Complexation of lipids with cyclodextrin carriers for fully defined supplementation of cell culture
Corbin, Elizabeth Dale
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Induced Pluripotent Stem Cells (iPSCs) hold great promise for revolutionizing medicine and research. Scientists are currently able to reprogram adult cells of almost any type to a genetically 'open' state, pluripotency, wherein they lose the characteristics of their original cell type, and revert to a state that can reproduce indefinitely, and can be differentiated to many different cell types. IPSCs are currently grown in 'chemically defined' media that contains no animal derived components. This media eliminates animal and human sera because these tend to be quite variable and confound the reprogramming process, but the chemically defined media in use has almost no lipid content. The central goal of this project was to develop methods for chemically defined addition of lipids to cell culture media. The methods developed promise to be an advance in stem cell and general cell culture methodology, providing more reproducible experimental results, and supporting cells in culture with optimized lipid contents. In order to facilitate the addition of lipids to cell culture media without animal serum or serum albumin, complexation of individual lipids with a methyl beta-cyclodextrin starch was accomplished without addition of other molecules or oxidation of delicate lipids at a 1:1 stoichiometry. The lipid/MBCD complexes are soluble in aqueous media, and can be added individually or as a mixture to cell cultures. Application of complexed lipids to stem cells and fibroblasts revealed significant differences in lipid responses. Supplementation of human fibroblasts with a mixture of complexed lipids and other elements caused a 60% increase in proliferation over a 10 day period. Supplementation of stem cells with complexed lipids significantly increased proliferation, without reduction of pluripotency. Differences in lipid responses were also found between iPSC and embryonic stem cells, that may help elucidate differences between genetic or metabolic states which may point the way for more effective reprogramming of adult cells to pluripotency.