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A nine-layered skin tissue model is newly developed for the Monte Carlo simulation of spectral reflectance. The derivation of the necessary parameters for each of the nine layers in the simulation is presented, in which the parameters used in the conventional three-layered model are modified on the basis of some histological findings on skin and reported examples. Using appropriate optical and geometrical parameters, simulated spectra can be produced that agree well with measured spectra. This approach provides a flexible means of spectral fitting to measured results and of estimating changes in the parameters of skin tissue.

Background/purpose: It is crucial to establish an accurate method for measuring skin pigmentation in cosmetic science and clinical dermatology. Here, we report a non-invasive precise method for measuring skin melanin content. Methods: In order to determine the concentrations of melanin and hemoglobin in skin, we used the spectrum resolution (SR) method. In brief, the absorption spectrum of the skin was calculated from the reflection spectrum using a spectrophotometer. The concentrations of melanin and hemoglobin were then determined using a multiple regression analysis, assuming that the absorption spectrum of the skin is expressed as a linear summation of the absorptions of melanin and hemoglobin according to the Lambert–Beer law. The skin changes in the volar forearm, which had been irradiated by ultraviolet rays (UV), were observed daily by the SR method. Results: A multiple regression analysis with an absorption spectrum of 500–700 nm was performed. The multiple correlation coefficient was 0.993, resulting in a satisfactory precise estimate of the concentrations of melanin. After UV irradiation, the concentration of melanin monitored by the SR method increased until 8 days and decreased gradually afterwards. Conclusions: The SR method allows the evaluation of the changes of epidermal melanin induced by UV irradiation.

Stratum corneum thickness was estimated from water concentration profiles of the skin measured by a confocal Raman spectrometer. Stratum corneum apparent thickness (SCAT) was defined as the depth where the water content reached an almost constant value. Site variations were determined using 15 healthy Japanese subjects (6 males, 9 females), and age variations at the cheek and forearm were examined using 27 female Japanese subjects. There were marked site variations in mean SCAT; 16.8 microm for cheek, 22.6 microm for volar forearm, 29.3 microm for back of the hand, and 173.0 microm for palm. These variations were similar to reported stratum corneum thickness values obtained in biopsy tissues. The SCAT tended to become age-dependently thicker at the forearm, but not at the cheek. In addition, SCAT was increased up to two-fold by hydration for 90 min, while lesser increases were seen with shorter hydration periods.

皮膚の生理学的性質を非侵襲的に測定する方法は皮膚計測工学の分野において数多く開発されてきた．それらは角層水分量，肌理，TEWL（経表皮水分蒸散量），皮膚色，血流，皮脂などであるが，最近，無侵襲で皮膚内部を観察する生体顕微鏡が開発され，物質の経皮吸収測定に活用されている．本稿では，in vivo共焦点ラマン顕微鏡と多光子顕微鏡を用いた測定例について述べる．

Background/aims: Recently, there has been increasing awareness regardless keeping nails in good condition. Maintenance of an optimum water level in the nail plate is very important since water content affects the nail condition. However, a simple method for in vivo measurement of water in the nail plate quantitatively has not been reported. In this study, we applied Near infrared spectroscopy (NIR)-based prediction of water content of the cut nail plate to in vivo measurement to obtain information about the seasonal change of water in the fingernail. Methods: Diffused reflectance spectra of the nail plate of the fingernail were collected in the NIR region with an NIR spectrometer in vivo both in summer and winter from the same 15 Japanese female subjects. The water content of the nail plate in vivo was calculated from the NIR spectrum using partial least square regression analysis. Results: It was possible to apply NIR-based prediction of water content of the cut nail plate to in vivo measurement. We also found that the water content of the nail plate varied from person to person but not with the finger of the individual. The average water content of the nail plate was significantly lower in winter than in summer (P

Chemical peeling with salicylic acid in polyethylene glycol vehicle (SA-PEG), which specifically acts on the stratum corneum, suppresses the development of skin tumors in UVB-irradiated hairless mice. To elucidate the mechanism through which chemical peeling with SA-PEG suppresses skin tumor development, the effects of chemical peeling on photodamaged keratinocytes and cornified envelopes (CEs) were evaluated in vivo. Among UVB-irradiated hairless mice, the structural atypia and expression of p53 protein in keratinocytes induced by UVB irradiation were intensely suppressed in the SA-PEG-treated mice 28 days after the start of weekly SA-PEG treatments when compared to that in the control UVB-irradiated mice. Incomplete expression of filaggrin and loricrin in keratinocytes from the control mice was also improved in keratinocytes from the SA-PEG-treated mice. In photo-exposed human facial skin, immature CEs were replaced with mature CEs 4 weeks after treatment with SA-PEG. Restoration of photodamaged stratum corneum by treatment with SA-PEG, which may affect remodeling of the structural environment of the keratinocytes, involved the normalization of keratinocyte differentiation and suppression of skin tumor development. These results suggest that the stratum corneum plays a protective role against carcinogenesis, and provide a novel strategy for the prevention of photo-induced skin tumors.

