The resveratrol analogue, 3,4,5,4'­trans-tetramethoxystilbene, inhibits the growth of A375 melanoma cells through multiple anticancer modes of action.

Resveratrol is a natural dietary product that has demonstrated multifaceted anticancer activity. Several analogues of resveratrol have been synthesized in an effort to enhance the pharmacological potency and improve the pharmacokinetic properties of the compound. 3,4,5,4'­trans­tetramethoxystilbene (3,4,5,4'­TMS) is a methoxylated analogue of resveratrol that has demonstrated anti-proliferative activity in vitro (in cancer cell lines) and in vivo (in xenograft models). In the present study, the anticancer effects of 3,4,5,4'­TMS in A375 human melanoma cells were examined. 3,4,5,4'­TMS markedly inhibited the proliferation of A375 cells (IC50=0.7 µM), via a mechanism involving mitotic arrest at the prometaphase stage of cell division. This effect was accompanied by the upregulation of the expression of the mitogen activated protein kinases, JNK and p38, and the concomitant activation of p38, that was verified by the nuclear translocation of the phoshorylated form of the protein. The pharmacological inhibition of p38 by SB203580 (4 µM) attenuated the effects of 3,4,5,4'­TMS, as demonstrated by decreased cell cycle progression at the mitotic phase. Furthermore, 3,4,5,4'­TMS increased the total levels of Aurora A, while it inhibited the localization of the protein to the spindle poles. Finally, 3,4,5,4'­TMS exhibited anti-metastatic activity, inhibiting A375 cell migration and the attachment of the cells to a collagen type IV-coated surface. Collectively, the data suggest that 3,4,5,4'­TMS is an effective chemotherapeutic drug for the treatment of human melanoma and that it exerts its effects through multiple anticancer modes of action.

Transporter associated with antigen processing deficiency syndrome: case report of an adolescent with chronic perforated granulomatous skin lesions due to TAP2 mutation.

A previously unreported case of transporter associated with antigen processing (TAP) deficiency syndrome (with no parental consanguinity) due to a homozygous TAP2 mutation is presented. Characteristic nonhealing, chronic, ulcerative granulomatous leg lesions combined with recurrent otitis media and sinopulmonary infections led to this boy being diagnosed at 15 years old. The role of the dermatologist was crucial in making the correct diagnosis and thereby positively influencing the quality of life and life expectancy of this boy.

Development of a three-dimensional surface imaging system for melanocytic skin lesion evaluation.

Even though surface morphology is always taken into account when assessing clinically pigmented skin lesions, it is not captured by most modern imaging systems using digital imaging. Our aim is to develop a novel three-dimensional (3D) imaging technique to record detailed information of the surface anatomy of melanocytic lesions that will enable improved classification through digital imaging. The apparatus consists of three high-resolution cameras, a light source, and accompanying software. Volume measurements of specific phantoms using volumetric tubes render slightly lower values than those obtained by our 3D imaging system (mean%± SD, 3.8%± 0.98, P<0.05). To examine the reproducibility of the method, sequential imaging of melanocytic lesions is carried out. The mean%± SD differences of area, major axis length, volume, and maximum height are 2.1%± 1.1, 0.9%± 0.8, 3.8%± 2.9, and 2.5%± 3.5, respectively. Thirty melanocytic lesions are assessed, including common and dysplastic nevi and melanomas. There is a significant difference between nevi and melanomas in terms of variance in height and boundary asymmetry (P<0.001). Moreover, dysplastic nevi have significantly higher variances in pigment density values than common nevi (P<0.001). Preliminary data suggest that our instrument has great potential in the evaluation of the melanocytic lesions. However, these findings should be confirmed in larger-scale studies.

Prehistological evaluation of benign and malignant pigmented skin lesions with optical computed tomography.

