Journal
INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES
Volume 19, Issue 8, Pages -Publisher
MDPI
DOI: 10.3390/ijms19082174
Keywords
keratinocyte; feeder layer; cholera toxin; isoproterenol; cell culture; statistical model
Funding
- Fondation des Pompiers du Quebec pour les Grands Brules (FPQGB)
- Canadian Institutes of Health Research (CIHR) [MOP-12087, FDN-143213]
- Quebec Network for Cell, Tissue and Gene Therapy-TheCell (a thematic network - Fonds de recherche du Quebec-Sante)
- Fonds de Recherche du Quebec en Sante (FRQS)
- Fondation du CHU de Quebec-Universite Laval
- Universite Laval Research Center on Organogenesis/LOEX
- Faculty of Medicine of the Universite Laval
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Human keratinocyte culture has provided the means to treat burns, wounds and skin pathologies. To date, to efficiently culture keratinocytes, cells are cultured on an irradiated feeder layer (iFL), either comprising human (iHFL) or murine (i3T3FL) fibroblasts, and the culture medium is supplemented with a cyclic adenosine monophosphate (cAMP) accumulation inducing agent such as isoproterenol (ISO) or cholera toxin (CT). Previous studies have characterized how the feeder layer type and the cAMP inducer type influence epithelial cells' phenotype independently from one another, but it is still unknown if an optimal combination of feeder layer and cAMP inducer types exists. We used sophisticated statistical models to search for a synergetic effect of feeder layer and cAMP inducer types on human keratinocytes' proliferative potential. Our data suggests that, when culturing human keratinocytes, using iHFL over i3T3FL increases population doublings and colony-forming efficiency through signaling pathways involving Ak mouse strain thymoma (Akt, also known as protein kinase B) isoforms 1 to 3, signal transducer and activator of transcription 5 (STAT5), p53, and adenosine monophosphate activated protein kinase alpha 1 (AMPK alpha 1). Both tested cAMP inducers ISO and CT yielded comparable outcomes. However, no significant synergy between feeder layer and cAMP inducer types was detected. We conclude that, to promote human keratinocyte growth in the early passages of culture, co-culturing them with a human feeder layer is preferable to a murine feeder layer.
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