Barrier Function Of Human Skin Research Articles

P19 Defining the mechanism of skin barrier restoration by Lactobacillus rhamnosus

Abstract Introduction and aims We aim to investigate the lysate of Lactobacillus rhamnosus GG (LGG), a bacterium previously established as a gut probiotic, and to examine its postulated ability to enhance and restore human skin barrier function. Compromised skin barriers are implicated in various skin conditions including atopic dermatitis. Furthermore, skin barrier function decreases with age leading to dryness and sensitivity. Previous research has shown promising effects of LGG lysates on keratinocytes, including increasing transepidermal electrical resistance, increasing tight junction protein expression, cell migration promotion and mitigation of Staphylococcus aureus toxicity. This study aims to take a closer look at the molecular mechanisms behind the effect of LGG lysates on the skin, including terminal differentiation, lamin dispersal and nuclear degradation in keratinocytes. Methods Normal human epithelial keratinocytes and rat epithelial keratinocytes were grown and treated with the LGG lysate. Western blot analysis was used to determine the relative concentrations of tight junction proteins and other differentiation markers. A phosphoproteomic study was performed to identify key pathways activated by the LGG lysate. A Staphylococcus epidermidis lysate control was used to filter out any nonspecific bacterial effects. Immunostaining of nuclear proteins was used to visualize the effects of LGG lysate on lamin dispersal and nuclear degradation. short hairpin RNA filaggrin knockdown normal human keratinocytes were developed to create a ‘barrier impaired’ model to assess ability of the lysate to ‘rescue’ the skin barrier. Results The LGG lysate was shown to increase tight junction protein expression in normal human keratinocytes and increase keratinocyte terminal differentiation markers. We present Results that show decreased tight junction protein expression in filaggrin knockdown cells. Conclusions These Results show the potential for LGG lysate to enhance and restore skin barrier function and begin to suggest potential mechanisms for this effect.

Natural Moisturizing Factor-Enriched Formulations Compared to a Ceramide-Based Cream.

We aimed to investigate the effects of 2 ceramide plus natural moisturizing factor-enriched formulations compared to a ceramide-based cream on skin moisturization. Two double-blinded comparative studies were conducted, which enrolled 35 (n=29 females, n=6 males) and 33 (n=21 females, n=12 males) participants, respectively. Participants applied ceramide plus natural moisturizer cream or ceramide-based cream (study 1) or applied ceramide plus natural moisturizing factor lotion or ceramide-based cream (study 2) to each of their lower legs for 10 days with a 5-day regression period (no moisturizer applied). Skin hydration by corneometry after bilateral application was conducted once daily for each leg in both groups.   Results: An increase in corneometer units vs baseline for the ceramide plus natural moisturizing factor-enriched cream and natural moisturizing factor-enriched lotion were greater than the increase vs baseline for the ceramide-based cream at days 10 and 15; with an overall statistical significance in favor of the ceramide plus natural moisturizing factor-enriched formulations at day 10.  Conclusions: The marked improvement in skin moisturization following utilization of the ceramide plus natural moisturizing  factor-enriched cream and lotion compared to the ceramide-based cream can be attributed to the inherent properties of the natural moisturizing factors. These properties are known to maintain the humectancy and intercellular lipid membrane of the stratum corneum, which directly improves the permeability barrier function of human skin in reducing transepidermal water loss. J Drugs Dermatol. 2024;23(3):     doi:10.36849/JDD.8172.

Jonzac Thermal Spring Water Reinforces Skin Barrier Function of Human Skin and Presents a Soothing and Regenerating Effect

Jonzac Thermal Spring Water Reinforces Skin Barrier Function of Human Skin and Presents a Soothing and Regenerating Effect

Protective Effect of Potentilla glabra in UVB-Induced Photoaging Process.