Diffuse reflectance (DF) spectra in the 1250–2500 nm region were measured in vivo for the skin of the forehead, cheek, jaw, elbow, volar forearm, palm, knee, and heel of seven healthy volunteers, using a Fourier transform near-infrared (FT-NIR) spectrophotometer with a fiber-optic probe. Apparent regional differences of water content in the skin, as estimated from the diffuse reflectance NIR spectra, are discussed in relation to the influence of measurement depth. The NIR spectra were collected with or without a 300 μm gap between the fiber-optic probe and the skin surface. For comparison, in vitro NIR spectra of stratum corneum sheets equilibrated at 41, 50, 63, and 81% relative humidity, at 25 °C, were also obtained. There was a difference in the ratio of the two water bands centered near 1450 nm and 1900 nm between the contact and non-contact measurements. In addition, regional differences of water content calculated from the peak height of the 1900 nm water band, which was normalized to the peak height of the 2175 nm amide band, were compared. The results of Monte Carlo simulation indicated that the apparent regional differences arise at least in part from differences in the measurement depth due to differences in specular reflection at the skin surface and in the thickness of the stratum corneum.

頸部は顔面に次いで露出されることも多く光老化が現れやすい部位と考えられるが, その皮膚生理に関する研究知見はほとんど報告されていない。そこで, われわれは18-69歳の女性61名を対象に頸部皮膚生理を測定し, 顔面との比較および加齢変化について解析を行った。その結果, 頸部は顔面に比べ角層水分量は多く, TEWLは低く角層バリア能は高かった。角層細胞面積は大きく, 角層ターンオーバーは遅いことが示された。また, 皮膚厚は薄く, 皮膚伸展性が高く, 皮溝は深く均一であった。顔面よりも少ないが, 頸部角層細胞中にもゼラチナーゼ活性が認められ, 日光の影響を受けていることがわかった。加齢変化に関しては, 顔面と同様に弾力性の低下が認められるが, 皮溝の異方性が増加し, 頸部特有の加齢現象を呈することがわかった。頸部は露光部である顔面と非露光部である体幹の間を示している皮膚生理パラメータが多く, 自然老化と光老化の影響を受けていることが示唆された。この結果を基に設計した首専用化粧品のプロトタイプ化粧品について4週間の連用テストを実施した結果, 連用前後で角層水分量は有意に増加し, ハリや皮膚の明るさ向上に高い実効感があった。

Transparency of the skin is an important factor for realizing beautiful skin, but there is only a visual sensory evaluation method by a beauty specialist. It is unclear how skin physiological characteristics can give transparency. Our purpose is to establish an objective method for evaluating the transparency of the skin and to develop a special cosmetics product which can enhance skin transparency, after investigating the mechanism of the skin characteristics which yield transparency. We developed an optical instrument that could independently measure a diffuse reflection as well as a specular reflection by assembling a polarizing filter in a gonio-spectrum photometer. We analyzed the relationship among skin transparency, specular reflection light and diffuse reflection light. It was shown that there was a strong positive correlation between the skin transparency and the intensity of the diffuse reflection light (r = 0.699) and no correlation between the skin transparency and the intensity of the specular reflection light (r = 0.190). The results of partial least squares (PLS) analysis of the skin transparency and the skin physiological characteristics showed that a highly transparent skin has a fine skin surface texture with a high hydration of stratum corneum, a low melanin content and a low hemoglobin content. On the basis of these results, we developed an essence that can improve skin texture and blood flow, reduce melanin content, and increase the moisture in the stratum corneum. After eight weeks’ usage of this product by consumer panels (n = 30), it was confirmed that the skin transparency was improved by the increase of the intensity of the diffuse reflection light by gonio-spectrum photometer measurement and by consumers’ self-assessment.

In vivo visualization of epidermal melanin was performed by reflectance-mode confocal microscopy (RCM). Firstly, we examined the distribution of epidermal melanin in pigmented animals and compared with that of the human skin. Melanocytes in the skin of pigmented animals were found to accumulate a large amount of melanin that can be easily visualized because of its brightness. Their RCM images correlated well with the Fontana-Masson-stained sections for melanin. In contrast, in the human skin, typical dendritic melanocytes were hardly observed even in pigmented lesions, although supranuclear melanin caps were easily visible. These results suggested that human melanocytes rapidly transfer the produced melanin to keratinocytes and do not accumulate it. Secondly, to elucidate the production of melanin by human melanocytes, we evaluated the changes of melanin after a single ultraviolet (UV) exposure. The melanin-accumulating melanocytes were clearly visualized during the skin pigmentation process. The RCM images showed the brightness because of melanin gradually increased from day 4, then dendrite-elongated melanocytes appearing from day 8, and finally melanin caps formed from day 29. In conclusion, RCM successfully evidenced the difference in melanin distribution between the pigmented animals and humans, and the UV-induced pigmentation process in vivo as well.

