Relative Radiosensitivity Of Human Inducible Pluripotent Stem Cells (Hipsc) Compared To Parent Fibroblast Line And Cells Of Differentiated Neural Rosettes

We sought to determine the radiosensitivity of human inducible pluripotent stem cells (hiPSC) compared to parent fibroblasts and cells in differentiated neural rosettes, Fibroblasts over-expressing 4 transfectants (OCT4, SOX2, NANOG and LIN28) were selected for hiPSC sublines. The radiosensitivity of hiPSC was compared to both the parent fibroblast progenitor cell line and cells from neural rosettes obtained following differentiation of hiPSCs. A human fibroblast parent cell line was grown in DMEM media with 10% FBS. The hiPSC cells were grown using mTeSR1 medium (Stem Cells Technology) on matrigel-coated 6-well plates in a humidified, 37oC incubator supplemented with 5% CO2. Neural rosettes were obtained by incubating the hiPSCs in neural precursor (NP) selection medium then transfer to NP expansion medium for 7 days which leads to the formation of spherical neural masses The hiPSC colonies, neural rosettes and the parent fibroblast cell line were irradiated to 0, 0.5, 1.0, 1.5, 2.0 or 2.5 Gy using a linear accelerator, dose rate of 300 cGy/min and then analyzed for apoptosis, cell cycle arrest and viability at 24 and 48 hr post-irradiation. Viability of the three starting cell populations was 92.1 ± 1.6% for hiPSC, 97.1 ± 0.7% for the fibroblasts and 83.8 ± 0.2% for the neural rosettes. Twenty-four hours after 2.5 Gy irradiation, the percent apoptosis of hiPSC increased from nonirradiated control levels of 7.4 ± 1.7% to 43.6 ± 8.0% (p = 0.0016). Cells from neural rosettes had an increased percent apoptosis from control of 16.2 ± 0.2% to 35.1 ± 3.9% (p = 0.0413). The parent fibroblast line had increased apoptosis from 2.6 ± 1.4% for control to 1.9 ± 0.5 after 2.5 Gy (p = 0.6841). There was a decreased viability of irradiated hiPSC cells at 24 hours after doses as low as 1.5 Gy (24.4 ± 6.2%) compared to nonirradiated control cells (49.3 ± 5.4%, p = 0.0250). In contrast, no significant decrease in viability after doses up to 2.5 Gy was detected in neural rosette cells (73.9 ± 1.7% for control nonirradiated compared to 67.7 ± 0.2% after 2.5 Gy, p = 0.0659) or parent fibroblasts viability (nonirradiated 78.2 ± 4.2% compared to 76.6 ± 1.4% after 2.5 Gy, p = 0.1118). Cell cycle arrest of irradiated hiPSC cells showed G1 phase pile-up at 48 hr after 2.0 or 2.5 Gy s (32.5 ± 3.9% for nonirradiated compared to 47.0 ± 3.6% and 45.1 ± 4.4% (p = 0.0253 or 0.0275, respectively). In contrast cells from neural rosettes or parent fibroblasts showed no significant G1 arrest at 24 or 48 hr after either irradiation dose. Human iPSCs are more radiosensitive than either parent fibroblasts or differentiated neural rosettes. These three cell populations are potentially of value to identify the molecular signaling pathways controlling induction and release from radiosensitivity.