Cutaneous Specimens Research Articles

Immunolocalization of Glioma-Associated Oncogene Homolog 1 in Non Melanoma Skin Cancer

Glioma-associated oncogene homolog (GLI)1 is involved in controlling cell proliferation and angiogenesis. The aim of this work was to explore its possible role in non-melanoma skin cancer pathogenesis through its immunohistochemical (IHC) expression in skin biopsies of these diseases and correlating this expression with the clinico-pathological parameters of the studied cases. Seventy-six cutaneous specimens were studied; 30 cases with basal cell carcinoma (BCC), 30 cases with squamous cell carcinoma (SCC) and 16 normal skin samples, from age- and gender-matched subjects, as a control group. GLI1 was expressed in all BCC cases and in 60% of SCC cases. All SCC cases showed cytoplasmic, while 70% of BCC cases showed nucleocytoplasmic immunoreactivity. It was over expressed in BCC and SCC compared to normal skin (p = 0.01 and 0.0006, respectively). Higher Histo (H) score in BCC cases was significantly associated with female gender (p = 0.04), multiple lesions, desmoplastic stromal reaction and stromal angiogenesis (p < 0.001 for all). Higher H score in SCC cases was significantly associated with scalp location, nodular type, recurrent lesions, high tumor grade, lymphovascular invasion (p = 0.004 for all), inflammatory stromal reaction (p = 0.01), lymph node involvement and absence of calcification (p = 0.001 for both). In conclusion, GLI1 may play a role in BCC pathogenesis through its role in cell proliferation, migration, and angiogenesis. Its upregulation and cytoplasmic localization in SCC may suggest that its role in tumor pathogenesis is through mechanisms other than Hedgehog pathway activation. Further studies are needed to clarify the exact molecular basis of its oncogenic action.

Clinical validation of the Lyra direct HSV 1+2/VZV assay for simultaneous detection and differentiation of three herpesviruses in cutaneous and mucocutaneous lesions.

We evaluated the Lyra Direct HSV 1+2/VZV multiplex real-time PCR assay for the detection and differentiation of herpes simplex virus 1 (HSV-1), HSV-2, and varicella-zoster virus (VZV) on 695 consecutive cutaneous and mucocutaneous lesion specimens. The intra-assay and interassay coefficient of variation values for the Lyra assay were 0.29 to 1.30% and 2.33 to 2.61%, respectively. The sensitivities, specificities, and positive and negative predictive values were 93.4 to 95.0%, 96.1 to 96.8%, 78.0 to 80.3%, and 99.0 to 99.1%, respectively, in comparison to those of viral culture. The values were further improved when a resolution analysis was performed with a laboratory-developed PCR assay.

IMPORTÂNCIA DA DERMATOSCOPIA NO DIAGNÓSTICO DE TUMORES COMPOSTOS – UM CASO CLÍNICO

A presença simultânea na mesma lesão de neoplasias cutâneas distintas designa-se por tumor composto ou de colisão, sendo considerada pouco frequente. Os autores descrevem o caso de um homem de 66 anos, observado por placa acastanhada de superfície verrucosa. Observou-se na região malar esquerda uma placa com 8 mm de maior eixo, limites mal definidos, coloração acastanhada e de superfície verrucosa, com bordo parcialmente eritematoso, com presença de múltiplas telangiectasias. A observação dermatoscópica evidenciou uma lesão não-melanocítica, distinguindo-se a combinação de múltiplos pseudo-quistos de milia (mais evidentes com luz não-polarizada), eritema e vasos arboriformes, com ramificações de menor calibre, na periferia (mais evidentes com luz polarizada). A primeira estrutura é característica de queratose seborreica, enquanto que as estruturas vasculares apresentam elevado valor preditivo positivo para o diagnóstico de carcinoma basocelular. A avaliação histopatológica confirmou os diagnósticos dermatoscópicos, com achados comuns às referidas neoplasias. A raridade deste tumor combinado contrasta com a prevalência das queratoses seborreicas. A dermatoscopia poderá constituir um complemento fundamental na sua abordagem, porque permite a observação de padrões de diagnóstico nem sempre evidentes do ponto de vista clínico, constituindo uma informação adicional à avaliação histopatológica.

