Abstract
Efforts to develop effective therapeutic treatments for promoting fast wound healing after injury to the epidermis are hindered by a lack of understanding of the factors involved. Re-epithelialization is an essential step of wound healing involving the migration of epidermal keratinocytes over the wound site. Here, we examine genetic variants in the keratin-1 (KRT1) locus for association with migration rates of human epidermal keratinocytes (HEK) isolated from different individuals. Although the role of intermediate filament genes, including KRT1, in wound activated keratinocytes is well established, this is the first study to examine if genetic variants in humans contribute to differences in the migration rates of these cells. Using an in vitro scratch wound assay we observe quantifiable variation in HEK migration rates in two independent sets of samples; 24 samples in the first set and 17 samples in the second set. We analyze genetic variants in the KRT1 interval and identify SNPs significantly associated with HEK migration rates in both samples sets. Additionally, we show in the first set of samples that the average migration rate of HEK cells homozygous for one common haplotype pattern in the KRT1 interval is significantly faster than that of HEK cells homozygous for a second common haplotype pattern. Our study demonstrates that genetic variants in the KRT1 interval contribute to quantifiable differences in the migration rates of keratinocytes isolated from different individuals. Furthermore we show that in vitro cell assays can successfully be used to deconstruct complex traits into simple biological model systems for genetic association studies.
Highlights
When the outer covering of the skin, the epidermis, is injured a wound healing response is initiated
We examined cultured primary human epidermal keratinocytes (HEK) cells derived from the epidermal layer of normal neonatal foreskin to determine whether they could be used as a model system to test our hypothesis that genetic variants in the KRT1 interval are associated with variation in cell migration rates
To use the epidermal cells to test this hypothesis it would be necessary for them to express KRT1 mRNA and translate it into protein at a sufficient level. Both mouse and human primary epidermal keratinocytes, including those derived from neonatal human foreskin, have been reported to express KRT1 mRNA and protein when grown to confluence in permissive calcium concentrations (0.10–0.16 mM) [12,13,14]
Summary
When the outer covering of the skin, the epidermis, is injured a wound healing response is initiated. The expression of keratin 6 (KRT6), KRT16, and KRT17 is induced while the expression of KRT1 and KRT10 is down-regulated [2,3] These alterations in intermediate filament gene and protein regulation are thought to be necessary for the keratinocytes to make the morphological changes required for migration [2,3]. Studies in the mouse show that inbred strains differ significantly in wound healing rates, indicating that in mammals there are genetic factors contributing to this trait [4,5,6,7]. These results suggest that humans may have DNA variants associated with wound healing rates. Using in vitro model systems of the various stages of wound healing, such as the re-epithelialization step, would enable the identification of genetic associations using considerably fewer individual samples
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