exosomes in the management of cancers, especially leukemia, holds great significance. In this article, we extracted exosomes from NALM6 cells and assessed their regulatory effects on proliferation and apoptosis in mesenchymal stem cells (MSCs). Method and result We first verified the exosomes using various techniques, including flow cytometry, transient electron microscopy, dynamic light scattering (DLS), and BCA protein assay. Then MTT analysis and flowcytometry (apoptosis and cell cycle assay) besides gene expressions were employed to determine the state of MSC proliferations. The results indicated that exosome-specific pan markers like CD9, CD63, and CD81 were present. Through DLS, we found out that the mean size of the exosomes was 89.68 nm. The protein content was determined to be 956.292 µg/ml. Analysis of MTT, flow cytometry (cell cycle and apoptosis assay), and RT-qPCR showed that in the dose of 50 µg/ml the proliferation of MSCs was increased significantly (p-value < 0.05). Conclusion All these data showed that exosomes use several signaling pathways to increase the MSCs' proliferation and drug resistance, ultimately leading to high mortalities and morbidities of acute lymphoblastic leukemia.

Identification of Prognostic Genes in Acute Myeloid Leukemia Microenvironment: A Bioinformatic and Experimental Analysis.

Acute myeloid leukemia (AML) is a lethal hematologic malignancy with a variable prognosis that is highly dependent on the bone marrow microenvironment. Consequently, a better understanding of the AML microenvironment is crucial for early diagnosis, risk stratification, and personalized therapy. In recent years, the role of bioinformatics as a powerful tool in clarifying the complexities of cancer has become more prominent. Gene expression profile and clinical data of 173 AML patients were downloaded from the TCGA database, and the xCell algorithm was applied to calculate the microenvironment score (MS). Then, the correlation of MS with FAB classification, and CALGB cytogenetic risk category was investigated. Differentially expressed genes (DEGs) were identified, and the correlation analysis of DEGs with patient survival was done using univariate cox. The prognostic value of candidate prognostic DEGs was confirmed based on the GEO database. In the last step, real-time PCR was used to compare the expression of the top three prognostic genes between patients and the control group. During TCGA data analysis, 716 DEGs were identified, and survival analysis results showed that 152 DEGs had survival-related changes. In addition, the prognostic value of 31 candidate prognostic genes was confirmed by GEO data analysis. Finally, the expression analysis of FLVCR2, SMO, and CREB5 genes, the most related genes to patients' survival, was significantly different between patients and control groups. In summary, we identified key microenvironment-related genes that influence the survival of AML patients and may serve as prognostic and therapeutic targets.

Expression Changes of SIRT1 and FOXO3a Significantly Correlate with Oxidative Stress Resistance Genes in AML Patients.

The increased metabolism in acute myeloid leukemia (AML) malignant cells resulted in the production of high levels of free radicals, called oxidative stress conditions. To avoid this situation, malignant cells produce a considerable amount of antioxidant agents, which will lead to the release of a continuous low level of reactive oxygen species (ROS), causing genomic damage and subsequent clonal evolution. SIRT1 has a key role in driving the adaptation to this condition, mainly through the deacetylation of FOXO3a that affects the expression of oxidative stress resistance target genes such as Catalase and Manganese superoxide dismutase (MnSOD). The aim of this study is to simultaneously investigate the expression of SIRT1, FOXO3a, and free radical-neutralizing enzymes such as Catalase and MnSOD in AML patients and measure their simultaneous change in relation to each other. The gene expression was analyzed using Real Time-PCR in 65 AML patients and 10 healthy controls. Our finding revealed that expression of SIRT1, FOXO3a, MnSOD and Catalase was significantly higher in AML patients in comparison to healthy controls. Also, there was a significant correlation between the expression of SIRT1 and FOXO3a, as well as among the expression of FOXO3a, MnSOD and Catalase genes in patients. According to the results, the expression of genes involved in oxidative stress resistance was higher in AML patients, which possibly contributed to the development of malignant clones. Also, the correlation between the expression of SIRT1 and FOXO3a gene reflects the importance of these two genes in increased oxidative stress resistance of cancer cells.

