Poster #094, Children's Hospital Los Angeles
Hypoxic and Normoxic effects on HM19 and H9 iPSCs
Mentors: Kevin B. Passi, MS , Narine Harutyunyan, and Stefano Da Sacco, PhD
Induced pluripotent stem cells (iPSCs) are unspecialized cells that can be differentiated into all other cells in the body. These cells are routinely grown in the lab using normoxia conditions which reflect the atmospheric oxygen (20.8%). However, this condition is not physiological. Infact, conditions within tissues of the human body are hypoxic, with concentration of oxygen often well below 10%. For this reason, culturing pluripotent cells at normoxic conditions might not reflect their physiological state. Our research question is how hypoxic and normoxic conditions affect cell lines growth and markers of undifferentiated state. HM19 male and H9 female iPSCs lines were cultured in the incubator at 5.8% and 20.8% oxygen concentrations. To check for pluripotency and cell growth we used Immunofluorescence and qPCR. Immunofluorescence was used to tag the antibodies to visualize markers of undifferentiated states like Sox2, Oct4,Nanog,c-Myc, KlF4 and the growth. qPCR was used to see if genes Sox2, Oct4, Nanog, c-Myc, KLF4 (markers of undifferentiated state) are modulated. Confluency of both the cell lines HM19 and H9 in the 5% incubator was found to be higher compared to those cultured in the 20% incubator, suggesting that cells grow faster in the 5% oxygen condition. We expect that the cell lines in the 5% incubator will show more active genes and markers of the undifferentiated state, since they appear to replicate faster. Based on the qPCR and immunofluorescence we can assess the effects of hypoxic and normoxia condition on markers of undifferentiated state in the cell lines HM19 and H9 by looking at Sox2, Oct4, Nanog, c-Myc, KLF4.