Open AccessPathway‐specific reporter genes to study stem cell biology
Author(s) -
Peterson Karen M.,
Franchi Federico,
Olthoff Michaela,
Chen Ian Y.,
Paulmurugan Ramasamy,
RodriguezPorcel Martin
Publication year - 2020
Publication title -
stem cells
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 2.159
H-Index - 229
eISSN - 1549-4918
pISSN - 1066-5099
DOI - 10.1002/stem.3167
Subject(s) - reporter gene , biology , mesenchymal stem cell , phenotype , microbiology and biotechnology , transfection , luciferase , stem cell , gene , gene expression , genetics
Little is known on the phenotypic characteristics of stem cells (SCs) after they are transplanted to the myocardium, in part due to lack of noninvasive platforms to study SCs directly in the living subject. Reporter gene imaging has played a valuable role in the noninvasive assessment of cell fate in vivo. In this study, we validated a pathway‐specific reporter gene that can be used to noninvasively image the phenotype of SCs transplanted to the myocardium. Rat mesenchymal SCs (MSCs) were studied for phenotypic evidence of myogenic characteristics under in vitro conditions. After markers of myogenic characteristics were identified, we constructed a reporter gene sensor, comprising the firefly luciferase ( Fluc ) reporter gene driven by the troponin T ( TnT ) promoter (cardio MSCs had threefold expression in polymerase chain reaction compared to control MSCs) using a two‐step signal amplification strategy. MSCs transfected with TnT‐Fluc were studied and validated under in vitro conditions, showing a strong signal after MSCs acquired myogenic characteristics. Lastly, we observed that cardio MSCs had higher expression of the reporter sensor compared to control cells (0.005 ± 0.0005 vs 0.0025 ± 0.0008 Tnt‐Fluc/ubiquitin‐Fluc, P < .05), and that this novel sensor can detect the change in the phenotype of MSCs directly in the living subject. Pathway‐specific reporter gene imaging allows assessment of changes in the phenotype of MSCs after delivery to the ischemic myocardium, providing important information on the phenotype of these cells. Imaging sensors like the one described here are critical to better understanding of the changes that SCs undergo after transplantation.