Exploration of a quantitative dansyl chloride technique for measurement of the rate of desquamation

A new fluorescence comparator has been used to measure dansyl chloride-induced fluorescence from the skin surface. By taking readings twice daily, it was possible to ideality a circadian rhythm in which there was relatively depressed desquamation at night, compared with during the day. The study also demonstrated that the rate of desquamution was considerably reduced from protected sites, compared with normal unprotected sites. Other experiments demonstrated the ability of the technique to determine the enhanced rate of desquamation from uninvolved areas of skin near lesions of psoriasis, and the keratolvtic effect of preparations containing salicylic acid. It is considered that the technique adds objectivity and quantitative ability to the dansyl chloride method for characterizing desquamation.     

Pre-operative diagnosis of pigmented skin lesions: in vivo dermoscopy performs better than dermoscopy on photographic images

BACKGROUND: Epiluminescence microscopy (ELM) (dermoscopy, dermatoscopy) is a technique for non-invasive diagnosis of pigmented skin lesions that improves the diagnostic performance of dermatologists. Little is known about the possible influence of associated clinical features on the reliability of dermoscopic diagnosis during in vivo examination. OBJECTIVE: To compare diagnostic performance of in vivo dermoscopy (combined clinical and dermoscopic examination) with that of dermoscopy performed on photographic slides (pure dermoscopy). DESIGN: This case series comprised 256 pigmented skin lesions consecutively identified as suspicious or equivocal during examination in a general dermatological clinic. Clinical examination and in vivo dermoscopy were performed before excision by two trained dermatologists. The same observers carried out dermoscopy on photographic slides at a later time, and these three diagnostic classifications were reviewed together with the histological findings for the individual lesions. This was carried out in a university hospital. RESULTS: In vivo dermoscopy performed better than dermoscopy on photographic slides for classification of pigmented skin lesions compared with histological diagnosis, and both performed better than general clinical diagnosis. In vivo dermoscopic diagnosis of melanoma showed 98.1% sensitivity, 95.5% specificity and 96.1% diagnostic accuracy while dermoscopic diagnosis of melanoma on photographic slides was less reliable with 81.5% sensitivity, 86.7% specificity and 85.2% diagnostic accuracy. In particular, diagnosis of melanoma based on photographic slides led to nine false negative cases (three in situ, six invasive; thickness ranges 0.2-1.5 mm). CONCLUSIONS: In vivo dermoscopy, i.e. combined clinical and dermoscopic examination, is more reliable than dermoscopy on photographic slides. In clinical practice, therefore, in vivo dermoscopy cannot be considered independent from associated clinical characteristics of the lesions, which help the trained observer to reach a more precise classification. This may have implications on the reliability of ELM diagnosis made by an observer not fully trained in the clinical diagnosis of pigmented skin lesions or by a remote observer during digital ELM teleconsultation.     

The dansyl chloride technique for stratum corneum renewal as an indicator of changes in epidermal mitotic activity following topical treatment

Using a hypomitotic agent, triamcinolone acetonide, and a hypermitotic agent, retinyl propionate, we investigated the relationship between epidermal mitotic activity and stratum corneum renewal time of topically treated skin as determined by the dansyl chloride staining technique. Treatment with the base cream resulted in a reduction in renewal time compared with an untreated control site. The predicted increase in renewal time with the hypomitotic agent and reduction with the hypermitotic agent was only observed when daily treatment was commenced 2 weeks prior to and continued after dansyl chloride staining and not when treatment was started after staining. These results indicate that in order to use cell renewal methods to demonstrate changes in mitotic activity brought about by topical treatments, it is necessary to pre-treat the skin with the test material to establish full epidermal equilibrium at the changed mitotic state before labelling with dansyl chloride. Meaningful claims for effects on cell renewal of specific cosmetic ingredients should only be made after comparison with a base cream treated site, both having been allowed to equilibrate, rather than on the basis of comparison with untreated skin.     

