We have previously established a treatment that allows us to efficiently and conditionally immortalize primary human keratinocytes and other epithelial cells by culturing them in the presence of a ROCK inhibitor and with feeder fibroblasts. The technique is invaluable for the efficient procurement of cells from individuals with specific genetic and infectious diseases. In FY2018 we have isolated keratinocytes and fibroblasts from individuals at the NIH clinical center to assess whether melorheostosis is associated with somatic mutations in these tissues of the epidermis. We have also isolated and cultured keratinocytes from different anatomical regions to assess whether they can efficiently support HPV infection. We are also assessing whether ROCK inhibitor treatment can allow us to generate immortalized cell lines expressing transgene or with CRISPR generated mutations.

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Stepp, Wesley H; Stamos, James D; Khurana, Simran et al. (2017) Sp100 colocalizes with HPV replication foci and restricts the productive stage of the infectious cycle. PLoS Pathog 13:e1006660
Chapman, Sandra; McDermott, David H; Shen, Kui et al. (2014) The effect of Rho kinase inhibition on long-term keratinocyte proliferation is rapid and conditional. Stem Cell Res Ther 5:60
Liu, Xuefeng; Ory, Virginie; Chapman, Sandra et al. (2012) ROCK inhibitor and feeder cells induce the conditional reprogramming of epithelial cells. Am J Pathol 180:599-607
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Chapman, Sandra; Liu, Xuefeng; Meyers, Craig et al. (2010) Human keratinocytes are efficiently immortalized by a Rho kinase inhibitor. J Clin Invest 120:2619-26