Analysis of the morphometric parameters of pig ear hair follicles.

BACKGROUND: Porcine ear skin is used in studies of percutaneous penetration as a substitute for human skin. The objective of the present study was to determine the structure of the hair follicles on the dorsal area of porcine ear skin and make a morphometric comparison with the hair follicles of human skin. MATERIALS AND METHODS: Sections of frozen biopsies were cut vertically to the skin surface in longitudinal sections using a cryotome and were investigated using microscopy. For each hair follicle, various parameters were determined. RESULTS: The follicular density in porcine ear skin varies according to the area studied, and the length of most of the follicles was approximately 1458 ± 286 µm. The size of the follicular orifice was also determined in a total of 305 follicles. It showed a diameter of roughly 113 ± 43 µm. CONCLUSION: The results showed a very good similarity between human and pig hair follicles. Therefore, porcine ear skin can be considered as a very suitable model of human skin in dermal and especially follicular penetration studies.

A Dual Fluorescence-Spin Label Probe for Visualization and Quantification of Target Molecules in Tissue by Multiplexed FLIM-EPR Spectroscopy.

Simultaneous visualization and concentration quantification of molecules in biological tissue is an important though challenging goal. The advantages of fluorescence lifetime imaging microscopy (FLIM) for visualization, and electron paramagnetic resonance (EPR) spectroscopy for quantification are complementary. Their combination in a multiplexed approach promises a successful but ambitious strategy because of spin label-mediated fluorescence quenching. Here, we solved this problem and present the molecular design of a dual label (DL) compound comprising a highly fluorescent dye together with an EPR spin probe, which also renders the fluorescence lifetime to be concentration sensitive. The DL can easily be coupled to the biomolecule of choice, enabling in vivo and in vitro applications. This novel approach paves the way for elegant studies ranging from fundamental biological investigations to preclinical drug research, as shown in proof-of-principle penetration experiments in human skin ex vivo.

Application of parelectric spectroscopy to detect skin cancer-A pilot study.

BACKGROUND: The early detection of skin cancer is still challenging and calls for objective, fast diagnostic, and ideally non-invasive methods in order to leave the potentially malignant tumor cells unaltered. In this paper, the parelectric spectroscopy was applied to evaluate the potential of a non-invasive detection of basal cell carcinoma (BCC) and malignant melanoma. MATERIALS AND METHODS: A prototype of parelectric spectroscopy was used to investigate non-invasively dipole density and mobility of suspicious skin lesions. The differences in investigated tissue were analyzed compared to pathohistological findings in a clinical study on 51 patients with suspected BCC and malignant melanoma. RESULTS: The non-invasive parelectric spectroscopy could differentiate between normal skin, BCC, and melanoma but failed to distinguish between different types of skin cancer. The data were normalized to unsuspected nearby skin because the different skin locations influence dipole density and mobility. CONCLUSION: The results of the pilot study indicate that the parelectric spectroscopy might be an additional, useful non-invasive diagnostic procedure to distinguish between normal skin and skin cancer.

Laser scanning microscopy for control of skin decontamination efficacy from airborne particulates using highly absorbent textile nanofiber material in combination with PEG-12 dimethicone.

BACKGROUND: The decontamination of the skin is indispensable if airborne particulate contaminants deposit on the skin surface. Skin washing can have adverse effects as by skin rubbing the particles can be transferred deeply into the hair follicles, where they can be entrapped for a period of more than 10 days. Thus, alternative skin decontamination strategies are necessary. MATERIALS AND METHODS: For imaging the contaminants in the skin, sodium fluorescein-labeled soot particles of submicron size (≈600 nm) were visualized using laser scanning microscopy. RESULTS: In the present ex vivo pilot study on porcine ear skin, it was shown that sodium fluorescein-labeled soot particles of submicron size (≈600 nm) could be efficiently removed from the skin with highly absorbent textile nanofiber material, whose efficacy could be further increased by spraying the contaminated skin area with the viscous fluid PEG-12 dimethicone before textile application. CONCLUSION: In case of skin contamination with particulates, the contact washing should be avoided due to rubbing particles deeply into the hair follicles, where they can accumulate for a long time and induce negative consequences. Efficient skin decontamination could include pretreatment of skin surface with the viscous fluid PEG-12 dimethicone and subsequent application of highly absorbent textile nanofiber material.

