Background: Recent studies have demonstrated that adipose-derived mesenchymal stem cells (MSCs) can attenuate ischemic tissue injury, possibly by their paracrine effects or differentiation into local cell types. However, homing of transplanted cells either transfused systematically or via direct intra-lesional injection, is reported to be very low and transient, leading to reduced therapeutic effects. CXCR4 is identified as a crucial factor for homing and engraftment of stem cells. In this study, we overexpressed CXCR4 in adipose-derived MSCs to enhance their level of engraftment and perhaps improve their translational effectiveness.
Materials and Methods: Wild type and WHIM-type variants of CXCR4 gene were chemically synthesized and cloned in pCDH-513b lentiviral vector. Viral particles were generated and their concentrations were determined according to Trono lab protocol after transfection of HEK293 cells. Adipose-derived mesenchymal stem cells in passage 3 and 4 were transduced with both variants of CXCR4. 72 h post transduction, cells were selected with puromycin for 3 days. Migration ability of transduced cells was then assessed by transwell chamber assay.
Results: Molecular cloning of the synthetic gene was confirmed by DNA sequencing. The presence of GFP positive MSCs confirmed the high efficiency of transduction. Our qPCR results showed overexpression of CXCR4 at mRNA level in transduced MSCs. Genetically modified MSCs (wild and WHIM types) had significantly enhanced migration capabilities in comparison with control group (p<0.05).
Conclusion: Overexpression of mutant CXCR4 resulted in improved homing of MSCs, which is very important in better success in cell therapy.