Multiple Sclerosis Gene Therapy Using Recombinant Viral Vectors: Overexpression of IL-4, IL-10 and Leukemia Inhibitory Factor in Wharton’s Jelly Stem Cells in The EAE Mice Model
Objective: 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 (EA...
| Main Authors: | , , , , , , , |
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| Format: | Article |
| Language: | English |
| Published: |
Royan Institute (ACECR), Tehran
2017-08-01
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| Series: | Cell Journal |
| Subjects: | |
| Online Access: | http://celljournal.org/journal/article/14336/download |
| Summary: | Objective: 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.
Conclusion: 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. |
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| ISSN: | 2228-5806 2228-5814 |