Bispectral fluorescence imaging of aggressive basal cell carcinoma combined with histopathological mapping: a preliminary study indicating a possible adjunct to Mohs micrographic surgery

BACKGROUND: Fluorescence imaging is an attractive diagnostic technique for skin tumour demarcation with potential to move to clinical use. Bispectral fluorescence imaging combines skin autofluorescence with delta-aminolaevulinic acid-induced fluorescence. To evaluate the technique, fluorescence data must be compared with the histopathological extent of the tumour, which is the purpose of the current study. OBJECTIVES: To investigate the agreement between bispectral fluorescence images and the histopathological tumour boundary of ill-defined basal cell carcinomas (BCCs). After fluorescence imaging the tumours were removed using Mohs micrographic surgery (MMS) to obtain histopathological maps of the tumour boundaries. METHODS: Twelve patients with aggressive BCC of mean diameter 16 mm (range 5-32) in the face were included in the study. The patients were subjected to bispectral fluorescence imaging within the 2 months prior to MMS. The fluorescence images and histopathological maps were aligned using image warping. RESULTS: Five patients (42%) showed good agreement with the histopathological mapping and the remaining seven patients (58%) showed partial agreement. Bispectral investigation combining autofluorescence with protoporphyrin IX (PpIX) fluorescence generally yielded better agreement with the histopathological boundaries of the tumours compared with using only the PpIX fluorescence. CONCLUSIONS: In this preliminary study the fluorescence has been compared with the histopathological tumour boundaries. The result implies that the technique can be applied as a useful tool for indicating tumour boundary of aggressive BCCs. Further refinement is needed to be able to indicate the exact tumour border.     

Assessment of skin surface by scanning densitometry of macrophotographs

There are no established techniques for the measurement of scaliness and skin smoothness. In this paper a new photographic technique for assessment of the skin surface is described. Photographic negatives of the skin surface taken under standardized conditions arc scanned on a densitometer and the scan tracing length used as a measure of roughness. The technique has been used to assess (a) the roughening effect of adhesive tape stripping, (b) the smoothing effect of emollient preparations, and (c) the alterations in roughness of scaling dermatoses during treatment. It has been found to be reproducible and to correlate well with assessment of skin surface contour.     

High-fluence fractional treatment of photodamaged facial skin using a 2940 nm erbium:yttrium-aluminum-garnet laser

Abstract

Introduction: Fractional resurfacing with an Erbium:yttrium-aluminum-garnet (Er:YAG) 2940 nm laser is an increasingly popular option for the treatment of the signs of facial photoaging, which include wrinkles as well as pigmentation issues and unwanted textural changes. Fractional treatment has produced favorable clinical responses, but with less complications and shorter recovery times than traditional laser resurfacing. This study was conducted to evaluate a fractionated Er:YAG treatment regimen of 1–2 higher fluence sessions with a multiple-pass technique. Materials and methods: Eight subjects with moderate to severely photodamaged facial skin received one to two full-face laser treatments. Multiple-pass (MP) treatment results were evaluated in terms of procedure time, discomfort, social downtime and effectiveness. A photographic evaluation, subject improvement assessments and a subject satisfaction assessment were performed. Results: An investigator's photographic review showed a 26–75% improvement in the signs of overall photoaging. Subjects treated with the MP technique exhibited a relatively short 3–4 day downtime and ratings of mostly moderate discomfort with the use of topical anesthetic cream only. Subjects treated with higher fluences demonstrated the highest average improvement in specific features of photoaging. Two laser treatments resulted in substantially higher improvement scores than those received just one laser treatment.     

Effect of pdo facelift threads on facial skin tissues: An ultrasonographic analysis

Background

Recent studies demonstrate that PDO threads, after being introduced into the facial skin, provide benefits due to collagen formation around the thread and improved vascularization.

Aims

This work aims to report the ultrasound and photographic follow-up after the execution of the facelift thread technique, thus evaluating the anti-aging treatment that PDO threads will provide for human facial skin.

Methods

The facelift will be performed using PDO thread 18G cannulated Sculpt i- Thread FML 100 × 185 mm, and 29G PDO thread Needled Flat i-Thread 38 × 50 mm. A total of 10 patients were selected for the study; all of them had medium-grade facial ptosis and aged 40–50 years old. In order to control tissue changes after thread, insertion ultrasound measurements will be made with Philips ultrasound 12 MHz linear probe at maximum resolution, initial evaluations and control of 30, 60, 90, and 120 days, measuring the skin layers: epidermis\dermis and hypodermis. Photographic control was performed before and after 120 days of intrusion of the PDO threads.

