中间丝聚合蛋白在特应性皮炎患者皮损中的表达

目的：探讨中间丝聚合蛋白（Filaggrin,FLG）在特应性皮炎（Atopic dermatitis,AD）患者皮损的表达及其临床意义。方法：采用sP免疫组化方法检测16例AD患者皮损及14例对照组皮肤组织内FLG的表达,用图像分析软件Image-ProPlus（IPP）判定FLG在AD患者皮损及健康者皮肤组织中阳性染色面积百分比。结果：FLG在AD患者皮损及健康者皮肤角质层及颗粒层均有表达,胞质染色多见;阳性染色面积百分比分别为（18．92±4．40）％及（23．00±2．28）％;FLG在AD患者皮损阳性染色面积明显低于健康人皮肤（t=－3．11,P=0．004）。结论：FLG在AD患者皮损中表达降低,可能是导致AD患者皮肤屏障功能障碍的原因。     

Defective barrier function in melasma skin

Background Melasma is characterized by increased pigmentation and photodamaged features, which include solar elastosis. Recently, we detected the downregulation of the genes most associated with lipid metabolism using microarray analysis in melasma. These findings suggested that lesional skin may have different biophysical characteristics, and, in particular, an altered skin barrier function. Objective To determine the cutaneous biophysical characteristics of melasma. Methods The melanin index, erythema index, stratum corneum hydration, sebum content and transepidermal water loss (TEWL) were measured for lesional and perilesional normal skin of 16 melasma patients and then compared. In addition, a skin biopsy was performed on 11 of the 16 study subjects to measure stratum corneum thickness and to study the protein expressions of PPAR‐α and ALOX15B. Results Melanin index, erythema index and stratum corneum hydration were significantly higher in lesional skin than in perilesional normal skin. No significant difference was found between lesional and normal skin in terms of basal TEWL level or sebum content. However, the rate of TEWL after barrier perturbation was significantly higher for lesional skin, and the barrier recovery rate was significantly delayed. Furthermore, a trend towards thinned stratum corneum was observed for lesional skin, and this was correlated with barrier recovery rate. The expressions of PPAR‐α and ALOX15B were variable in the samples. Conclusions Melasma skin is characterized by impaired stratum corneum integrity and a delayed barrier recovery rate.     

Distinct barrier integrity phenotypes in filaggrin-related atopic eczema following sequential tape stripping and lipid profiling

Background Filaggrin gene (FLG) loss‐of‐function mutations have been shown to represent the strongest so far known genetic risk factor for atopic dermatitis (AD). Whereas the barrier characteristics in FLG mutation carriers under baseline conditions have been investigated, there are only limited data on the permeability barrier function in filaggrin‐AD under compromised conditions. Aim We investigated: (i) stratum corneum (SC) integrity/cohesion; (ii) barrier recovery after controlled mechanical and irritant‐induced barrier abrogation; and (iii) the lipid composition of the non‐lesional and lesional skin of AD patients harbouring the European R501X, 2282del4, 3702delG, R2447X or S3247X FLG variants. Methods Thirty‐seven AD patients (14 FLG mutation carriers and 23 non‐carriers) and 20 healthy controls participated in the study. Stratum corneum integrity/cohesion was assessed by measurement of transepidermal water loss (TEWL) and amount of removed protein following sequential tape stripping. Barrier recovery was monitored by repeated measurements of TEWL and erythema up to 96 h after barrier abrogation. Samples for lipid analysis were obtained from non‐lesional and lesional skin using the cyanoacrylate method. Results Tape stripping revealed distinct genotype‐related impairment of the SC integrity/cohesion. No differences in the rate of barrier recovery among the groups were found. The SC lipid analysis revealed significant differences regarding the percentage amount of cholesterol, ceramide/cholesterol ratio and triglycerides in the uninvolved skin as well as the amounts of free fatty acids, CER[EOH] and triglycerides in the skin lesions of the AD FLG mutation carriers. Conclusions Our results provide evidence for discernible FLG‐related barrier integrity phenotypes in atopic eczema.     

