Activation of Peroxisome Proliferator-Activated Receptor Alpha Improves Aged and UV-Irradiated Skin by Catalase Induction.

Peroxisome proliferator-activated receptor alpha (PPARα) is a nuclear hormone receptor involved in the transcriptional regulation of lipid metabolism, fatty acid oxidation, and glucose homeostasis. Its activation stimulates antioxidant enzymes such as catalase, whose expression is decreased in aged human skin. Here we investigated the expression of PPARα in aged and ultraviolet (UV)-irradiated skin, and whether PPARα activation can modulate expressions of matrix metalloproteinase (MMP)-1 and procollagen through catalase regulation. We found that PPARα mRNA level was significantly decreased in intrinsically aged and photoaged human skin as well as in UV-irradiated skin. A PPARα activator, Wy14643, inhibited UV-induced increase of MMP-1 and decrease of procollagen expression and caused marked increase in catalase expression. Furthermore, production of reactive oxygen species (ROS) was suppressed by Wy14643 in UV-irradiated and aged dermal fibroblasts, suggesting that the PPARα activation-induced upregulation of catalase leads to scavenging of ROS produced due to UV irradiation or aging. PPARα knockdown decreased catalase expression and abolished the beneficial effects of Wy14643. Topical application of Wy14643 on hairless mice restored catalase activity and prevented MMP-13 and inflammatory responses in skin. Our findings indicate that PPARα activation triggers catalase expression and ROS scavenging, thereby protecting skin from UV-induced damage and intrinsic aging.

Supplementating with dietary astaxanthin combined with collagen hydrolysate improves facial elasticity and decreases matrix metalloproteinase-1 and -12 expression: a comparative study with placebo.

Photoaging accounts for most age-related changes in skin appearance. It has been suggested that both astaxanthin, a potent antioxidant, and collagen hydrolysate can be used as antiaging modalities in photoaged skin. However, there is no clinical study using astaxanthin combined with collagen hydrolysate. We investigated the effects of using a combination of dietary astaxanthin and collagen hydrolysate supplementation on moderately photoaged skin in humans. A total of 44 healthy subjects were recruited and treated with astaxanthin (2 mg/day) combined with collagen hydrolysate (3 g/day) or placebos, which were identical in appearance and taste to the active supplementation for 12 weeks. The elasticity and hydration properties of facial skin were evaluated using noninvasive objective devices. In addition, we also evaluated the expression of procollagen type I, fibrillin-1, matrix metalloproteinase-1 (MMP-1) and -12, and ultraviolet (UV)-induced DNA damage in artificially UV-irradiated buttock skin before and after treatment. The supplement group showed significant improvements in skin elasticity and transepidermal water loss in photoaged facial skin after 12 weeks compared with the placebo group. In the supplement group, expression of procollagen type I mRNA increased and expression of MMP-1 and -12 mRNA decreased compared with those in the placebo group. In contrast, there was no significant difference in UV-induced DNA damage between groups. These results demonstrate that dietary astaxanthin combined with collagen hydrolysate can improve elasticity and barrier integrity in photoaged human facial skin, and such treatment is well tolerated.

Long-term topical oestrogen treatment of sun-exposed facial skin in post-menopausal women does not improve facial wrinkles or skin elasticity, but induces matrix metalloproteinase-1 expression.

It is controversial whether treatment with oestrogen stimulates collagen production or accumulation in sun-exposed skin. The aim of this study was to determine the effect of long-term treatment with topical oestrogen on photoaged facial skin, with regard to wrinkle severity, and expression of procollagen and matrix metalloproteinase-1 enzyme. Two groups of 40 post-menopausal women applied either 1 g of 1% oestrone or vehicle cream once daily to the face for 24 weeks. Visiometer R1-R5 values (skin wrinkles) and Cutometer values (skin elasticity) were not significantly improved in the oestrone group after 24 weeks of treatment. Type I procollagen immunostaining did not increase in the oestrone group compared with the control group. However, levels of matrix metalloproteinase-1 mRNA increased robustly (10.3 times) in oestrone-treated skin compared with vehicle-treated skin. Thus, treatment with topical oestrogen may be deleterious in ultraviolet-induced skin ageing, at least in part, through induction of matrix metalloproteinase-1 (MMP-1) expression in human skin.

Inhibitory effects of TRPV1 blocker on UV-induced responses in the hairless mice.

