Effects of Vitamin D on Antimicrobial Host Defense in Human Keratinocyte Cells

Abstract

Epithelial tissues are constantly exposed to a variety of microbial challenges and previous findings have shown that their function as a mechanical barrier provides the main means of resistance to bacterial infection. In addition, antimicrobial peptides function as chemical barriers in that resistance as innate immunity factors. Many keratinocyte functions are regulated by 1α,25-dihydroxyvitamin D3 (VD3), the active form of vitamin D, which stimulates differentiation including promotion of innate and adaptive immunity. However, the mechanisms that regulate the interaction between antimicrobial peptides and VD3 are poorly understood. We investigated the effects of VD3 on expression of antimicrobial peptides by keratinocytes.HaCaT cells from a human keratinocyte cell line were grown in DMEM supplemented with 10% fetal bovine serum, then incubated with VD3 at 0 (control), 1, 10 or 100 nM for 4, 8, 24, 48, or 72 hours. Expressions of antimicrobial peptides, human β-defensin (hBD)-1, -2, -3, and LL-37/hCAP18 (LL-37), and mRNAs in HaCaT cells were observed by quantitative RT-PCR using TaqMan^Ⓡprobes. The relative expression of each mRNA was calculated as the ΔΔCt using a formula. Expressions of hBD-1, -2, -3, and LL-37 peptides in HaCaT cells were examined using ELISA assays. Obtained data were analyzed using ANOVA and Scheffe's test. Differences between experimental groups were considered statistically significant at p＜0.05.Expression levels of hBD-1 and -2 mRNA, and their peptides in HaCaT cells cultured with VD3 were significantly higher as compared to the non-treated control, whereas those of hBD-3 mRNA and its peptide did not change when exposed to VD3. Keratinocytes incubated with VD3 showed upregulated expressions of LL-37 mRNA and its peptide in both dose- and time-dependent manners.Our results indicate that VD3 plays crucial roles in the chemical barriers present in epithelium.