Genji Imokawa’s research while affiliated with Utsunomiya University and other places

Little is known about the anti-pigmenting effects of skin-whitening agents on solar lentigos (SLs). To characterize the anti-pigmenting effects of a newly designed derivative ascorbyl glucoside–arginine complex (AGAC) on SLs, lotions with or without 28% AGAC were applied twice daily for 24 weeks in a double-blind half-face study of 27 Japanese females with SLs. The pigmentation scores and skin colors of previously selected SLs on the right and left sides of the faces of the subjects were evaluated using a photo-scale, a color difference meter and a Mexameter. Treatment with the test lotion elicited a significant decrease in pigmentation scores at 24 weeks compared to week 0, with a significant decrease in pigmentation scores at 24 weeks compared to the placebo lotion. In the test lotion-treated SLs, the lightness (L) and melanin index (MI) values that reflect the pigmentation level significantly increased and decreased, respectively, at 12 and 24 weeks of treatment compared to week 0. Comparisons of increased L values or decreased MI values between the test and placebo lotion-treated SLs demonstrated that the test lotion-treated SLs had significantly higher increased L or decreased MI values than the placebo lotion-treated SLs both at 12 and 24 weeks of treatment. The sum of our results strongly indicates that AGAC is distinctly effective in ameliorating the hyperpigmentation levels of SLs at a level visibly recognizable by the subjects, without any hypo-pigmenting effects or skin problems.

Neprilysin is a skin wrinkle‐inducing membrane bound elastase that is expressed abundantly in UV‐exposed and in aged dermal fibroblasts. The overexpression of neprilysin is closely associated with enhanced epithelial‐mesenchymal cytokine interactions mainly via interleukin (IL)‐1α, which has the distinct potential to stimulate the expression of neprilysin by human dermal fibroblasts (HDFs). The over‐expression of neprilysin also accelerates the formation of wrinkles, accompanied by disruptions of the three‐dimensional architecture of dermal elastic fibers that are responsible for the loss of skin elasticity. Because the signaling pathway(s) that lead to the IL‐1α‐stimulated expression of neprilysin in HDFs remain unclear, we characterized the signaling pathway involved, including their related transcription factors, in IL‐1α‐treated HDFs. Since qRT‐PCR analysis revealed that the mRNA expression level of neprilysin is stimulated to a stronger extent in adult HDFs (aHDFs) by IL‐1α than in neonatal HDFs, we used aHDFs for the signaling analysis. Western blotting analysis of the phosphorylation of signaling factors revealed that IL‐1α significantly stimulated the phosphorylation of ERK1/2, RSK, JNK, p38, MSK1, NFkB, c‐Jun, ATF‐2, CREB, and STAT3. Analysis using various signaling inhibitors demonstrated that inhibiting ERK and JNK but not p38, MSK1, NFkB, or STAT3 significantly abrogated the IL‐1α stimulated expression of neprilysin at the mRNA, protein, and enzyme activity levels. Furthermore, silencing c‐Fos significantly down‐regulated the IL‐1α‐increased expression of neprilysin at the protein and enzyme activity levels. These findings strongly suggest that the IL‐1α‐stimulated expression of neprilysin in aHDFs is mediated via the intracellular signaling axis of ERK/JNK/c‐Jun/c‐Fos/AP‐1.

Little is known about anti-pigmenting effects of whitening agents on solar lentigo (SLs). To characterize the anti-pigmenting effects of newly designed ascorbyl glucoside arginine complex (AGAC) on SLs, lotions with or without 28% AGAC were applied twice daily for 24 weeks in a double-blind half-face study of 27 Japanese females with SLs. Pigmentation scores were evaluated using a photo-scale and skin colors were assessed using a color difference meter and a Mexameter. Treatment with the test lotion elicited a significant decrease in pigmentation scores at 24 weeks com-pared to week 0, with a significant decrease in pigmentation scores at 24 weeks compared to the placebo lotion-treated SLs. In the test lotion-treated SLs, the lightness (L) values and melanin index (MI) reflecting pigmentation level significantly (p<0.0001) increased and decreased, respectively, at 12 and 24 weeks of treatment compared to week 0. Comparisons of increased L values or decreased MI values between the test and placebo lotion-treated SLs demonstrated that the test lotion-treated SLs had significantly higher increased L or decreased MI values than the placebo lotion-treated SLs both at 12 and 24 weeks of treatment. The sum of our results strongly indicates that AGAC is distinctly effective in ameliorating the hyperpigmentation levels of SLs at a visibly recognizable level by the subjects without any hypo-pigmenting effects or skin irritation.

