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Human keratinocytes are efficiently immortalized by a Rho kinase inhibitor
Sandra Chapman, … , Richard Schlegel, Alison A. McBride
Sandra Chapman, … , Richard Schlegel, Alison A. McBride
Published June 1, 2010
Citation Information: J Clin Invest. 2010;120(7):2619-2626. https://doi.org/10.1172/JCI42297.
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Technical Advance Dermatology

Human keratinocytes are efficiently immortalized by a Rho kinase inhibitor

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Abstract

Primary human keratinocytes are useful for studying the pathogenesis of many different diseases of the cutaneous and mucosal epithelia. In addition, they can form organotypic tissue equivalents in culture that can be used as epidermal autografts for wound repair as well as for the delivery of gene therapy. However, primary keratinocytes have a finite lifespan in culture that limits their proliferative capacity and clinical use. Here, we report that treatment of primary keratinocytes (originating from 3 different anatomical sites) with Y-27632, a Rho kinase inhibitor, greatly increased their proliferative capacity and resulted in efficient immortalization without detectable cell crisis. More importantly, the immortalized cells displayed characteristics typical of primary keratinocytes; they had a normal karyotype and an intact DNA damage response and were able to differentiate into a stratified epithelium. This is the first example to our knowledge of a defined chemical compound mediating efficient cell immortalization, and this finding could have wide-ranging and profound investigational and medical applications.

Authors

Sandra Chapman, Xuefeng Liu, Craig Meyers, Richard Schlegel, Alison A. McBride

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Figure 3

Telomerase expression increases over time, and the length of telomere ends stabilizes after culture with Y-27632.

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Telomerase expression increases over time, and the length of telomere en...
(A) Relative levels of TERT mRNA from HFK strain a, cultured in the absence or presence of 10 μM Y-27632 at the pass indicated, as quantitated by real-time PCR. Each bar represents the mean of replicated samples ± SD. (B) Relative length of telomeres in HFK strain a, cultured in the absence or presence of 10 μM Y-27632 at the pass indicated, as quantitated by real-time PCR. Each bar represents the mean of replicated samples ± SD.

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ISSN: 0021-9738 (print), 1558-8238 (online)

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