Figure 1.

Induced pluripotent stem cells (iPSCs) are rejected by CD4⁺ T cells. (a) To determine whether iPSCs are rejected in syngeneic mice, luciferase‐expressing 129x1/SvJ iPS or embryonic stem cells (ESCs) were injected into 129x1/SvJ mice, n = 6. Mice were imaged regularly to determine the engraftment of the cells. iPSCs could not be detected after 14 days. (b) ESCs () were not rejected in syngeneic mice over the 40 days of observation. In contrast iPSCs () were rejected after a mean of 12 days. Furthermore, mice challenged for a second time with iPSCs () rejected those iPSCs within 5–6 days. For statistical analysis, the Log rank test was used. *P < 0·05, **P < 0·01. (c) To prove that both iPSCs and ESCs were pluripotent, the teratoma assay was performed in NOD‐SCID mice. In both cases, large teratomas developed. This is a representative result for the 129SvJ cells. (d) To determine the mechanism of iPSC rejection, splenocytes of mice that had rejected iPSCs were collected and CD4⁺ and CD8⁺ cells were sorted. The cells were exposed to iPS‐embryoid body (EB) cells in a proliferation assay. iPSCs, but not ESCs, stimulated CD4⁺ T cells derived from animals that had rejected iPSCs. In contrast, CD8⁺ T cells minimally proliferated to stimulation by iPS‐EB cells (e). Bothe CD4⁺ and CD8⁺ T cells from naive animals proliferated minimally. iPS‐EB cells induce T‐cell stimulation much more than ES‐EBs. These experiments were performed in triplicates in three mice and repeated twice. **P < 0·01 and *P < 0·05. (f) iPSCs are pluripotent and iPS‐EB cells poorly express MHC I and MHC II molecules. iPS‐EB cells do not express MHC antigens. EBs were harvested on day 7 and the cells were used to measure MHC class I and class II expression. 129x1/SvJ splenocytes were used as controls. EB cells hardly express any class I or class II antigens. Open histograms indicate class I or class II positive population and filled histograms indicate isotype control staining. [Colour figure can be viewed at wileyonlinelibrary.com]