Solvent-Containing Closure Material Can Be Used to Prevent Follicular Penetration of Caffeine and Fluorescein Sodium Salt on Porcine Ear Skin.

AIM: The skin represents a drug delivery portal. The establishment of a skin model capable of distinguishing between the follicular and intercellular penetration pathways remains a challenge. The study described herein was aimed at showing the influence of two nail varnishes as closure material and four application techniques to spread the active pharmaceutical ingredient (API) on a successful follicular closure without inducing penetration-enhancing effects. MATERIALS AND METHODS: For all experiments, ex vivo porcine ear skin was used. In study design A, a standard and a solvent-free nail varnish were compared. It was tested whether the different application techniques (spreading with pipette, careful finger massage, 5-Hz finger massage, 5-Hz automatic massage) potentially destroy an intact follicular closure. Laser scanning microscopy imaging was used to measure if the model drug (fluorescein sodium salt) penetrated into the hair follicles. Study design B investigated how the penetration is affected when applying standard nail varnish containing solvents to skin. It was tested if the varnish blocks the API (caffeine) on completely covered areas and if adjacent areas show increased penetration. Furthermore, lateral diffusion of the API was investigated. After 20 h, the skin layers were separated by tape stripping and heat separation. The tissue samples were homogenized. Caffeine was quantified by chromatography. RESULTS: In study design A, the standard nail varnish showed a secure follicular closure, while the solvent-free nail varnish was not able to prevent follicular penetration. Moreover, rapid application techniques were found to destroy an intact follicular closure. Only the two most gentle application techniques kept the follicular closing intact. In study design B, no caffeine was detected in both skin areas that were completely covered. Since no significant difference in caffeine penetration between the two uncovered groups was found, any influence of the applied closure material on adjacent areas was excluded. CONCLUSION: This study clearly demonstrates that a standard nail varnish in combination with a gentle application technique of the API provides a secure follicular closure. The presented study only investigated the closure for the substances caffeine and fluorescein sodium salt. The results might not be transferable to all kinds of APIs.

Determination of the pH Gradient in Hair Follicles of Human Volunteers Using pH-Sensitive Melamine Formaldehyde-Pyranine Nile Blue Microparticles.

Nanoparticles can be applied to the hair follicles, which can serve as reservoirs for triggered drug release. A valid measurement method for the determination of the pH within the hair follicle in vivo has not been shown yet. Here, melamine formaldehyde particles up to 9 µm in size were applied on 40 freshly plucked scalp hairs of eight individuals to determine the pH along the hair shaft down to the root area of the hair. For fluorescent pH indicators, pyranine and Nile blue were incorporated into the particles. Measurements were conducted using confocal laser scanning microscopy. A pH decay gradient could be found from the hair sheath towards the external hair shaft (p = 0.012) with pH values at the hair sheath of 6.63 ± 0.09, at the hair sheath end at 6.33 ± 0.11, and at the external hair shaft at 6.17 ± 0.09 (mean ± SE). The pH difference between the hair sheath end and the external hair shaft was found to be significant (p = 0.036). The results might be comparable with the pH within the hair follicle in vivo indicating a pH increase towards the hair root.

Influence of Storage and Preservation Techniques on Egg-Derived Carotenoids: A Substantial Source for Cutaneous Antioxidants.

