Stimulation of the Extracellular Matrix Production in Dermal Fibroblasts by Velvet Antler Extract

Background

Fibroblasts produce many components of the extracellular matrix (ECM) and so they contribute to the maintenance of connective tissue integrity.

Objective

The aim of this study is to evaluate the effect of velvet antler extract (VAE) on the ECM production of dermal fibroblasts cultured in vitro.

Methods

Primary cultured human dermal fibroblasts were treated with VAE, and then the ECM production was determined by RT-PCR, ELISA and Western blot analysis. Furthermore, the change of gene expression according to VAE treatment was evaluated by cDNA microarray.

Results

VAE accelerated the growth of fibroblasts in a dose-dependent manner. VAE increased the production of several ECM components, including type 1 collagen, fibronectin and elastin. In line with these results, the phosphorylations of p42/44 ERK and p38 mitogen-activated protein kinase were markedly increased by VAE, suggesting that the enhancement of ECM production may be linked to the activation of intracellular signaling cascades. VAE also significantly increased cell migration on an in vitro scratch wound test. In cDNA microarray, many genes related with connective tissue integrity were identified to be up-regulated by VAE.

Conclusion

These results suggest that VAE has a potential to stimulate ECM production, and VAE may be applicable for maintaining the skin''s texture.     

Effects of Human Adipose-derived Stem Cells on Cutaneous Wound Healing in Nude Mice

Background

Despite numerous treatments available for deteriorated cutaneous wound healing such as a diabetic foot, there is still the need for more effective therapy. Adipose-derived stem cells (ASCs) are mesenchymal stem cells, which are self-renewing and multipotent. Mesenchymal stem cells have the potential for tissue repair and regeneration.

Objective

To investigate the effects of human ASCs on the healing of cutaneous wounds in nude mice.

Methods

15-mm round full-thickness skin defects were generated on the back of BALB/c nude mice. The mice were divided into three groups for wound coverage: (i) human ASCs-populated collagen gel, (ii) human dermal fibroblasts-populated collagen gel, and (iii) collagen gel alone. Wound contraction was prevented with a splint method. Wound size was measured 10 days after injury. At 28 days histological analysis was performed.

Results

Both ASCs and dermal fibroblasts accelerated wound closure, but dermal fibroblasts were more effective than ASCs. At 28 days, the dermal portion of ASCs or dermal fibroblasts wound scars were thicker than collagen gel wound scars.

Conclusion

ASCs and dermal fibroblasts stimulate cutaneous wound healing and improve scar thickness.     

Effects of Xanthium stramarium and Psoralea corylifolia Extracts Combined with UVA1 Irradiation on the Cell Proliferation and TGF-β1 Expression of Keloid Fibroblasts

Background

Xanthium stramarium (XAS) and Psoralea corylifolia (PSC), phototoxic oriental medicinal plants, has been used in traditional medicines in Asian countries.

Objective

The effects of highly purified XAS or PSC extract combined with ultraviolet A1 (UVA1) irradiation on cell proliferation and transforming growth factor-beta1 (TGF-β1) expression of the keloid fibroblast were being investigated to define potential therapeutic uses for keloid treatments.

Methods

The keloid fibroblasts were treated with XAS or PSC alone or in the combination with UVA1 irradiation. The cell viability, apoptosis, and expression of TGF-β1 and collagen I were investigated.

Results

XAS and PSC in combination with UVA1 irradiation suppressed cell proliferation and induced apoptosis of keloid fibroblasts. Furthermore, the XAS and PSC in combination with UVA1 irradiation inhibited TGF-β1 expression and collagen synthesis in keloid fibroblasts.

Conclusion

These findings may open up the possibility of clinically used XAS or PSC in combination with UVA1 irradiation for keloid treatments.     

Cholesterol, a Major Component of Caveolae, Down-regulates Matrix Metalloproteinase-1 Expression through ERK/JNK Pathway in Cultured Human Dermal Fibroblasts

Background

Cholesterol is a major component of specialized membrane microdomains known as lipid rafts or caveolae, which modulate the fluidity of biological membranes. Membrane cholesterol therefore plays an important role in cell signaling and vesicular transport.

Objective

In this study, we investigated the effects of cholesterol on matrix metalloproteinase-1 (MMP-1) expression in human dermal fibroblasts.

Methods

MMP-1 mRNA and protein expression were determined by RT-PCR and Western blotting, respectively. AP-1 DNA binding activity was detected by electrophoretic mobility shift assays. The amount of cholesterol was analyzed by cholesterol assay kit.