Maintaining skin homeostasis is one of the most important factors for skin health. UVB-induced skin photoaging is a difficult problem that has negative impacts on skin homeostasis. So far, a number of compounds have been discovered that improve human skin barrier function and hydration, and are thought to be effective ways to protect skin homeostasis. Potentilla glabra var. mandshurica (Maxim.) Hand.-Mazz. Ethanol Extract (Pg-EE) is a compound that has noteworthy anti-inflammatory properties. However, its skin-protective effects are poorly understood. Therefore, we evaluated the capacity of Pg-EE to strengthen the skin barrier and improve skin hydration. Pg-EE can enhance the expression of filaggrin (FLG), transglutaminase (TGM)-1, hyaluronic acid synthase (HAS)-1, and HAS-2 in human keratinocytes. Moreover, Pg-EE down-regulated the expression of pro-inflammatory cytokines and up-regulated the production of FLG, HAS-1, and HAS-2 suppressed by UVB through inhibition of p38 mitogen-activated protein kinase (MAPK) and extracellular signal-regulated kinase (ERK) pathways. Given the above, since Pg-EE can improve skin barrier, hydration and reduce the UVB-induced inflammation on skin, it could therefore be a valuable natural ingredient for cosmetics or pharmaceuticals to treat skin disorders.

Keratinocyte Proliferative and Wound Healing Effects of Edible Bird's Nest Extract on Human Skin

Edible bird’s nest is a delicacy of Chinese cuisine. Its major components include mucin and sialic acid in addition to epidermal growth factor-like substances. However, its effects on skin cells and skin barrier function currently remain unknown. Therefore, we herein examined the effects of edible bird’s nest extract (EBN) on skin cell proliferation, wound healing, and tight junction proteins as well as human skin barrier function. The results obtained showed that EBN (0.1%) enhanced the proliferation of keratinocytes and fibroblasts as well as the migration and tight junction protein expression by keratinocytes. The oral ingestion of EBN (70 mg/day) standardized by sialic acid for 1 month reduced transepidermal water loss, the shallow wrinkle area, and dermal thickness. These results suggest that EBN exerts epidermal protective and smoothing effects through keratinocytes. Thus, EBN is a valuable ingredient for maintaining healthy skin.

Skin-Derived SPINK9 Kills Escherichia coli

Antimicrobial peptides play a critical role in the barrier function of human skin. They offer a fast response to invading microorganisms and protect from external microbial infection. Here we show the isolation of the kallikrein-related peptidase inhibitor SPINK9 as a major antibacterial factor from healthy stratum corneum. In total, six N-terminal SPINK9 variants were identified in the stratum corneum. Whereas all variants exhibited similar inhibition activities against kallikrein-related peptidase, only three variants with either lysine or glutamine as their first N-terminal residues were able to kill various Escherichia coli strains, but not other bacteria or fungi. The killing activity also depended on the sequence essential for kallikrein-related peptidase inhibition. Ultrastructural electron microscopy analyses suggested that SPINK9 entered the cell and killed growing bacteria. A bacterial chaperone, SKP, was identified as the major SPINK9 interacting partner in E.coli cells. The Skp-deleted mutant was more sensitive to SPINK9 than the wild-type control, suggesting that the bactericidal activity of SPINK9 should first overcome the resistance from the bacterial chaperone SKP. Thus, SPINK9 is a member of epidermal antimicrobial peptides for selective killing of E.coli, which might contribute to the innate barrier function of human skin.

LB1529 Repeated exposure to hard water and surfactant impairs epidermal barrier development in vitro

The development and organization of human skin barrier function after birth is a delicate adaptive process leading to a mature stratum corneum (SC) at the end of infancy. Before SC complete formation, skin is considered as immature and clinical data have demonstrated its fragility and sensitivity towards the domestic water hardness increasing the atopic dermatitis risk due to surfactants deposition. An experimental in vitro model recapitulating immature skin features has been developed based on a Reconstructed Human Epidermis (RHE) used at early stage of differentiation (day 12). In order to mimic a daily hygiene procedure according to standard surfactants exposure levels for cosmetics, the RHEs have been exposed since day 12 to a daily application (10 μL twice) of the following solutions: ultrapure water, hard water (HW) (400 mg/L calcium carbonate, equivalent to 40 F), Sodium Dodecyl Sulfate 0,05% (SDS) in ultrapure and hard water and compared to untreated RHEs. Transepithelial electrical resistance values, corresponding to barrier integrity, decreased in the RHE treated with SDS/HW compared to HW alone at day 14 (after 4 exposures) and 17 (after 3 days of recovery). Immunolabeling of K10 and K14 revealed significant modifications of their localization and expression levels in response to SDS/HW compared to HW, suggesting a delayed differentiation of the RHE. Involucrin, filaggrin and claudin-1 completely disappeared in the RHE treated with SDS/HW compared to HW at day 17, confirming clinical findings of barrier function impairment. The immature RHE model has shown a higher sensitivity compared to fully differentiated RHE and appears to be a reproducible and robust model to assess new ingredients or formulations intended to reduce the impact of surfactant in presence of hard water on the epidermal development and barrier formation in infancy.