Discrimination of benign and malignant melanocytic lesions is a major issue in clinical dermatology. Assessment of the thickness of melanoma is critical for prognosis and treatment selection. We aimed to evaluate a novel optical computed tomography (optical-CT) system as a tool for three-dimensional (3-D) imaging of melanocytic lesions and its ability to discriminate benign from malignant melanocytic lesions while simultaneously determining the thickness of invasive melanoma. Seventeen melanocytic lesions, one hemangioma, and normal skin were assessed immediately after their excision by optical-CT and subsequently underwent histopathological examination. Tomographic reconstructions were performed with a back-propagation algorithm calculating a 3-D map of the total attenuation coefficient (AC). There was a statistically significant difference between melanomas, dysplastic nevi, and non-dysplastic nevi, as indicated by Kruskal-Wallis test. Median AC values were higher for melanomas compared with dysplastic and non-dysplastic nevi. No statistically significant difference was observed when thickness values obtained by optical-CT were compared with histological thickness using a Wilcoxon sighed rank test. Our results suggest that optical-CT can be important for the immediate prehistological evaluation of biopsies, assisting the physician for a rapid assessment of malignancy and of the thickness of a melanocytic lesion.

Pharmacogenetic analysis of TNF, TNFRSF1A, and TNFRSF1B gene polymorphisms and prediction of response to anti-TNF therapy in psoriasis patients in the Greek population.

BACKGROUND: Although biologic therapies have revolutionized the treatment of psoriasis, patients exhibit a substantial heterogeneous response that could be due to complex genetic heterogeneity. OBJECTIVE: The aim of this study was to investigate the possible influence of tumor necrosis factor-α (TNF), TNF receptor I (TNFRSF1A), and TNF receptor II (TNFRSF1B) gene polymorphisms on anti-TNF treatment responsiveness in psoriasis patients. METHODS: A Greek multicenter collaboration was established to recruit a cohort of patients (n = 80) with psoriasis treated with anti-TNF drugs. Single nucleotide polymorphisms (SNPs) in TNF (-238G>A, -308G>A, -857C>T), TNFRSF1A (36A>G), and TNFRSF1B (676T>G) were genotyped by PCR-restriction fragment length polymorphism assays. SNPs and haplotypes, including stratification by comorbidity status, were analyzed for association with treatment response after 6 months of therapy, defined as a reduction in the Psoriasis Area and Severity Index (PASI) score by >75% (responders) or ≤50% (nonresponders). RESULTS: Sixty-three patients (78.8%) were defined as responders (PASI score reduction >75%) and 17 patients (21.2%) were defined as nonresponders (PASI score reduction ≤50%). Carriage of TNF -857C or TNFRSF1B 676T alleles was associated with positive response to drug treatment in patients treated with etanercept (p = 0.002 and p = 0.001, respectively). None of the genotyped SNPs were associated with responsiveness to treatment with infliximab or adalimumab. Additionally, when patients were stratified by comorbidity status, none of the genotyped SNPs were alone associated with responsiveness to drug treatment. CONCLUSION: This study is the first in the field of psoriasis demonstrating a strong association between genetic markers and positive response to drug treatment. Validation of this result in larger studies, as well as analysis of other drug treatments, could provide the basis for individually tailored treatment, along with increased cost effectiveness and reduced unnecessary exposure to toxicity.

Successful treatment of cutaneous leishmaniasis with intralesional aminolevulinic acid photodynamic therapy.

Leishmaniasis is a protozoan infectious disease that often affects the skin and may acquire a chronic and difficult to treat course. Topical photodynamic therapy (PDT) is a novel treatment which involves the selective uptake of a photosensitizing agent. Exposure to an appropriate light source in the presence of oxygen leads to formation of reactive oxygen species and destruction of the target cells. We report on the successful treatment of a 69-year-old patient with a relapse of long-standing cutaneous leishmaniasis using intralesional aminolevulinic acid-PDT.

Tailor-made three-dimensional hybrid scaffolds for cell cultures.