Antioxidants like carotenoids play a major role in the prevention of the destructive influence of free radicals in our skin. Carotenoids, as well as all other antioxidants, are substantial substances which must be supplied by nutrition. Resonance Raman spectroscopy (RRS) allows measurement of the carotenoid content of eggs, representing a rich carotenoid source in our nutrition. A previous study showed that eggs from organic production contain higher carotenoid levels in contrast to eggs from conventionally housed chicken. The uptake of these organically produced eggs led to an increased antioxidant concentration in the skin. In this study, the effects of different storage modalities, conservation techniques, and the effects of food processing on the carotenoid levels in eggs were investigated with RRS. Common storage modalities and preservation techniques showed only a limited influence on egg-derived carotenoid concentrations. However, a colder environment (at least for shell eggs) and high-pressure preservation had the best preservative influence on the carotenoid content. Surprisingly, food processing such as boiling increased the carotenoid concentration in eggs, whilst broiling destroyed the carotenoids almost completely. In conclusion, RRS is suitable for monitoring egg-derived carotenoid levels, and carotenoid levels in eggs are generally stable under common storage and preservation modalities. Boiling in contrast to broiling of eggs might be superior in terms of carotenoid preservation within food processing.

Laser Scanning Microscopic Investigations of the Decontamination of Soot Nanoparticles from the Skin.

BACKGROUND/AIMS: Airborne pollutants, such as nano-sized soot particles, are increasingly being released into the environment as a result of growing population densities and industrialization. They can absorb organic and metal compounds with potential biological activity, such as polycyclic aromatic hydrocarbons and airborne pollen allergens. Local and systemic toxicities may be induced in the skin if the particulates release their harmful components upon dermal contact. METHODS: In the present study, skin pretreatments with serum and/or shield as barrier formulations prior to exposure and washing with a cleanser subsequent to exposure were evaluated as a protection and decontamination strategy using laser scanning microscopy. RESULTS: The results indicate that while the application of serum and a cleanser was insufficient for decontamination, the pretreatment with shield prior to nanoparticle exposure followed by washing led to the removal of a considerable amount of the carbon black particles. The combined application of serum and shield before the administration of carbon black particles and subsequent washing led to their elimination from the skin samples. CONCLUSION: The application of barrier-enhancing formulations in combination with a cleanser may reduce the penetration of harmful airborne particulates by preventing their adhesion to the skin and facilitating their removal by subsequent washing with the cleanser.

From UV Protection to Protection in the Whole Spectral Range of the Solar Radiation: New Aspects of Sunscreen Development.

Sunscreens have been constantly improving in the past few years. Today, they provide an efficient protection not only in the UVB but also in the UVA spectral region of the solar radiation. Recently it could be demonstrated that 50% of all free radicals induced in the skin due to solar radiation are formed in the visible and infrared spectral region. The good protective efficacy of sunscreens in the UV region prompts people to stay much longer in the sun than if they had left their skin unprotected. However, as no protection in the visible and infrared spectral region is provided, high amounts of free radicals are induced here that could easily exceed the critical radical concentration. This chapter describes how the effect of sunscreens can be extended to cover also the visible and infrared spectral region of the solar radiation by adding pigments and antioxidants with high radical protection factors to the sunscreen formulations.

Prevention of Cutaneous Penetration and CD1c+ Uptake of Pollen Allergens by a Barrier-Enhancing Formulation.

Recent studies have shown that pollen proteins can penetrate the impaired skin barrier of atopic patients and exacerbate their disease. In the presented study the effect of a topically applied barrier-enhancing formulation was investigated for its preventive effect on the uptake of pollen allergens into CD1c+ epidermal cells. The pollen proteins were fluorescence labelled and applied on barrier-disrupted excised human skin. CD1c+ cells were selected after magnetic cell sorting and analysed using laser scanning microscopy. In untreated disrupted skin, 81% of the CD1c+ cells contained the fluorescence-labelled pollen allergens. In formulation-pretreated skin only 12% of the CD1c+ cells showed an uptake of pollen allergens. These results encourage the treatment of atopic patients with barrier-enhancing formulations to reduce the impact of pollen on air-exposed skin areas and hence the exacerbation of cutaneous symptoms.