Results

We observed that MMP-1 mRNA and protein expression was dose-dependently decreased by cholesterol treatment. In contrast, cholesterol depletion by a cholesterol depletion agent, methyl-beta-cyclodextrin (MβCD) in human dermal fibroblasts, increased MMP-1 mRNA and protein expression in a dose-dependent manner. Also, we investigated the regulatory mechanism of MβCD-induced MMP-1 expression: cholesterol depletion by MβCD, activated ERK1/2 and JNK, but not p38 MAPK cascade, and it also significantly increased c-Jun phosphorylation, c-Fos expression and activator protein-1 binding activity. Furthermore, the inhibition of ERK or JNK with specific chemical inhibitors prevented MβCD-induced MMP-1 expression, which indicates that ERK and JNK play an important role in cholesterol depletion-mediated MMP-1 induction. In addition, MβCD-induced phosphorylation of ERK and JNK and MMP-1 expression were suppressed by cholesterol repletion.

Conclusion

Our results suggest that cholesterol regulates MMP-1 expression through the control of ERK and JNK activity in human dermal fibroblasts.     

Comparison of Gene Expression Profiles between Keratinocytes,Melanocytes and Fibroblasts

Background

The skin has many important functions such as protection, preservation, temperature regulation, and vitamin D synthesis. It is composed of a variety of cell types including keratinocytes, melanocytes and fibroblasts.

Objective

We attempted to compare the gene expression profiles between keratinocytes, melanocytes and fibroblast, using cDNA microarray.

Methods

Keratinocytes, melanocytes and fibroblasts were primary cultured from five foreskin specimens. Total RNAs were extracted and pooled to reduce the individual variations, and then used for cDNA microarray.

Results

Total 12,028 genes were selected as the reliable genes whose expression was detected in at least one of the three cell types. By comparing the relative expression levels with cutoff limitation as a fourfold change, we obtained 126 fibroblast-specific, 179 keratinocyte-specific and 173 melanocyte-specific genes, many of which are known to be characteristically expressed in each cell type. In addition, we identified many genes whose skin-specific functions have not yet been determined.

Conclusion

Our data provide important information on which to base further investigation into the specification of skin cell types.     

Tissue immunostaining for factor XIIIa in dermal dendrocytes of pityriasis alba skin lesions

BACKGROUND

Pityriasis alba affects 1% of the world population and about 9.9% of the children in Brazil. However, its etiology remains uncertain.

OBJECTIVE

The objective of the present study was to evaluate the immunoexpression of factor XIIIa in dermal dendrocytes of skin lesions of pityriasis alba.

METHOD

Twenty patients with pityriasis alba and 20 patients with atopic dermatitis underwent biopsy. The dermal dendrocytes marked by factor XIIIa were counted by means of immunohistochemical analysis.

RESULTS

The mean amount of dermal dendrocytes found in the patients with pityriasis alba was 2, whereas in the patients with atopic dermatitis it was 4, with a statistically significant difference between them. A cutoff point of 3 cells/square inch was established to differentiate pityriasis alba from atopic dermatitis, with 80% sensibility and 90% specificity.

CONCLUSION

We believe that pityriasis alba and atopic dermatitis should be considered different clinical forms within the spectrum of atopic disease, in which sun radiation plays an important role by modulating the progression of the disease.     

Notch Intracellular Domain Expression in Various Skin Fibroproliferative Diseases

Background

The effects of the Notch signaling pathway in fibroproliferative skin diseases have not been fully elucidated.

Objective

The aim of this study was to investigate the expression of activated Notch signaling molecules in various skin fibroproliferative diseases.

Methods

Immunohistochemical analysis of Notch intracellular domain (NICD) expression in keloid, hypertrophic scar, morphea, dermatofibroma, and normal control skin specimens was performed, and the clinical characteristics of patients with various skin fibroproliferative diseases were analyzed.

Results

NICD was highly expressed in fibroblasts of keloids and moderately to highly expressed in hypertrophic scars and dermatofibromas, whereas low or no expression was detected in the fibroblasts of normal skin specimens and morpheas. NICD was constitutively expressed in keratinocytes, endothelial cells, and immune cells in normal skin specimens.

Conclusion

NICD was significantly expressed in human fibroproliferative skin disorders, especially keloids, suggesting that an activated Notch signaling pathway is involved in the pathogenesis of skin fibrosis.     