The transient receptor potential vanilloid 1 (TRPV1) channel can be activated by vanilloids, exposure to ultraviolet (UV) irradiation, heat, or protons, and conditions that occur during tissue injury. In the present study, we investigated whether or not TRPV1-specific blocker, 5'-iodoresiniferatoxin (I-RTX), can reduce UV-induced matrix metalloproteinases (MMPs), pro-inflammatory cytokines, cyclooxygenase (COX)-2, and p53 expression in the skin of hairless mice. Our results showed that I-RTX inhibited UV-induced skin thickening, as measured by a caliper, or in hematoxylin and eosin (H&E)-stained sections. UV-induced mRNA and protein expression of MMP-13, MMP-9, MMP-3, and MMP-2 was significantly reduced by I-RTX. We also observed the inhibitory effects of I-RTX on UV-induced mRNA expression of the pro-inflammatory cytokines, interleukin (IL)-1ß, IL-2, IL-4, and tumor necrosis factor-α. UV-induced COX-2 and p53 protein expression was also significantly decreased by I-RTX. From the above results, we suggest that TRPV1-specific blocker, I-RTX, could prevent UV-induced skin responses, and provide new insight into development of effective therapeutic methods for photoaging.

Modulation of elastin exon 26A mRNA and protein expression in human skin in vivo.

Photoaged skin contains elastotic materials in the upper reticular dermis, a result of a commonly known as solar elastosis. It is known that the primary transcript of elastin undergoes extensive alternative splicing and that this results in the translation of multiple heterogeneous protein isoforms. In this study, we found that UV irradiation and heat treatment increased the levels of elastin transcript containing exon 26A and its encoded elastin isoform in the epidermis of human skin in vivo and in cultured human keratinocytes in vitro. We also found that the elastin transcript containing exon 26A was upregulated in photoaged forearm skin compared with intrinsically aged buttock skin in the same elderly individuals. We observed that topical retinoic acid treatment to human skin did not increase the expression of exon 26A mRNA, but that tropoelastin mRNA expression was increased by this treatment. These findings suggest that the production of the elastin isoform containing exon 26A peptide is increased by UV exposure and heat treatment in human skin in vivo and that it may play an important role in the development of solar elastosis in photoaged human skin.

Photoaging-associated changes in epidermal proliferative cell fractions in vivo.

The epidermis is a dynamic epithelium with constant renewal throughout life. Epidermal homeostasis depends on two types of proliferative cells, keratinocyte stem cells (KSCs), and transit amplifying (TA) cells. In the case of chronologic aging, levels of KSCs tend to decrease and change functionally. However, little is known about the effect of photoaging on epidermal proliferative subtype populations. The aim of this study was to validate involucrin/beta1-integrin ratio as a molecular marker of epidermal photoaging, and to investigate the effects of photoaging caused by chronic UV exposure on the proliferative subtype populations. A total of 15 male volunteers (age range 20-24 and 77-85 years, Fitzpatrick skin phototype III-IV) provided sun-exposed and sun-protected skin samples for real-time RT-PCR, Western blot analysis and immunostaining. Fractional changes in proliferative subtype populations in photoaged and chronologically aged skins were analyzed by flow cytometry. The expression of beta1-integrin was found to be significantly reduced in photoaged skin and ratios of the expressions of involucrin to beta1-integrin were increased 2.6-fold only in elderly subjects. Interestingly, immunostaining of the sun-exposed skins of elderly subjects showed aberrant beta1-integrin expression over the basal layer and greater numbers of Ki-67-positive cells than in sun-protected buttock skin. Flow cytometric analysis revealed that the proportion of KSCs to TA cells was reversed in sun-exposed and sun-protected skins of elderly subjects. Our results suggest that KSC numbers may be lower in photoaged skin than in chronologically aged skin and could be applied to hyperplastic pattern of photoaging. These findings suggest that the epidermis of photoaged skin is impaired in terms of its proliferative potential by attempting to repair chronic UV exposure and that photoaging may be associated with alteration in the two proliferative cell fractions.

Promotive effect of minoxidil combined with all-trans retinoic acid (tretinoin) on human hair growth in vitro.

Minoxidil induces hair growth in male pattern baldness and prolongs the anagen phase. All-trans retinoic acid (ATRA) has been reported to act synergistically with minoxidil in vivo: they can enhance more dense hair regrowth than either compound alone. We evaluated the effect of minoxidil combined with ATRA on hair growth in vitro. The effect of co-treatment of minoxidil and ATRA on hair growth was studied in hair follicle organ culture. In cultured human dermal papilla cells (DPCs) and normal human epidermal keratinocytes, the expressions of Erk, Akt, Bcl-2, Bax, P53 and P21 were evaluated by immunoblot analysis. Minoxidil plus ATRA additively promoted hair growth in vitro, compared with minoxidil alone. In addition, minoxidil plus ATRA elevated phosphorylated Erk, phosphorylated Akt and the ratio of Bcl-2/Bax, but decreased the expressions of P53 and P21 more effectively than by minoxidil alone. Our results suggest that minoxidil plus ATRA would additively enhance hair growth by mediating dual functions: 1) the prolongation of cell survival by activating the Erk and Akt signaling pathways, and 2) the prevention of apoptosis of DPCs and epithelial cells by increasing the ratio of Bcl-2/Bax and downregulating the expressions of P53 and P21.

