Primary evaluation of induced pluripotent stem cells using flow cytometry

Abstract

Induced pluripotent stem (iPS) cells are an attractive cell source in regenerative medicine; however, some problems must be overcome to improve clinical applications. iPS cells generated using the genomic integration method increase the risk of tumor generation because of transgene reactivation and the disruption of endogenous genes. The somatic cell sources of iPS cells also affect teratoma formation. Therefore, it is important to select a suitable cell source from among adult somatic cells, to generate iPS cells using a transgene insertion-free method, and to screen for good iPS clones that do not contain any differentiation-resistant cells after differentiation induction. Recently, we reported a method for obtaining high-quality iPS cells using purified mesenchymal stem cells (MSCs). In this report, we produced genomic integration-free iPS cells from adult tissues, purified MSCs, and tail tip fibroblasts using the Sendai virus. Then, we evaluated the residual undifferentiated cells in secondary neurospheres generated from retroviral induction iPS cell lines and non-integration iPS cell lines derived from adult MSCs. As a result, we could generate integration-free iPS cells only from MSCs. Nevertheless, some iPS cell lines generated by the non-integration method contained undifferentiated cells. Interestingly, the integration-free iPS cells that could not differentiate correctly showed a higher side scatter (SSC) intensity than the other ES/iPS cells. Some somatic cell-derived iPS cells had a higher SSC intensity, and these cells also could not differentiate normally. Our findings suggested that an SSC intensity analysis may be efficient method for evaluating individual iPS cells before their use in therapies.