Specific barrier response profiles after experimentally induced skin irritation in vivo.

BackgroundRecently, natural moisturizing factors (NMFs) and corneocyte surface topography were suggested as biomarkers for irritant dermatitis.ObjectivesTo investigate how exposure to different irritants influences corneocyte surface topography, NMF levels and the barrier function of human skin in vivo.MethodsEight healthy adult volunteers were exposed to aqueous solutions of 60% n‐propanol, 0.5% sodium lauryl sulfate (SLS), 0.15% sodium hydroxide, and 2.0% acetic acid, and distilled water, in a repeated irritation test over a period of 96 hours. Erythema, transepidermal water loss (TEWL), skin hydration, the dermal texture index (DTI) and NMF levels were measured at baseline, and after 24 and 96 hours.ResultsSLS and sodium hydroxide had the most pronounced effects on erythema and TEWL. Although n‐propanol caused only slight changes in TEWL and erythema, it showed pronounced effects on skin hydration, NMF levels, and the DTI. NMF was the only parameter that was significantly altered by all investigated irritants. The changes in the DTI were inversely associated with NMF levels and skin hydration.ConclusionSkin barrier impairment and the inflammatory response are irritant‐specific, emphasizing the need for a multiparametric approach to the study of skin irritation. NMF levels seem to be the most sensitive parameter in detecting irritant‐induced skin barrier alterations.

Diabetic and sympathetic influences on the water permeability barrier function of human skin as measured using transepidermal water loss: A case-control study.

The presence of long-standing hyperglycemic conditions has been suggested to lead to many skin problems associated with an impaired skin barrier function. However, the relationship between impaired skin barrier status and altered peripheral nervous system function has not yet been determined. The purpose of this study was to investigate the water evaporation rate as a measure of the permeability barrier function of diabetic skin and its relationship to diabetic sensorimotor polyneuropathy (DSPN) and peripheral autonomic neuropathy (PAN) using well-controlled confounding variables.This case-control study included 42 participants with chronic diabetes and 43 matched healthy controls. The diabetic group underwent a nerve conduction study and sympathetic skin response (SSR) test to confirm the presence of DSPN and PAN, respectively. Different skin regions were analyzed using the noninvasive Tewameter instrument (Courage + Khazaka Electronic GmbH, Cologne, Germany). The impacts of PAN, DSPN, age, and diabetes duration on the values of transepidermal water loss (TEWL) were each analyzed and compared between the groups.Regardless of the presence of DSPN or PAN, the TEWL values as measured on the distal extremities were significantly lower in the diabetic group than in the control group. In the diabetic group, participants with abnormal SSR test results showed decreased TEWL values in the finger, sole, and first toe, as compared with participants with normal SSR test results. In the control group, age showed a negative correlation with the TEWL values with respect to some measured regions. However, in the diabetic group, there was no significant correlation between either patient age or diabetes duration and TEWL values.The presence of a long-term hyperglycemic state can reduce the permeability barrier function of the skin, a phenomenon that might be related to the presence of an impaired peripheral sympathetic nervous system, rather than peripheral sensorimotor denervation.

Sixteen novel mutations in PNPLA1 in patients with autosomal recessive congenital ichthyosis reveal the importance of an extended patatin domain in PNPLA1 that is essential for proper human skin barrier function.

