|Year : 2017 | Volume
| Issue : 3 | Page : 95-98
The role of the tumor-suppressive microRNA-370 and microRNA-29b in acute myeloid leukemia
Marwa Saied1, Fatma Mohamed2, Mohamed Eissa3, Dalia Naefae2
1 Department of Clinical Pathology, Faculty of Medicine, Alexandria University, Alexandria, Egypt
2 Department of Hematology, Faculty of Medicine, Alexandria University, Alexandria, Egypt
3 Department of Clinical Pathology, Faculty of Medicine, Zagazig University, Zagazig, Egypt
|Date of Submission||07-Jul-2017|
|Date of Acceptance||18-Jul-2017|
|Date of Web Publication||9-Nov-2017|
Clinical Pathology Department, Faculty of Medicine, Alexandria University, Alexandria, 21512
Source of Support: None, Conflict of Interest: None
Background MicroRNAs (miRNAs) are small noncoding RNAs that play important roles as diagnostic and prognostic markers in malignancy. They have diverse effects in acute myeloid leukemia (AML), mainly as tumor suppressors and occasionally provoke the disease.
Aim The aim of this research was to study the expressions of both miRNA-370 and miRNA-29b in de-novo AML Egyptian patients and correlate their expression with clinical and laboratory parameters.
Patients and methods This is a case–control study that was conducted in Alexandria Main University Hospital, Alexandria University. MiRNA quantification by real-time PCR was conducted on 30 de-novo AML patients admitted to the hematology unit of internal medicine department. Their age ranged between 13 and 63 years. Their results were compared with 10 age-matched and sex-matched controls. Descriptive statistics was performed using SPSS statistics software.
Results Both miRNA-370 and miRNA-29b were significantly downregulated in AML patients (P=0.011 and 0.015, respectively). There was significant positive correlation between the expressions of miRNA-370 and miRNA-29b in AML patients (P=0.003). Neither miRNAs showed significant correlation with FAB subtypes. Moreover, miRNA-370 expression was significantly negatively correlated with the age of the patients (P=0.034) − that is, the older the patient the lower the expression level of miRNA-370.
Conclusion There was a significant down-regulation of both miRNA-370 and miRNA-29b in de-novo AML patients. The positive correlation between both miRNA expressions might point to their potential roles as tumor suppressors.
Keywords: acute myeloid leukemia, microRNA-29b, microRNA-370
|How to cite this article:|
Saied M, Mohamed F, Eissa M, Naefae D. The role of the tumor-suppressive microRNA-370 and microRNA-29b in acute myeloid leukemia. Egypt J Haematol 2017;42:95-8
|How to cite this URL:|
Saied M, Mohamed F, Eissa M, Naefae D. The role of the tumor-suppressive microRNA-370 and microRNA-29b in acute myeloid leukemia. Egypt J Haematol [serial online] 2017 [cited 2018 Jul 22];42:95-8. Available from: http://www.ehj.eg.net/text.asp?2017/42/3/95/217880
| Introduction|| |
Acute myeloid leukemia (AML) is a disease of heterogeneous disorders resulting from proliferation or arrest of myoblasts at different stages of maturation ,. Many pathways are involved in the pathogenesis of AML, which are controlled by various genes. Remarkably, small noncoding RNAs (920–922 nucleotides) were shown to target genes regulating the leukemogenesis , as upregulation or downregulation of microRNAs (miRNAs) can initiate or propagate AML . Interestingly, some miRNAs have controversial effects in cancer − for example, they can act as apoptotic or oncogenic factors. Examples of those miRNAs are miRNA-370 and miRNA-29b .
miRNA-370 is found to be downregulated in different tumors ,. In AML, miRNA-370 can act as a tumor suppressor gene through targeting FOXM1 , resulting in AML initiation and progression. Similarly, miRNA-29b expression is deregulated in AML. Restoration of the function of suppressed miRNA-29b results in apoptosis of AML cell lines . In addition, deregulation of miRNA-29b causes overexpression of KIT, resulting in AML propagation .