We previously reported that a pathogenic abnormality in the barrier and water-holding functions of the stratum corneum (SC) in the skin of patients with atopic dermatitis (AD) is mainly attributable to significantly decreased levels of total ceramides in the SC. That decrease is mediated by the abnormal expression of a novel ceramide-reducing enzyme, sphingomyelin/glucosylceramide deacylase (SGDase), which is the β-subunit (ASAH1b) of acid ceramidase. In this study, we determined whether mice overexpressing ASAH1b in their epidermis develop AD-like skin symptoms. We generated transgenic (TG) mice overexpressing ASAH1b, regulated by the involucrin promoter, to localize its expression in the upper epidermis. After hair removal using a depilatory cream containing glycolic acid, the TG mice without any visible skin inflammation at 8 weeks of age had increased levels of ASAH1b and decreased levels of SC ceramide, with disrupted barrier functions measured by trans-epidermal water loss compared to the wild-type (WT) mice. Interestingly, enzymatic assays revealed that SGDase activity was not detectable in the skin of the TG mice compared to WT mice. Immunological staining revealed that there was an increased expression level of IL-33 in the epidermis and an accumulation of macrophages in the dermis of TG mice compared to WT mice, which are phenotypic characteristics of AD, that were exacerbated by tape-stripping of the skin. In the skin of the TG mice, the mRNA levels of IL-5, CCL11, IL-22, CXCL10, and IFNγ were significantly upregulated compared to the WT mice, and tape-stripping significantly increased the mRNA levels of IL-4, IL-33, CXCL1, CXCL12, TLR9, and CD163 compared to WT mice. These findings strongly indicate that the skin of the depilatory cream-treated TG mice exists in an atopic dry skin condition that is highly sensitive to various environmental stimuli. The sum of our results suggests that ASAH1b itself, even in the absence of its enzymatic activity, is a major etiologic factor for atopic dry skin symptoms via an unknown mechanism.

Lotions with or without 28% ascorbyl glucoside arginine complex (AGAC) were applied twice daily for 24 weeks in a double‐blind half‐face study of 27 Japanese females with solar lentigos (SLs). Pigmentation scores were evaluated using a photo‐scale and skin colors were assessed using a color difference meter and a Mexameter. Treatment with the test lotion elicited a significant decrease in pigment scores at 24 weeks compared to week 0; with a significant decrease in pigment scores at 24 weeks compared to the placebo lotion‐treated SLs. In the test lotion‐treated SLs; the L values and melanin index (MI) significantly (p<0.0001) increased and decreased; respectively; at 12 and 24 weeks of treatment compared to week 0. Comparisons of increased L (△L) values or decreased MI (△MI) values between test and placebo lotion‐treated SLs demonstrated that the test lotion‐treated SLs had significantly higher △L or △MI values than the placebo lotion‐treated SLs both at 12 and 24 weeks of treatment. The sum of our results strongly indicates that AGAC is distinctly effective in ameliorating the hyperpigmentation levels of SLs at a visibly recognizable level by the subjects without any hypo‐pigmenting effects or skin irritation.