The construction of the ideal three-dimensional scaffold for cell culture is one of the most intriguing topics in tissue engineering. It has been shown that cells can be cultured on most organic biomimetic materials, which now are losing popularity in favour of novel, hybrid systems. In this study, a series of photosensitive sol-gel hybrid materials, based on silicon-zirconium and silicon-titanium oxides, have been investigated for their suitability in three-dimensional scaffold fabrication. These materials can be structured by two-photon polymerization, a laser-based technique allowing the fabrication of micrometre-size structures with submicron resolution. The work presented here examined the effect of the organic/inorganic composition of the materials on cell behaviour and the establishment of a 'cell-culture friendly' environment. This is vital for cell adhesion, growth and differentiation, as the organic part of the material provides the soft matrix for cell growth, whereas the inorganic component gives the mechanical stability and rigidity of the three-dimensional structures. In addition, the use of femtosecond laser structuring permits the fabrication of a wide range of mechanically stable scaffolds of different sizes and shapes to be tested in terms of cell viability, proliferation and orientation.

Factors associated with the prevalence of atypical nevus in a Mediterranean pigmented skin lesion clinic.

Atypical melanocytic nevi constitute central risk factor and are precursor lesions for cutaneous melanoma. Data regarding factors associated with their prevalence are mainly derived from fair-skinned populations, whereas little is known regarding their epidemiological associations in darker-skinned, chronically sun-exposed populations. The aim of this study was to identify risk factors for the occurrence of at least one atypical nevus on Crete, the southernmost island of Greece. This hospital-based case-control study included 143 patients and 189 controls with at least one atypical nevus presented at the pigmented skin lesion clinic of the University of Crete. All participants were interviewed and underwent complete skin examination by the same two experienced dermatologists. Multivariate logistic regression analysis was used to adjust for potential confounders. In the multivariate analysis, common melanocytic nevi [odds ratio (OR): of 2.2, 7.5, and 58.9 for the presence of 11-25, 26-100, and >100 common nevi, respectively] and recreational sun exposure (OR: 4.4) increased significantly the risk of the presence of atypical nevus. A decreased risk for atypical nevi was related to an increasing age (OR: 0.96/age), and professional sun exposure (OR: 0.5). Intermittent, recreational sun exposure is mainly associated with the prevalence of atypical nevi in our sample and this effect does not depend on skin phototype. Promotion of sun protection, especially in patients with high numbers of common nevi, might serve as a measure to prevent the development of atypical nevi.

A CD40 and an NCOA5 gene polymorphism confer susceptibility to psoriasis in a Southern European population: a case-control study.

Recent genome-wide association studies of many complex diseases have successfully identified novel susceptibility loci, with many of them shared by multiple disease-associated pathways. The genes CD40 and nuclear receptor coactivator 5 (NCOA5), located in a 400-kb region surrounding CD40, have been reported to be associated with increased risk for rheumatoid arthritis and other autoimmune diseases. We hypothesized that those genes may also have a role in psoriasis (PS), an autoimmune, chronic inflammatory skin disease. In a case-control study, 198 patients with PS and 400 controls were genotyped for 2 single nucleotide polymorphisms (SNPs) of the CD40 and NCOA5 genes located on chromosome 20q.12-q13.12. Here, we demonstrate for the first time the association of both SNPs with susceptibility to PS, thus suggesting a putative key role of both genes in multiple autoimmune diseases. Alleles G and C of the CD40 rs4810485 and NCOA5 rs2903908 SNPs, respectively, were more common in individuals with PS than in controls (p = 0.03, odds ratio [OR] = 1.42, 95% confidence interval [95% CI] 1.05-1.95 and p = 0.000 003, OR = 1.93, 95% CI 1.47-2.55, respectively). The identification of shared genetic susceptibility loci may provide insight into our understanding of the pathophysiology of autoimmune diseases.

Identification of the M541L sequence variation of the transmembrane KIT domain in Merkel cell carcinoma.