Results

The formation of collagen around the PDO thread was evidenced through ultrasound images in all patients in this study, as well as improvement in the appearance of the skin and sagging, evaluated by photographic control.

Conclusions

We can define that the PDO threads used for the facelift are effective in the formation of collagen around the thread (observed through the increase in the dermal layer), and improvement of the condition of the anti-aging factor through sagging skin, vascularization and thinning of the superficial fat layer (observed through the reduction of the subcutaneous or hypodermis layer).     

Hematoxylin stainable epidermal protein of the newborn rat. IV. The change of antigenicity in situ by transglutaminase as determined by an immunofluorescent study

Based on the activity of transglutaminase, the change of antigenicity in situ of hematoxylin stainable protein (HSP) purified from 3-day-old rat epidermis and located on the cell membrane region of the stratum corneum was investigated by indirect immunofluorescent technique using a polyclonal antibody against hematoxylin stainable protein. Three different techniques were employed: (i) the skin section was incubated with a commercial guinea pig liver transglutaminase (GLT), (ii) the skin section was incubated with an epidermal extract of Tris-HCl (EX), and (iii) the skin section was incubated with an epidermal extract of 1% Triton X-100 (EXT). The sections incubated in Tris-HCl or Triton X-100 were used as controls. Each incubation was done both in the presence and absence of Ca2+ ions. After these incubations, an indirect immunofluorescent technique using a polyclonal antibody was performed. In the presence of Ca2+ ions, the specific fluorescence of the cell membrane region of the entire stratum corneum cells disappeared after the preincubation with GLT. That of the lower one third of the stratum corneum disappeared after the incubation with EXT. In contrast, in the absence of Ca2+ ions, no preincubation with GLT, EX, or EXT showed any disappearance of the fluorescence anywhere in the stratum corneum. There was also no disappearance of the fluorescence in the control sections. These findings suggest that the antigenicity of HSP in situ could be lost by the activity of transglutaminase.     

A fluorescence photographic photometric technique to assess stratum corneum turnover rate and barrier function in vivo

A photographic photometric method for objective measurement of fluorescence in skin is described. This method has been used to (a) improve the dansyl chloride method for determining stratum corneum turnover time by making assessments objective and quantitative, and (b) examine the barrier function of the stratum corneum by measuring the concentration of a fluorescent material at different levels within the stratum corneum at different times.     

Antigen specificities of antibasement membrane zone antibodies: immunofluorescence and Western immunoblotting studies

Summary To clarify the antigen specificities of autoantibodies in sera and blister fluids from patients diagnosed as bullous pemphigoid (BP) by routine histology and immunofluorescence (IF) methods, indirect IF studies using the salt split-skin technique were performed. In addition, to detect the BP antigen(s) in human epidermal extracts, Western immunoblotting analyses were carried out. Of 41 sera, 39 (95%) showed a linear pattern of fluorescence along the epidermal side of the separation. Two (5%) sera showed a linear pattern of fluorescence along the dermal side. Blister fluids produced IF staining patterns identical with those of serum samples. These fluorescence patterns were not related to the BP antigen expression of the skin used as substrates. In Western immunoblotting analyses, selected sera showing an epidermal pattern on separated skin primarily reacted with 240 kD, 220 kD, 180 kD, and 150 kD proteins extracted from normal human epidermis. Two sera showing a dermal pattern on separated skin revealed no specific bands. The protein bands recognized by blister fluids were indentical with those of serum samples. These results indicated that blister fluids are also available in immunological analysis, and that BP antibodies have more than one antigenic specificity. Moreover, it is suggested that differential diagnosis between BP and other bullous diseases may be more important than previously recognized, particularly in patients with epidermolysis bullosa acquisita (EBA).     

Evaluation of salt split technique of immunofluorescence in bullous pemphigoid

Recent studies suggest that salt split skin is a more sensitive substrate than intact skin for immunofluorescence diagnosis of bullous pemphigoid. We undertook this study to define the role of salt split technique of immunofluorescence findings in 32 clinical and histopathology confirmed cases of bullous pemphigoid. Both direct and indirect immunofluorescences were performed using normal and split skin. Direct immunofluorescence positivity of 100% was noted with both routine and salt split method. Additional immunoreactant deposition was noted with direct method on split skin in 5 cases. Patterns of fluorescence in the latter were roof (40.60%), floor (9.4%) and combined roof and floor (50%). On indirect immunofluorescence, positivity was almost doubled with salt split technique ( 68%) as compared to routine method (36%). Thus, salt split technique was equivalent to routine on direct method in positivity with additional immunoreactant deposits noted in some and had double the sensitivity of the indirect method in detecting immunofluorescence in bullous pemphigoid.     