Autosomal recessive congenital ichthyosis (ARCI) is a genetically heterogeneous group of rare Mendelian skin disorders characterized by cornification and differentiation defects of keratinocytes. Mutations in nine genes including PNPLA1 are known to cause nonsyndromic forms of ARCI. To date, only 10 distinct pathogenic mutations in PNPLA1 have been reported. To identify new causative PNPLA1 mutations. We screened genetically unresolved cases, including our ARCI collection, comprising more than 700 families. Screening for mutations was performed either by direct Sanger sequencing or in combination with a multigene panel, followed by sequence and mutation analysis. Here we report on 16 novel mutations present in patients from 17 families. While all previously reported mutations and most of our novel mutations are located within the core patatin domain, we report five novel PNPLA1 mutations that are downstream of this domain. Thus, as recently described for PNPLA2, we hypothesize that a region larger than the core domain is required for full enzymatic activity of PNPLA1 in human skin barrier formation. We estimate the frequency of PNPLA1 mutations among patients with ARCI to be around 3%. Most of our patients were born as collodion babies and showed a relatively mild ichthyosis phenotype. In four unrelated patients we observed a cyclic scaling course, which seems to be a potential phenotypic variation in a small percentage of patients with PNPLA1 mutations. The variability of the clinical manifestations and the lack of typical clinical features are specific for patients with PNPLA1 mutations, and emphasize the importance of DNA sequencing for differential diagnosis of ARCIs.

Data-based modeling of drug penetration relates human skin barrier function to the interplay of diffusivity and free-energy profiles

Based on experimental concentration depth profiles of the antiinflammatory drug dexamethasone in human skin, we model the time-dependent drug penetration by the 1D general diffusion equation that accounts for spatial variations in the diffusivity and free energy. For this, we numerically invert the diffusion equation and thereby obtain the diffusivity and the free-energy profiles of the drug as a function of skin depth without further model assumptions. As the only input, drug concentration profiles derived from X-ray microscopy at three consecutive times are used. For dexamethasone, skin barrier function is shown to rely on the combination of a substantially reduced drug diffusivity in the stratum corneum (the outermost epidermal layer), dominant at short times, and a pronounced free-energy barrier at the transition from the epidermis to the dermis underneath, which determines the drug distribution in the long-time limit. Our modeling approach, which is generally applicable to all kinds of barriers and diffusors, allows us to disentangle diffusivity from free-energetic effects. Thereby we can predict short-time drug penetration, where experimental measurements are not feasible, as well as long-time permeation, where ex vivo samples deteriorate, and thus span the entire timescales of biological barrier functioning.

Assessment of skin barrier function and biochemical changes of ex vivo human skin in response to physical and chemical barrier disruption

Topical dermatotherapy is intended to be used on diseased skin. Novel drug delivery systems even address differences between intact and diseased skin underlining the need for pre-clinical assessment of different states of barrier disruption.Herein, we studied how short-term incubation in culture media compared to incubation in humidified chambers affects human skin barrier function and viability. On both models we assessed different types and intensities of physical and chemical barrier disruption methods with regard to structural integrity, biophysical parameters and cytokine levels.Tissue degeneration and proliferative activity limited the use of tissue cultures to 48h. Viability is better preserved in cultured tissue. Tape-stripping (50×TS) and 4h sodium lauryl sulfate (SLS) pre-treatment were identified as highly reproducible and effective procedures for barrier disruption. Transepidermal water loss (TEWL) values reproducibly increased with the intensity of disruption while sebum content and skin surface pH were of limited value. Interleukin (IL)-6/8 and various chemokines and proteases were increased in tape-stripped skin which was more pronounced in SLS-treated skin tissue extracts.Thus, albeit limited to 48h, cultured full-thickness skin maintained several barrier characteristics and responded to different intensities of barrier disruption. Potentially, these models can be used to assess pre-clinically the efficacy and penetration of anti-inflammatory compounds.

Japanese Cedar (Cryptomeria japonica) pollen allergen induces elevation of intracellular calcium in human keratinocytes and impairs epidermal barrier function of human skin ex vivo.

Cry j1 is the major peptide allergen of Japanese cedar (Sugi), Cryptomeria japonica. Since some allergens disrupt epidermal permeability barrier homeostasis, we hypothesized that Cry j1 might have a similar effect. Intracellular calcium level in cultured human keratinocytes was measured with a ratiometric fluorescent probe, Fura-2 AM. Application of Cry j1 significantly increased the intracellular calcium level of keratinocytes, and this increase was inhibited by trypsin inhibitor or a protease-activated receptor 2 (PAR-2) antagonist. We found that Cry j1 itself did not show protease activity, but application of Cry j1 to cultured keratinocytes induced a rapid (within 30s) and transient increase of protease activity in the medium. This transient increase was blocked by trypsin inhibitor or PAR-2 antagonist. The effect of Cry j1 on transepidermal water loss (TEWL) of cultured human skin was measured in the presence and absence of a trypsin inhibitor and PAR-2 antagonist. Cry j1 significantly impaired the barrier function of human skin ex vivo, and this action was blocked by co-application of trypsin inhibitor or PAR-2 antagonist. Our results suggested that interaction of Cry j1 with epidermal keratinocytes leads to the activation of PAR-2, which induces elevation of intracellular calcium and disruption of barrier function. Blocking the interaction of Cry j1 with epidermal keratinocytes might ameliorate allergic reaction and prevent disruption of epidermal permeability barrier homeostasis.