BACKGROUND: Merkel cell carcinoma (MCC) is an aggressive KIT-positive cutaneous tumor. KIT mutations are considered to play a key role in the pathogenesis of various neoplasms, but have not been found so far in MCC. The aim of the present study was therefore to investigate the presence of KIT mutations in MCC. MATERIALS AND METHODS: The entire coding region of KIT in the MCC cell line MCC-1 was sequenced. KIT exon 10 was amplified from archival paraffin-embedded MCC specimens by PCR and sequenced. RESULTS: Exon 10 M541L KIT sequence variation, which confers increased sensitivity to KIT ligand stem cell factor, was detected in the MCC-1 cell line. Sequencing of KIT exon 10 in six archival MCC specimens revealed the wild-type sequence. CONCLUSION: The presence of the M541L KIT variation in MCC warrants further studies for its role in the pathogenesis of this tumor.

KIT receptor activation by autocrine and paracrine stem cell factor stimulates growth of merkel cell carcinoma in vitro.

The co-expression of KIT receptor and its ligand stem cell factor (SCF) has been reported in biopsy specimens of Merkel cell carcinoma (MCC). However, the functional role of SCF/KIT in the pathogenesis of this aggressive tumor has not been elucidated. The present study reports expression and effects of SCF and KIT in the Merkel cell carcinoma cell line MCC-1 in vitro. SCF and KIT were endogenously co-expressed in MCC-1 cells. Exogenous soluble SCF modulated KIT receptor mRNA and protein expression, stimulated growth of MCC-1 cells, upregulated endogenous activation of KIT, AKT, and of extracellular signal-regulated kinase (ERK) 1/2 signaling pathway. On the contrary, an inhibitory antibody that neutralized the KIT ligand binding site, reduced growth of MCC-1 cells, as did high doses of the KIT kinase inhibitors imatinib and nilotinib. Also, inhibitors of KIT downstream effectors, U0126 that blocks MEK1/2 as well as wortmannin and LY294002 that inhibit phosphatidylinositol 3-kinase-dependent AKT phosphorylation, inhibited the proliferation of MCC-1 cells. These data support the hypothesis that KIT is activatable by paracrine or autocrine tumor cell-derived SCF and stimulates growth of Merkel cell carcinoma in vitro. Blockade of KIT and the downstream signaling cascade at various levels results in inhibition of Merkel cell carcinoma growth in vitro, suggesting targets for therapy of this cancer.

Significant differences in skin irritation by common suture materials assessed by a comparative computerized objective method.

BACKGROUND: Erythema can be described only through subjective evaluation, except when it is quantified by digital image analysis software. Using such software, the authors performed comparisons of the erythema produced after skin closure of clean surgical wounds. Five suture materials were compared with respect to the local skin irritation that was caused. Different quantities of erythema are produced by suture material after the skin closure of clean surgical wounds. The authors present an objective method of measuring how unreactive a suture material is in comparison with another when applied to the skin. METHODS: The suture materials polydioxanone, polypropylene blue, polyamide 6, metallic clips, and polyglactin were compared in the present study. Digital photographs of 100 patients were compared by means of software, evaluating red color superiority (mean value of red color) in the region surrounding the wound. RESULTS: The least to most irritation caused to the skin by different suture materials was established for paired data. The Kolmogorov-Smirnov criterion and the Wilcoxon signed rank test were used. Polydioxanone was found to have the best performance, followed in order by polyglactin, polyamide, polypropylene, and metallic clips. Immediately after suture removal, differences between the effects of suture materials were statistically significant on postoperative day 10. CONCLUSIONS: Absorbable sutures can be used in skin closure of clean surgical wounds and can produce less erythematous reaction than nonabsorbable ones. Digital image analysis is a reliable method of quantitative evaluation of skin erythema resulting after skin closure of surgical wounds.

Analysis of epidemiology, clinical features and management of erysipelas.