An overview of the molecular and clinical significance of the angiopoietin system in leukemia.

The angiogenesis efficacy in solid tumors and hematological malignancies has been identified for more than twenty years. Although the exact role of angiogenesis in leukemia as a common hematological malignancy has not yet been extensively studied, its effect is demonstrated on the initiation and maintenance of a favorable microenvironment for leukemia cell proliferation. The angiopoietin family is a defined molecular mediator for angiogenesis, which contributes to vascular permeability and angiogenesis initiation. They participate in the angiogenesis process by binding to tyrosine kinase receptors (Tie) on endothelial cells. Considering the role of angiogenesis in leukemia development and the crucial effects of the Ang-Tie system in angiogenesis regulation, many studies have focused on the correlation between the Ang-Tie system and leukemia diagnosis, monitoring, and treatment. In this study, we reviewed the Ang-Tie system's potential diagnostic and therapeutic effects in different types of leukemia in the gene expression level analysis approach. The angiopoietin family context-dependent manner prevents us from defining its actual function in leukemia, emphasizing the need for more comprehensive studies.

The effect of microvesicles derived from K562 cells on proliferation and apoptosis of human bone marrow mesenchymal stem cells.

Objectives: Microvesicles (MVs) are small membrane-bound particles that act as a vehicle to transfer their contents, such as proteins, RNAs, and miRNAs, to the target cells, making them undergo several changes. Depending on the origin and the target cell, MVs may cause cell survival or apoptosis. This study investigated the effects of MVs released from the leukemic K562 cell line on the human bone marrow mesenchymal stem cells (hBM-MSCs) to evaluate changes in the survival or apoptosis of the cells in an in vitro system. Materials and Methods: In this experimental study, we added the isolated MVs from the K562 cell line to hBM-MSCs, and after three and then seven days, subsequently cell count, cell viability, transmission electron microscopy, tracing MVs by carboxyfluorescein diacetate, succinimidyl ester (CFSE) solution, flow cytometry analysis for Annexin-V/PI staining and qPCR for the evaluation of BCL-2, KI67, and BAX expression were carried out. On the 10th day of the culture, hBM-MSCs were examined by Oil red O and Alizarin Red staining to evaluate their differentiation into adipocytes and osteoblasts. Results: There was a significant decrease in cell viability and KI67 and BCL-2 expression; however, BAX was significantly upregulated in the hBM-MSCs compared to control groups. Annexin-V/PI staining results also showed the apoptotic effects of K562-MVs on hBM-MSCs. Moreover, the differentiation of hBM-MSCs into adipocytes and osteoblasts was not observed. Conclusion: MVs from the leukemic cell line could affect the viability of normal hBM-MSCs and induce cell apoptosis.

AML-derived extracellular vesicles negatively regulate stem cell pool size: A step toward bone marrow failure.

PURPOSE OF THE STUDY: Long-term repopulating hematopoietic stem cells (LTR-HSCs) have been previously shown to reside in close proximity to osteoblasts, where they take shelter in the bone marrow (BM) microenvironment against cytotoxic and apoptotic stimuli. Nevertheless, the function of the HSC niche is believed to undergo an adaptive evolutionary modification during leukemogenesis. Recent studies have demonstrated that leukemic clones can impact BM homing through extracellular vesicle (EV) secretion. However, the exact mechanism driving BM conversion is still unclear. In the present study, the human osteoblast cell line (MG-63) were subjected to various concentration of sera-derived EVs of patients with acute myeloid leukemia (AML) and healthy volunteers to assess if they are associated strongly enough to alter the expression pattern of cross-talk molecules involved in niche interactions. METHOD: To gain a brief insight into the EVs secretion criteria, we first conducted a comparative analysis of sera-derived EVs by dynamic light scattering (DLS), transmission electron microscopy (TEM), and Bradford assay. After incubating MG-63 cell lines with increasing concentrations of the EVs, Trypan-blue and microculture tetrazolium test (MTT) assays were used to evaluate the cell survival, logarithmic growth, and metabolic activity. Finally, the expression levels of OPN, ANGPT-1, and JAG-1 transcripts were evaluated through the qRT-PCR technique. RESULTS: Here, we report that AML-derived EVs can affect the viability, cell growth, and metabolic activity of the human osteoblasts cell line (MG-63) compared to those that received healthy-derived EVs. We also found that leukemic EVs tend to induce overexpression of OPN but reduce the expression of ANGPT-1 and JAG-1 genes in the osteoblast transcriptome, which may provide a potential context imposing selective suppression of HSC pool size. CONCLUSION: These findings extend the general concept of a novel mechanism in which leukemic EVs would make it possible to create a specialized pre-metastatic microenvironment in the interest of tumor expansion, allowing leukemic clones to overcome their HSCs counterparts.

