Poster #025, CHLA LA-HIP
Effects of Hypoxic Culture Conditions on the Maintenance of Pluripotency in Induced Pluripotent Stem Cells
Mentors: Narine Harutyunyan, Stem Cell Core Manager and Kevin Passi, Research Specialist
Background: Induced pluripotent stem cells (iPSCs) are adult cells that have been reprogrammed to act like embryonic stem cells. Stem cells are usually grown under normal oxygen conditions in the laboratory settings, called normoxia (20.8% oxygen). However, in the human body, including during development, cells are often exposed to lower oxygen levels, called hypoxia (below 10% oxygen). The purpose of this study is to compare how iPSCs grow under normoxic and hypoxic conditions. We hypothesize that the different oxygen levels affect the markers of the undifferentiated state.
Experimental Approach: To test this hypothesis, we used matrigel and extracellular matrix to culture the cells. iPSC were seeded onto matrigel coated dishes to promote cell attachment. Cultured with mTESR media to maintain pluripotency. Over the course of 5 weeks we recorded two different time points. One in the middle (around week 3) and another at the end of the experiment. Quantitative polymerase chain reaction (qPCR) was used to measure the expression of genes related to pluripotency and differentiation: Sox2, Oct4, Nanog, Cyn-c, and Klf4. Immunofluorescence was used to detect and visualize proteins that are associated with the undifferentiated state: Sox2, Oct4, Nanog, Cyn-c, and Klf4.
Key Finding: We expect that iPSCs cultured under hypoxic conditions will maintain or enhance the expression of pluripotency markers while exhibiting reduced expression of differentiation markers compared with cells cultured under normoxic conditions.
Conclusion: This study will determine whether hypoxic culture conditions provide a more accurate environment for maintaining iPSC pluripotency and may contribute to improved stem
cell culture methods for regenerative medicine and future research.