ABSTRACT
BACKGROUND: Growing evidence has demonstrated that microRNAs have a major effect on development of different types of cancer including AML. The overexpression of miR-625 could decrease tumorgenesis of acute myeloid leukemia cell lines through Integrin-linked kinase signaling pathway and reducing the associated oncogenes. The aim of this study is to evaluate the effect of hsa-miR-625 upregulation on apoptosis and proliferation of KG1 cell line via ILK signaling pathway. METHODS: The KG-1 cell line was transfected with pLenti-III-premir625-GFP through viral method. Then, expression of miR-625 and genes were analyzed by quantitative PCR. Western blotting was used to evaluate for the protein level. Apoptosis was investigated by flow cytometry. Cell cycle analysis with PI and CCK-8 assay were performed to evaluate proliferation. RESULTS: KG-1 cells transfected with pLenti-III-pre mir625-GFP construct showed a significant increase in cell apoptosis. Gene expression of ILK and NF-κB were downregulated and AKT, c-fos, Caspase3, cyclin D1, KLF-4, OCT-4 and Nanog were upregulated but no alteration in GSK3 expression profile was observed. Downregulation of NF-κB and upregulation of Caspase 3 and p-ß-catenin protein levels were indicated (p<0.05). CONCLUSION: MiR-625 can be a promising approach to aid in the treatment of AML. However, further studies are required in this respect.
Subject(s)
Leukemia, Myeloid, Acute , MicroRNAs , Apoptosis/genetics , Cell Line, Tumor , Cell Proliferation/genetics , Glycogen Synthase Kinase 3/genetics , Glycogen Synthase Kinase 3/metabolism , Humans , Leukemia, Myeloid, Acute/genetics , Leukemia, Myeloid, Acute/metabolism , MicroRNAs/genetics , MicroRNAs/metabolism , NF-kappa B/metabolism , Protein Serine-Threonine Kinases , Up-RegulationABSTRACT
Objective: Acute myeloid leukemia (AML) is characterized by abnormalities of differentiation and growth of primary hematopoietic stem cells (HSCs) in the blood and bone marrow. In many studies, miR-625-5p has been shown to inhibit downstream pathways from affecting the metastasis and invasion of the integrin-linked kinase (ILK) signaling pathway. It has been proved that the expression of miR-625-5p decreases in AML cell lines. This study aimed to investigate the effect of miR-625-5p upregulation on the invasion of KG1 ell line in vitro. Materials and Methods: In this experimental study, we investigated the impact of upregulation of miR-625-5p on invasion via the ILK/AKT pathway in the KG1 cell line. After transfection using the viral method, the cellular invasion was assessed by invasion assay and the levels of miR-625-5p genes and protein were evaluated by quantitative polymerase chain reaction (qPCR) and western blotting. Moreover, CXCR4 level was assessed by flow cytometry. Results: The invasion significantly reduced in MiR-625-5p-transfected KG1 cells (P<0.01) that was concomitant with remarkably decreasing in the expression levels of ILK, NF-κB, and COX2 genes compare with the control group (P<0.01). Incontrast, MMP9, AP1, and AKT significantly increased (P<0.01, P<0.001 and P<0.01, respectively) and GSK3ß did not change significantly in MiR-625-5p-transfected KG1 cells. The protein level of NF-κB decreased (P<0.01) and MMP9 increased, however it was not significant. Moreoever, the expression of CXCR4 was significantly lower (P<0.01) in comparison with the control group. Conclusion: miR-625-5p leads to a reduction in cell invasion in the AML cell line through ILK pathway. Therefore, it could be a breakthrough in future AML-related research. However, further studies are needed to support this argument.