A Randomized, Open-Label, Multicenter Trial of Topical Tacrolimus for the Treatment of Pruritis in Patients with Atopic Dermatitis

Background

Pruritis caused by atopic dermatitis (AD) is not always well controlled by topical corticosteroid therapy, but use of tacrolimus often helps to soothe such intractable pruritis in clinical settings.

Objective

To determine the anti-pruritic efficacy of topical tacrolimus in treating AD in induction and maintenance therapy.

Methods

Prior to the study, patients were randomly allocated into two groups, induction therapy followed by tacrolimus monotherapy maintenance, and induction therapy followed by emollient-only maintenance. In the induction therapy, the patients were allowed to use topical tacrolimus and emollients in addition to a low dose (<10 g/week) of topical steroids. Patients showing relief from pruritis were allowed to proceed to maintenance therapy. Recurrence of pruritis in maintenance therapy was examined as a major endpoint.

Results

Two-thirds of patients (44/68; 64.7%) showed relief from pruritis after induction therapy. Pruritis recurred in 23.8% (5/21) of the tacrolimus monotherapy group and in 100% (21/21) of the emollient group during maintenance period, a difference that was statistically significant.

Conclusion

Use of topical tacrolimus is effective in controlling pruritis of AD compared to emollient.     

Oxidative Damage and Nuclear Factor Erythroid 2-Related Factor 2 Protein Expression in Normal Skin and Keloid Tissue

Background

Reactive oxygen species (ROS) play an important role in the induction of apoptosis under pathological conditions. Recently, a significant increase in ROS production and disrupted apoptosis mechanisms in keloids have been reported. Nuclear factor erythroid 2-related factor 2 (Nrf2) represents one of the most important cellular defense mechanisms against oxidative stress and is implicated in the regulation of apoptosis. Recently, it has been reported that Nrf2 upregulates Bcl-2, an anti-apoptotic protein.

Objective

To compare Nrf2 protein expression in normal skin tissues to keloid tissues.

Methods

ROS generation in keloid tissues was evaluated with OxyBlot analysis. Western blotting and/or immunohistochemical staining approaches were used to study expression of Nrf2 or Bcl-2 in keloid and normal skin tissues. Cellular fractionation was performed to examine subcellular distribution of Nrf2. Transfection of fibroblasts with Nrf2-specific small interfering RNA (siRNA) was conducted to understand the relationship between Nrf2 expression and apoptosis induction.

Results

Protein oxidation, a marker of oxidative stress, is increased in keloid tissues. Western blot analysis clearly showed that Nrf2 and Bcl-2 are downregulated in keloid tissues. Immunohistochemical staining of Nrf2 confirmed the results of the western blot analysis. Transfection of fibroblasts with the Nrf2-specific siRNA results in increased apoptosis and decreased cell viability.

Conclusion

Collectively, our data indicate that Nrf2 expression is downregulated in keloid tissues, and that Nrf2 is involved in the development of apoptosis in Nrf2 siRNA-transfected fibroblasts. We propose that a defective antioxidant system and apoptotic dysregulation may participate in keloid pathogenesis.     

Research of primary hyperhidrosis in students of medicine of the State of Sergipe,Brazil

BACKGROUND

Hyperhidrosis or excessive sweat production occurs at 2.9-9% of the population.

OBJECTIVE

To estimate the prevalence and disorders due to primary hyperhidrosis (HP) in medicine students in the state of Sergipe.

METHODS

Cross-sectional study using individual interviews.

RESULTS

Hyperhidrosis was found in 14.76% of subjects, the most affected regions were palmar, plantar and axillary, causing prejudice in daily activities. Family history occurred in 45% and 22.72% was diagnosed by a physician.

CONCLUSION

The prevalence of hyperhidrosis in medicine students of Sergipe was high, with strong family and a small portion of diagnoses made by medical professionals.     

Aging decreases collagen IV expression <Emphasis Type="Italic">in vivo</Emphasis> in the dermo-epidermal junction and <Emphasis Type="Italic">in vitro</Emphasis> in dermal fibroblasts: possible involvement of TGF-β1

Background

Collagen IV is a major component of the dermo-epidermal junction (DEJ).

Objectives

To study expression of collagen IV upon aging in the DEJ and dermal fibroblasts isolated from the same patients. A model of senescent fibroblasts was developed in order to identify biological compounds that might restore the level of collagen IV.

Materials & methods

Skin fragments of women (30 to 70 years old) were collected. Localisation of collagen IV expression in the DEJ was studied by immunofluorescence. Fibroblast collagen IV expression was studied by real-time PCR, ELISA, and western blotting. Premature senescence was simulated by exposing fibroblasts to subcytotoxic H₂O₂ concentrations.