BACKGROUND: Erysipelas is a superficial form of cellulitis affecting the upper dermis and superficial lymphatics. The widespread use of antibiotics may affect clinical findings and response to therapy of infectious disorders. The purpose of the study was to investigate the epidemiological, clinical, and laboratory features of erysipelas and to compare the results of treatment with penicillin vs. other antibiotic regimens. METHODS: All charts of erysipelas patients treated at the University Hospital of Heraklion, Crete, Greece from 1994 to 2002 were retrospectively studied. RESULTS: Median age of the 99 patients was 54.5 years; 59% were females. The most frequent site involved was the lower extremity (76%), followed by the face (17%) and upper extremity (6%). In 61 patients (62%), a possible entry portal was identified. The most common manifestation of erysipelas was local symptoms and signs (pain, erythema, and swelling) in all patients, together with elevated erythrocyte sedimentation rate (ESR) (60%). Fever was present in 25% of patients. The most commonly used antibiotic was intravenous penicillin G (64%). In the penicillin group, mean duration of fever after treatment initiation was shorter than in the nonpenicillin group (1.7 vs. 4.5 days, P = 0.002). Both treatment failures and recurrences were the same between the two groups. DISCUSSION: The diagnosis of erysipelas can be based on careful examination for local signs and symptoms. The role of ESR in primary diagnosis needs further investigation. Penicillin seems to preserve its fundamental role in the treatment of disease.

The protein tyrosine phosphatase, non-receptor type 22 R620W polymorphism does not confer susceptibility to psoriasis in the genetic homogeneous population of Crete.

Recent whole-genome and candidate-gene association studies in patients with psoriasis (PS) have identified a number of single-nucleotide polymorphisms (SNPs) that predispose to disease with moderate risk. Predisposition to PS is known to be affected by genetic variation in human leukocyte antigen-C as well as other non-human leukocyte antigen genes. We recently reported for the first time as a PS-associated SNP the signal transducer and activator of transcription-4 (STAT4) rs7574865 polymorphism, which is also associated with several autoimmune diseases. The aim of this study was to assess whether the functional R620W polymorphism of protein tyrosine phosphatase, non-receptor type 22 (PTPN22) gene encoding the lymphoid-specific tyrosine phosphatase, which is known to be associated with various autoimmune diseases, also confers increased risk for PS in the genetic homogeneous population of Crete. A case-control study was performed with 173 PS patients consecutively recruited and 348 healthy controls, all of them from the island of Crete. We found that the mutated T allele of the PTPN22 1858T SNP was more common in control individuals than in patients with PS (odds ratio = 0.39, 95% confidence interval = 0.11-1.04, p = 0.09). No considerable difference was observed in terms of sex, age of onset, or clinical presentation of psoriatic arthritis. Our results provide evidence that the PTPN22 1858T allele is not a susceptibility factor for PS in the Cretan population.

STAT4 gene polymorphism is associated with psoriasis in the genetically homogeneous population of Crete, Greece.

Recent genome-wide association studies (GWAS) of many complex diseases have successfully identified novel susceptibility loci, with many of them being associated with more than one condition. Taking into consideration that different autoimmune diseases may share some common pathogenetic pathways, we hypothesized that STAT4, a susceptibility gene found to be associated with increased risk for systemic lupus erythematosus, rheumatoid arthritis, type 1 diabetes, Sjögren's syndrome, Wegener's granulomatosis, Crohn's disease, and ulcerative colitis may also have a role in psoriasis. Psoriasis is an autoimmune, chronic inflammatory skin disease. Here we performed a case-control study in the population of island of Crete and demonstrated for the first time the association of a STAT4 single nucleotide polymorphism (SNP) with susceptibility to psoriasis, thus suggesting a putative key role of STAT4 in multiple autoimmune diseases. We found that mutated allele T of the STAT4 rs7574865 SNP, which previously was implicated in the predisposition to many autoimmune diseases, were more common in individuals with psoriasis than in controls (p = 0.045, odds ratio = 1.42, 95% confidence interval 1.01-2.00), thus concluding that the polymorphism examined is associated with the development of psoriasis in our population.