Disruption of barrier function in dermatophytosis and pityriasis versicolor

Dermatophytes have the ability to form molecular attachments to keratin and use it as a source of nutrients, colonizing keratinized tissues, including the stratum corneum of the skin. Malassezia species also affect the stratum corneum of the skin. Therefore, dermatophytosis and pityriasis versicolor of the skin are thought to be important factors of profound changes in skin barrier structure and function. We aimed to describe the changes in transepidermal water loss (TEWL), stratum corneum hydration, and skin pH in the lesions of the dermatophytosis and pityriasis versicolor. Thirty-six patients with dermatophytosis (14 with tinea cruris, 13 with tinea corporis and nine with tinea pedis or tinea manus) and 11 patients with pityriasis versicolor were included in this study. TEWL, stratum corneum conductance and skin pH were determined by biophysical methods to examine whether our patients exhibited changes in barrier function. Dermatophytosis and pityriasis versicolor except tinea pedis and tinea manus showed highly significant increase in TEWL compared with adjacent infection-free skin. Hydration was significantly reduced in lesional skin compared with adjacent infection-free skin. From this study, infections with dermatophytes and Malassezia species on the body can alter biophysical properties of the skin, especially the function of stratum corneum as a barrier to water loss. On the contrary, infections with dermatophytes on the palms and soles little affect the barrier function of the skin.     

Impaired sphingomyelinase activity and epidermal differentiation in atopic dermatitis

A defective permeability barrier leads to the penetration of environmental allergens into the skin and initiates immunological reactions and inflammation crucially involved in the pathogenesis of atopic dermatitis (AD). Decreased stratum corneum ceramide content may cause the defect in permeability barrier function consistently found in AD. Acid and neutral sphingomyelinase (A- and N-SMase) generate ceramides with structural and signal transduction functions in epidermal proliferation and differentiation. We determined epidermal SMase activities, DNA synthesis, involucrin, loricrin, filaggrin, and keratin expression in lesional and non-lesional skin of AD patients. We found decreased epidermal A-SMase activity in lesional and non-lesional skin, correlating with reduced stratum corneum ceramide content and disturbed barrier function. N-SMase activity was reduced in non-lesional skin and more significantly reduced in lesional skin, correlating with impaired expression of cornified envelope proteins and keratins, important for skin barrier function. Changes in involucrin, loricrin, filaggrin, keratin K 5 (basal) and K 16 (proliferation associated) were noticed in non-lesional and lesional skin, whereas changes in K 10 (suprabasal), K 6 (proliferation associated), and K 17 (inflammation associated) were found only in lesional skin. In summary, reduction in SMase-generating ceramides and impaired differentiation are involved in the defective barrier function found in AD.     

High-expression of sphingomyelin deacylase is an important determinant of ceramide deficiency leading to barrier disruption in atopic dermatitis

We have previously demonstrated that there is abnormal expression of sphingomyelin (SM) deacylase-like enzyme in the epidermis of patients with atopic dermatitis (AD), which results in decreased levels of ceramides in their involved and uninvolved stratum corneum. For quantitation of the expression of SM deacylase in AD, we synthesized 16-(9-anthroyloxy) hexadecanoylsphingosylphosphorylcholine or [palmitic acid-14C] SM and used them as substrates to directly measure the activity of SM deacylase by detecting the release of labeled free fatty acid. Direct enzymatic measurements demonstrated that stratum corneum from lesional forearm skin (volar side) of AD patients has an extremely high SM deacylase activity that is at least five times higher than in the stratum corneum from healthy controls. In stratum corneum from nonlesional skin of AD patients, SM deacylase activity is still at least three times higher than in healthy controls. In contrast, stratum corneum from contact dermatitis patients shows levels of SM deacylase similar to healthy controls. In extracts of whole epidermis biopsies from AD patients, SM deacylase activities are significantly (3-fold) increased over healthy controls in the particulate fraction, whereas there is no significant difference in the activity of sphingomyelinase between AD and healthy control. In peripheral blood lymphocytes of AD patients, there is no increase in activity compared with healthy controls, indicating a possibility that the high expression of SM deacylase is highly associated with the skin of AD patients. These findings suggest that, in contrast to changes in sphingolipid metabolism due to aging, the hitherto undiscovered enzyme SM deacylase, is highly expressed in the epidermis of AD patients, and competes with sphingomyelinase or beta-glucocerebrosidase for the common substrate SM or glucosylceramide, which leads to the ceramide deficiency of the stratum corneum in AD.     