美容上問題となる「目の下のくま」の発生要因として, 一般的に血流の停滞が言われているが, 実際にくまと血流の関係について調査した報告は少なく, また, くまについて皮膚生理学的に論じた報告もほとんどない。そこで本研究では, 非侵襲的な方法を用いくまを皮膚科学的に解析し, その要因を明らかにするとともに, その対応を考え, 「目の下のくま」改善効果の高い商品の開発を行った。くまの発生要因を明らかにするために, くまのある女性とない女性を対象に, 眼下部の皮膚メラニン量, ヘモグロビン量およびヘモグロビン酸素飽和度, 血流速度の計測を実施した。くまのある女性の眼下部では, ヘモグロビン量の増加およびヘモグロビン酸素飽和度の減少が観察された。くま部位では頬と比較して血流速度の低下が見られたことから, 上記結果は血流の停滞により引き起こされたと考えられる。またメラニン量の増加も同時に観測され, その傾向は高年齢層でより顕著であった。以上の結果から, くま部位の明度低下は, 血流速度の低下による皮膚毛細血管内の還元ヘモグロビンの増加と, 皮膚メラニン量の増加によるものと考えられた。以上の検討をもとに, くま発生の主要因である「鬱血」および「色素沈着」, くまを目立たせる小じわ, キメの悪化の各々に対処した「血行促進剤」「美白剤」「保湿剤」配合プロトタイプ製剤を処方し, 3週間の連用効果試験を実施した。その結果, 美容技術者による目視評価, 被験者のアンケートにおいてプロトタイプ製剤の, 高いくま改善効果が確認された。また, 機器測定によってもヘモグロビン酸素飽和度の上昇とメラニン量の低下が確認され, 「くま」の改善に本処方は効果的であることが示された。

We observed the internal structure of an oil/wax/water/surfactant system using a scanning electron microscope to investigate the relationship between its hardness and state of wax crystal. The molten wax (hydrogenated jojoba oil, ceresin, polyethylene wax, carnauba wax, or microcrystalline wax) was cast in a home-made spiral mold of aluminum foil for preparing the test specimen for SEM observation. In hydrogenated jojoba oil a fine frame-like crystal structure, the so-called ‘card-house structure,’ was observed but not in other waxes. The mixture of hydrogenated jojoba oil and water showed a few small droplets deposited on the roundish wax frame-like structure. On the other hand, waxes other than hydrogenated jojoba oil did not change their internal structure when they were mixed with water. This result suggested that hydrogenated jojoba oil showed uniquely high affinity for water. In the system of oil, water, surfactant, and various kinds of waxes, their crystal structure, hardness, and the shape of dispersed water particles were remarkably changed with the combination of waxes. In the system with ceresin and carnauba wax, the hardness measured by a card-tension meter was high, and the internal crystal structure was fine and amorphous. The water particle in the ceresin and carnauba wax system had a smaller diameter than that in the system containing hydrogenated jojoba oil. The system containing hydrogenated jojoba oil showed a card house-like wax crystal structure without high hardness. It was considered that the wax crystal structure played an important role in providing hardness and in contributing to the water distribution in the oil/wax/water/surfactant system.

Analysis of the dermo-epidermal surface in three-dimensions has great value in evaluating cosmetics. One approach is based on the active contour model, which is used extensively in computer vision and image processing applications, particularly for local object boundaries with closed curve form. The dermo-epidermal surface, however, is a plane with open form. We have developed a method of automatically extracting the dermo-epidermal surface from volumetric confocal microscopic images, as well as constructing a 3D visual model of the surface by using the geometric information contained in the control points. Our method is a 3D extension of the active contour model, so we call it the active open surface model (AOSM). The initial surface for AOSM is an open curve plane, guided by a 3D internal force, a 3D external constraint force, and a 3D image force, which pull it towards the objective surface. The proposed tecnique has been applied to extract actual dermo-epidermal surface in the given volumetric confocal microscopic images.

We have previously established a non-invasive method to evaluate the maturity of cornified envelopes (CEs), and have reported the appearance of immature CEs in the stratum corneum (SC) with poor barrier function, such as the SC of the face. The purpose of the present study was to evaluate CEs in inflammatory skin disorders, and to clarify the relationship between the appearance of the immature CEs and parakeratosis, which is often used as a marker for defective keratinization in inflammatory skin disorders. Cornified envelopes in the outermost SC of involved areas of psoriasis vulgaris (PV) and atopic dermatitis (AD) were strikingly heterogeneous, and consisted of immature CEs stained with anti-involucrin and mature CEs stained with Nile red, whereas CEs of the uninvolved areas were relatively homogeneous, exhibiting mature phenotype. The ratio of immature CEs was significantly higher in the involved areas of PV and AD than that in the corresponding uninvolved areas, suggesting that defective CE maturation may, at least in part, account for the inflammatory disorders. Simultaneous evaluation of CE maturity and parakeratosis was carried out by a combination of involucrin immunostaining and nuclear staining of detergent-dissociated corneocytes. In the involved area of PV, four types of corneocytes in regard to the combination of involucrin staining and nuclear remnant were observed, while both immature CEs and parakeratosis were more often detected in the involved areas of PV than in the uninvolved areas or the upper arm of healthy subjects as a normal control. Thus, corneocytes with involucrin-positive immature CEs were not always associated with parakeratosis at the cellular level. In the involved areas of PV, the ratio of immature CEs and that of parakeratosis were heterogeneous, depending on the cases, and no correlation between the ratios was observed. Inter-individual and intraindividual variations in CE maturity were also suggested by the heterogeneous localization of involucrin in the psoriatic epidermis as examined by immunohistochemistry. In addition, in the face of healthy subjects, four types of corneocytes were similarly detected, and the ratio of immature CEs was significantly higher than that of parakeratosis. These results obviously suggest that the maturation of CEs and disappearance of nuclei are differentially regulated in the epidermis.

