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• Rosacea epidermis show high HSD11B1 expression. • TLR3 agonist Poly(I:C) increases HSD11B1 expression in human epidermal keratinocytes. • Differentiated keratinocytes augments glucocorticoid synthetases induction by Poly(I:C). • The knockdown of TLR3 suppress the by Poly(I:C) stimuli. • TLR3 transduction augments HSD11B1 induction by Poly(I:C) in keratinocytes. While most skin diseases benefit from topical steroids, rosacea symptoms are exacerbated by topical steroids. In the rosacea pathogenesis, abnormal innate immune mechanisms including overexpression of the Toll-like receptor (TLR) have been proposed. However, the links between glucocorticoid metabolism and innate immunity in the epidermis have not been elucidated. In order to understand the pathology by which rosacea symptoms are exacerbated by steroids and environment stimuli, we examined the molecular links between the innate immune system and glucocorticoid synthesis in epidermis. We examined the expression of glucocorticoid-synthetic enzymes in rosacea skin. We stimulated epidermal keratinocytes by TLR ligands and examined the regulation of glucocorticoid-synthetic enzymes. We also employed siRNA and adenovirus vectors to knockdown and transduce TLR molecules, respectively. Rosacea epidermis showed high HSD11B1 in the granular layer. Among TLR ligands, TLR3 ligand Poly(I:C) enhanced the expression of multiple glucocorticoid-synthetic enzymes including HSD11B1 and CYP11A1, and increased cortisol in the cultured media. Induction of HSD11B1 by Poly(I:C) was abolished by pretreatment with TLR3 siRNA. Transfection with an adenoviral vector incorporating TLR3 enhanced HSD11B1 and CYP11A1 protein expression by Poly(I:C). In addition, cell staining revealed increased expression of HSD11B1 and CYP11A1 proteins in the group transfected with TLR3 under the same conditions. TLR3-stimulated epidermal keratinocytes and rosacea epidermis enhance the expression of glucocorticoid-synthetic enzymes, which would promote cortisol activation in the epidermis. The innate immunity modulates glucocorticoid-synthetic enzymes expression via the TLR3 pathway in epidermal keratinocytes. [ABSTRACT FROM AUTHOR]

• TLR3 agonist Poly(I:C) increases uptake of melanosome-rich globules, melanosomes, and melanocores by keratinocytes. • Poly(I:C) increases protease-activated receptor-2 (PAR-2) expression. • The knockdown of PAR-2 does not completely suppress endo/phagocytosis of melanin by Poly(I:C) stimuli. • Poly(I:C) increases the expressions and activation of RHOA and CDC42. • The knockdown of RHOA and CDC42 suppress endo/phagocytosis of melanin by Poly(I:C) stimuli. Keratinocytes and melanocytes in human epidermis express Toll-like receptors (TLR) and induce immune responses. We previously reported that TLR3 stimulation increases melanosome transport from perinuclear to cell membrane in melanocytes and enhanced release of melanosome from melanocytes, which were followed by increase in melanosome uptake into keratinocytes. In this study, we investigated whether TLR3 stimuli directly affect keratinocytes to enhance melanosome uptake. To observe keratinocyte's melanosome uptake ability precisely without melanocytes influences, we isolated melanosomes from human melanocytes and applied isolated melanosomes to keratinocytes stimulated by Poly(I:C). Poly(I:C)-stimulated keratinocytes enhanced uptake of isolated melanosome-rich globules five-times as much as control. Poly(I:C) increases the RNA and protein expressions of RHOA and CDC42, which are small GTP-binding proteins inducing the endocytosis. Pull-down assay showed that Poly(I:C) increased the GTP-binding RHOA and CDC42, suggesting TLR3 stimulation activated RHOA and CDC42. The knockdown of TLR3 suppressed RHOA and CDC42 induction by Poly(I:C). Consistently, the knockdown of RHOA and CDC42 significantly suppressed the melanosome-rich globules uptake by Poly(I:C)-stimulated keratinocytes. Because RHOA and CDC42 activation induces endocytosis by modification of actin stress fiber and filopodia formation, respectively, these results suggested that TLR3 stimulation enhances melanosome uptake into keratinocytes through endocytosis mechanisms. Combining with the data of our previous publications, TLR3, which signal is activated by sensing viral molecules, enhance pigmentation by controlling both melanin transport system by RAB GTPases induction in melanocytes and uptake system by RHOA and CDC42 in keratinocytes. [ABSTRACT FROM AUTHOR]

Highlights from the article: To examine the TRIF and MAVS involvement in RAB27 A induction by Poly(I:C), we employed siRNA for TRIF (siTRIF) and MAVS (siMAVS). Thus, the perinuclear accumulation of TRIF and MAVS by Poly(I:C) suggested that Poly(I:C) activates TRIF and MAVS in melanocytes.

Highlights from the article: RAB17-dependent filopodia formation serves as conduits for melanosome transfer and the knockdown of RAB17 or RAB11 induces melanosome accumulation in melanocytes and decreased melanosome release [[4]]. These data indicate that RAB11A is most abundant among RE-associated RABs, and TLR2 agonist HKLM enhances RAB11A expression and localization in normal human epidermal melanocytes. In conclusion, TLR2 stimuli augment RAB11A expression in human epidermal melanocytes and facilitate melanosome transfer to neighboring keratinocytes through RAB11A-associated melanosome transportation.