In the current study, we investigate the expression of both miRNAs − miRNA-370 and miRNA-29b − in Egyptian de-novo AML and compare their expression with healthy controls.
| Patients and methods|| |
The study was performed on 30 peripheral mononuclear cells (PBMNCs) from de-novo AML patients admitted to the Haematology Unit at Main University Hospital Alexandria University, Egypt, from January 2013 to June 2013 (group 1), and 10 healthy volunteers matched for age and sex as controls (group 2). The study was approved by the local ethics committee. Total RNA including miRNA was extracted and purified from PBMNCs from de-novo AML patients using Qiagen miRNeasy Mini Kit (Qiagen, Catalogue Number 217004, Hilden, Germany) according to the manufacturer’s instructions. RNA was reverse-transcribed using the M-MuLV Reverse Transcription Kit (Fermentas, St. Leon-ROT, Germany) and the miScript reverse transcription Kit (Qiagen) for mRNA and miRNA, respectively. Real-time PCR was performed using Quantitect SYBR Green PCR reagents on a Rotor-Gene Q. All samples were normalized to the internal control (β-actin) and fold changes were calculated with the 2△△Ct method. All experiments were performed in accordance with approved guidelines and regulations.
RT-PCR primers were purchased from Qiagen Quantitect collection; β-actin (catalog number QT00095431) and the primers for miRNAs were also purchased from Qiagen: miR-29b (catalog number 4440886) and miR-370 (catalog number 4426961).
Statistical analysis was performed using the F-test (analysis of variance), Spearman’s r correlation, and the nonparametric tests unpaired χ2, Mann–Whitney U, and Kruskal–Wallis test. Results were considered significant if the P value was less than 0.05. Statistical analysis was carried out using the software package SPSS version 20.0 (SPSS Inc., Chicago, Illinois, USA).
| Results|| |
Descriptive data of AML patients and controls are shown in [Table 1]. There was significant difference in the platelet count between the patients and controls (P=0.035, [Table 1]). Study of the FAB classification among AML patients revealed 13 patients with M5 morphology (43%), nine patients with M4 morphology (30%), and the remaining patients were divided equally between M1 and M2 (13.3% each) ([Table 2]).
|Table 1 Comparison between the acute myeloid leukemia patients and controls according to different clinical parameters|
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|Table 2 Distribution of the studied patients according to FAB classification|
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Downregulation of both microRNA-370 and microRNA-29b expressions in peripheral mononuclear cells from de-novo acute myeloid leukemia patients
We analyzed miRNA-370 and miRNA-29b expressions in PBMNCs samples from 30 de-novo AML patients at the time of diagnosis using quantitative RT-PCR. As shown in [Figure 1] and [Figure 2], the miR-370 and miR-29 expressions in patients’ samples were significantly reduced (P=0.011 and 0.015, Mann–Whitney U-test, respectively) compared with that from healthy controls. Additionally, there was no association between the miR-370 or miR-29 expressions with the FAB subtypes (P=0.899 and 0.418, respectively).
|Figure 1 Differential microRNA-370 (miRNA-370) expression in acute myeloid leukemia (AML) compared with controls. MiRNA-370 expression was significantly lower in AML patients compared with controls (Mann–Whitney U-test, P=0.011).|
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|Figure 2 Differential microRNA-29-b (miRNA-29b) expression in acute myeloid leukemia (AML) compared with controls. MiRNA-29b expression was significantly lower in AML patients compared with controls (Mann–Whitney U-test, P=0.015).|
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MicroRNA-370 expression shows different correlation with clinical parameters versus microRNA-29b expression
Next, we investigated whether there was a significant correlation between the miRNA-370 and miRNA-29b expressions and different studied clinical laboratory parameters ([Table 3]). There was a significant positive correlation between the expression of both miRNA-370 and miRNA-29b in AML patients (Spearman’s r, P=0.003). In addition, there was a significant negative correlation between miRNA-370 expression and the age of the patients, whereas there was no such correlation regarding miRNA-29b (P=0.067). In addition, miRNA-370 expression was significantly negatively correlated with both serum alkaline phosphatase (P=0.013) and prothrombin time (P=0.004). We did not find similar correlations as regards miRNA-29b. However, miRNA-29b expression was significantly negatively correlated with aspartate aminotransferase (AST) liver enzyme (P=0.033) ([Table 3]).
