Cellular adhesion on collagen: a simple method to select human basal keratinocytes which preserves their high growth capacity

The regenerative capacity of human interfollicular epidermis is closely linked to the potential of immature keratinocytes present within its basal layer. The availability of selection methods and culture systems allowing precise assessment of basal keratinocyte characteristics is critical for increasing our knowledge of this cellular compartment. This report presents a multi-parametric comparative study of basal keratinocytes selected according to two different principles: 1) high adhesion capacity on a type-I collagen-coated substrate [Adh(+++)], 2) high cell-surface expression of alpha 6-integrin [Itg-alpha 6(high)]. Importantly, analysis performed at the single-cell level revealed similar primary clone-forming efficiency values of 45.5% +/- 6.7% [Itg-alpha 6(high)] and 43.7% +/- 7.4% [Adh(+++)], which were markedly higher than those previously reported. In addition, both methods selected keratinocytes exhibiting an extensive long-term growth potential exceeding 100 cell doublings and the capacity for generating a pluristratified epidermis. Our study also included a global transcriptome comparison. Genome-wide profiling indicated a strong similarity between [Adh(+++)] and [Itg-alpha 6(high)] keratinocytes, and revealed a common basal-associated transcriptional signature. In summary, cross-analysis of [Adh(+++)] and [Itg-alpha 6(high)] keratinocyte characteristics showed that these criteria identified highly equivalent cellular populations, both characterized by unexpectedly high growth capacities. These results may have broad impacts in the tissue engineering and cell therapy fields.