Horse-derived ceramide (HC), which contains galactosylceramides as its main component, significantly improves skin symptoms when applied topically to patients with atopic dermatitis. We speculated that efficacy resulted from the amelioration of epidermal ceramide metabolism, and we characterized those effects using reconstructed human epidermal equivalents. Lipid analysis, RT-PCR and Western blotting revealed that HC significantly increased the total ceramide content of the stratum corneum (SC), accompanied by significantly increased gene and/or protein expression levels of ceramide synthase (CERS) 3, fatty acid elongase (ELOVL) 4, glucosylceramide synthase (GCS), β-glucocerebrosidase, sphingomyelin synthase and acid sphingomyelinase. Mechanistic analyses using cultures of primary human keratinocytes revealed the marked stimulatory effects of HC on the mRNA expression levels of CERS3, ELOVL4 and GCS under high calcium-derived differentiation conditions. Signaling analyses demonstrated that an antagonist of PPARβ/δ significantly abrogated the HC-stimulated mRNA expression levels of GCS, CERS3 and ELOVL4. GW9662, an antagonist of PPARγ, significantly abolished the HC-up-regulated mRNA expression levels of GCS and ELOVL4, but not of CERS3. These findings suggest that HC has the distinct potential to accentuate the expression of GCS, CERS3 and ELOVL4 via the activation of PPARβ/δ and/or PPARγ to accelerate ceramide synthesis in the SC.

Because ceramide-like lipo-amino acid cholesteryl derivatives can exert a bound water-holding function due to their lamellae-forming properties, in this study, we determined if topical application of those derivatives to atopic dry skin would elicit an ameliorative effect on skin symptoms, at least on its water-holding function. In this clinical study, daily treatment with a nano-emulsion containing 10% phytosteryl/octyldodecyl lauroyl glutamate (POLG) significantly (p < 0.0001) improved skin symptoms, including dryness/scaling, itchiness and stimulus sensations, in the non-lesional skin of patients with atopic dermatitis (AD) at 3 and at 6 weeks compared with week 0. Those significant improvements in skin symptoms were accompanied by a significantly enhanced water content (conductance) and a significant improvement of roughness (SESC) and smoothness (SESM) values measured using a Visioscan at 3 and 6 weeks. Those effects appeared concomitant with a significantly increased corneocyte size, a significantly down-regulated degree of thick abrasions, and a significant impairment of the corneocyte lipid envelope at 6 weeks. Thus, our clinical study suggests, for the first time, that topical application of the POLG nano-emulsion has the distinct potential to ameliorate atopic dry skin symptoms, particularly scaling and itchiness, in the skin of patients with AD. Those effects result from alleviation of the disrupted water-holding function probably due to the increased supply of lamellae structures into the stratum corneum despite the failure to improve barrier function.

β-Sitosterol 3-O-d-glucoside (BSG) is known to act as an agonist by binding to estrogen receptors, and estrogen has been reported to enhance the activity of β-glucocerebrosidase, an epidermal ceramide metabolizing enzyme. In this study, we determined whether BSG up-regulates ceramide levels in the stratum corneum (SC) of a reconstructed human epidermal keratinization (RHEK) model. Treatment with BSG significantly increased the total ceramide content by 1.2-fold compared to that in the control in the SC of the RHEK model, accompanied by a significant increase of the ceramide species, Cer[EOS] by 2.1-fold compared to that in the control. RT-PCR analysis demonstrated that BSG significantly up-regulated the mRNA expression levels of serine palmitoyltransferase (SPT)2, ceramide synthase (CerS)3, glucosylceramide synthase (GCS) and acid sphingomyelinase by 1.41–1.89, 1.35–1.44, 1.19 and 2.06-fold, respectively, compared to that in the control in the RHEK model. Meanwhile, BSG significantly down-regulated the mRNA expression levels of sphingomyelin synthase (SMS)2 by 0.87–0.89-fold. RT-PCR analysis also demonstrated that BSG significantly up-regulated the mRNA expression levels of CerS3 and GCS by 1.19–1.55 and 1.20-fold, respectively, but not of SPT2 and significantly down-regulated that of SMS2 by 0.74-fold in HaCaT keratinocytes. Western blotting analysis revealed that BSG significantly increased the protein expression levels of CerS3 and GCS by 1.78 and 1.28–1.32-fold, respectively, compared to that in the control in HaCaT cells. These findings indicate that BSG stimulates ceramide synthesis via the up-regulated expression levels of CerS3 and GCS in the glucosylceramide pathway, which results in a significantly increased level of total ceramides in the SC accompanied by significantly increased levels of acylceramide species such as Cer[EOS].