Noninvasive Monitoring of Oxidative Stress in Transplanted Mesenchymal Stromal Cells

• Published in: JACC. Cardiovascular imaging Vol. 6; no. 7; pp. 795 - 802
• Main Authors: Psaltis, Peter J., Peterson, Karen M., Xu, Rende, Franchi, Federico, Witt, Tyra, Chen, Ian Y., Lerman, Amir, Simari, Robert D., Gambhir, Sanjiv S., Rodriguez-Porcel, Martin
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 01.07.2013
• Subjects: Animals; Antioxidants - pharmacology; bioluminescence; Biosensing Techniques; Cardiovascular; Cell Survival; Cell Tracking - methods; Cells, Cultured; Cyclic N-Oxides - pharmacology; Cytomegalovirus - genetics; Disease Models, Animal; Genes, Reporter; Luciferases, Firefly - biosynthesis; Luciferases, Firefly - genetics; Luciferases, Renilla - biosynthesis; Luciferases, Renilla - genetics; Luminescent Measurements; Mesenchymal Stem Cell Transplantation; mesenchymal stem cells; Mesenchymal Stem Cells - metabolism; Molecular Imaging; Myocardial Reperfusion Injury - genetics; Myocardial Reperfusion Injury - metabolism; Myocardial Reperfusion Injury - surgery; NAD(P)H oxidase; oxidative stress; Oxidative Stress - drug effects; Phenotype; Phosphoproteins - genetics; Promoter Regions, Genetic; Rats; reporter gene; Spin Labels; Time Factors; Transfection; IR; LV; TSTA; CMV; NAD(P)H oxidase; bioluminescence; Fluc; mesenchymal stem cells; FBS; ROS; NAD(P)H; reporter gene; MSC; oxidative stress; BLI; PET; bioluminescence imaging; ischemia/reperfusion; cytomegalovirus; reactive oxygen species; left ventricular; nicotinamide adenine dinucleotide phosphate; firefly luciferase; fetal bovine serum; mesenchymal stromal cell; positron emission tomography; 2-step transcriptional amplification
• ISSN: 1936-878X; 1876-7591; 1876-7591
• DOI: 10.1016/j.jcmg.2012.11.018

The goal of this study was to validate a pathway-specific reporter gene that could be used to noninvasively image the oxidative status of progenitor cells. In cell therapy studies, reporter gene imaging plays a valuable role in the assessment of cell fate in living subjects. After myocardial injury, noxious stimuli in the host tissue confer oxidative stress to transplanted cells that may influence their survival and reparative function. Rat mesenchymal stromal cells (MSCs) were studied for phenotypic evidence of increased oxidative stress under in vitro stress. On the basis of their up-regulation of the pro-oxidant enzyme p67phox subunit of nicotinamide adenine dinucleotide phosphate (NAD[P]H oxidase p67phox), an oxidative stress sensor was constructed, comprising the firefly luciferase (Fluc) reporter gene driven by the NAD(P)H p67phox promoter. MSCs cotransfected with NAD(P)H p67phox–Fluc and a cell viability reporter gene (cytomegalovirus–Renilla luciferase) were studied under in vitro and in vivo pro-oxidant conditions. After in vitro validation of the sensor during low-serum culture, transfected MSCs were transplanted into a rat model of myocardial ischemia/reperfusion (IR) and monitored by using bioluminescence imaging. Compared with sham controls (no IR), cardiac Fluc intensity was significantly higher in IR rats (3.5-fold at 6 h, 2.6-fold at 24 h, 5.4-fold at 48 h; p < 0.01), indicating increased cellular oxidative stress. This finding was corroborated by ex vivo luminometry after correcting for Renilla luciferase activity as a measure of viable MSC number (Fluc:Renilla luciferase ratio 0.011 ± 0.003 for sham vs. 0.026 ± 0.004 for IR at 48 h; p < 0.05). Furthermore, in IR animals that received MSCs preconditioned with an antioxidant agent (tempol), Fluc signal was strongly attenuated, substantiating the specificity of the oxidative stress sensor. Pathway-specific reporter gene imaging allows assessment of changes in the oxidative status of MSCs after delivery to ischemic myocardium, providing a template to monitor key biological interactions between transplanted cells and their host environment in living subjects.