br Results br Discussion Pluripotent stem cells

Results
Discussion Pluripotent stem csf-1r offer unprecedented potential for regenerative medicine as they can be differentiated by a specific sequence of signals to become any cell type of the body. At the same time, any cell product generated from them confers a substantial risk for neoplastic growth or teratoma formation (Cunningham et al., 2012). This risk may come from contamination with undifferentiated cells, culture-acquired mutations, or de-differentiation of cells following transplantation (Baker et al., 2007; Lee et al., 2013; Mayshar et al., 2010). While several studies have been undertaken to determine the required number of pluripotent cells in a cell preparation needed to cause teratoma formation (Cao et al., 2007; Lee et al., 2009; Nussbaum et al., 2007), little is known about the sensitivity of potential imaging or serum biomarkers for teratoma detection following cell transplantation. First-generation iPSC lines were made using lentivirus-based reprogramming due to their efficiency (Takahashi and Yamanaka, 2006). However, the risk for random integration of delivered genes is well known and led to the eventual development of non-integrative reprogramming methods (Aoi et al., 2008; Fusaki et al., 2009). Here we generated a lentiviral-based iPSC line (line T) that continued to express reprogramming factors after differentiation. We reasoned that line T would be an appropriate model for a cell product contaminated with pluripotent cells or for cells with culture-acquired oncogenic transformations. The transcription factors used to induce pluripotency are master regulators of cell states and are accordingly strictly regulated and normally silenced during development (Jaenisch and Young, 2008). Standard safety screening of any cell product derived from pluripotent cells will include expression analysis of pluripotency genes; such a screen did easily detect the presence of pluripotent cells in our T-CMs. However, if only a small population expresses these genes or if expression is transiently repressed, detection will become much more difficult. While the risk of aberrant expression of reprogramming factors will be much smaller for iPSC lines generated by non-integrative methods, the risk for contamination of a cell product with undifferentiated cells or de-differentiation will remain. Current in vitro screening methods can detect down to 1 undifferentiated cell in 100,000 differentiated cells (Tano et al., 2014). Given that regenerative cell therapies for MI may administer up to one billion cells (Chong et al., 2014) and preclinical studies indicate that 1 × 104 to 1 × 105 cells are sufficient to induce teratoma growth (Lee et al., 2009), current procedures for cell purification and characterization cannot ensure that the cell product is free of undifferentiated cells. With limited data on ESC and iPSC-based cell therapies, the clinical risk for teratoma formation is currently unknown. However, the risk for tumor growth from stem cells has been highlighted previously by a patient who developed a brain tumor after a controversial non-US Food and Drug Administration (FDA) approved neuronal stem cell transplantation (Amariglio et al., 2009). Clinical trials designed to assess the efficacy and safety profile of cardiomyocytes derived from pluripotent cell sources are likely to involve imaging modalities such as MRI or ultrasound to assess cardiac function (Menasche et al., 2015). We therefore decided to assess the suitability of these modalities for safety screening. We were able to detect neoplastic growth (teratoma) reliably using T2w imaging and LGE once masses had reached a volume of 8 mm3. Although this sensitivity will decrease with the lower resolution of clinical MRI systems, it is likely to be much lower than average teratoma sizes at the time of detection currently reported (Chang and Lin, 2014; Coleman et al., 2014). Bland-Altman plots showed no systematic offset for teratoma volume estimates based on T2w imaging or LGE. LGE might be particularly convenient for potential teratoma or neoplastic growth detection since it is frequently included in clinical studies to assess changes in scar size (infarct size). Teratomas were poorly vascularized and surrounded by a hypointense rim on T2w and T2∗w images. This easy detectable feature was caused by hemorrhage around the teratoma leading to hemosiderin accumulation, which we confirmed histologically. Although short T2 values have been described previously for human ovarian teratomas, these were primarily due to the formation of bone or fat tissues in mature teratomas (Saba et al., 2009).