Transcription analysis of a histones modifiers panel coupled with critical tumor suppressor genes displayed frequent changes in patients with AML.: mRNA levels of histones modifiers and TSGs in AML.

Epigenetic alterations could cause leukemia through the activation of normally silent loci or silencing of normally active loci. We herein aimed to compare the expression patterns of a histone modifiers panel consisted of SUV39H1, PRDM16, UHRF2, KDM2B, and KDM3C between acute myeloid leukemia(AML) cells and normal cells and to assess the correlation of these genes with the expression of vital tumor suppressor genes, including p16INK4A and p53. Bone marrow or peripheral blood samples of 50 AML patients at diagnosis and also 18 subjects with a normal hematopoietic system as a control group were obtained after informed consent. Then, qRT-PCR was performed to determine the expression levels of the aforementioned genes. We found a broad alteration in the expression signature of five out of seven studied genes in AML patients as compared with the control group. UHRF2 and p53 were remarkably downregulated in AML patients (P<0.001), while SUV39H1, PRDM16, and KDM3C were significantly overexpressed (P<0.01). Based on the Spearman rank correlation, SUV39H1 and KDM2B negatively regulated both p16INK4A and p53 expression. Taken together, our findings provided preliminary evidence regarding the pervasive mRNA perturbation of histone modifiers and their plausible influences on critical tumor suppressor genes. Future studies in this area would be required to assist in establishing these results in the clinical practice of AML patients.

Evaluation of Sestrin 2, Adiponectin, AMPK, and mTOR Genes Expression in Acute Myeloid Leukemia Patients.

BACKGROUND: Effective treatment of acute myeloid leukemia (AML) is still controversial, therefore; a comprehensive understanding regarding the impaired cellular signaling pathways in AML can be useful in designing new therapeutic approaches. Among signaling pathways involved in AML, the mammalian target of rapamycin (mTOR) signaling pathway is of particular importance. While dysregulation of mTOR signaling has been reported in a wide range of patients with AML, but most studies have focused on mTOR downstream targets, and mTOR upstream targets have been overlooked. OBJECTIVE: In this study, expression of mTOR genes and three upstream targets (5' adenosine monophosphate-activated protein kinase (AMPK, adiponectin, and sestrin 2) involved in mTOR signaling was investigated. MATERIALS AND METHODS: In this study, expression of mTOR, AMPK, sestrin 2, and adiponectin genes in 60 patients with AML were evaluated compared to those of 30 healthy individuals as controls using the Real-Time polymerase chain reaction (Real-Time RT-PCR) method. RESULTS: According to the results, there was a significant difference in the expression of all the studied genes in patients in comparison to the normal control group (P <0.05). Expression of the mTOR gene was increased, while expression of AMPK, sestrin 2, and adiponectin genes was decreased in the patients with AML. Mean expression of the genes (2-ΔCt) (AMPK, sestrin 2, adiponectin, and mTOR) was equal to 7.9, 3.2, 3.74, and 1.49 for controls and 6, 2.1, 2.83, and 2.64 for patients with AML, respectively. CONCLUSIONS: Given the decreased expression levels of sestrin 2, adiponectin, and AMPK genes as tumor inhibitors and the increased expression level of the mTOR gene as an oncogene in the patients with AML in our study, it is thought that disruption of this pathway may be involved in leukemogenesis and can be considered as an effective factor in the progression of cancer.