The water content of human nail plates was determined using a portable near-infrared (NIR) spectrometer with an InGaAs photodiode array detector. NIR diffuse reflectance (DR) spectra were collected from 108 cut nail plates with different relative humidity and in vivo from fingernails. Partial least-squares (PLS) regression was applied to the NIR spectra in the 1115–1645 nm region to develop calibration models that determine the water content in the cut nail plates and fingernails. A good correlation was obtained between the NIR spectra and the water content measured by nuclear magnetic resonance (NMR) for the NIR measurement of both cut nail plates and fingernails. The results indicate that the water content in the nails can be determined very rapidly (1 s) by means of the portable NIR spectrometer and PLS regression.

官能評価は, 化粧品を開発する上で重要な評価項目である。しかしながら, 評価者の好みや体調, 心理状態によって変動することも多く, 客観的で再現性の良い機器による評価が望まれている。これまでにわれわれは, ヒト皮膚表面でのすべり摩擦測定を行い, 摩擦係数と皮膚生理パラメータとの関連性を検討してきた。本報告では, 化粧品塗布後の皮膚表面摩擦と官能評価との関係を検討することを目的とした。実験は, 市販の表面摩擦測定機器, KES-SETM (カトーテック (株)) にアームホルダーを取り付けて行い, 摩擦特性値として, 平均摩擦係数 (MIU) とその平均偏差 (MMD) を用いた。皮膚摩擦特性に大きく影響する角層水分量はComeometer CM 825TMを用いて測定した。使用感触の異なる9種類の乳液を用いて, 塗布直後, 1.5, 4, 5.5, 7時間後のMIU, MMDを測定し, 官能評価は6名の専門家パネルおよび149名の一般パネルにより行い, 機器測定の結果と対応させた。その結果, MIUと「しっとり」「べたつき」とは正の相関, 「つるつる感」との間には負の相関が認められた。さらに, 化粧品塗布直後からのMIUの経時変化のパターンは, 化粧品のなじみの良さと関係あることを見出した。また, MIUとMMDの値から「油っぽさ」と「しっとり」を分けて評価できることがわかった。以上の結果から, 本機器は, 化粧品を皮膚に塗布した後の感触評価に活用可能であることが示された。

The surface contours of lesional skin of certain skin diseases, such as parapsoriasis en plaque (PEP) and extramammary Paget's disease (EMPD), in which there is a massive infiltration by non-epidermal cells, looks somewhat different from that of the adjacent normal skin, needless to state that they are apparently different from that of ordinary chronic inflammatory dermatoses where we found acanthotic epidermis accompanied by hyperkeratosis. We attempted to objectively characterize these unique skin surface changes qualitatively and quantitatively using non-invasive methods. Negative impression replicas were taken from the lesional skin of patients with EMPD or PEP as well as from the adjacent uninvolved skin. The findings were confirmed histologically. The replicas were examined by using computerized image analysis. Several parameters were analyzed that correlate with the changes in the anisotropy of the skin furrows (VC1), average skin roughness (KSD), average length of skin furrows (LEN), and number of skin furrows (NUM). There were significant decreases in KSD and NUM in EMPD, indicating a smoother skin surface in the lesional skin than in the adjacent normal skin. In contrast, the PEP lesion had an increase in VC1 and LEN and a decrease in NUM, which suggests larger skin ridges in the lesional skin than in the uninvolved skin. Thus the unique skin surface of the cutaneous disorders accompanied by epidermal invasion by non-epidermal cells, such as EMPD and PEP, was characterized both qualitatively and quantitatively using computerized image analysis of negative impression replicas.

Background/aims: Sensory evaluation is an important factor for cosmetic products. Several devices for the measurement of sensory properties have been developed in recent years. The objective here is to measure skin surface friction using these devices and examine the correlation with other physiological parameters in order to evaluate the potential of physical measurement of tactile sensation. Methods: A KES-SE Frictional Analyzer, a commercial device for measurement of surface frictional characteristics, was used in this study. An arm holder was added to this device for measurement on the human forearm. The frictional coefficient (MIU) and its mean deviation (MMD) were used as the parameter to indicate surface friction. The moisture content in the stratum corneum was measured with a Corneometer CM825, the transepidermal water loss with a Tewameter TM210, the viscoelastic properties of the skin with a Cutometer SEM575 and the skin surface pattern by observing the negative replica using silicon rubber. Results: The MIU was not influenced by load; however, it was increased due to water application on the skin. The relationship between MIU and the moisture content in the stratum corneum, between MMD and skin surface pattern and between MMD and viscosity of both normal human forearm skin and SDS (sodium dodecyl sulfate)-induced dry skin were confirmed by statistical analysis in a test on human subjects. There was also a correlation between either MIU or MMD and sensory evaluation in the morning after the application of moisturizing products. Conclusion: Human skin surface friction was measured by using a KES-SE Frictional Analyzer. Judging from the correlation between either MIU or MMD and sensory evaluation, we considered this instrumental analysis to be useful for evaluating the tactile impression of human skin.

