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Cell J. 2017 Oct;19(3):361-374. doi: 10.22074/cellj.2017.4497. Epub 2017 Aug 19.

Multiple Sclerosis Gene Therapy with Recombinant Viral Vectors: Overexpression of IL-4, Leukemia Inhibitory Factor, and IL-10 in Wharton's Jelly Stem Cells Used in EAE Mice Model.

Author information

1
Cellular and Molecular Biology Research Center, Shahid Beheshti University of Medical Sciences, Tehran, Iran.
2
Department of Cell Biology and Anatomical Science, Faculty of Medicine, Shahid Beheshti University of Medical Sciences, Tehran, Iran. Electronic address: Prof_hosseini@yahoo.com.
3
Department of Molecular Medicine, School of Advanced Technologies in Medicine, Tehran University of Medical Sciences, Tehran, Iran.
4
Iranian Institute of Cell and Gene Therapy, Tehran, Iran.
5
Cellular and Molecular Biology Research Center, Babol University of Medical Sciences, Babol, Iran.
6
Department of Genetics, Faculty of Medicine, Babol University of Medical Sciences, Babol, Iran.
7
Cellular and Molecular Research Center, Shahrekord University of Medical Sciences, Shahrekord, Iran.
8
Urogenital Stem Cell Research, Shahid Beheshti University of Medical Sciences, Tehran, Iran.
9
Department of Biotechnology, Fasa University of Medical Sciences, Fasa, Iran.
10
BioViva USA Inc, Bainbridge Island WA, USA.

Abstract

OBJECTIVES:

Immunotherapy and gene therapy play important roles in modern medicine. The aim of this study is to evaluate the overexpression of interleukin-4 (IL-4), IL-10 and leukemia inhibitory factor (LIF) in Wharton's jelly stem cells (WJSCs) in the experimental autoimmune encephalomyelitis (EAE) mice model.

MATERIALS AND METHODS:

In this experimental study, a DNA construction containing IL- 4, IL-10 and LIF was assembled to make a polycistronic vector (as the transfer vector). Transfer and control vectors were co-transfected into Human Embryonic Kidney 293 (HEK-293T) cells with helper plasmids which produced recombinant lentiviral viruses (rLV). WJSCs were transduced with rLV to make recombinant WJSC (rWJSC). In vitro protein and mRNA overexpression of IL-4, LIF, and IL-10 were evaluated using quantitative polymerase chain reaction (qPCR), enzyme-linked immunosorbent assay (ELISA) and western blot (WB) analysis. EAE was induced in mice by MOG-CFA and pertussis toxin. EAE mice were injected twice with 2×105 rWJSCs. The in vivo level of IL-4, LIF, IL-10 cytokines and IL-17 were measured by ELISA. Brain tissues were analyzed histologically for evaluation of EAE lesions.

RESULTS:

Isolated WJSCs were performed to characterize by in vitro differentiation and surface markers were analyzed by flow cytometry method. Cloning of a single lentiviral vector with five genes was done successfully. Transfection of transfer and control vectors were processed based on CaPO4 method with >90% efficiency. Recombinant viruses were produced and results of titration showed 2-3×107 infection-unit/ml. WJSCs were transduced using recombinant viruses. IL-4, IL-10 and LIF overexpression were confirmed by ELISA, WB and qPCR. The EAE mice treated with rWJSC showed reduction of Il-17, and brain lesions as well as brain cellular infiltration, in vivo. Weights and physical activity were improved in gene-treated group.

CONCLUSIONS:

These results showed that gene therapy using anti-inflammatory cytokines can be a promising approach against multiple sclerosis (MS). In addition, considering the immunomodulatory potential of WJSCs, an approach using a combination of WJSCs and gene therapy will enhance the treatment efficacy.

KEYWORDS:

Cytokines; Gene Therapy; Multiple Sclerosis; Wharton’s jelly stem cells

Conflict of interest statement

There is no conflict of interest in this study.

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