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:: Volume 17, Issue 2 (Iranian South Medical journal 2014) ::
Iran South Med J 2014, 17(2): 130-140 Back to browse issues page
Expression of the human coagulation factor IX in the bone marrow mesenchymal stem cells
Azadehsadat Azadbakhsh1, Mohammad Reza Sam2, Farah Farrokhi3, Alireza Zomorodipour4, Ali Akbar Haddad Mashahrizeh5, Aram Mokarizadeh6
1- Department of Biology, School of Science, Urmia University, Urmia, IRAN
Department of Cellular and Molecular Biotechnology, Institute of Biotechnology, Urmia University, Urmia, IRAN
2- Department of Biology, School of Science, Urmia University, Urmia, IRAN
Department of Cellular and Molecular Biotechnology, Institute of Biotechnology, Urmia University, Urmia, IRAN , m.sam@urmia.ac.ir
3- Department of Biology, School of Science, Urmia University, Urmia, IRAN
4- Department of Molecular Genetics, National Institute of Genetic Engineering and Biotechnology, Tehran, IRAN
5- Department of Biology, School of Science, Ferdowsi University of Mashad, Mashad, IRAN
6- Department of Immunology, School of Veterinary Medicine, Urmia University, Urmia, IRAN
Abstract:   (6886 Views)

Background: Mesenchymal stem cells (MSCs) are appropriate target for gene and cell-based therapy of hemophilia B patients. MSCs possess several unique properties such as capability of differentiating into multiple lineages and lower immunogenecity in transplant procedure that make them attractive candidates for cell and gene therapy. One of the challenges in the gene therapy is the low expression level of transgene. To improve expression, strong regulatory elements in the context of vectors could contribute to improve efficacy of gene therapy strategies. In this study four human factor IX (hFIX)-expressing plasmids equipped with various combination of human -globin (hBG) introns and Kozak sequence were transfected into the MSCs and expression of the hFIX was evaluated in vitro. Material and Methods: MSCs were obtained from tibias and the femora of rats and phenotypic characterization of the MSCs was determined by flow cytometry. Four hFIX-expressing plasmids were introduced into the culture-expanded MSCs using transfection agent. 48 hours after transfection, ability of the MSCs for expression of the hFIX and efficacies of the plasmids were evaluated by performing sandwich ELISA on cultured media as well as semi-quantitative RT-PCR. All analyses were performed with One-way ANOVA using SPSS software. Results:The highest expression level of the hFIX was obtained from intron-less and hBG intron-I containing construct. The highest biological activity was obtained from hBG intron-I,II containing construct. Conclusion:Successful expression of the hFIX was obtained from recombinant MSCs. MSCs were able to splice heterologous hBG intron-I from the hFIX-cDNA. Application of thehBG introns reduced the hFIX expression levels, probably due to improper splicing of the hBG introns.

Keywords: mesenchymal stem cells, hemophilia B, human factor IX, plasmid, human β-globin introns
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Type of Study: Original | Subject: General
Received: 2012/12/20 | Accepted: 2013/02/4 | Published: 2014/04/14
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Azadbakhsh A, Sam M R, Farrokhi F, Zomorodipour A, Haddad Mashahrizeh A A, Mokarizadeh A. Expression of the human coagulation factor IX in the bone marrow mesenchymal stem cells. Iran South Med J. 2014; 17 (2) :130-140
URL: http://ismj.bpums.ac.ir/article-1-519-en.html


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Volume 17, Issue 2 (Iranian South Medical journal 2014) Back to browse issues page
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