Microanalysis of an antimicrobial peptide, beta-defensin-2, in the stratum corneum from patients with atopic dermatitis

Background  Antimicrobial peptides, such as defensin and cathelicidin, have recently been reported to play important roles in host defence and in cutaneous innate immunity. Although β-defensin-2 has been reported to be downregulated in the skin of patients with atopic dermatitis (AD), little is known about its role in the colonization of Staphylococcus aureus in the stratum corneum of patients with AD. A precise evaluation of these peptides in the stratum corneum as an antimicrobial barrier against S. aureus colonization has not yet been performed.
Objectives  To compare β-defensin-2 levels in the skin of patients with AD and healthy controls.
Methods  We developed a microanalytical technique to measure β-defensin-2 in the stratum corneum using a combination of immunoprecipitation and Western blotting.
Results  β-Defensin-2 in the stratum corneum was significantly higher in AD lesional skin compared with healthy control skin. The β-defensin-2 content in AD lesional skin also increased in proportion to the severity of the disease. Counting bacterial colonies revealed higher populations of S. aureus on lesional and nonlesional skin surfaces of patients with AD compared with healthy controls. Comparison of S. aureus colony numbers and β-defensin-2 levels demonstrated a positive correlation ( r  =   0·342, P  =   0·004, n  =   67) between both factors.
Conclusions  Collectively, these findings suggest that β-defensin-2 is induced in response to bacteria, injury or inflammatory stimuli and is not associated with vulnerability to S. aureus colonization in the skin of patients with AD.     

上海两社区特应性皮炎患儿的皮肤屏障功能研究

【摘要】 目的 探讨上海两个社区特应性皮炎（AD）患儿及健康对照儿童皮肤屏障功能及AD皮损严重程度与皮肤屏障功能的相关性。方法 3 ~ 12岁AD患儿169例和健康对照儿童142例来自上海长宁新泾社区和嘉定菊园社区,检测前臂伸侧、屈侧及脸颊、胫前4个部位非皮损区的角质层含水量和经皮失水量（TEWL）。并用欧洲AD评分标准（SCORAD）对AD患儿临床严重程度进行评分。结果 AD患儿前臂伸侧、屈侧及脸颊、胫前四个部位的TEWL值均高于健康对照儿童（P ＜ 0.05）,角质层含水量在前臂伸侧和胫前均显著低于健康对照儿童（P ＜ 0.05）。AD患儿SCORAD与TEWL均值呈正相关,与角质层含水量均值呈负相关。结论 皮肤屏障功能可以作为评价AD临床严重程度的指标之一。     

Defective barrier function accompanied by structural changes of psoriatic stratum corneum

Although barrier function of psoriatic skin is shown to be decreased by measuring transepidermal water loss (TEWL), few reports exist examining other physical skin properties and components including stratum corneum hydration, natural moisturizing factor (NMF), free fatty acids (FFA), β‐sheet and α‐helix ratio of structural protein(s), and sebum content. We compared the skin properties and components of normal, involved and uninvolved skin of psoriasis. Using a corneometer and attenuated total reflection‐infrared spectrometer, we measured TEWL, stratum corneum hydration, NMF, FFA, β/α ratio and sebum in psoriasis vulgaris patients and healthy controls. TEWL and β/α ratio of involved psoriatic skin were significantly increased compared with uninvolved skin and normal control skin. In contrast, stratum corneum hydration , NMF and FFA, but not sebum, are significantly decreased in the involved skin compared with uninvolved skin and normal skin. TEWL and stratum corneum hydration returned to the normal levels following clinical improvement of the lesion. Barrier function and hydration of psoriatic skin are defective and secondary structure in stratum corneum protein is altered in the involved psoriatic skin.     

