The Effect of Mir-451 Upregulation on Erythroid Lineage Differentiation of Murine Embryonic Stem Cells


Narges Obeidi, Ph.D, 1,2Ali Akbar Pourfathollah, Ph.D, 3Masoud Soleimani, Ph.D, 4Mahin Nikougoftar Zarif, Ph.D, 1,*Fatemeh Kouhkan, Ph.D, 5
Blood Transfusion Research Center, High Institute for Education and Research in Transfusion Medicine, Tehran, Iran
Department of Hematology, School of Para Medicine, Bushehr University of Medical Sciences, Bushehr, Iran
Department of Immunology, School of Medicine, Tarbiat Modares University, Tehran, Iran
Department of Hematology, School of Medicine, Tarbiat Modares University, Tehran, Iran
Stem Cell Technology Research Center, Tehran, Iran
Blood Transfusion Research Center, High Institute for Education and Research in Transfusion Medicine, Tehran, Iran
Department of Hematology, School of Para Medicine, Bushehr University of Medical Sciences, Bushehr, Iran
Department of Immunology, School of Medicine, Tarbiat Modares University, Tehran, Iran
Department of Hematology, School of Medicine, Tarbiat Modares University, Tehran, Iran
Stem Cell Technology Research Center, Tehran, Iran
*Corresponding Address: P.O.Box: 146651157 Blood Transfusion Research Center High Institute for Education and Research in Transfusion Medicine Tehran Iran Email:nikougoftar@ibto.ir
Any use, distribution, reproduction or abstract of this publication in any medium, with the exception of commercial purposes, is permitted provided the original work is properly cited This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
Obeidi Narges, Pourfathollah Ali Akbar, Soleimani Masoud, Nikougoftar Zarif Mahin, Kouhkan Fatemeh. The Effect of Mir-451 Upregulation on Erythroid Lineage Differentiation of Murine Embryonic Stem Cells. Cell J. 2016; 18(2): 165-178.

Abstract

Objective

MicroRNAs (miRNAs) are small endogenous non-coding regulatory RNAs that control mRNAs post-transcriptionally. Several mouse stem cells miRNAs are cloned differentially regulated in different hematopoietic lineages, suggesting their possible role in hematopoietic lineage differentiation. Recent studies have shown that specific miRNAs such as Mir-451 have key roles in erythropoiesis.

Materials and Methods

In this experimental study, murine embryonic stem cells (mESCs) were infected with lentiviruses containing pCDH-Mir-451. Erythroid differentiation was assessed based on the expression level of transcriptional factors (Gata-1, Klf-1, Epor) and hemoglobin chains (α, β, γ , ε and ζ) genes using quantitative reverse transcriptase-polymerase chain reaction (qRT-PCR) and presence of erythroid surface antigens (TER-119 and CD235a) using flow cytometery. Colony-forming unit (CFU) assay was also on days 14thand 21thafter transduction.

Results

Mature Mir-451 expression level increased by 3.434-fold relative to the untreated mESCs on day 4 after transduction (P<0.001). Mir-451 up-regulation correlated with the induction of transcriptional factor (Gata-1, Klf-1, Epor) and hemoglobin chain (α, β, γ, ε and ζ) genes in mESCs (P<0.001) and also showed a strong correlation with presence of CD235a and Ter- 119 markers in these cells (13.084and 13.327-fold increse, respectively) (P<0.05). Moreover, mESCs treated with pCDH-Mir-451 showed a significant raise in CFU-erythroid (CFU-E) colonies (5.2-fold) compared with untreated control group (P<0.05).

Conclusion

Our results showed that Mir-451 up-regulation strongly induces erythroid differentiation and maturation of mESCs. Overexpression of Mir-451 may have the potential to produce artificial red blood cells (RBCs) without the presence of any stimulatory cytokines.