Generation of Novel Reporter Stem Cells and Their Application for Molecular Imaging of Cardiac-Differentiated Stem Cells In Vivo

Abstract

Stem cell therapies offer the potential for repair and regeneration of cardiac tissue. To facilitate evaluation of stem cell activity in vivo, we created novel dual-reporter mouse embryonic stem (mES) cell lines that express the firefly luciferase (LUC) reporter gene under the control of the cardiac sodium-calcium exchanger-1 (Ncx-1) promoter in the background of the 7AC5-EYFP mES cell line that constitutively expresses the enhanced yellow fluorescent protein (EYFP). We compared the ability of recombinant clonal cell lines to express LUC before and after induction of cardiac differentiation in vitro. In particular, one of the clonal cell lines (Ncx-1-43LUC mES cells) showed markedly enhanced LUC expression (45-fold increase) upon induction of cardiac differentiation in vitro. Further, cardiac differentiation in these cells was perpetuated over a period of 2-4 weeks after transplantation in a neonatal mouse heart model, as monitored by noninvasive bioluminescence imaging (BLI) and confirmed via postmortem immunofluorescence and histological assessments. In contrast, transplantation of undifferentiated pluripotent Ncx-1-43LUC mES cells in neonatal hearts did not result in detectable levels of cardiac differentiation in these cells in vivo. These results suggest that prior induction of cardiac differentiation in vitro enhances development and maintenance of a cardiomyocyte-like phenotype for mES cells following transplantation into neonatal mouse hearts in vivo. We conclude that the Ncx-1-43LUC mES cell line is a novel tool for monitoring early cardiac differentiation in vivo using noninvasive BLI.

In vivo bioluminescence for tracking cell fate and function

“…This provides exciting opportunities for transcriptional targeting, double reporter labeling for BLI monitoring, as well as gene therapy. For example, by linking the expression of luciferase to the cardiac-specific promoter myosin light chain 2v, it is possible to monitor cells undergoing cardiac differentiation over time via the detection of reporter gene expression by BLI (27,35).…”

Current standing and future prospects for the technologies proposed to transform toxicity testing in the 21st century

“…In vivo imaging has been used to study disease progress such as infection (Luker and Luker, 2010) and cancer (Lee et al, 2010). Moreover, bioluminescence imaging was used to study the viability, proliferation, migration, and differentiation of stems cells in vitro (Kammili et al, 2010;Logeart-Avramoglou et al, 2010) and after transplantation in vivo (Wilson et al, 2008). The applications for toxicity testing have been limited so far.…”

Molecular imaging for In vivo tracking of stem cell fate

“…[1][2][3][4][5][6] Stem cells isolated from various sources have been tested in early-phase clinical trials in humans, and many are now being examined in longterm clinical trials. 7,8 Indeed, a few adult stem cell therapies are already in clinical use.…”