对称性肢端角化病皮损中脂肪酸结合蛋白5及二氢硫辛酰胺脱氢酶表达

目的 探讨脂肪酸结合蛋白5（FABP5）及二氢硫辛酰胺脱氢酶（DLD）在对称性肢端角化病中的表达和意义。 方法 收集9例对称性肢端角化病患者腕部皮损及其周围皮肤活检标本,9例健康人腕部皮肤为对照,用逆转录PCR（RT-PCR）及免疫组化法检测FABP5及DLD表达水平。 结果 RT-PCR显示,FABP5 mRNA及DLD mRNA表达在对称性肢端角化病患者腕部皮损、皮损周围皮肤、健康人腕部皮肤3组间差异均无统计学意义（P > 0.05）。免疫组化显示,对称性肢端角化病皮损中FABP5 阳性染色范围和强度增加,角质层、颗粒层、棘层均有强染色;而皮损周围皮肤中角质层、颗粒层及棘层弱阳性染色,健康对照皮肤中仅有棘层和基底层弱阳性染色。对称性肢端角化病皮损中DLD 表达减少,仅在角质层和少数病例棘层表达;皮损周围及健康人皮肤表皮全层均有DLD阳性染色,胞核、胞质均呈深棕色。 结论 对称性肢端角化病皮损中FABP5高表达可能参与角质形成细胞异常分化过程,而DLD表达下调提示存在能量代谢失衡。     

寻常型银屑病患者皮肤屏障功能相关指标检测分析

目的:探讨寻常型银屑病患者皮损区与非皮损区、患者与正常人皮肤屏障功能相关指标之间的差异。方法:1.收集寻常型银屑病患者35例,健康志愿者36例,使用皮肤屏障功能测试仪测定患者皮损区、非皮损区、健康志愿者的皮肤角质层含水量、PH值及经皮水分流失(TEWL)。2.收集30例寻常型银屑病患者与34例健康志愿者的血清,分别采用压板法、分光光度法测定NO、VitE水平。结果:寻常型银屑病患者皮损区PH值为(5.69±0.61)高于非皮损区PH值和健康对照组PH值;皮损区含水量为(17.90±6.75),低于非皮损区及健康对照组;皮损区TEWL为(18.58±11.55)g/m²h,高于非皮损区和健康对照组;然而非皮损区与健康对照组的PH值、角质层含水量及TEWL无统计学差异;寻常型银屑病患者血清维生素E含量(7.27±0.96)μg/mL低于健康对照组,银屑病组血清NO含量(5.29±1.91)μg/mL高于健康对照组。结论:寻常型银屑病患者皮损区,皮肤屏障功能指标异常,表现在皮肤表面PH值升高、角质层含水量下降、TEWL值升高、血清VitE下降、血清NO升高。     

Cutaneous pH in children affected by atopic dermatitis and in healthy children: a multicenter study

Background/aims: Several studies have been conducted in order to assess, objectively and non-invasively, atopic skin parameters. Since little is known about clinically uninvolved, perilesional and lesional skin pH in atopic children at different stages of the disease, we studied cutaneous pH in atopic and healthy children and investigated possible relationships among different sites, phases of the disease and main symptoms.
Methods: Two hundred and eighty-four children of both sexes, 143 atopic and 141 healthy children, aged 3 months to 11 years, examined in 10 different Italian dermatological centres were included in the study and subjected to clinical examinations and pH measurements.
Results: Atopic children presented statistically significant higher pH values than controls, not only at the level of eczematous lesions (P=0.004) and of perilesional skin (P=0.003) but also at the level of apparently healthy skin (P=0.002). A progressive statistically significant increase in pH values was observed from healthy to perilesional to lesional skin. In acute dermatitis, children affected by moderate itching (considering itching complaints and scratching lesions) presented, at the level of lesional and perilesional skin, statistically significant higher pH values than children affected by mild itching.
Conclusion: The correlation found in this study between severity of itch and skin surface alkalinity, far from demonstrating a therapeutic role of acidic substances against itching in atopic children, deserves new epidemiological studies on a large scale in order to identify new strategies of treatment of atopic skin.     

Altered penetration of polyethylene glycols into uninvolved skin of atopic dermatitis patients

Involved regions of the skin in atopic dermatitis (AD) patients have an altered barrier function. Whether uninvolved skin also has a diminished barrier is controversial. To assess the barrier function of uninvolved skin in AD patients, the percutaneous penetration of polyethylene glycols (PEGs) of various molecular sizes was determined in vivo in AD patients and control subjects using tape stripping of the stratum corneum (SC). The diffusion and partition coefficients were determined using Fick's second law of diffusion. The SC thickness was similar in both groups; however, the trans-epidermal water loss was higher in atopic skin. The apparent diffusion coefficient of PEGs through atopic skin was twice as high as through normal skin, and decreased with increasing molecular weight (MW) in both groups. The partition coefficient in the skin of AD patients was half of that for normal skin but as for normal skin, there was no MW dependency. Although atopic skin exhibited altered barrier with respect to diffusion and partitioning, the permeability coefficients were nearly the same for atopic and normal skin. The results support the assumption of altered skin barrier of AD patients even in the skin that is visibly unaffected by disease.     