Rad6 is a Potential Early Marker of Melanoma Development

Melanoma is the leading cause of death from skin cancer in industrialized countries. Several melanoma-related biomarkers and signaling pathways have been identified; however, their relevance to melanoma development/progression or to clinical outcome remains to be established. Aberrant activation of Wnt/β-catenin pathway is implicated in various cancers including melanoma. We have previously demonstrated Rad6, an ubiquitin-conjugating enzyme, as an important mediator of β-catenin stability in breast cancer cells. Similar to breast cancer, β-catenin-activating mutations are rare in melanomas, and since β-catenin signaling is implicated in melanoma, we examined the relationship between β-catenin levels/activity and expression of β-catenin transcriptional targets Rad6 and microphthalmia-associated transcription factor-M (Mitf-M) in melanoma cell models, and expression of Rad6, β-catenin, and Melan-A in nevi and cutaneous melanoma tissue specimens. Our data show that Rad6 is only weakly expressed in normal human melanocytes but is overexpressed in melanoma lines. Unlike Mitf-M, Rad6 overexpression in melanoma lines is positively associated with high molecular weight β-catenin protein levels and β-catenin transcriptional activity. Double-immunofluorescence staining of Rad6 and Melan-A in melanoma tissue microarray showed that histological diagnosis of melanoma is significantly associated with Rad6/Melan-A dual positivity in the melanoma group compared to the nevi group (P=.0029). In contrast to strong β-catenin expression in normal and tumor areas of superficial spreading malignant melanoma (SSMM), Rad6 expression is undetectable in normal areas and Rad6 expression increases coincide with increased Melan-A in the transformed regions of SSMM. These data suggest a role for Rad6 in melanoma pathogenesis and that Rad6 expression status may serve as an early marker for melanoma development.

Detection ofRickettsia sibirica mongolitimonaeby Using Cutaneous Swab Samples and Quantitative PCR

Detection of<i>Rickettsia sibirica mongolitimonae</i>by Using Cutaneous Swab Samples and Quantitative PCR

Inverse correlation between microtubule‐associated protein 1A/1B‐light chain 3 and p62/sequestosome‐1 expression in the progression of cutaneous squamous cell carcinoma

The expression of autophagy-related markers has occasionally been reported to correlate with the clinical stage of disease in patients with solid cancer, indicating autophagy activation. However, there have been no such reports for cutaneous squamous cell carcinoma. In this study, we investigated the expression levels of two autophagy-related markers, microtubule-associated protein IA/IB light chain 3 (LC3) and p62/sequestosome-1 (p62), in cutaneous squamous cell carcinoma specimens and assessed their correlation to clinicopathological factors in patients with this type of cancer. As a marker of the autophagosome, LC3 expression increases with autophagosome formation/accumulation, whereas p62 expression decreases due to selective degradation via autophagy. We performed immunostaining for LC3 and p62 in 50 cutaneous squamous cell carcinoma specimens obtained from patients treated by surgical resection, counted the number of cells that showed positive staining, and calculated the percentage of positive cells per low-power microscopic field. We next investigated the correlations between the expression levels of these markers and various clinicopathological factors. The results indicated that LC3 expression increased significantly with advanced clinical stage (P<0.001) and increased tumor diameter (P=0.046). By contrast, the expression of p62 decreased significantly with advanced clinical stage (P<0.001) and increased tumor diameter (P=0.001). These results suggest that autophagy becomes activated during disease progression in patients with cutaneous squamous cell carcinoma.

Histiocytoid Sweet syndrome may indicate leukemia cutis: A novel application of fluorescence in situ hybridization

In patients with malignancy-associated Sweet syndrome, a thorough evaluation for leukemia cutis should be considered. We sought to describe the clinicopathologic characteristics of histiocytoid Sweet syndrome. We retrospectively identified patients with histiocytoid Sweet syndrome at our institution from January 1992 through December 2010. We evaluated the underlying cutaneous infiltrate using immunohistochemistry and fluorescence in situ hybridization. We re-evaluated all 22 patients with hematologic malignancy-associated Sweet syndrome. Six patients had a monocytoid infiltrate that was consistent with histiocytoid Sweet syndrome; subsequent evaluation of these patients demonstrated cytogenetic abnormalities on prior bone-marrow biopsy specimens. Fluorescence in situ hybridization analysis was feasible in cutaneous specimens from 5 of the 6 patients and demonstrated the same cytogenetic abnormalities that were identified on prior bone-marrow biopsy specimens in 4 patients. Therefore, these 4 patients may have had a form of leukemia cutis. This was a retrospective study. For patients with histiocytoid Sweet syndrome, an underlying hematologic malignancy, and a monocytoid infiltrate on biopsy specimen, fluorescence in situ hybridization of the cutaneous infiltrate may be beneficial to identify cytogenetic abnormalities that may indicate leukemia cutis.