Influence of lidocaine hydrochloride and penetration enhancers on the barrier function of human skin

Skin penetration enhancers (SPEs) are commonly employed in pharmaceutical and personal care products. These compounds transiently alter the barrier properties of the skin and we have previously investigated the effects of specific SPEs on skin barrier function in vivo. In the present study the effects of incorporation of an active pharmaceutical ingredient (API), lidocaine hydrochloride (LID HCl) in the SPEs previously studied were investigated. Solutions of LID HCl were prepared and applied to the volar forearm of human subjects with occlusion for 24h. Subsequently, tape stripping and trans epidermal water loss (TEWL) measurements were conducted for treated and control sites. The activities of the desquamatory proteases, kallikrein 5 (KLK 5) and kallikrein 7 (KLK 7) and API content were also measured from the tape strips. The propylene glycol (PG) formulation increased TEWL significantly (p<0.05) compared with the other SPEs and a mixture of the SPEs. However, only the isopropyl myristate (IPM) solution altered protease activity with a significant observed increase in kallikrein 5 (KLK 5). Incorporation of LID HCl appeared to ameliorate the effects of some of the SPEs on TEWL measurements compared with our previous study. Overall uptake of LID HCl into skin from the various formulations correlated very well with changes in TEWL. The findings should have implications for the choice of SPEs in topical and transdermal formulations, particularly where the skin barrier function of patients is already impaired for example in atopic eczema or psoriasis.

Solar UV radiation reduces the barrier function of human skin.

The ubiquitous presence of solar UV radiation in human life is essential for vitamin D production but also leads to skin photoaging, damage, and malignancies. Photoaging and skin cancer have been extensively studied, but the effects of UV on the critical mechanical barrier function of the outermost layer of the epidermis, the stratum corneum (SC), are not understood. The SC is the first line of defense against environmental exposures like solar UV radiation, and its effects on UV targets within the SC and subsequent alterations in the mechanical properties and related barrier function are unclear. Alteration of the SC's mechanical properties can lead to severe macroscopic skin damage such as chapping and cracking and associated inflammation, infection, scarring, and abnormal desquamation. Here, we show that UV exposure has dramatic effects on cell cohesion and mechanical integrity that are related to its effects on the SC's intercellular components, including intercellular lipids and corneodesmosomes. We found that, although the keratin-controlled stiffness remained surprisingly constant with UV exposure, the intercellular strength, strain, and cohesion decreased markedly. We further show that solar UV radiation poses a double threat to skin by both increasing the biomechanical driving force for damage while simultaneously decreasing the skin's natural ability to resist, compromising the critical barrier function of the skin.

Quantification of human skin barrier function and susceptibility to quantum dot skin penetration

In this study, we utilize in vivo human skin and a viable ex-vivo human skin model to investigate the effect of a commercial depilatory agent on barrier function. Tape stripping was used as a positive control. The magnitude of skin barrier was quantified by measuring transepidermal water loss values on in vivo human skin and transepithelial electrical resistance measurements and tissue histology on ex vivo skin. The susceptibility to carboxylated quantum dot penetration through ex vivo skin was investigated using fluorescent microcopy analysis of microtomed skin sections and flow cytometry to quantify quantum dot association with live epidermal cells. Results show that depilatory treatment modifies the outside-in barrier sufficiently to allow quantum dots to penetrate the stratum corneum but to a lesser extent than tape stripping. The implications of these finding are discussed.