Normal recovery of the stratum corneum barrier function following damage induced by tape stripping in patients with atopic dermatitis

Patients with atopic dermatitis (AD) constantly inflict mechanical damage to their skin by scratching induced by pruritus. On excoriated lesions of the cheek we found exceedingly high levels of transepidermal water loss (TEWL) as compared to those in the normal skin of healthy subjects. However, it is not clear whether the skin of patients with AD also shows an abnormally slow recovery after mechanical damage. We compared the recovery of the barrier function of the stratum corneum (SC). after its complete removal by tape stripping, in patients with AD and age-matched healthy control subjects. On the normal-looking skin of the flexor forearm, we found no difference in the recovery process of the water barrier function of the SC between the two groups. This suggests that ability to reconstruct SC barrier function after mechanical damage is not impaired in AD patients.     

Ceramide profiles of the uninvolved skin in atopic dermatitis and psoriasis are comparable to those of healthy skin

Ceramides are sphingolipids consisting of sphingoidbases, which are amide-linked to fatty acids. In the stratum corneum, they represent the major constituent of the free extractable intercellular lipids and play a significant role in maintaining and structuring the water permeability barrier of the skin. Using thin layer chromatography, which represents the method of the first choice in analyzing the stratum corneum ceramides, at least seven classes can be distinguished. Each ceramide class contains various species, which have the same head group and different chain lengths. As in many other skin disorders, atopic dermatitis and psoriasis show derangements in content and profile of the ceramides. Such derangements were reported for both the lesional involved as well as for the normal-appearing uninvolved skin. In this study, we focused on investigating the stratum corneum ceramides of the uninvolved skin in atopic dermatitis and psoriasis patients compared to healthy skin. The aim of the investigations was to explore possible significant and specific differences which can be accomplished for purposes of early diagnostics. The skin lipids were collected by means of an in vivo topical extraction procedure using an extraction mixture consisting of n-hexane and ethanol, (2:1). An automated multiple development-high performance thin layer chromatography (AMD-HPTLC) method with photodensitometric detection were applied to separate the ceramides and to estimate their contents. For studying their molecular profile within each ceramide class, a new method of normal phase HPLC with atmospheric pressure chemical ionization mass spectrometry were used. The results obtained by AMD-HPTLC exposed no significant alterations regarding the relative composition of the major stratum corneum lipids and primarily the ceramides. In addition, the mass spectrometric profiles within each ceramide class were similar in the patients and the healthy control subjects. In conclusion, this study revealed that the normal-appearing uninvolved skin of atopic dermatitis and psoriasis patients does not prove significant or specific deficiencies with respect to the free extractable major stratum corneum lipids and mainly the ceramides, when compared to healthy skin. Thus, they cannot be used for diagnostic purposes. Furthermore, our data are not consistent with the concept that impairments in the ceramide composition represent an obligate etiologic factor for both diseases.     

Percutaneous penetration of sodium lauryl sulphate is increased in uninvolved skin of patients with atopic dermatitis compared with control subjects

BACKGROUND: Involved regions of the skin in patients with atopic dermatitis (AD) have been shown to have higher transepidermal water loss (TEWL), indicating a compromised skin barrier. Whether uninvolved skin also has diminished barrier characteristics is controversial. OBJECTIVES: To study the penetration of sodium lauryl sulphate (SLS) into uninvolved skin of patients with AD compared with the skin of control subjects. METHODS: Percutaneous penetration was assessed using the tape stripping technique on the stratum corneum (SC). Twenty patients with AD and 20 healthy subjects were exposed to 1% SLS for 4 h on the mid-volar forearm. After the end of exposure the SC was removed by adhesive tape. The amount of SLS was determined in each consecutive strip. Fick's second law of diffusion was used to deduce the diffusivity and the partition coefficient of SLS between water and the SC. RESULTS: The SC thickness was similar in both groups; however, the TEWL was higher in patients with AD compared with that of the control group (mean+/-SD 8.4+/-4.3 and 6.3+/-2.0 g m-2 h-1, respectively). There was a correlation between SC thickness and TEWL in control subjects but no correlation was found in patients with AD. The diffusivity of SLS through uninvolved AD skin was higher compared with normal skin (mean+/-SD 12.7+/-5.8x10(-9) and 6.2+/-3.0x10(-9) cm-2 h-1, respectively), while the partition coefficient between SC and water was lower (mean+/-SD 137+/-64 and 196+/-107, respectively). CONCLUSIONS: The results show a different penetration profile of SLS into the SC of patients with AD compared with control subjects. This indicates that even noninvolved skin in patients with AD has altered barrier characteristics, emphasizing the importance of skin protection and prevention of skin contact with chemicals.     

