High Calcium, ATP, and Poly(I:C) Augment the Immune Response to β-Glucan in Normal Human Epidermal Keratinocytes

Abstract

β-Glucans are pathogen-associated molecular patterns of fungi such as Candida albicans. Here, we studied their effects on normal human epidermal keratinocytes (NHEKs) from neonatal foreskin, and with high calcium to induce keratinocyte differentiation, danger signals, and pathogen-associated compounds such as adenosine 5'-triphosphate (ATP), poly(I:C), and lipopolysaccharide (LPS). β-Glucan stimulation significantly increased IL-8, IL-6, and IL-1α production by NHEKs. Well-differentiated NHEKs produced elevated IL-8 levels, whereas ATP, a danger signal, significantly increased IL-8 and IL-6 production, and the pathogen-associated compound, poly(I:C), augmented IL-1α production by β-glucan-stimulated NHEKs. No response to LPS from Escherichia coli was seen. Dectin-1 is known as the major receptor for β-glucans on phagocytes and dendritic cells. Dectin-1 mRNA was detected in NHEKs by reverse transcription-PCR. Flow-cytometric analyses confirmed the NHEK cell surface expression of dectin-1. Immunoblotting showed that β-glucan induced dual phosphorylation of p44/42 mitogen-activated protein kinase (MAPK) (extracellular signal-regulated kinase (ERK)1/2), and p38 MAPK in NHEKs; these signaling pathways are known to be associated with dectin-1. Treatment with the ERK inhibitor PD98059 and with the p38 kinase inhibitor SB203580 effectively suppressed β-glucan-induced IL-8 production by NHEKs. Thus, high calcium, ATP, and poly(I:C) augment the cytokine and chemokine production by β-glucan-stimulated NHEKs. Dectin-1 is present on NHEKs and may have an important role in cell response to β-glucan.