Effects of microneedle length, density, insertion time and multiple applications on human skin barrier function: Assessments by transepidermal water loss

Microneedle (MN) arrays have attracted considerable attention in recent years due to their ability to facilitate effective transdermal drug delivery. Despite appreciable research, there is still debate about how different MN dimensions or application modes influence permeabilization. This study aimed to investigate this issue by taking transepidermal water-loss measurements of dermatomed human skin samples following the insertion of solid polymeric MNs. Insertions caused an initial sharp drop in barrier function followed by a slower incomplete recovery – a paradigm consistent with MN-generation of microchannels that subsequently contract due to skin elasticity. While 600 μm-long MNs were more skin-perturbing than 400 μm MNs, insertion of 1000 μm-long MNs caused a smaller initial drop in integrity followed by a degree of long term permeabilization. This is explainable by the longest needles compacting the tissue, which then decompresses over subsequent hours. Multiple insertions had a similar effect as increasing MN length. There was some evidence that increasing MN density suppressed the partial barrier recovery caused by tissue contraction. Leaving MNs embedded in skin seemed to reduce the initial post-insertion drop in barrier function. Our results suggest that this in vitro TEWL approach can be used to rapidly screen MN-effects on skin.

Inter- and intra-individual variability in human skin barrier function: A large scale retrospective study

In vitro transepidermal tritiated water flux measurements are frequently used to evaluate skin barrier integrity for quality control purposes. However, research in this area to date has been largely based upon small-scale studies, each involving relatively few skin permeation measurements. In order to enhance our understanding in this area, we have conducted a much larger scale retrospective statistical analysis of tritiated water kp values. These values reflected the permeability of 2400 skin samples that were derived from 112 female volunteers over a 4 year period. It was found that the population of tritiated water kp values constituted a positively skewed, non-Normal distribution. Mean kp was 2.04 x 10(-3)cm/h while the 95th percentile was 4.50 x 10(-3)cm/h. Both values are higher than those reported in previous smaller studies. Hence, our study indicates that previously suggested upper limits for tritiated water flux are too low and that they be revised upwards to a value of 4.5 x 10(-3)cm/h. Analysis was also performed on smaller data subsets allowing inter-individual and intra-individual comparisons. For intra-individual kp variability, site-related differences yielded a non-Normal, positively skewed pattern in most individuals. Inter-individual variability was Normally-distributed and showed scatter that was much smaller in magnitude.

Routes and Modes of Administration of Resorcinol and Their Relationship to Potential Manifestations of Thyroid Gland Toxicity in Animals and Man

Medical case reports published in the 20th century over the course of several decades show that resorcinol caused reversible adverse effects on the human thyroid gland (TG) manifested as hypothyroidism. Affected patients had ulcerating leg varicosities and underwent prolonged treatment with ointments containing high concentrations of resorcinol. In animal studies resorcinol failed to induce TG toxicity, unless pharmacokinetic/toxicokinetic (PK/TK) conditions were manipulated (e.g., injection of resorcinol in oil or application in a slow release formulation). A recently completed two-generation reproductive toxicity study in rats did not detect any adverse effects on either reproductive or TG end points (Welsch, Nemec, and Lawrence, 2008, Int. J. Toxicol. 37, this issue). Resorcinol intake via drinking water up to the palatability limit had resulted in average daily intakes (mg/kg) of 233 in F0 and F1 males and 304 (premating/gestation) or 660 (lactation) in females. Free resorcinol in blood plasma was barely detectable in a few parental animals, indicating rapid metabolism. This short review communication offers a perspective on compromised human skin barrier function as a likely cause of drastic increases in resorcinol absorption. In conjunction with multiple daily applications over many months to hyperemic, inflamed, and lesioned human skin much higher absorption was likely responsible for the reported human TG toxicity.

A New Method of Measuring the Transepidermal Water Loss (TEWL) of Dog Skin

Human skin barrier function is evaluated by measuring transepidermal water loss (TEWL). However, this conventional method has not been applied to assess canine skin barrier function because the equipment is not suitable for dogs due to the effects of air turbulence resulting from movement of the subject and vapor from the subject's hair coat. The TEWL analyzer CC-01 was developed as a closed-chamber method device; this means that instead of using the open-chamber method, it has a ventilated chamber that uses dry air. TEWL values measured by CC-01 show less variability than those measured by the conventional method. An ambient temperature of 20-26 degrees C is optimal for measurement with the CC-01, and humidity affects the length of measurement but not the values. The CC-01 may be more reliable for measurement of TEWL than the conventional methods and may give new insights in the evaluation of skin barrier function in dogs.