Bioluminescent Imaging of Genetically Selected Induced Pluripotent Stem Cell-Derived Cardiomyocytes after Transplantation into Infarcted Heart of Syngeneic Recipients

Abstract

Cell loss after transplantation is a major limitation for cell replacement approaches in regenerative medicine. To assess the survival kinetics of induced pluripotent stem cell (iPSC)-derived cardiomyocytes (CM) we generated transgenic murine iPSC lines which, in addition to CM-specific expression of puromycin N-acetyl-transferase and enhanced green fluorescent protein (EGFP), also constitutively express firefly luciferase (FLuc) for bioluminescence (BL) in vivo imaging. While undifferentiated iPSC lines generated by random integration of the transgene into the genome retained stable FLuc activity over many passages, the BL signal intensity was strongly decreased in purified iPS-CM compared to undifferentiated iPSC. Targeted integration of FLuc-expression cassette into the ROSA26 genomic locus using zinc finger nuclease (ZFN) technology strongly reduced transgene silencing in iPS-CM, leading to a several-fold higher BL compared to iPS-CM expressing FLuc from random genomic loci. To investigate the survival kinetics of iPS-CM in vivo, purified CM obtained from iPSC lines expressing FLuc from a random or the ROSA26 locus were transplanted into cryoinfarcted hearts of syngeneic mice. Engraftment of viable cells was monitored by BL imaging over 4 weeks. Transplanted iPS-CM were poorly retained in the myocardium independently of the cell line used. However, up to 8% of cells survived for 28 days at the site of injection, which was confirmed by immunohistological detection of EGFP-positive iPS-CM in the host tissue. Transplantation of iPS-CM did not affect the scar formation or capillary density in the periinfarct region of host myocardium. This report is the first to determine the survival kinetics of drug-selected iPS-CM in the infarcted heart using BL imaging and demonstrates that transgene silencing in the course of iPSC differentiation can be greatly reduced by employing genome editing technology. FLuc-expressing iPS-CM generated in this study will enable further studies to reduce their loss, increase long-term survival and functional integration upon transplantation.

Uterine cells—an immunoprivileged cell source for therapy—but are they for everyone?

“…The newly formed myocytes, however, showed incomplete maturation and arrhythmic complications were observed. Our group and others have shown beneficial effects of iPS-derived cardiomyocytes in rodent models [17,38,39] with retention of the injected cells in the host myocardium, improved cardiac function, and decreased cardiac fibrosis. However, the generation of patient-specific iPS cells and their differentiation is long-winded and expensive.…”

Stem Cells in Treatment of Coronary Heart Disease and Its Monitoring: Tissue Engineering and Clinical Evaluation

“…These pluripotent cells form regenerative myocardial tissue, smooth muscle, or endothelial vascular cells in situ to repair myocardial infarction in heart or increased ventricular wall thickness and electrical stability [29]. Recently, different clinical centers claim their success differently to transplant pluripotent stem cells in remodeling myocardial muscle or endothelial vascular cells [10,[28][29][30][31][32][33][34][35]. In fact, stem cell treatment centers follow the strategy that pluripotent stem cells may be stable rather than terminally differentiated as meta-analysis of randomized controlled clinical trials on stem cell therapy also indicated clearly that intracoronary adult bone marrow stem cells improve left ventricular function and reduce the risk of recurrent heart failure soon after acute myocardial infarction (AMI) [36].…”

“…The light-emitting capability of BLI illustrates the insights of cardiac regeneration [80]. Recently, survival kinetics of induced pluripotent stem cell and engraftment of viable cells was monitored by BLI imaging by visualizing the retention of bioluminescent agents in adult stem cells as shown in Figure 6 to monitor stem cells [34]. Efforts are still continuing for regenerating the heart and using myocardial stem cells in cardiovascular system in treatment of heart disease or remodeling [81].…”

Tracking and Quantification of Magnetically Labeled Stem Cells Using Magnetic Resonance Imaging

“…[54] In addition, the survival of therapeutic stem cells has been investigated with a bioluminescence approach after local transplantation for tissues such as bone, heart, and muscle. [55] However, it is not quantitative and a large number of cells are required for detection in deep tissues. Radionuclear methods, including single-photon emission computed tomography (SPECT) and positron emission tomography (PET), are extremely sensitive and can detect pico-molar (or even lower) concentrations of radiolabels.…”