Human Induced Pluripotent Stem Cell–Derived Keratinocyte-Like Cells for Research on Protease-Activated Receptor 2 in Nonhistaminergic Cascades of Atopic Dermatitis

Keratinocytes are the most abundant cells in the epidermis, and as part of the frontline immunologic defense system, keratinocytes function as a barrier to exogenous attacks. Protease-activated receptor 2 (PAR2) is expressed in human keratinocytes and activated in several inflammatory conditions, such as atopic dermatitis (AD). In this study, we demonstrated the differentiation of human induced pluripotent stem cell into keratinocytes by the improved, robust differentiation procedure and confirmed that human induced pluripotent stem cell–derived keratinocyte-like cells (iKera) express PAR2, which is activated by external addition of the ligand peptide and trypsin. The activation of PAR2 led to the release of calcium from intracellular calcium storage, followed by the release of the proinflammatory cytokine tumor necrosis factor α. Moreover, PAR2 antagonist I-191 (CAS No. 1690172-25-8) inhibited calcium release in a dose-dependent manner. This is the first study to demonstrate that iKera expresses a functional PAR2 protein. Furthermore, our results indicate crosstalk between the PAR2- and IL-4–mediated inflammatory axes in iKera, suggesting that iKera can be used as a platform for a broad range of mechanism-targeted drug screening in AD.

SIGNIFICANCE STATEMENT

This is the first study to provide evidence that human induced pluripotent stem cell–derived keratinocyte-like cells (iKera) express functional protease-activated receptor 2 (PAR2). Furthermore, this study demonstrated in iKera that the IL-4 inflammatory axis can crosstalk with the PAR2-mediated inflammatory axis in keratinocytes. To the best of our knowledge, this is the first report to indicate that iKera can be used for research and as a drug screening platform for atopic dermatitis.