Jang-Hee Oh

Changes in glycosaminoglycans and related proteoglycans in intrinsically aged human skin in vivo.

Abstract:  Glycosaminoglycans (GAGs) and proteoglycans (PGs) are involved in various structural functions and physiological regulations in the skin. To investigate the intrinsic ageing‐dependent GAG and PG changes and their gender‐specific difference, immunohistochemical stains of several GAGs and PGs were performed in sun‐protected buttock skin tissues of young and old, male and female (total n = 32) human skin. Stains of alcian blue, hyaluronic acid (HA) and heparan sulphate were reduced in aged skin in both genders, whereas chondroitin sulphate stain was decreased only in female. Stains of HA synthase‐2, CD44, CD44v3, syndecan‐1 and decorin were decreased in aged skin in both genders, whereas perlecan stain was reduced only in female, and syndecan‐4 stain did not change. Versican stain was increased in male aged skin, but not in female. Age‐ and gender‐related changes in GAGs and PGs in intrinsically aged buttock skin elucidated in this study may play important roles in intrinsic skin ageing process.

Intrinsic aging- and photoaging-dependent level changes of glycosaminoglycans and their correlation with water content in human skin

BACKGROUND Glycosaminoglycans (GAGs) have various structural and physiological regulatory functions in skin, including tissue water maintenance, due to their high water-holding capacity. OBJECTIVE To investigate changes of GAGs during intrinsic aging and photoaging of human skin and their correlations with water content. METHODS Samples of sun-protected buttock and sun-exposed forearm skin were obtained from young male (21-30 years, n=8) and female (20-33 years, n=8) subjects, as well as old male (70-78 years, n=8) and female (70-80 years, n=8) subjects, and their epidermal and dermal contents of hyaluronic acid (HA), total sulfated GAG (tsGAG), total uronic acid (tUA), and tissue water were measured. HA content was determined by enzyme-linked immunosorbent assay using HA-binding protein, tsGAG by the sulfated GAG assay kit using 1,9-dimethylmethylene blue, tUA by carbazole reaction, and tissue water by subtraction of tissue dry weight from wet weight. RESULTS In the buttock, HA was higher in dermis than in epidermis, while tsGAG and tUA were higher in epidermis. In intrinsically aged buttock, epidermal HA and dermal tsGAG and tUA decreased. However, when analyzed for each gender, epidermal tsGAG, tUA, and tissue water decreased only in females. Forearm/buttock ratios of each molecule were compared for determination of photoaging-dependent changes. Forearm/buttock ratios of HA, tsGAG, tUA, and tissue water increased in aged dermis, but showed no change in aged epidermis. When analyzed for each gender, ratios of epidermal HA and tissue water increased only in aged females, while ratios of epidermal tsGAG, tUA, and tissue water decreased only in aged males. Correlations of water content with HA, tsGAG, and tUA were found in epidermis, but not with tsGAG in dermis. CONCLUSION These intrinsic aging- and photoaging-dependent GAG changes and their correlations with water content provide new insights into the pathophysiology of dry skin in the elderly.

Relationship between skin phototype and MED in Korean, brown skin

The Fitzpatrick skin classification has been a useful method to categorize cutaneous sensitivity to ultraviolet radiation (UVR), although it was based originally on responses in white skin. Because the relevance of this phototype in brown skin is in question, we investigated skin phototypes of university students by a self‐reporting questionnaire and measured their MEDs in Korean, brown skin. After studying our explanation of the definition of Fitzpatrick skin types, 707 Korean university students answered the questionnaire. We then measured UVB MEDs in 156 randomly selected male students. The order of frequency of skin type was type III (55.0%), IV (29.0%), and V (12.3%) by the questionnaire, with the sun sensitive categories (types I and II) reported only for 3.7%. There was no significant difference in MEDs between types IV and V, and the mean MED of each skin type did not show a monotonic increase with increasing skin type. Subjects with MEDs of 70–90 mJ/cm2 (corresponding to the MED of skin type V, as proposed by Pathak & Fitzpatrick) represented about half or more of the subjects in all categories, even types II and III. Subjects with MEDs lower than 60 mJ/cm2 were more prevalent in types II and III compared with types IV and V We suggest that there is at best a weak relationship between the skin types, by the Fitzpatrick method, and MEDs in Korean, brown skin.