Triangle collaboration assessment of autophagy, ER stress and hypoxia in leukemogenesis: a bright perspective on the molecular recognition of B-ALL.

B-lineage acute lymphoblastic leukemia (B-ALL) is the most common acute leukemia in childhood and adults, which caused by many various crystalline and unclear agents. Owning to this matter, no significant progress has been made in the patients-recovery. Recently, autophagy pathway is considered as an ambiguous agent in leukemia evaluation. We aim to discover the expression levels of upstream autophagy-regulating genes in newly diagnosed B-ALL patients. In B-ALL group, BECN1, HIF1A and ERN1 expressions were significantly down-regulated, while BCL2 expression was up-regulated compared to the control group (p < .05). Moreover, there was significant positive correlation between the decreased BECN1 compared with Hypoxia and endoplasmic reticulum (ER) stress-related genes expression in the patients (p < .05). Our findings revealed that, ERN1 and ER stress pathway-related genes could be effective regulators of autophagy in B-ALL. More investigation is recommended to gain a deeper understanding into molecular pathophysiology of B-ALL to improve treatment and monitoring approaches in affected patients.

The protective effect of oleuropein against radiation-induced cytotoxicity, apoptosis, and genetic damage in cultured human lymphocytes.

PURPOSE: The aim of this study was to evaluate the effects of oleuropein radiation protection and to find an effective radioprotector. MATERIALS AND METHOD: Human mononuclear cells were treated with oleuropein at the concentration of 100 µM (optimum concentration), incubated for 24 h, and then exposed to 2 Gy gamma-rays. The anti-radiation effect of oleuropein was assessed by MTT assay, flow cytometry, comet assay, and micronucleus (MN) assay. RESULTS: It was found that pretreatment with oleuropein (25, 50, 75, 100, 200, 400, and 800 nM, and 1, 5, 10, 15, 20, 25, 30, 40, 50, 75, 100, 125, 150, 175, and 200 µM) significantly increased the percentage of cell viability compared to the irradiated group (p < .001). Moreover, oleuropein treatment with the above concentrations defined without gamma-ray did not show any cytotoxicity effect in human mononuclear cells. The LD50/24h dose was calculated as 2.9 Gy, whereas by 200, 150, 50, and 100 µM oleuropein prior to radiation (1, 2,and 4 Gy), radiation LD50/24h increased to 3.36, 3.54, 3.81, and >4 Gy, in that order. A very noticeable dose-modifying factor (DMF) of 1.16, 1.23, 1.31, and 1.72 was observed for 200, 150, 50, and 100 µM, in order. Therefore, 100 µM of oleuropein was selected as the desirable dose for radio-protection trial, and 2 Gy gamma-rays were used for further research. Human mononuclear cells treatment with oleuropein (100 µM) prior to 2 Gy gamma-rays significantly decreased apoptosis, genomic damage, and MN occurrence in human mononuclear caused by gamma-radiation (p < .001). Furthermore, treatment with oleuropein (100 µM) without radiation did not lead to apoptosis, genotoxicity, or clastogenic effects caused by oleuropein in human mononuclear cells. CONCLUSION: The results revealed that oleuropein is able to significantly reduce cytotoxicity, apoptosis, genotoxic, and clastogenic effects of gamma-rays.

IGF family effects on development, stability, and treatment of hematological malignancies.

Multiple factors, including growth factors, are shown to be culprits of cancer outset and persistence. Among growth factors, insulin-like growth factors (IGFs) family are of more importance in the prognosis of blood malignancies. After binding to their corresponding receptor, IGFs initiate PI3K/AKT signaling pathway and increase the translation of intracellular proteins, such as cell division-related proteins. They also stimulate the transcription of cell division-related genes using the Ras-GTP pathway. In addition to organs such as the liver, IGFs are secreted by tumor cells and can cause growth and proliferation of self or tumor cells via autocrine and paracrine methods. Current studies indicate that decreasing the effects of IGF by blocking them, their receptors, or PI3K/AKT pathway using various drugs could help to suppress the division of tumor cells. Here, we delineate the role of the IGF family in hematologic malignancies and their potential mechanisms.