Relationship between Histological and Clinical Course of Psoriasis: A Pilot Investigation by Reflectance Confocal Microscopy during Goeckerman Treatment.

Alterations of the skin microvasculature are known to play an important role in the development and maintenance of psoriatic skin lesions. In this study, we investigated lesional skin in 11 psoriatic patients during a modified Goeckerman treatment using reflectance confocal microscopy (RCM) to study the relationship between clinical clearance and histological normalization of psoriatic skin and the significance of histological abnormalities on the course of disease. The treatment regimen resulted in a significant reduction of the Psoriasis Area and Severity Index (PASI) as well as capillary and papillary diameters (p < 0.0001). The capillary and papillary diameters were still enlarged when compared to those in normal skin (p < 0.001). Capillary and papillary diameters correlated with each other prior to and after treatment (correlation coefficient = 0.63 and 0.64, p = 0.01 and 0.002, respectively) but not with the PASI. Capillary and papillary diameters after treatment and percentage reduction of the PASI during treatment seemed to be better predictors for the clinical course of relapse than the PASI after treatment. These findings make the subclinical changes of psoriatic skin vessels and dermal papillae a legitimate target for treatment. Further investigations of a large group of patients are needed to evaluate the potential of RCM findings as successor of the PASI in the monitoring of psoriasis.

Influence of Topical, Systemic and Combined Application of Antioxidants on the Barrier Properties of the Human Skin.

BACKGROUND: The formation of free radicals in human skin by solar ultraviolet radiation is considered to be the main reason for extrinsic skin aging. The antioxidants in human tissue represent an efficient protection system against the destructive action of these reactive free radicals. In this study, the parameters of the skin, epidermal thickness, stratum corneum moisture, elasticity and wrinkle volume, were determined before and after the treatment with antioxidant- or placebo-containing tablets and creams. METHODS: The study included 5 groups of 15 volunteers each, who were treated for 2 months with antioxidant-containing or placebo tablets, creams or a combination of antioxidant-containing tablets and cream. The skin parameters were measured at time point 0 and at week 8 utilizing ultrasound for the determination of epidermal thickness, a corneometer for stratum corneum moisture measurements, skin profilometry for quantifying the wrinkle volume and a cutometer for determining the elasticity. RESULTS: The verum cream had a positive influence on epidermal thickness, elasticity and skin moisture, but the verum tablets improved the epidermal thickness only. The combined application of verum tablets and creams led to a significant improvement of all investigated skin parameters, whereas the application of placebo tablets or cream did not influence any parameters. CONCLUSION: The topical and oral supplementation of antioxidants can be an instrument to improve several skin parameters and potentially counteract or decelerate the process of extrinsic skin aging.

Penetration of topically applied nanocarriers into the hair follicles of dog and rat dorsal skin and porcine ear skin.

BACKGROUND: In humans, topically applied nanocarriers penetrate effectively into the hair follicles where they can be exploited for the localized and targeted treatment of skin disorders. OBJECTIVE: The objective of the present study was to examine the applicability of particle-based systems for follicular drug delivery in companion animals and livestock, which have a large follicular reservoir. ANIMALS: Skin samples from 10 beagle dogs, 14 Wistar rats and four ears from freshly slaughtered cross-bred pigs were used. METHODS: Fluoresceinamine labelled poly (L-lactide-co-glycolide) nanocarriers (256 or 430 nm) were applied on the different skin samples. After penetration, skin biopsies were removed and cryohistological cross sections prepared and investigated with regard to the follicular penetration depths (in µm ± standard deviation) of the nanocarriers using confocal laser scanning microscopy. RESULTS: In canine, rat and porcine hair follicles, the smaller nanoparticles were detected at mean follicular penetration depths of 630.16 ± 135.75 µm, 253.55 ± 47.36 µm and 653.40 ± 94.71 µm, respectively. The larger particles were observed at average follicular depths of 604.79 ± 132.42 µm; 262.87 ± 55.25 µm and 786.81 ± 121.73 µm, respectively, in canine, rat and porcine hair follicles. Statistically significant differences (P < 0.05) in the mean follicular penetration depths of the differently sized nanocarriers could be determined for the canine and porcine skin samples. CONCLUSION: The mean follicular penetration depths of the differently sized nanocarriers were mostly significantly different between the different species, which might be due to different species-specific follicular dimensions. This issue needs to be addressed specifically in further studies.

