Abstract 4937: A Novel Molecular Imaging Sensor of Cellular Oxidative Stress
Increased oxidative stress has been suggested to play an important role in the biology of mesenchymal stem cells (MSCs). However, little is known of the effect of increased oxidative stress in-vivo, partly due to the lack of appropriate imaging technologies for monitoring of stem cell biology in the living subject. Recent developments in molecular imaging permit, for the first time, the longitudinal monitoring of stem cell biology in vivo. We hypothesize that changes in oxidative stress can be monitored using a reporter gene strategy (molecular imaging).
Methods: We have developed a novel oxidative stress sensor, using a two-step transcriptional amplification strategy to enhance the expression of firefly luciferase (Fluc), driven by the pro-oxidant enzyme NAD(P)H promoter, and used to monitor oxidative stress in rMSCs. rMSCs was induced by hypoxia (1% O2 for 24 hours) and serum deprivation (SD) and confirmed using a dichlorofluorescein colorimetric assay (measures the production of hydrogen peroxide-H2O2-) and NAD(P)H protein expression.
Results: Both serum deprivation and hypoxia led to a pro-oxidant state (Fig. A–B⇓). Most importantly, increased oxidative stress led to increased activity of the novel reporter gene sensor (Fig. C⇓) which was decreased in the presence of antioxidants (superoxide dismuatse mimetic Tempol, 5mM, SD+T, Fig D⇓). Results were reproduced in a different cell type (rat cardiomyoblast, data not shown).
Conclusion: Changes in oxidative stress in MSCs can be monitored using molecular imaging. Reporter Gene-based Molecular Imaging, a strategy that is clinically applicable, provides the opportunity to monitor stem cell survival in the living subject.