ABSTRACT
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 2×10[5] 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 2-3×10[7] 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