[6]-Shogaol inhibits melanogenesis in B16 mouse melanoma cells through activation of the ERK pathway

Aim:To investigate the effect of [6]-shogaol, an active ingredient in ginger, on melanogenesis and the underlying mechanisms.Methods:B16F10 mouse melanoma cells were tested. Cell viability was determined with the MTT assay. Melanin content and tyrosinase activity were analyzed with a spectrophotometer. The protein expression of tyrosinase and microphthalmia associated transcription factor (MITF), as well as phosphorylated or total ERK1/2 and Akt were measured using Western blot.Results:Treatment of the cells with [6]-shogaol (1, 5, 10 μmol/L) reduced the melanin content in a concentration-dependent manner. [6]-Shogaol (5 and 10 μmol/L) significantly decreased the intracellular tyrosinase activity, and markedly suppressed the expression levels of tyrosinase and MITF proteins in the cells. Furthermore, [6]-shogaol (10 μmol/L) activated ERK, which was known to negatively regulate melanin synthesis in these cells. Pretreatment with the specific ERK pathway inhibitor PD98059 (20 μmol/L) greatly attenuated the inhibition of melanin synthesis by [6]-shogaol (10 μmol/L).Conclusion:The results demonstrate that [6]-shogaol inhibits melanogenesis in B16F10 mouse melanoma cells via activating the ERK pathway.

Glycosaminoglycan and proteoglycan in skin aging

Glycosaminoglycans (GAGs) and proteoglycans (PGs) are abundant structural components of the extracellular matrix in addition to collagen fibers. Hyaluronic acid (HA), one of GAGs, forms proteoglycan aggregates, which are large complexes of HA and HA-binding PGs. Their crosslinking to other matrix proteins such as the collagen network results in the formation of supermolecular structures and functions to increase tissue stiffness. Skin aging can be classified as intrinsic aging and photoaging based on the phenotypes and putative mechanism. While intrinsic aging is characterized by a thinned epidermis and fine wrinkles caused by advancing age, photoaging is characterized by deep wrinkles, skin laxity, telangiectasias, and appearance of lentigines and is mainly caused by chronic sun exposure. The major molecular mechanism governing skin aging processes has been attributed to the loss of mature collagen and increased matrix metalloproteinase expression. However, various strategies focusing on collagen turnover remain unsatisfactory for the reversal or prevention of skin aging. Although the expression of GAGs and PGs in the skin and their regulatory mechanisms are not fully understood, we and others have elucidated various changes in GAGs and PGs in aged skin, suggesting that these molecules are important contributors to skin aging. In this review, we focus on skin-abundant GAGs and PGs and their changes in human skin during the skin aging process.

Serum amyloid A1 secreted from UV‐irradiated keratinocytes induces matrix metalloproteinase‐1 in fibroblasts through toll‐like receptor 4

Ultraviolet (UV) irradiation on skin triggers photoageing‐related phenotypes such as formation of wrinkles. UV ray upregulates matrix metalloproteinase‐1 (MMP‐1), which in turn degrades extracellular matrix proteins, mostly collagens. Serum amyloid A1 (SAA1) is an acute‐phase protein of which plasma concentration increases in response to inflammation. Although the expression of SAA1 in the skin was reported, its function in the skin is yet to be studied. In this research, we found that the expression of SAA1 was increased in acute UV‐irradiated buttock skin and photoaged forearm skin in vivo. UV irradiation also increased SAA1 in normal human epidermal keratinocytes (NHEK), and treatment of recombinant human SAA1 (rhSAA1) induced MMP‐1 in normal human dermal fibroblasts (NHDF) but not in NHEK. Next, we demonstrated that NHDF treated with UV‐irradiated keratinocyte‐conditioned media showed the increased MMP‐1 expression; however, this increase of MMP‐1 in NHDF was inhibited by knockdown of SAA1 in NHEK. In addition, knockdown of Toll‐like receptor 4 (TLR4) inhibited rhSAA1‐induced MMP‐1 expression in NHDF. Taken together, our data showed that UV‐induced SAA1 production in NHEK, and this secreted SAA1 induced MMP‐1 expression in NHDF in a paracrine manner through TLR4 signalling pathway. Therefore, our results suggest that SAA1 can be a potential mediator for UV‐induced MMP‐1 expression in human skin.

Toll-like receptor family members in skin fibroblasts are functional and have a higher expression compared to skin keratinocytes

Toll-like receptors (TLRs) are known to recognize not only pathogen-associated molecular patterns but also danger-associated molecular patterns. Recent studies have characterized the expression levels and functions of TLRs in human epidermal cells. However, the characteristics of TLR family members in human dermal fibroblasts have not been thoroughly studied. Therefore, the present study systematically investigated the expression levels of TLRs and their functional responses to each ligand in skin fibroblasts. All 10 TLRs are expressed in skin fibroblasts. Stimulation of skin fibroblasts with each TLR ligand resulted in an increase of the interleukin-6 (IL-6), IL-8 and matrix metalloproteinase-1 proteins, indicating that ≥ 9 TLRs in skin fibroblasts are functionally active. Furthermore, stimulating skin fibroblasts with TLR1/2, 3 and 4 ligands induced the phosphorylation of inhibitor of nuclear factor κBα and the active phosphorylation of extracellular-signal regulated kinase 1/2. The expression level of each TLR was much higher in fibroblasts compared to keratinocytes. In particular, the fold-increase in IL-6 and IL-8 mRNA levels upon exposure to a TLR1/2 ligand was much higher in fibroblasts compared to keratinocytes, which appears to reflect the difference in expression levels of TLR1 and 2 between fibroblasts and keratinocytes. Taken together, these results show that all 10 TLRs are constitutively expressed and functional (except TLR10) in skin fibroblasts and suggest that TLRs in skin fibroblasts may play an important role in the detection of and response to different classes of pathogens and danger signals.