In vivo study for the discrimination of cancerous and normal skin using fibre probe-based Raman spectroscopy.

Raman spectroscopy has proved its capability as an objective, non-invasive tool for the detection of various melanoma and non-melanoma skin cancers (NMSC) in a number of studies. Most publications are based on a Raman microspectroscopic ex vivo approach. In this in vivo clinical evaluation, we apply Raman spectroscopy using a fibre-coupled probe that allows access to a multitude of affected body sites. The probe design is optimized for epithelial sensitivity, whereby a large part of the detected signal originates from within the epidermal layer's depth down to the basal membrane where early stages of skin cancer develop. Data analysis was performed on measurements of 104 subjects scheduled for excision of lesions suspected of being malignant melanoma (MM) (n = 36), basal cell carcinoma (BCC) (n = 39) and squamous cell carcinoma (SCC) (n = 29). NMSC were discriminated from normal skin with a balanced accuracy of 73% (BCC) and 85% (SCC) using partial least squares discriminant analysis (PLS-DA). Discriminating MM and pigmented nevi (PN) resulted in a balanced accuracy of 91%. These results lie within the range of comparable in vivo studies and the accuracies achieved by trained dermatologists using dermoscopy. Discrimination proved to be unsuccessful between cancerous lesions and suspicious lesions that had been histopathologically verified as benign by dermoscopy.

Comparative study of hair follicle morphology in eight mammalian species and humans.

BACKGROUND: The objective of the present study was the investigation of hair follicle morphology in eight mammalian species in order to evaluate the species-specific contribution of hair follicles to skin penetration particularly with regard to the utilization of the different animal species as skin models for human skin. METHODS: Cyanoacrylate skin surface biopsy method (CSSB), light microscopy and also digital photography were used for the measurements of hair follicle morphology. RESULTS: The results revealed species-specific differences regarding the pattern of hair follicle distribution and also differences with regard to hair follicle parameters and characteristics. The results also showed that hair follicles generally possess enormous reservoir capacities, regarding the follicular volume. In all examined species, hair follicles reached at least one-fifth of stratum corneum storage capacity. The results were compared with human data obtained in a previous study. CONCLUSION: With regard to hair follicle morphology and skin structure, the porcine skin seems to be the most appropriate skin model for human skin analog to previous investigations, whereas the skin of dog, cat, and rabbit showed the most significant differences.

New strategies for preoperative skin antisepsis.

During the past decades, encouraging progress has been made in the prevention of surgical site infections (SSI). However, as SSI still occur today, strategic prevention measures such as standardized skin antisepsis must be implemented and rigorously promoted. Recent discoveries in skin physiology necessitate the development of novel antiseptic agents and procedures in order to ameliorate their efficacy. In particular, alternate target structures in the skin need to be taken into consideration for the development of the next generation of antiseptics. Recent investigations have shown that a high number of microorganisms are located within and in the close vicinity of the hair follicles. This suggests that these structures are an important reservoir of bacterial growth and activity in human skin. To date, it has not been fully elucidated to what extent conventional liquid antiseptics sufficiently target the hair follicle-related microbial population. Modern technologies such as tissue-tolerable plasma (TTP) have been tested for their potential antiseptic efficiency by reducing the bacterial load in the skin and in the hair follicles. First experiments using liposomes to deliver antiseptics into the hair follicles have been evaluated for their potential clinical application. The present review evaluates these two innovative methods for their efficacy and applicability in preoperative skin antiseptics.