The Expression of P53, MDM2, c-myc, and P14ARF Genes in Newly Diagnosed Acute Lymphoblastic Leukemia Patients.

The treatment response of acute lymphoblastic leukemia (ALL) depends on the percentage of lymphoblasts, cytogenetic aberrations, and altered gene expression. The analysis of the gene expression is applicable for determination of risk stratification and prognosis of cancers. c-MYC, P14ARF, MDM2, and P53 play a vital role in cell survival through a functional network. This study aimed to investigate the expression of these genes, also their correlation with immunophenotypic subtypes of ALL and the percentage of blasts. Real-time PCR was performed for the expression analysis of P53, MDM2, c-MYC, and P14ARF in the bone marrow or peripheral blood samples of 52 ALL patients and 13 normal samples as controls. The morphological analysis and flow cytometry were carried out to examine the phenotypes and percentage of lymphoblasts. The decreased expression levels of P53 and MDM2 were seen in ALL patients compared with control group. In T cell subgroup of ALL the expression of P14ARF gene was more decreased among other subgroups. The expression of MDM2 was decreased in ALL patients who were under the age of 16. Based on our study, the interaction between P53 and MDM2 might be more complex and different from reports published in previous studies. Our findings showed that MDM2 is not negatively correlated with P53, at least in our samples. It can be very effective on the current and future studies to use different techniques for analysis of genome, transcriptome, and proteome in definitive risk stratification and prognosis determination.

Biodecomposition of Phenanthrene and Pyrene by a Genetically Engineered Escherichia coli.

BACKGROUND: Genetically engineered microorganisms (GEMs) can be used for bioremediation of the biological pollutants into nonhazardous or less-hazardous substances, at lower cost. Polycyclic aromatic hydrocarbons (PAHs) are one of these contaminants that associated with a risk of human cancer development. Genetically engineered E. coli that encoded catechol 2,3- dioxygenase (C230) was created and investigated its ability to biodecomposition of phenanthrene and pyrene in spiked soil using high-performance liquid chromatography (HPLC) measurement. We revised patents documents relating to the use of GEMs for bioremediation. This approach have already been done in others studies although using other genes codifying for same catechol degradation approach. OBJECTIVE: In this study, we investigated biodecomposition of phenanthrene and pyrene by a genetically engineered Escherichia coli. METHODS: Briefly, following the cloning of C230 gene (nahH) into pUC18 vector and transformation into E. coli Top10F, the complementary tests, including catalase, oxidase and PCR were used as on isolated bacteria from spiked soil. RESULTS: The results of HPLC measurement showed that in spiked soil containing engineered E. coli, biodegradation of phenanthrene and pyrene comparing to autoclaved soil that inoculated by wild type of E. coli and normal soil group with natural microbial flora, were statistically significant (p<0.05). Moreover, catalase test was positive while the oxidase tests were negative. CONCLUSION: These findings indicated that genetically manipulated E. coli can provide an effective clean-up process on PAH compounds and it is useful for bioremediation of environmental pollution with petrochemical products.

The Expression of Interferon Gamma (IFN-γ) and Interleukin 6 (IL6) in Patients with Acute Lymphoblastic Leukemia (ALL).

Interferon gamma (IFN-γ) and interleukin 6 (IL-6) are including the most important cytokines which have been associated with the biological behavioral and immune responses in malignancies. Based on the critical roles which these two cytokines play against tumor cells, the present study was aimed to investigate the genes expression level of IL6 and IFN-γ in patients with Acute Lymphoblastic Leukemia and compare with normal controls. Fifty-two patients with ALL and 13 healthy volunteer were under studied. The peripheral blood mononuclear cells of all patients and normal controls were separated by ficoll. The expression of interferon gamma and interleukin 6 genes were determined by RQ-PCR. Finally all data were analyzes using T student, one way ANOVA and Mann-Whitney tests were use to analyze all samples data. Our finding showed that the level of IFN-γ gene expression was significant decreased in patients with All as compared with healthy controls (83 change fold, p < 0.0001). The level of IL-6 Gene expression was not changeable in B-ALL patients as compared with healthy control (p = 0.4), but in T-ALL patients, was significantly reduced (p < 0.01). The results of present study indicated that IFN-γ gene expression reduced in ALL patients. It provides a valuable insight that immune system may disrupted in patients with ALL, which cause tumor cells escape from immune surveillance.