|Table 3 Correlation of microRNA-370 and microRNA-29b expressions and different studied clinical laboratory parameters|
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| Discussion|| |
MiRNAs are considered effective regulators of gene expression, and thus they have important roles in different cellular processes. Dysregulation of miRNAs is usually accompanied with cancer, and even miRNA expression can discriminate cancer subtypes ,. The current study is the first report on investigating both miRNA-370 and miRNA-29b in Egyptian patients with de-novo AML. We observed a decreased expression of miR-370 in the PBMNCs derived from 30 AML patients compared with 10 healthy controls. In addition, miR-370 expression had no correlation with FAB AML subtypes as previously reported in a study on 48 newly diagnosed AML . In contrast, overexpression of miRNA-370 was stated in M5 AML patients . These contradicting results of the correlation between the miRNA-370 expression and FAB subtypes could not be related to the number of the patients, as studies with quite low number of patients − for example, 48 patients  and 27 patients  − or high number of patients − for example, 215  − did not show such correlation. However, some miRNAs (e.g. miRNA-370) can differentiate between two distinct AML subtypes (e.g. M1 and M5) . Yet, there is evidence of distinctive miRNA-370 expression in acute promyelocytic leukemia with t(15;17) . miRNA-370 is described to be downregulated in AML through promoter methylation, thus silencing the gene .
In the present study, miRNA-29b was significantly downregulated in de-novo AML. MiRNA-29b is part of a cluster of miRNAs localized on chromosome 7q32 in a region frequently deleted in therapy-related AML. As miRNA-29b was downregulated in many AML subtypes , miRNA-29b could act as a tumor suppressor miRNA in AML. Similar to miRNA-370, we did not find a correlation between miRNA-29b and the AML FAB subtypes. However, miRNA-29b expression is suggested to be correlated with AML cytogenetic changes [e.g. t(11q23)/Mixed lineage leukemia (MLL)] versus all other AML patients . The mechanism of downregulation of miR-29b is through activation of a KIT–MYC axis . In addition, miRNA-29b is involved in the regulation of DNA methylation by targeting DNA methyltransferases 3A (DNMT3A) and 3B (DNMT3B) . This miRNA-29b overexpression is repressed by histone deacetylases and can be restored by the application of histone deacetylase inhibitors, which might suggest a role of miR-29b as an epigenetic modulator .
Moreover, miRNA-370 expression was significantly correlated with miRNA-29b expression in AML patients, which supports our finding that both miRNAs may act as tumor suppressor genes. Additionally, there was a significant negative correlation between miR-370 expression and the age of the patients. Interestingly, Lin et al.  reported that serum miR-370 level was significantly low in pediatric AML patients.
In addition, miR-370 expression was significantly negatively correlated with both serum alkaline phosphatase (P=0.013) and prothrombin time (P=0.004), which might indicate hepatic involvement . Also, miRNA-29b expression was inversely correlated with AST. The association between miRNA-29b downregulation and elevated level of AST might be an indicator of liver fibrotic damage .
In conclusion, the present study demonstrated the possible roles of both miRNAs as tumor suppressors in de-novo AML patients. Remarkably, the negative correlation of miRNA-370 expression with age supports the previous assumption of miRNA-370 as possible diagnostic and prognostic markers in pediatric AML.
