Several recent reports have demonstrated that photoreceptors are expressed in human skin. The rod and cone photoreceptor-like proteins are expressed in human skin and rhodopsin, long wavelength-opsin, and short wavelength-opsin are also present in cultured murine melanocytes. Furthermore, the photopigment rhodopsin is expressed in human melanocytes and is involved in ultraviolet A phototransduction which induces early melanin synthesis. In this study, we investigated whether rhodopsin is expressed and plays any physiological roles in the normal human epidermal keratinocytes (NHEKs). We found that rhodopsin was expressed and localized on the plasma membrane in NHEKs, and only violet light among several wavelengths within the visible range significantly increased the expression of rhodopsin mRNA. We further found that rhodopsin over-expression decreased the mRNA expression levels of keratinocyte differentiation markers, such as keratin-1 and keratin-10, and violet light also decreased the mRNA expression levels of keratinocyte differentiation markers and these decreased expression levels were recovered by a rhodopsin-directed siRNA. Moreover, we further demonstrated that violet light significantly decreased the phosphorylation levels of cAMP responsive element-binding protein (CREB) and that it more effectively decreased the phosphorylation of CREB when rhodopsin was over-expressed. In addition, we observed that pertussis toxin, a Gαi protein inhibitor, restored the rhodopsin-induced decrease in the differentiation markers in NHEKs. Taken together, these results suggest that rhodopsin down-regulates the expression levels of specific keratinocyte differentiation markers via the Gαi signaling pathway in NHEKs.

As the outermost physical barrier of an organism, the skin is diurnally exposed to UV radiation (UVR). Recent studies have revealed that the skin exhibits a circadian rhythm in various functions, and this oscillation is disturbed and reset via a strong environmental cue, the UVR. However, a molecular link between circadian perturbation by UVR and UVR-induced cellular responses has not been investigated. We identified tissue inhibitor of metalloproteinase (TIMP)-3 as a novel circadian locomotor output cycles kaput (CLOCK)–dependent diurnal gene by using a CLOCK-knockdown strategy in human keratinocytes. Among dozens of identified transcripts down-regulated by CLOCK knockdown, TIMP3 displayed a rhythmic expression in a CLOCK-dependent manner, in which the expression of matrix metalloproteinase (MMP)-1 and inflammatory cytokines, such as TNF-α, chemokine (C-X-C motif) ligand (CXCL)-1, and IL-8, were inversely regulated. Upon UVB exposure, the expression of CLOCK and TIMP3 was down-regulated, which led to an up-regulation of secretion of MMP1 and TNF-α proteins and in the transcription of CXCL1 and IL-8 via CCAAT-enhancer binding protein (C/EBP)-α. UVB-induced TNF-α secretion increased further or decreased by knockdown or overexpression of TIMP3, respectively, as well as by CLOCK. As a novel CLOCK-dependent diurnal gene, TIMP3 inhibits the expression of inflammatory cytokines that are up-regulated by UV irradiation in human keratinocytes. Thus, our work suggests a molecular link between circadian perturbation by UVR and UVR-induced inflammation.—Park, S., Kim, K., Bae, I.-H., Lee, S. H., Jung, J., Lee, T. R., Cho, E.-G. TIMP3 is a CLOCK-dependent diurnal gene that inhibits the expression of UVB-induced inflammatory cytokines in human keratinocytes.

Various kinds of glycosaminoglycans (GAGs) and proteoglycans (PGs) have been known to be involved in structural and space-filling functions, as well as many physiological regulations in skin. To investigate ultraviolet (UV) radiation-mediated regulation of GAGs and PGs in cultured human dermal fibroblasts, transcriptional changes of many types of PGs and GAG chain-synthesizing enzymes at 18 hr after 75 mJ/cm2 of UV irradiation were examined using quantitative real-time polymerase chain reaction methods. Hyaluronic acid synthase (HAS)-1, -2, and -3 and hyaluronidase-2 mRNA expressions were significantly increased by UV irradiation. Expressions of lumican, fibromodulin, osteoglycin, syndecan-2, perlecan, agrin, versican, decorin, and biglycan were significantly decreased by UV irradiation, while syndecan-1 was increased. Expressions of GAG chain-synthesizing glycosyltransferases, xylosyltransferase-1, β1,3-glucuronyltransferase-1, β1,4-galactosyltransferase-2, -4, exostosin-1, chondroitin polymerizing factor, and chondroitin sulfate synthase-3 were significantly reduced, whereas those of β1,3-galactosyltransferase-6, β1,4-galactosyltransferase-3, -7, β-1,3-N-acetylglucosaminyltran sferase-2, and -7 were increased by UV irradiation. Heparanase-1 mRNA expression was increased, but that of heparanase-2 was reduced by UV irradiation. Time-course investigation of representative genes showed consistent results. In conclusion, UV irradiation may increase hyaluronic acid production through HAS induction, and decrease other GAG productions through downregulation of PG core proteins and GAG chain-synthesizing glycosyltransferases in cultured human dermal fibroblasts.

Allan CM, Kupper TS. Human epidermal Langerhans cells maintain immune homeostasis in skin by activating skin resident regulatory T cells. Immunity 2012;36:873e84. Tewary P, de la Rosa G, Sharma N, Rodriguez LG, Tarasov SG, Howard OM, et al. beta-Defensin 2 and 3 promote the uptake of self or CpG DNA, enhance IFNalpha production by human plasmacytoid dendritic cells, and promote inflammation.

Autophagy is an essential process degrading damaged components. Although resveratrol has various beneficial activities for health, little is known about the effects of resveratrol on autophagy in skin. We investigated whether resveratrol affects autophagy in human dermal fibroblasts grown in complete medium. We found that after the resveratrol treatment, LC3‐II reached a maximum level at 8 h and then gradually decreased. By PCR array analysis, we identified death‐associated protein kinase 1 (DAPK1) as a new target of resveratrol, and we confirmed that the expression level of DAPK1 was enhanced by resveratrol. We also demonstrated that DAPK1 knock‐down by siRNA was sufficient to reduce resveratrol‐induced autophagy but did not affect the phosphorylation level of AMP‐activated kinase (AMPK), a well‐known target of resveratrol. These data indicate that resveratrol‐induced autophagy can be mediated by DAPK1, raising the possibility that some of the beneficial effects of resveratrol may be due to its regulation of DAPK1.