Standardization of Plasma Rich in Growth Factors (PRGF) and its effects on androgenic hair loss.

Androgenic alopecia (AGA), as the most common cause of hair loss, is a chronic process that affects 80% of men and 50% of women throughout life. Existing and approved treatments for this condition are limited, and unfortunately, the length of treatment is long, while its efficacy is not much suitable. Plasma rich in growth factors (PRGF) autologous therapy is based on the delivery of a pool of bioactive molecules impressive for the treatment of AGA.Thirteen patients were included in this study. Our patients were evaluated in two groups: the first group was injected once and the second group was injected thrice, then evaluated for the number and diameter of the hair.Both groups of patients showed hopeful results so that in the first group hairs number and thickness increased by 9-54% and 11-76% respectively (p < .01). For patients who underwent PRGF injection thrice, the increases in hairs number and thickness were remarkably higher with an average of 211 and 221 respectively (p < .001). No adverse effect was reported in any patient.Our results revealed that PRGF platelet concentration using a higher volume of blood compared to previous protocols has higher effectiveness in treating AGA. However, more randomized clinical studies with longer follow up courses as well as larger sample sizes are needed to standardize an optimum protocol for PRGF based treatments.

Evaluation of ATG7 and Light Chain 3 (LC3) Autophagy Genes Expression in AML Patients.

BACKGROUND AND AIM: Autophagy, known as cell death type II, is a housekeeping pathway that currently has been worked on in matters of tumorigenesis and leukomogenesis. Therefore, expression levels of ATG7 and LC3 as two key genes in AML patients are targeted in this study. MATERIAL AND METHOD: This study was performed on 55 de novo AML patients against 17 healthy volunteers, acquired samples from bone marrow (BM) and peripheral blood (PB) sources in different ages and gender. The evaluation was executed by mRNA extraction, cDNA synthesis, real-time PCR and data was analyzed by SPSS. RESULTS: Analyzed data indicate a significant decrease between expression of ATG7 and LC3 in AML patients against control (Pv < 0.05). Decrease in both genes expression was detected in most of the patients, 81.81% and 75.55%, respectively. Also LC3 overexpression was detected in 11.33% of AML patients. Moreover, a positive significant correlation between ATG7 and LC3 genes was detected (r = 0.481; Pv = 0.001). CONCLUSION: This study showed that significant reduction of autophagy genes in de novo AML patients is important to overcome this system and initiate leukomogenesis. It seems a new insight is required for new achievements in diagnosis, prognosis, treatment and monitoring AML patients.

Hypoxia-Inducible Factor1-Α (HIF1α) and Vascular Endothelial Growth Factor-A (VEGF-A) Expression in De Novo AML Patients

Background: Bone marrow hypoxia can promote leukemia progression in human cases of acute myeloid leukemia (AML). In addition, low oxygen tension is able to regulate the expression of different genes involved in malignancy. In this study, we hypoxia-inducible factor-1α (HIF-1α) and vascular endothelial growth factor (VEGF-A) genes were assessed as principal regulators of hypoxia in do novo AML patients. Methods: Peripheral blood and bone marrow samples were collected from 57 AML patients and 17 normal control subjects with informed consent. Expression of HIF1α and VEGF-A was then evaluated using quantitative real-time PCR (Q-Real time PCR) and data were analyzed with SPSS 16. Result: HIF1α and VEGF-A showed overexpression in AML patients compared to normal controls (P <0.0001 and P<0.005, respectively). The expression level of HIF1α was significantly higher in AML-M3 cases versus AML-non M3 cases. Furthermore, there was a positive correlation between HIF1α and VEGF-A ( P <0.0001 and r = 0.497). Conclusion: Adding to the many studies on the role of hypoxia in solid tumors, our data indicate that HIF1a and VEGF-A overexpression also occurs in AML patients. We consider that this is possibly involved in leukemic cell growth and therefore could be a promising target for clinical control.