Financial support and sponsorship
Conflicts of interest
There are no conflicts of interest.
| References|| |
Estey E, Döhner H. Acute myeloid leukaemia. Lancet
Estey EH. Treatment of acute myeloid leukemia. Haematologica
Altuvia Y, Landgraf P, Lithwick G, Elefant N, Pfeffer S, Aravin A et al.
Clustering and conservation patterns of human microRNAs. Nucleic Acids Res
Calin GA, Croce CM. MicroRNA signatures in human cancers. Nat Rev Cancer
Starczynowski DT, Morin R, McPherson A, Lam J, Chari R, Wegrzyn J et al.
Genome-wide identification of human microRNAs located in leukemia-associated genomic alterations. Blood
Bandrés E, Cubedo E, Agirre X, Malumbres R, Zárate R, Ramirez N et al.
Identification by real-time PCR of 13 mature microRNAs differentially expressed in colorectal cancer and non-tumoral tissues. Mol Cancer
Meng F, Wehbe-Janek H, Henson R, Smith H, Patel T. Epigenetic regulation of microRNA-370 by interleukin-6 in malignant human cholangiocytes. Oncogene
Zhang X, Zeng J, Zhou M, Li B, Zhang Y, Huang T et al.
The tumor suppressive role of miRNA-370 by targeting FoxM1 in acute myeloid leukemia. Mol Cancer
Garzon R, Fabbri M, Liu S, Heaphy CE, Callegari E, Schwind S et al.
MicroRNA-29b induces global DNA hypomethylation and tumor suppressor gene reexpression in acute myeloid leukemia by targeting directly DNMT3A and 3B and indirectly DNMT1. Blood
Liu S, Wu L-C, Pang J, Santhanam R, Schwind S, Wu YZ et al.
Sp1/NFkappaB/ HDAC/miR-29b regulatory network in KIT-driven myeloid leukemia. Cancer Cell
Iliopoulos D, Drosatos K, Hiyama Y, Goldberg IJ, Zannis VI. MicroRNA-370 controls the expression of MicroRNA-122 and Cpt1α and affects lipid metabolism. J Lipid Res
Lutherborrow M, Bryant A, Jayaswal V, Agapiou D, Palma C, Yang YH, Ma DD. Expression profiling of cytogenetically normal AML identifies microRNAs that target genes involved in monocytic differentiation. Am J Hematol
Jongen-Lavrencic M, Sun SM, Dijkstra MK, Valk PJ, Lowenberg B. MicroRNA expression profiling in relation to the genetic heterogeneity of acute myeloid leukemia. Blood
Marcucci G, Mrózek K, Radmacher MD, Garzon R, Bloomfield CD. The prognostic and functional role of microRNAs in acute myeloid leukemia. Blood
Havelange V. MicroRNAs: new actors in acute myeloid leukaemia. P Belg Roy Acad Med
Zebisch A, Hatzl S, Pichler M, Wolfler A, Sill H. Therapeutic resistance in acute myeloid leukemia: the role of non-coding RNAs. Int J Mol Sci
Tu J, Ng SH, Luk AC, Liao J, Jiang X, Feng B et al.
MicroRNA-29b/Tet1 regulatory axis epigenetically modulates mesendoderm differentiation in mouse embryonic stem cells. Nucleic Acids Res
Lin X, Wang Z, Wang Y, Feng W. Serum microRNA-370 as a potential diagnostic and prognostic biomarker for pediatric acute myeloid leukemia. Int J Clin Exp Pathol
Munoz-Garrido P, Fernandez-Barrena MG, Hijona E, Carracedo M, Marín JJG, Bujanda L, Banales JM. MicroRNAs in biliary diseases. World J Gastroenterol
Zhang Y, Wu L, Wang Y, Zhang M, Li L, Zhu D et al.
Protective role of estrogen-induced miRNA-29b expression in carbon tetrachloride-induced mouse liver injury. J Biol Chem
[Figure 1], [Figure 2]
[Table 1], [Table 2], [Table 3]