特应性皮炎患者皮肤屏障与水闸蛋白1表达的相关性研究

目的 研究特应性皮炎（AD）患者皮肤屏障功能情况,并分析其与水闸蛋白1（claudin-1）表达的相关性。 方法 纳入AD患者和健康人各11例。应用皮肤经表皮失水率测定仪和皮肤高频超声检测仪测定受试者经表皮失水率、表皮厚度与表皮致密度,并用双抗体夹心ELISA法定量检测血清中脱落claudin-1表达量。应用单因素方差分析和t检验比较不同组别之间相关参数的差异;应用Pearson相关系数分析不同参数之间的相关性。 结果 AD患者皮疹部位经表皮失水率为（36.9 ± 34.2） g·m-2·h-1,非皮疹部位为（9.1 ± 6.0） g·m-2·h-1,均高于健康对照［（4.4 ± 3.1） g·m-2·h-1］;AD患者皮疹部位表皮厚度（0.23 ± 0.04） mm,显著高于非皮疹部位［（0.18 ± 0.03） mm］和健康对照［（0.18 ± 0.02） mm］。AD患者皮疹部位有其特征性表皮下低回声带。AD患者claudin-1表达量为（0.80 ± 0.88） ng/ml,显著低于健康人［（1.73 ± 1.85） ng/ml］;claudin-1与表皮厚度显著负相关（r = -0.61）,与经表皮失水率的倒数显著正相关（r = 0.44）。 结论 AD患者损伤的皮肤屏障功能与claudin-1表达相关,屏障功能状态可用经表皮失水率、经表皮失水率倒数和表皮厚度进行定量表述。     

Evaluation of skin surface hydration in Korean psoriasis patients: a possible factor influencing psoriasis

Psoriasis is recognized as a chronic, relapsing disorder of the skin, and its clinical course is influenced by various environmental factors. However, only a few articles have addressed the effect of low epidermal hydration status as an exacerbating factor of psoriasis. Our study was performed to investigate the relationship between subjective assessment and quantified data regarding dryness parameters in psoriasis patients. Seventy psoriasis patients were enrolled in our study. Subjective assessments were undertaken by evaluating the extent of involvement, erythema, scale and infiltration of individual lesions. Functional status of stratum corneum was evaluated by measuring electrical capacitance and conductance of the lesional and uninvolved skin of psoriatic patients. In addition, measurement of trans-epidermal water loss was performed. The results showed poor hydration status and barrier function in involved psoriatic epidermis compared with the uninvolved skin. Subjective assessment showed fairly good correlation with the quantified values. In conclusion, the degree of dryness in psoriatic skin lesions--which we presume to be one of the aggravating factors of psoriasis--seemed to be related to disease severity.     

麻风患者愈后表皮某些生理功能障碍研究

目的：了解麻风愈后者其原受累及部位是否存在角质层生理功能障碍。方法：利用多功能皮肤生理仪对麻风患者痊愈后前臂原皮损处角质层含水量、酸碱度、透过皮肤水分丢失率及屏障功能的恢复速度进行测量。结果：与正常对照组相比，麻风愈后者原皮损处角质层含水量明显降低（P〈0．002），以瘤型麻风患者降低幅度最大；皮肤表面的pH明显增高（P〈0．0001），界线类偏结核型患者pH改变不甚明显；麻风愈后者原皮损处的基础表皮通透屏障功能好于正常人，以界线类偏瘤型患者水分丢失量降低最明显：麻风愈后者原皮损处表皮通透屏障功能的恢复速度无明显改变。结论：麻风患者痊愈后仍然有角质层生理功能障碍。