Optimizing reporter constructs for in vivo bioluminescence imaging of interferon-γ stimulated mesenchymal stromal cells

Abstract

Mesenchymal stromal cells (MSCs) are a promising treatment modality for a variety of diseases. Strategies to investigate the fate of MSCs in vivo are important to unravel their therapeutic mechanisms. However, currently available techniques are hampered by their low sensitivity. We therefore aimed to optimize in vivo bioluminescence imaging of MSCs. We compared MSCs transduced with firefly luciferase (Fluc) and transmembrane-bound Gaussia luciferase driven by the human cytomegalovirus, spleen focus-forming virus (SFFV), and elongation factor 1-α (EF1α) promoters. Although cytomegalovirus-transmembrane-bound Gaussia luciferase-transduced MSCs showed the highest light intensity in vitro, the signal was almost undetectable in vivo. Spleen focus-forming virus-Fluc-transduced MSCs revealed a bright signal in vivo, but transgene expression was silenced upon in vitro stimulation with interferon (IFN)-γ. Therefore, the SFFV promoter was replaced by the EF1α promoter. Light emission of Fluc under the control of EF1α was similar to SFFV-Fluc. Although EF1α-Fluc light emission was decreased tenfold in the presence of IFN-γ when compared with unstimulated MSCs, the bioluminescent signal could still be detected and was clearly distinguishable from untransduced MSCs. Furthermore, stimulation of MSCs with tumor necrosis factor-α hardly affected transgene expression in EF1α-Fluc-transduced MSCs. Thus, the use of the EF1α promoter partially overcomes silencing and allows in vivo bioluminescence imaging of IFN-γ-stimulated MSCs.