Palmoplantar erythrodysesthesia-like skin symptoms in patients under various chemotherapeutics: preventive and therapeutic options.

BACKGROUND/AIMS: The palmoplantar erythrodysaesthesia (PPE) is an inflammatory cutaneous side effect in patients under chemotherapy with pegylated liposomal doxorubicin (PLD), with indications that also other chemotherapeutics induce similar side effects. Recently, it has been demonstrated that PLD escapes with the sweat onto the skin inducing radical-forming processes that damage the skin. The topical application of antioxidants with a high radical protection factor has proven to be a very efficient prevention strategy for PLD-treated patients. METHODS: 68 patients, who had been treated with 12 different chemotherapeutics and experienced side effects similar to PPE, were treated with a meanwhile commercially available ointment. RESULTS: At the beginning of the therapy, 46 patients suffered from a PPE of severity grade III, while in 22 patients a PPE of severity grade II was diagnosed. The application of the ointment resulted in a significant improvement of the clinical symptoms and the skin status in all these patients; their chemotherapies could be continued. CONCLUSION: The obtained results suggest that radical-forming processes play an essential role in a great number of chemotherapeutics which induce dermal side effects. The topical application of the antioxidant-containing ointment proved to be a good therapeutic option which needs further evaluation.

Efficient prevention strategy against the development of a palmar-plantar erythrodysesthesia during chemotherapy.

BACKGROUND: Pegylated liposomal doxorubicin (PLD) is a highly efficient chemotherapeutic; however, it induces dermal side effects such as palmar-plantar erythrodysesthesia (PPE) in up to 80% of cases, probably by being emitted with the sweat onto the skin surface. AIM: The aim of the present study was to examine whether a topically applied ointment containing antioxidants with a high radical protection factor is able to prevent the formation of PPE. METHODS: Twenty patients suffering from ovarian carcinoma and treated with PLD were observed. RESULTS: 60% of the patients tolerated the regular application of the cream and developed no PPE. The remaining 40% interrupted the application. Six of them developed PPE and resumed ointment application thereafter. In these cases the PPE disappeared or was strongly reduced. CONCLUSION: The results of the observation clearly demonstrate that topical application of the ointment is an efficient strategy against the development of PPE during chemotherapy with PLD.

Antioxidants in Asian-Korean and caucasian skin: the influence of nutrition and stress.

BACKGROUND: The antioxidant status of the human skin provides protection against the destructive action of free radicals. Most antioxidants cannot be synthesized by the human organism itself, but have to be ingested with a healthy nutrition rich in fruit and vegetables. The Korean cuisine is known to be one of the healthiest worldwide. This binational study investigated the cutaneous carotenoid concentrations in German subjects, South Korean subjects and immigrant Korean subjects resident in Germany and examined whether dietary- and lifestyle-related differences are reflected in the cutaneous carotenoid concentrations. METHODS: Measurements of the carotenoid concentrations of 714 healthy volunteers were performed using a non-invasive spectroscopic measurement system based on reflectance spectroscopy. RESULTS: In the present study South Korean residents showed a significantly higher antioxidant status than both native German residents and Korean immigrants living in Germany (p < 0.001). The first generation of Korean immigrants to Germany over the age of 50 mostly preserved Korean dietary habits, showing significantly higher concentrations (p < 0.001) than the German-born second and third Korean generations under the age of 50. CONCLUSION: The results of the study indicate that a healthy nutrition alone does not provide a high antioxidant status unless the stress exposure can be reduced simultaneously.

Approach towards developing a novel procedure to selectively quantify topically applied substances in the hair follicles of the model tissue porcine ear skin.

Hair follicles represent reservoirs for localized drug therapy and transport pathways for systemic drug delivery. This study describes an approach towards developing a novel procedure for quantifying topically applied substances located in the hair follicles of porcine ear skin, a model for human in vivo skin, using a fluorescent dye. Approximately 5% of the topically applied dye was recovered from the hair follicles, which is in accordance with a previous study.