Botulinum toxin type A--treatment of a patient with multiple cutaneous piloleiomyomas.

BACKGROUND: Treatment of multiple cutaneous piloleiomyomas which are rare, frequently painful, benign tumors originating from the arrector pilorum muscle of hair follicles is difficult. OBJECTIVE: To determine the efficiency of botulinum toxin type A (BT-A) treatment for pain relief of cutaneous piloleiomyomas. METHODS: A patient with multiple painful piloleiomyomas was treated with local injections of 200 units of BT-A. RESULTS: There was a rapid and sustained decrease in pain. Treatment was repeated every 3 months for 2 years with the same efficacy. CONCLUSION: BT-A may be a promising new treatment option for multiple painful cutaneous piloleiomyomas.

Co-expression of KIT receptor and its ligand stem cell factor in Merkel cell carcinoma.

BACKGROUND/AIMS: KIT receptor has been implicated in the pathogenesis of cancer, either by mutation or autocrine activation. Merkel cell carcinoma (MCC) is a rare KIT-positive cutaneous tumor. We investigated the co-expression of KIT and its ligand stem cell factor (SCF) in MCC. METHODS: Sixteen specimens from 13 MCC patients of various tumor stages were examined by immunohistochemistry for SCF, KIT, Ki67/MIB-1 and cleaved caspase 3 expression, and for apoptosis by TUNEL. RESULTS: KIT was expressed in 13 of 16 tumors, and SCF in 15 of 16 specimens. Co-expression of KIT and SCF was detected in 12 of 16 tumors. KIT and SCF immunoreactivity scores were independent of tumor stage. Ki67/MIB-1 proliferation rates were high, whereas apoptosis rates were low, and did not depend on KIT or SCF expression. CONCLUSION: Co-expression of KIT and SCF in a high percentage of MCC tumors hints to an autocrine mechanism. KIT and SCF expression in primary tumors and in metastases suggests an early event in Merkel cell transformation.

Interferon-alpha inhibits proliferation and induces apoptosis of merkel cell carcinoma in vitro.

Merkel cell carcinoma is a tumor with aggressive biological behavior and limited response to chemotherapy. The present study investigated the effect of interferon (IFN)-alpha on growth and apoptosis of Merkel carcinoma cells in vitro. Proliferation of MCC-1 cell line was reduced dose-dependently by IFN-alpha and diminished when higher IFN-alpha concentrations were used. Additionally, IFN-alpha potently decreased DNA-synthesis and Ki67/MIB-1 proliferation index of MCC-1 cultures. Furthermore, IFN-alpha induced dose-dependently apoptosis of MCC-1 cells as shown by caspase-3 activation, and detection of apoptotic DNA strand breaks and fragmented nuclei. These findings suggest that IFN-alpha may have antitumor activity against Merkel cell carcinoma.

From the protein to the graph: how to quantify immunohistochemistry staining of the skin using digital imaging.

Quantitative immunohistochemistry is needed in order to reliably and accurately assess the expression of cellular proteins in tissue. Skin is a difficult tissue for automated image analysis due to its heterogeneous composition and its architecture. In the present study we used a psoriatic skin model to compare the expression of p53 and bcl-2 before and after treatment with anti-tumor necrosis factor-alpha using digital image analysis. Digital photomicrographs were acquired and analyzed with Scion image software in order to obtain the fraction of p53 and bcl-2 immunoreactive cells' area out of the total area investigated. Statistical analysis with ANOVA revealed a significant increase of p53 expression and a decrease of bcl-2 expression in all 3 epidermal layers during the course of therapy (p<0.001). The results were in line with the conventional histopathological evaluation using an arbitrary scale to grade the extent and intensity of the staining. So, the estimation of volume fraction of immunohistochemically labelled cells in skin tissue can be performed easily and rapidly using commonly available image analysis software and provides reproducible and unbiased numerical estimations of the amount of cell labelling.