Poly(I:C) induces expressions of MMP-1, -2, and -3 through various signaling pathways including IRF3 in human skin fibroblasts

BACKGROUND Ultraviolet (UV) irradiation can result in premature skin aging (photoaging) which is characterized by decreased expression of collagen and increased expression of matrix metalloproteinases (MMPs). Double-stranded RNAs (dsRNAs) can be generated at various conditions including virally infected cells or UV-damaged skin cells. Recent studies have shown that a synthetic dsRNA, polyinosinic-polycytidylic acid (poly(I:C)), can reduce procollagen expression in human skin fibroblasts. However, little is known about the effect of poly(I:C) on the expression of MMPs in skin fibroblasts and its underlying mechanisms. OBJECTIVE We examined the effect of poly(I:C) on MMP-1, -2, and -3 expressions in human skin fibroblasts. Then, we further explored the underlying signaling pathways involved in the processes. METHODS Human skin fibroblasts were treated with poly(I:C) for the indicated times in the presence or the absence of various chemical inhibitors or small interfering RNAs (siRNAs) at the indicated concentrations. Protein and mRNA levels of various target molecules were examined by Western blotting and quantitative real-time PCR, respectively. RESULTS Poly(I:C) induced MMP-1, -2, and -3 expressions, which were dependent on TLR3. Poly(I:C) also induced activations of the mitogen-activated protein kinases (MAPKs), the nuclear factor-kappaB (NF-κB) and the interferon regulatory factor 3 (IRF3) pathways. By using specific inhibitors, we found that poly(I:C)-induced expressions of MMP-1, -2, and -3 were differentially regulated by these signaling pathways. In particular, we found that the inhibition of IRF3 signaling pathways attenuated poly(I:C)-induced expressions of all the three MMPs. CONCLUSION Our data show that the expressions of MMP-1, -2, and -3 are induced by poly(I:C) through various signaling pathways in human skin fibroblasts and suggest that TLR3 and/or IRF3 may be good targets for regulating the expressions of MMP-1, -2, and -3 induced by dsRNAs.

Vasoactive intestinal peptide stimulates melanogenesis in B16F10 mouse melanoma cells via CREB/MITF/tyrosinase signaling.

Vasoactive intestinal peptide (VIP), one of the major skin neuropeptides, has been suggested to have active roles in the pathogenesis of inflammatory skin disorders such as atopic dermatitis and psoriasis, which can commonly cause post-inflammatory hyperpigmentation. However, the effect of VIP on melanogenesis remains unknown. In this study, we showed that the melanin contents, tyrosinase activity, and gene expression of tyrosinase and microphthalmia-associated transcription factor (MITF) were significantly increased by treatment with VIP in B16F10 mouse melanoma cells and the stimulatory melanogenic effect was further examined in human epidermal melanocytes (HEMns). In addition, phosphorylated levels of CRE-binding protein (CREB) and protein kinase A (PKA) were markedly increased after VIP treatment, but not p38 mitogen-activated protein kinase (p38 MAPK), extracellular signal-regulated kinase (ERK), or Akt, indicating the possible PKA-CREB signaling pathway involved in VIP-induced melanogenesis. This result was further verified by the fact that VIP induced increased melanin synthesis, and protein levels of phosphorylated CREB, MITF, tyrosinase were significantly attenuated by H89 (a specific PKA inhibitor). These data suggest that VIP-induced upregulation of tyrosinase through the CREB-MITF signaling pathway plays an important role in finding new treatment strategy for skin inflammatory diseases related pigmentation disorders.

Src kinase mediates UV-induced TRPV1 trafficking into cell membrane in HaCaT keratinocytes

The skin is constantly exposed to harmful external stimuli such as pathogen, particulate matter, and sunlight. Solar radiation consists of various types of rays, among which ultraviolet (UV), visible light, and infrared can reach the earth and thereby affect human skin (1). Especially, UV has several detrimental effects on the skin including cutaneous cancer, inflammation, and photoaging (2). When skin cells are exposed to UV, it causes nuclear factor kappa B (NF-kB) signaling pathway leading to the increases of proinflammatory cytokines such as tumor necrosis factor-alpha (TNF-α), interleukin (IL)-1β, IL-6, and IL-8 (3). This article is protected by copyright. All rights reserved.