2001年4月より化粧品の効能が拡大され, ファンデーション類でもスキンケア効果を標榜することができるようになり, これまで以上に有用性評価法・技術に関心が高まっている。一方, 有用性評価と密接に関係する皮膚計測工学の進歩は著しく, 非侵襲的に皮膚内部構造や細胞形態まで観察できるようになってきた。本稿においては最近, 開発された技術のうち, 非接触キメ・シワ測定法, 美白評価に活用できる共焦点レーザー顕微鏡, 角層厚や表皮-真皮ジャンクションを調べる光干渉断層撮影法 (Optical Coherence Tomography: OCT), 角層水分の分布状態を調べる共焦点ラマン顕微鏡を取り上げ, その技術について概説するとともに化粧品の有用性評価への可能性を述べる。

皮膚観察用のin vivo共焦点顕微鏡が開発され, ヒト皮膚内部構造に関して非侵襲的に細胞レベルでの検討が可能になってきた。われわれは本測定機器 (Vivascope 100, Lucid社, USA) を導入し, 健常人を対象に皮膚深さ方向における構造・細胞形態変化の観察, SDS (Sodium Dodecyl Sulfite) やテープストリッピングによって惹起された実験的肌荒れ, 露光部位・非露光部位による表皮厚, 表皮-真皮界面の凹凸状態 (アンジュレーション) の変化について検討した。表皮突起に関しては得られた皮膚水平断面像を積み重ね三次元立体構築を行い, 縦断層像を作成し, Snakes法を用いて表皮-真皮界面を抽出した。その結果, 皮膚深さ方向において細胞形態は大きく変化することがわかった。基底細胞の大きさは約100μm2程度で角層表面での大きさは約1200μm2と分化・角化とともに細胞は偏平化し, 10倍以上大きくなることが示された。SDS, テープストリッピング処理によって表皮は著しく肥厚し (2倍以上), その影響は25日後でも認められた。一方, 表面形態は25日後ではほとんど処理前の状態に戻っていた。このように外的刺激に対して角層表面での回復は比較的早いものの, 皮膚内部への影響はそれよりも長く続くことが示唆された。露光部位 (頬, 前腕外側), 非露光部位 (上腕内側) における表皮厚, 表皮突起の状態を調べた結果, 頬では他の部位に比べ著しく表皮が肥厚しており, 表皮-真皮界面の凹凸が激しいことがわかった。以上の結果からin vivo共焦点顕微鏡はこれまで困難であった皮膚内部構造を非侵襲的に捉えることができ, 化粧品の有用性実証に役立つ機器と考えられた。

Cornified envelopes (CEs), rigid and insoluble structures in the stratum corneum, which are assembled by crosslinking of several precursor proteins by transglutaminases, provide a hydrophobic foundation for barrier function; omega-hydroxyceramides are covalently attached to the outer surface of CE components, and onto this hydrophobic assembly, lamellar layers of intercellular lipids are organized. Morphologically irregular, fragile CEs are found in the deep layer of the stratum corneum or in certain disorders, such as psoriasis, whereas most CEs from healthy subjects are rigid and polygonal. We have established a staining method to characterize such fragile CEs as immature and less hydrophobic CEs, and employed it to examine regional differences in the properties of CEs, especially in relation to the barrier function of the skin. CEs from the outermost stratum corneum of the trunk and extremities of healthy subjects were relatively uniform in morphology with larger shape, and were homogeneous in hydrophobicity as judged from the use of an environment-sensitive fluorescent dye, Nile red. However, CEs from the face were strikingly heterogeneous, and consisted of both rigid and fragile CEs. Rigid CEs were Nile red-positive and little stained by anti-involucrin. In contrast, fragile CEs were Nile red-negative but strongly stained with anti-involucrin, as detected by indirect immunofluorescence. Thus, CEs from the face were stained with Nile red or involucrin in a mutually exclusive manner. Fragile CEs were stained with antibodies against other CE components, including loricrin, envoplakin, filaggrin, and isopeptides. Such fragile, involucrin-positive CEs were detected not only in the face, but also in the deep layer of the stratum corneum of the arm. In addition, experimental barrier disruption resulted in the appearance of involucrin-positive CEs in the outermost stratum corneum. These results suggest that involucrin-positive, fragile CEs are immature and less hydrophobic, and that their occurrence is closely related to impairment of the barrier function of the skin.

皮膚の物理特性や生理特性を非侵襲で計測するために種々の機器が考案されており, 力学的, 電気的, 温熱的, 光学的, 音響的特性などの物性測定から発汗, 皮脂分泌, 血流, 代謝活性などの生理機能測定まで広範囲にわたっている。これらの諸特性は肌状態を調べたり, 肌質の分類や化粧品の有用性評価に応用されている。ここでは, スキンケア商品の有用性評価においてよく用いられる測定技術のうち皮膚表面形態, 角層水分量, 角層バリアー機能, 角層ターンオーバー, 不全角化度, 角層中遊離アミノ酸, しわ, しみなどの測定法, さらには皮膚内部の構築状態について測定する超音波断層撮影, in vivo共焦点顕微鏡などの測定法について原理・応用面から述べる。

来るべき高齢化社会において, 化粧品や医薬部外品は重要な役割を果たすと考えられているが, 我々は皮膚老化を防御する戦略として, 乾燥防御, 紫外線防御に加えて酸化防御を提唱してきた。本報告では, 特に酸化防御の重要性について論じた。今までに著者らは, CL-HPLCシステムを用いた皮表における脂質過酸化反応の検討から, ヒト皮膚においては一重項酸素がキーとなっている活性酸素種であることを報告してきたが, 本報告において, 皮表脂質とヒポタウリンの一重項酸素との反応速度定数を測定して明らかにした. さらに光酸化条件下において, ヒト皮表において第一の酸化ターゲットであるスクアレンの酸化防御効果をヒポタウリンが発現することを明らかにした。一方ヒポタウリンは, 哺乳類において含硫アミノ酸の代謝産物の一つであることが知られており, 皮膚, 眼球, 肝臓, 脳, 心臓, 血清中等でその存在が認められている。またその酸化ストレスへの生体防御機構としての存在意義も議論されている。皮膚において生体の酸化防御機構を再構築, 強化する有用性を化粧品における新たなコンセプトとして提案した。

