Non-Viral Gene Transfer Into Murine Adipose Derived Mesenchymal Stem Cells: A Comparative Study of Different Non-Viral Methods



Bahrambeigi V^{1, 2, 3} ; Ahmadi N² ; Ghafarizadeh AA³ ; Lotfi M⁴ ; Hashemibeni B⁵ ; Javanmard SH^{2, 6} Show Affiliations
Source: Gene Therapy and Molecular Biology Published:2013

Abstract

Murine mesenchymal stem cells (MSCs) are common tools in gene therapy experiments. In this study, we measured, optimized, and compared transfection efficiency of different non-viral methods into murine adipose derived MSCs (AD-MSCs). AD-MSCs were isolated from fat tissue of C57BL6 mice. Authenticity and surface markers expression of murine AD-MSCs were assessed. Transfection yields of different transfectants along with nucleofection in different conditions or programs were determined and compared. The isolated AD-MSCs were successfully differentiated into three lineages. AD-MSCs were positive for CD73, CD90, CD105, negative for CD34 and CD45 and lowly expressed CD24. The best transfection rates were 18.14 ± 3.41% (ExpressIn), 24.64 ± 4.37% (GenCarrier-2), 11.43 ± 3.17% (GeneTranTMII), 26.4 ± 4.17% (LipofectaminTM 2000), 11.51 ± 1.97% (MesenFectagen), 29.49 ± 5.31% (TransIT®-2020), and 73.62 ± 6.81 (nucleofection; with ~41% of toxicity). After selecting an appropriate method, we used PhiC31 (φC31) integrase system to overcome the problem of transient gene expression. Although, we reached only moderate transfection rates using transfection reagents, nucleofection was the most efficient method for gene transfer into murine AD-MSCs. Results also indicated that stably transfected AD-MSCs can be generated after φC31 integrase-mediated transgene integration of nucleofected AD-MSCs. Our results show non-viral gene delivery can be used for efficient, safe, and stable modification of MSCs.