Human cutaneous topography a new photographic technique: observations on normal skin.

We describe herein a convenient and inexpensive technique for photographic examination of the surface of the skin. With this method, the fine lines of the skin appear as furrows or folds. The furrows demarcate units of epidermal structure which we have designated polyangular units. These, in turn, are arranged in a mosaic pattern. This technique was used to define the topography of normal young adults. Surface features fall into four major patterns: 1) dermatoglyphic, 2) hair bearing, 3) frictional, and 4) flexural.     

Fluorescence spectroscopy: a rapid, noninvasive method for measurement of skin surface thickness of topical agents

We report the quantification of skin surface thickness of topical agents by in vivo fluorescence spectroscopy, and demonstrate its potential uses for assessment of application technique and substantivity. A series of studies were performed on forearm skin of eight normal subjects using three creams which have intrinsic fluorescence: a sunscreen (Neutrogena SPF15 waterproof cream), an antiseptic (Hewlett's cream) and a steroid (Trimovate (clobetasone butyrate) cream). Initially, the dose-response relationship was established for each agent by applying a series of five doses (0.5-8μ/cm²) and measuring cream fluorescence using appropriate excitation and emission wavelengths. Next, the influence of application technique was examined by comparing light application of cream with firm rubbing. Substantivity of the three ceams was assessed on dry skin by taking fluorescence measurements over 8 h. Finally, water resistance of 2 μ/cm² of sunscreen and antiseptic cream were compared by measuring fluorescence after each of four water immersions. The fluorescence intensity was strongly correlated with the logarithm of surface density, r = 1.0, 0.92 and 0.98 for sunscreen, antiseptic and steroid creams, respectively, allowing derivation of a simple expression for equivalent thickness. Surface thickness of each cream was lower following firm rubbing compared with light application (P<0.01). The rate constants for reduction of surface density of the three creams with time on dry skin were not significantly different. However, on washed skin, the rate constant was higher rate for Hewlett's than Neutrogena cream (0.503 and 0.243 h. respectively. P= 0.02), with a higher rate for each cream on wet compared with dry skin (P <0.001). Hence, fluorescence spectroscopy is a simple, rapid method for measurement of cream thickness in vivo. The many potential applications in dermatology include quantitative assessment of application technique and substantivity of topical agents.     

Fluorescence excitation spectroscopy provides information about human skin in vivo

Fluorescence spectroscopy of human skin has the potential to provide useful morphologic and biochemical information. The endogenous fluorescence of human skin has been investigated in vivo on normal human volunteers as well as on patients with psoriasis and it was found that characteristic bands can be identified in the fluorescence spectra that are associated with specific skin fluorophores. One epidermal band (295 nm excitation, attributed to tryptophan) and two dermal bands (335 and 370 nm excitation, attributed to collagen cross-links) were consistently present in all fluorescence spectra. In addition, the fluorescence spectra obtained from lesions and nonlesional sites of psoriatic patients differed from those obtained from healthy volunteers and the hyperproliferative state of the lesions was characterized by a significantly larger signal at 295 nm excitation. These results indicate that fluorescence spectroscopy is a promising technique for the investigation of human skin in vivo.     

抗凋亡基因Bcl-2在皮肤良恶性肿瘤中的表达

为探讨Ｂｃｌ－２蛋白与皮肤肿瘤的关系，作者用流式细胞免疫荧光技术，检测了３３例皮肤良恶性病变。结果显示：Ｂｃｌ－２蛋白的相对含量ＦＩ（ｆｌｕｏｒｅｓｃｅｎｃｅ ｉｎｄｅｘ）在恶性肿瘤蝇的表达明显高于良性肿瘤，Ｐ〈０．００１。提示细胞恨抑止作用在肿瘤的发生、发展中也起着重要的作用，同时研究下调Ｂｃｌ－２的机制，可能成为Ｂｃｌ－蛋白过度表达肿瘤治疗的一种途径。     