Cosmetics and quasi-drugs will play new important roles in the aging and stressful society. We have proposed 3 clinical concepts, i.e., anti-drying, anti-UV radiation and anti-oxidation to prevent the skin aging. Especially, the importance of anti-oxidation is discussed in this paper. The peroxidation of human skin surface lipids has been investigated by a CL-HPLC (chemiluminescence-high performance liquid chromatography) system. Squalene monohydroperoxide was produced at the forehead and the scalp as well as in the dandruff under daily life conditions in most of the cases. Therefore it was considered that squalene was the first target lipid on a human skin surface by an oxidative stress. In order to confirm the effect of hydroperoxides of the skin on cultured cells and the living skin equivalent, an authentic sample was prepared. We have also clarified the reaction rate constants of the skin surface lipids with singlet oxygen and free radicals. And we have also confirmed that the application of cosmetics and the ingestion of aliments containing antioxidants had a preventive effect against the lipid peroxidation on skin surface.

Sphingolipids are funcitionally important in the barrier function of the human stratum corneum. Free amino acids also play an important role in its water retention properties. To increase our understanding of the way in which these substances affect scaly skin, we investigated the change of sphingolipids and free amino acids in scaly skin induced by a surfactant, sodium lauryl sulfate (SLS). One week after the treatment, the total amount of amino acids was decreased and their composition was changed significantly. However, the total amount of sphingolipids did not differ from that in control skin. The order of the stratum corneum (SC) intercellular lipids evaluated by attenuated total reflection infrared spectroscopy (ATR-IR) did not change at all, and only the composition of five sphingolipids species changed significantly. These findings suggest that the abnormal morphology of the SC itself and the decline of amino acids are responsible for the dryness of the scaly skin.

Background The photo-aged facial skin is characterized by various unique features such as dark spots, wrinkles, and sagging. Elderly people, particularly Asians, tend to show a yellowish skin color change with photo-aging. However, there has been no analytical study conducted on this unique skin color change of the aged facial skin. Objective The purpose of the present study is to examine whether the carbonyl modification in the dermal protein is involved in the yellowish color change that occurs in the photo-aged skin. Methods Normal skin samples excised from the face, abdomen and buttock of variously aged Japanese were separated into the epidermal and the dermal portions. These skin samples were histologically examined for carbonyl modification. Moreover, an in vitro constructed dermis model composed of a contracted collagen gel was treated with acrolein or 4-hydroxynonenal. All these samples were also studied colorimetrically. Results The dermal samples obtained from the photo-aged facial skin exhibited an appearance of yellowish color, whereas neither the facial epidermis nor the dermis obtained from the abdomen or buttock showed such a yellowish discoloration. The upper layer of the dermis that revealed the yellowish color showed elastosis whose elastic fibers were found to colocalize with carbonyl protein as detected by a labeled hydrazide, as well as by an immunohistochemical examination using the antibody against acrolein adduct. Experimental induction of carbonyl modification in a dermis model in vitro by a long-term treatment with acrolein or 4-hydroxynonenal was found to show the appearance of the yellowish change which was also proven by an increase in b* value of colorimetry. It was more pronounced than that induced by glycation. Conclusion Our present results strongly suggest that carbonyl modification of the dermal protein is involved in the production of the yellowish color change that is noted in the photo-aged facial skin.

The peroxidation of human skin surface lipids was investigated by a CL-HPLC (chemiluminescence-high performance liquid chromatography) system. Squalene mono-hydroperoxide was produced at the forehead and the scalp as well as in the dandruff under daily life conditions in most of the cases. Therefore it was considered that squalene was the first target lipid on a human skin surface by an oxidative stress. In order to confirm the effect of squalene monohydroperoxide on proliferation of cultured cells and TESTSKINTM (Living Skin Equivalent), an authentic squalene mono-hydroperoxide was prepared by a dye-sensitized photooxidation and its chemical structure was identified by 1H-NMR, 13C-NMR and FAB-MS. Detection limit of squalene mono-hydroperoxide in CL-HPLC system was as little as 1 pmol. We also found that cosmetics containing antioxidants had a preventive effect against the lipid peroxidation on a Human Skin Surface.

The aim of this study was to determine differences in the functional properties of the stratum corneum of children and adults, focusing on the influence of approaching puberty. Biophysical measurements were made of the stratum corneum of 32 healthy Japanese children aged 10–14 years and their mothers in summer and the following winter. The children showed significantly lower skin surface hydration. Stratum corneum barrier function, evaluated in terms of trans-epidermal water loss, was poorer on the forearm in the children than in the adults regardless of season. By contrast, the stratum corneum barrier of the cheek, which was better in the children, tended to become poorer when the children reached puberty. Although the immaturity of the cornified envelopes of the superficial corneocytes, which ratio increased significantly in winter, was not different from that of adults, the corneocytes were significantly smaller in the children, suggesting a more rapid turnover of the stratum corneum. The amount of skin surface lipid, which was measured only on the cheek, remained low until 13 years of age, but at 14 years of age it increased remarkably, approaching adult levels. We conclude that, until puberty, most functional characteristics of the skin of children remain distinct from those of adults. Key words: children; puberty; skin surface hydration; skin surface lipids; trans-epidermal water loss.