CDK Blockade Using AT7519 Suppresses Acute Myeloid Leukemia Cell Survival through the Inhibition of Autophagy and Intensifies the Anti-leukemic Effect of Arsenic Trioxide.

The strong storyline behind the critical role of cyclin-dependent kinase (CDK) inhibitor proteins in natural defense against malignant transformation not only represents a heroic perspective for these proteins, but also provides a bright future for the application of small molecule inhibitors of CDKs in the novel cancer treatment strategies. The results of the present study revealed that the inhibition of CDKs using pan-CDK inhibitor AT7519, as revealed by the induction of G1 cell cycle arrest as well as the reduction of cyclins expression, resulted in decreased survival in acute myeloid leukemia (AML)-derived KG-1 cells, either in the context of single agent or in combination with arsenic trioxide (ATO). Apart from alterations in the expression of proliferation and apoptotic genes, the anti-survival property of AT7519 was coupled with the inhibition of autophagy-related genes. Notably, we found that the blockage of autophagy system in KG-1 cells resulted in a superior cytotoxic effect, introducing autophagy as a probable suppressor of cell death. As far as we are aware, to date, no study has reported the contributory mechanisms correlated with the less sensitivity of acute leukemia cells to AT7519 and our study suggested for the first time that the activation of both PI3K and c-Myc signaling pathways could overshadow, at least partly, the efficacy of this agent in KG-1 cells. Overall, due to the pharmacologic safety of AT7519, our study proposed this inhibitor as a promising agent for the treatment of AML either as a single agent or in a combined-modal strategy.

Comparison between the cultures of human induced pluripotent stem cells (hiPSCs) on feeder-and serum-free system (Matrigel matrix), MEF and HDF feeder cell lines.

Human induced pluripotent stem cells (hiPSCs) are a type of pluripotent stem cells artificially derived from an adult somatic cell (typically human fibroblast) by forced expression of specific genes. In recent years, different feeders like inactivated mouse embryonic fibroblasts (MEFs), human dermal fibroblasts (HDFs), and feeder free system have commonly been used for supporting the culture of stem cells in undifferentiated state. In the present work, the culture of hiPSCs and their characterizations on BD Matrigel (feeder-and serum-free system), MEF and HDF feeders using cell culture methods and molecular techniques were evaluated and compared. The isolated HDFs from foreskin samples were reprogrammed to hiPSCs using gene delivery system. Then, the pluripotency ability of hiPSCs cultured on each layer was determined by teratoma formation and immunohistochemical staining. After EBs generation the expression level of three germ layers genes were evaluated by Q-real-time PCR. Also, the cytogenetic stability of hiPSCs cultured on each condition was analyzed by karyotyping and comet assay. Then, the presence of pluripotency antigens were confirmed by Immunocytochemistry (ICC) test and alkaline phosphatase staining. This study were showed culturing of hiPSCs on BD Matrigel, MEF and HDF feeders had normal morphology and could maintain in undifferentiated state for prolonged expansion. The hiPSCs cultured in each system had normal karyotype without any chromosomal abnormalities and the DNA lesions were not observed by comet assay. Moreover, up-regulation in three germ layers genes in cultured hiPSCs on each layer (same to ESCs) compare to normal HDFs were observed (p < 0.05). The findings of the present work were showed in stem cells culturing especially hiPSCs both MEF and HDF feeders as well as feeder free system like Matrigel are proper despite benefits and disadvantages. Although, MEFs is suitable for supporting of stem cell culturing but it can animal pathogens transferring and inducing immune response. Furthermore, HDFs have homologous source with hiPSCs and can be used as feeder instead of MEF but in therapeutic approaches the cells contamination is a problem. So, this study were suggested feeder free culturing of hiPSCs on Matrigel in supplemented media (without using MEF conditioned medium) resolves these problems and could prepare easy applications of hiPSCs in therapeutic approaches of regenerative medicine such as stem-cell therapy and somatic cell nuclear in further researches.