Mechanobiology and force transduction in scars developed in darker skin types

BACKGROUND: Scarring is a complex process involving many cell types, cytokines and biological pathways including mechanobiology. Some subtle mechanical properties of skin can be assessed by measuring the speed of ultrasound shear wave propagation. The orientation of abnormal skin tension forces can be visualized, particularly in darker skin types, using dermoscopy showing distinct patterns of rete ridges' conformation. AIM: To assess some mechanobiological features of scars in darker skin types. PATIENTS AND METHODS: Large atrophic and hypertrophic surgical scars were examined on the trunk of 35 darker skin subjects. The surrounding skin was used as a comparator. Dermoscopic aspects were recorded. Resonance running time measurements (RRTM) were performed using a shear wave propagation device (Reviscometer). They were performed in four specific directions at given angles with regard to the long axis of the scar. The minimum, maximum and mean RRTM values were recorded at each site. RESULTS: Dermoscopy revealed patterns of melanin deposits in scars distinct from the normal honeycomb network seen in the surrounding skin. Hypertrophic scars showed a patchy pattern of large macular melanoderma dispersed in a lighter background. In these cases, low RRTM values were obtained with little variations according to the orientation of the measurements. By contrast, atrophic scars showed a streaky laddering melanotic pattern under dermoscopy. Higher RRTM values were often obtained, particularly in the transversal direction of the scars. Mechanical anisotropy was greater in the atrophic scars compared with the normal skin. DISCUSSION: Darker skin types represent a model for visualizing the main orientation of the epidermal rete ridges. A correlation was found between the pattern of melanized rete ridges of scars and the main orientation of the intrinsic forces in the skin.     

A pilot methodology study for the photographic assessment of post‐inflammatory hyperpigmentation in patients treated with tretinoin

Background Post‐inflammatory hyperpigmentation (PIH) is a common occurrence in patients with acne vulgaris, particularly in those with skin of colour. Aims A previous study has demonstrated the benefit of tretinoin (retinoic acid) in the treatment of PIH; however, there is currently no standard protocol to evaluate change in PIH following treatment. Based on these findings, we performed a pilot, exploratory, blinded, intraindividual‐controlled methodology study that consisted of a photographic assessment protocol with facial mapping. Materials and methods The study was based on a secondary analysis of a phase 4, community‐based trial of 544 acne patients who were treated with tretinoin gel microsphere 0.04% or 0.1%. Only patients with Fitzpatrick types III–V (skin of colour) were included in the study; subjects with Fitzpatrick skin type VI were excluded because the photographic assessment did not allow for proper evaluation. Results Despite the small number of subjects evaluated (n = 25), the results revealed consistent assessment of improvement in PIH between two independent graders (weighted κ = 0.84). Conclusion Further study with a larger population is recommended to validate the accuracy of this method.     

In vivo fluorescence of human skin. A potential marker of photoaging

Fluorescence is a feature of elastin and collagen, both major compounds of human dermis that are altered by age and photoexposure. We studied the intrinsic fluorescence of skin in vivo in 28 human volunteers to determine whether photoaging and chronologic aging of the skin could be evaluated by this noninvasive technique. We demonstrate that the excitation of skin autofluorescence by laser ultraviolet radiation yields characteristic tissue fluorescence spectra that are unrelated to age, pigmentation, or skin thickness. The differences in skin autofluorescence appear to be related to photoexposure. Thus, laser-induced fluorimetry, a noninvasive technique, may be adaptable as a marker of photoaging.     

Plantar hyperkeratosis: a study of callosities and normal plantar skin

Although callosities of the plantar skin are common and often disabling, little is known of their pathology or the reasons for their persistence. In this study plantar epidermal structure and cell renewal were investigated in patients with callosities and normal, age-, sex- and site-matched control subjects. Tritiated thymidine autoradiographic labeling indices were increased in the calluses but the dansyl chloride fluorescence clearance time was prolonged, reflecting the increased thickness of the stratum corneum. The number of corneocytes that could be removed from the surface of callosities by a standardized stimulus was considerably increased compared to controls but after adhesive tape stripping no such increase was observed. The density of corneocytes as measured on Percoll gradients was decreased in corneocytes from callus compared to normal plantar skin, and their volume was increased. These observations suggest that there are differences in epidermal differentiation due to an increased rate of epidermal cell production in plantar skin affected by callosity.