In the case of protein analysis using reversed-phase high-performance liquid chromatography(RP-HPLC), it became evident that metal impurities in silica gel give great influence on peak shapes of protein. High metal impuritie resulted in poor column efficiency. The influence of metal impurities was found larger than that of residual silanols.

We studied the change in skin thickness of forehead and cheek with age for healthy male and female subjects using ultrasound method. Skin thickness decreased with ageing in forehead and cheek in both sex groups, and it was thicker at every age group in males than in females.We also measured the skin thickness from twelve different sites at face in six healthy subjects. It was shown that skin thickness of eyelid was thinner than that of jaw or cheek.

Background/purpose: Stratum corneum (SC) is the interface of body and environment and is continuously exposed to oxidative stress, resulting in oxidative modification of proteins. Consequent carbonylated proteins (CPs) have so far been labeled with 2,4-dinitrophenyl (DNP) hydrazine and subsequently detected with anti-DNP antibody. We developed a simpler, non-invasive method to assess CP level in the SC and applied it to following research. Methods: SC was collected by adhesive tape stripping and its carbonyl groups were labeled with fluorescein-5-thiosemicarbazide (FTZ). The staining image was observed by fluorescence microscopy and the average fluorescence intensity of the SC extracted from the image was calculated as stratum corneum carbonylated protein (SCCP) level. Results: By reaction with FTZ, carbonyl groups in the SC could be detected easily. Relatively strong fluorescence was observed in exfoliating scales. Lipid removal from the SC in vitro or in vivo did not show any change in fluorescence intensity, suggesting that carbonyl groups were mainly derived from proteins, not from lipids. SCCP level was higher in the upper layer than the lower layer, and higher in the cheek (sun-exposed) than the inside of upper arm (unexposed), positively correlated with age especially in male cheek, positively correlated with transepidermal water loss, negatively correlated with water content, and showed a subtle correlation with sebum level. On the other hand, SC collected by cyanoacrylate resin and labeled with FTZ revealed strong fluorescence around the pores in the cheek and on the grooves in the upper arm, suggesting the role of sebum in the generation of SCCP. Conclusion: SCCP was assessed in a simple and non-invasive method, and suggested to be a novel indicator that reflects some aspect of skin condition.

We have previously established a non-invasive method to evaluate the maturity of cornified envelopes (CEs), and have reported the appearance of immature CEs in the stratum corneum (SC) with poor barrier function, such as the SC of the face. The purpose of the present study was to evaluate CEs in inflammatory skin disorders, and to clarify the relationship between the appearance of the immature CEs and parakeratosis, which is often used as a marker for defective keratinization in inflammatory skin disorders. Cornified envelopes in the outermost SC of involved areas of psoriasis vulgaris (PV) and atopic dermatitis (AD) were strikingly heterogeneous, and consisted of immature CEs stained with anti-involucrin and mature CEs stained with Nile red, whereas CEs of the uninvolved areas were relatively homogeneous, exhibiting mature phenotype. The ratio of immature CEs was significantly higher in the involved areas of PV and AD than that in the corresponding uninvolved areas, suggesting that defective CE maturation may, at least in part, account for the inflammatory disorders. Simultaneous evaluation of CE maturity and parakeratosis was carried out by a combination of involucrin immunostaining and nuclear staining of detergent-dissociated corneocytes. In the involved area of PV, four types of corneocytes in regard to the combination of involucrin staining and nuclear remnant were observed, while both immature CEs and parakeratosis were more often detected in the involved areas of PV than in the uninvolved areas or the upper arm of healthy subjects as a normal control. Thus, corneocytes with involucrin-positive immature CEs were not always associated with parakeratosis at the cellular level. In the involved areas of PV, the ratio of immature CEs and that of parakeratosis were heterogeneous, depending on the cases, and no correlation between the ratios was observed. Inter-individual and intraindividual variations in CE maturity were also suggested by the heterogeneous localization of involucrin in the psoriatic epidermis as examined by immunohistochemistry. In addition, in the face of healthy subjects, four types of corneocytes were similarly detected, and the ratio of immature CEs was significantly higher than that of parakeratosis. These results obviously suggest that the maturation of CEs and disappearance of nuclei are differentially regulated in the epidermis.

Analysis of the dermo-epidermal surface in three-dimensions is important for evaluating cosmetics. One approach is based on the active contour model, which is used for extracting local object boundaries with closed curve form. The dermo-epidermal surface, however, is a plane with open form. We have developed a method of automatically extracting the dermo-epidermal surface from volumetric confocal microscopic images, as well as constructing a 3-D visual model of the surface by using the geometric information contained in the control points. Our method is a 3-D extension of the active contour model, so we call it the active open surface model (AOSM). The initial surface for AOSM is an open curve plane, guided by a 3-D internal force, a 3-D external constraint force, and a 3-D image force, which pull it toward the objective surface. The proposed technique has been applied to extract actual dermo-epidermal surface in the given volumetric confocal microscopic images.