Results

Collagen IV decreased in the DEJ and fibroblasts relative to age. TGF-β1 treatment significantly increased collagen IV gene and protein expression in fibroblasts and restored expression in the model of senescence. Addition of TGF-β1-neutralizing antibody to fibroblast cultures decreased collagen IV expression.

Conclusion

Taken together, the results suggest that the decrease in collagen IV in the DEJ, relative to age, could be due to a decrease in collagen IV expression by senescent dermal fibroblasts and may involve TGF-β1 signalling.

     

Clinical and Immunohistochemical Characteristics of Mucoceles

Background

Mucoceles have diverse clinical and histological features. Their pathogenesis remains to be elucidated.

Objective

To determine the structural and/or pathogenic differences between two clinically different types of mucoceles.

Methods

Seventeen oral mucoceles were examined clinically and immunohistologically. The mucoceles were divided into two groups by their clinical manifestations: papular group (PG) and nodular group (NG).

Results

Histologically, granulation tissue formed more frequently in the NG group, while CD4 and CD8 positive cells were more abundant in the PG group. There were no significant differences in the tumor necrosis factor-alpha or the matrix metalloproteinases (MMP)-2 and 9, between the two groups.

Conclusion

There were significant differences in the depth of the lesions, granulation tissue formation and infiltrating T lymphocytes between the PG and NP type of mucoceles. These findings suggest that the clinical manifestations may be influenced by the type of inflammatory response and extracellular matrix remodeling.     

An Open, Randomized, Comparative Clinical and Histological Study of Imiquimod 5% Cream Versus 10% Potassium Hydroxide Solution in the Treatment of Molluscum Contagiosum

Background

Although molluscum contagiosum (MC) resolves spontaneously, there are several reasons to treat this dermatological disorder.

Objective

To evaluate the safety and efficacy of 5% imiquimod cream versus 10% potassium hydroxide (KOH) solution in treating MC, and to propose the mechanism of cure by observing the histological findings.

Methods

Imiquimod or KOH were applied by the patient or a parent 3 days per week until all lesions cleared. The number of MC lesions was counted and side effects were evaluated at 5 points during the treatment (the initial visit, week 2, week 4, week 8, and week 12). Histological changes were compared between 2 patients of each group, before and after the 2 weeks of application.

Results

In both group, the mean lesion counts decreased all through to week 12, and the reduction in number of lesions were statistically significant in both groups (p <0.005). Over 40% of each group developed local side effects, and no systemic side effects were noted in either group. Before treatment, histological findings showed little or no dermal infiltrates. After treatment, specimens showed dense lymphocytic infiltrates, especially T cells, around the lesions which had resolved.

Conclusion

Both 10% KOH solution and 5% imiquimod cream are effective and safe treatment of MC.     

Physiological and lifestyle factors contributing to risk and severity of peri-orbital dark circles in the Brazilian population

BACKGROUND

Peri-orbital dark circles are a cosmetic concern worldwide, and have been attributed to hyperpigmentation from allergy or atopic dermatitis, blood stasis, structural shadowing effects, and a thin epidermis/dermis under the eye. It is of interest to better understand lifestyle and demographic risk factors and the relative impact of melanin, blood and epidermal/dermal factors on the severity of Peri-orbital dark circles.

OBJECTIVE

To compare by non-invasive imaging the impact of biological factors to a visual grading scale for Peri-orbital dark circles, and test the correlation of various demographic factors with Peri-orbital dark circles.

METHODS

Subjects completed a lifestyle and health survey, and Peri-orbital dark circles severity was evaluated using standardized photographs. Hyperspectral image analysis was used to assess the contributions of melanin, blood volume, degree of blood oxygen saturation, and dermal scattering.

RESULTS

Family history was the most significant risk factor for Peri-orbital dark circles. The average age of onset was 24 years, and earlier onset correlated with higher severity scores. Asthma was significantly associated with Peri-orbital dark circles scores, but self-reported allergy was not. In this study, sleep was not correlated with Peri-orbital dark circles scores. Hyperspectral imaging indicated that melanin was the dominant correlate for Peri-orbital dark circles severity, while oxygen saturation was secondary. The difference between under-eye and cheek measurements for ∆L*and ∆E* were the most significant instrumental parameters correlated with visual assessment of Peri-orbital dark circles severity.

CONCLUSION

Although typically associated with lack of sleep, risk of Peri-orbital dark circles is primarily hereditary. The main factors contributing to the appearance of Peri-orbital dark circles are melanin and (deoxygenated) blood.