Expression of androgen and estrogen receptors in human scalp mesenchymal cells in vitro.

The expression levels of sex hormone receptors were identified to be different in human mesenchymal cells [dermal papilla cell (DPC), dermal sheath cell (DSC), dermal fibroblast and (DF)] from occipital scalps. Transcriptional and translational activities of androgen receptor (AR) and estrogen receptor beta (ERbeta) were most intensely expressed in DPC, followed by DSC and DF. On the contrary, estrogen receptor alpha (ERalpha) was shown with the strongest positivity in DSC, succeeded by DPC and DF subsequently. Immunocytochemical staining showed the similar expression to previous patterns. Our results suggest that the expression levels of ER subtypes and AR may be important for the regulation of follicular mesenchymal cells in human scalp. Further studies of the interactions of hormones and receptors in human hair follicles are required to promote our understanding of the effects of sex hormones on hair biology.

Perifollicular fibrosis: pathogenetic role in androgenetic alopecia.

Androgenetic alopecia (AGA) is a dihydrotestosterone (DHT)-mediated process, characterized by continuous miniaturization of androgen reactive hair follicles and accompanied by perifollicular fibrosis of follicular units in histological examination. Testosterone (T: 10(-9)-10(-7) M) treatment increased the expression of type I procollagen at mRNA and protein level. Pretreatment of finasteride (10(-8) M) inhibited the T-induced type I procollagen expression at mRNA (40.2%) and protein levels (24.9%). T treatment increased the expression of transforming growth factor-beta 1 (TGF-beta1) at protein levels by 81.9% in the human scalp dermal fibroblasts (DFs). Pretreatment of finasteride decreased the expression of TGF-beta1 protein induced by an average of T (30.4%). The type I procollagen expression after pretreatment of neutralizing TGF-beta1 antibody (10 microg/ml) was inhibited by an average of 54.3%. Our findings suggest that T-induced TGF-beta1 and type I procollagen expression may contribute to the development of perifollicular fibrosis in the AGA, and the inhibitory effects on T-induced procollagen and TGF-beta1 expression may explain another possible mechanism how finasteride works in AGA.

Augmentation of UV-induced skin wrinkling by infrared irradiation in hairless mice.

Skin aging can be divided into intrinsic aging and photoaging. Sunlight is a major cause of photoaging, and is composed of ultraviolet (UV) and infrared (IR) radiation. Although the effects of UV radiation on skin aging have been widely studied, little is known about the biological effects of IR on the photoaging process in human skin. We found that chronic IR treatment induced wrinkles in hairless mice, and augments UV-induced wrinkle formation and UV-induced skin thickening in hairless mice. Histologically, we found that IR treatment augments UV-induced epidermal and dermal thickening, and that UV-induced increases of collagen and elastic fibers in dermis. Moreover, chronic IR treatment increased MMP-3 and MMP-13 mRNA expressions significantly in hairless mouse skin and augmented UV-induced MMP-3 and MMP-13 mRNA expressions and UV-induced MMP-2 and MMP-9 activities. From these results, we demonstrate that IR alone induces skin wrinkling and augments UV-induced wrinkle formation. Taken together, we suggest that IR plays an important role in the development of photoaging.

Heat modulation of tropoelastin, fibrillin-1, and matrix metalloproteinase-12 in human skin in vivo.

Photoaged skin contains elastotic materials in the upper reticular dermis. This phenomenon is commonly known as solar elastosis. In this study, we investigated the effects of heat on the expression of tropoelastin and fibrillin-1, two main components of elastic fibers, and on matrix metalloproteinase (MMP)-12, the most active MMP against elastin, in human skin in vivo. Heat was found to increase tropoelastin mRNA and protein expression in the epidermis and in the dermis. Fibrillin-1 mRNA and protein expression were increased by heat in the epidermis, but were decreased in the dermis. We found that pre-treatment of skin with N-acetyl cysteine or genistein for 24 h prior to heat treatment inhibited the heat-induced expression of tropoelastin, but not of fibrillin-1. These data indicate that reactive oxygen species may play a role in tropoelastin expression by heat, but not in fibrillin-1 expression. We also found that heat treatment increases MMP-12 mRNA and protein expression in human skin. Our results suggest that the abnormal production of tropoelastin and fibrillin by heat in human skin and that their degradation by various MMP, such as MMP-12, may contribute to the accumulation of elastotic material in photoaged skin.