BACKGROUND/AIMS: Changes in the skin conditions after exposure to low humidity have been generally experienced in everyday life, but there have been few reports to approach it-especially in healthy skin. We have examined the effect of low humidity on healthy human skin by using noninvasive measurement devices. METHODS: Skin conditions on the ventral forearm and the cheek before and after 3 or 6 h exposure to low humidity were evaluated by measuring skin surface conductance, skin surface capacitance and transepidermal water loss. Skin surface replicas were also taken before and after exposure and analysed for roughness parameters--Ra (arithmetic mean roughness value), Rz (10-point height), Sm (mean value of the profile element) and VC1 (anisotropy of skin furrows). RESULTS: There was a significant decrease of water content of stratum corneum at both test sites from the time points 0 h to 3 h and 6 h (P < 0.01) and transepidermal water loss from the time point 0 h to 6 h (P < 0.05). Regarding the roughness parameters, a significant increase of Rz in the directions of 45 degrees/225 degrees and 90 degrees/270 degrees to the body axis and Sm in the directions of 0 degrees/180 degrees (P < 0.05) on the forearm and VC1 (P < 0.05) on the cheek. The parameter Rz also showed a tendency to increase in the directions of 45 degrees/225 degrees (P = 0.06) on the cheek. A specific pattern of the changes to be related to the Langer's lines in the surface morphology was observed. The changes of skin surface pattern in our experiment lead us to consider that exposure to low humidity even in such a short period would be related to inducing aggravation of skin texture and the formation of fine wrinkles. CONCLUSION: A short exposure of skin to a low-humidity environment induced changes in the moisture contents in the stratum corneum and skin surface pattern, which lead us to assume that a dry environment in our daily life would make fine wrinkles related to lack of water in the stratum corneum.

We attempted to analyze the skin color of women from the world using identical apparatuses and under identical conditions. The data was used for developing cosmetic products. The skin colors of a total of 1703 women over the world were measured to clarify the characteristics of skin color. Skin color measurement was done by using a spectrometric colorimeter. We modified the conventional erythema and melanin indices and established new respective indices which can reflect various spectral information of whole visible region. The present results were used to design new cosmetics in consideration of the optical properties of skin.

Sensory evaluation is an important factor for cosmetic products. Several devices for the measurement of sensory properties have been developed in recent years. The objective here is to measure skin surface friction using these devices and examine the correlation with other physiological parameters in order to evaluate the potential of physical measurement of tactile sensation.A KES-SE Frictional Analyzer, a commercial device for measurement of surface frictional characteristics, was used in this study. An arm holder was added to this device for measurement on the human forearm. The frictional coefficient (MIU) and its mean deviation (MMD) were used as the parameter to indicate surface friction. The moisture content in the stratum corneum was measured with a Corneometer CM825, the transepidermal water loss with a Tewameter TM210, the viscoelastic properties of the skin with a Cutometer SEM575 and the skin surface pattern by observing the negative replica using silicon rubber.The MIU was not influenced by load; however, it was increased due to water application on the skin. The relationship between MIU and the moisture content in the stratum comeum, between MMD and skin surface pattern and between MMD and viscosity of both normal human forearm skin and SDS (sodium dodecyl sulfate)-induced dry skin were confirmed by statistical analysis in a test on human subjects. There was also a correlation between either MIU or MMD and sensory evaluation in the morning after the application of moisturizing products.Human skin surface friction was measured by using a KES-SE Frictional Analyzer. Judging from the correlation between either MIU or MMD and sensory evaluation, we considered this instrumental analysis to be useful for evaluating the tactile impression of human skin.

As a depigmenting treatment, combined topical applications of all-trans retinoic acid (atRA) aqueous gel and 5% hydroquinone, 7% lactic acid ointment were used for Oriental patients with hyperpigmented skin lesions such as senile lentigines and nevus spilus. A narrow-band reflectance spectrophotometer and a tristimulus colorimeter were used to evaluate objectively the intensity of pigmentation and erythema at each clinical visit. L*, a*, and b* values measured with a tristimulus colorimeter (Chroma Meter CR-300) enabled the evaluation of erythema but not pigmentation. On the other hand, the melanin and hemoglobin values measured with a narrow-band reflectance spectrophotometer (Mexameter MX-16) expressed both erythema and pigmentation well. It was revealed that, in our bleaching protocol, the narrow-band reflectance spectrophotometer was quite useful for estimating accurately the intensity of pigmentation and erythema and determining the best time point for the cessation of atRA treatment.

The effects of four different magnesium salts on the cutaneous barrier recovery rate after barrier disruption were evaluated. We spread an aqueous solution of each salt on the flank skin of hairless mice, occluded the area with a plastic membrane for 20 min, and then left the skin surface to dry. All of the magnesium salts, except magnesium bis(dihydrogen phosphate), accelerated barrier repair. We next estimated the effects of magnesium chloride aqueous solutions which contained calcium chloride at different molar ratios. When the calcium to magnesium ratio was lower than 1, the mixture accelerated barrier repair. The application of an aqueous solution of 10 mM magnesium chloride and 10 mM calcium chloride was found to hasten the barrier recovery more effectively than a solution of 10 mM magnesium chloride. These results suggest that the effects of these metal ions are different depending on the counter ion and/or the method of application.