Impact of DNA methyltransferases on the epigenetic regulation of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) receptor expression in malignant melanoma

Aberrant promoter methylation and resultant silencing of TRAIL decoy receptors were reported in a variety of cancers, but to date little is known about the relevance of this epigenetic modification in melanoma. In this study, we examined the methylation and the expression status of TRAIL receptor genes in cutaneous and uveal melanoma cell lines and specimens and their interaction with DNA methyltransferases (DNMTs) DNMT1, DNMT3a, and DNMT3b. DR4 and DR5 methylation was not frequent in cutaneous melanoma but on the contrary it was very frequent in uveal melanoma. No correlation between methylation status of DR4 and DR5 and gene expression was found. DcR1 and DcR2 were hypermethylated with very high frequency in both cutaneous and uveal melanoma. The concordance between methylation and loss of gene expression ranged from 91% to 97%. Here we showed that DNMT1 was crucial for DcR2 hypermethylation and that DNMT1 and DNMT3a coregulate the methylation status of DcR1. Our work also revealed the critical relevance of DcR1 and DcR2 expression in cell growth and apoptosis either in cutaneous or uveal melanoma. In conclusion, the results presented here claim for a relevant impact of aberrant methylation of decoy receptors in melanoma and allow to understand how the silencing of DcR1 and DcR2 is related to melanomagenesis.

Epigenetic regulation of REG1A and chemosensitivity of cutaneous melanoma

Regenerating gene 1A (REG1A) plays an important role in tissue regeneration and in cell proliferation in epithelium origin tumors; however, its role in melanoma has not been explored in details. The objective of this study was to identify whether REG1A is expressed in cutaneous melanoma and if REG1A expression status can predict prognosis in cutaneous melanoma patients with metastasis. We also determined whether epigenetic regulation of the promoter region regulates REG1A expression. AJCC stage III cutaneous melanoma specimens with clinically well annotated stage III lymph node melanoma metastasis tissue microarray were assessed by IHC. MALDI-TOF-mass spectrometry and HM450K array were used to identify REG1A promoter region CpG site methylation. Chemotherapeutic agent response by melanoma cells as related to REG1A protein expression was assessed. Post-surgery melanoma patients followed by adjuvant chemotherapy with high REG1A expression had a significantly better prognosis (disease-specific survival) compared with patients with low REG1A expression (log rank test; p = 0.0013). The demethylating reagent 5-Aza-2′-deoxycytidine activated REG1A promoter region resulting in enhanced REG1A mRNA and protein expression in melanoma cell lines. Promoter region CpG methylation was shown to regulate REG1A expression in melanoma cells. Moreover, melanoma lines with high REG1A mRNA expression were more susceptible to Dacarbazine and Cisplatin, as compared with those with low REG1A mRNA expression. In conclusion, REG1A expression status may be useful as a biomarker in melanoma patients for sensitivity to these chemotherapeutic agents. The epigenetic regulation of the REG1A promoter region may offer a potential therapeutic approach to improve chemotherapy for metastatic melanoma patients.

Comparison of two multiplexed PCR assays for the detection of HSV-1, HSV-2, and VZV with extracted and unextracted cutaneous and mucosal specimens

BackgroundSeveral analyte specific reagents (ASRs) are available for the detection and differentiation of HSV-1, HSV-2, and VZV in clinical specimens. However, there is limited data on the test performance of these reagents used in multiplexed PCR assays. ObjectiveThis study compared the performance of two multiplexed ASR sets for detection of HSV-1, HSV-2, and VZV in dermal specimens. Study designTwo commercially available ASRs were combined to produce multiplexed PCR assays for simultaneous detection of HSV-1, HSV-2, and VZV. Seeded samples were used to determine the limit of detection (LOD) for each assay. Patient samples (n=156) were tested in duplicate and results for each method compared to the reference standard of culture. Both extracted and unextracted specimens were used in the study. ResultsBoth multiplexed PCR assays showed similar test performance, with minimal LOD differences observed. The LOD was 103copies/mL for HSV-1 and HSV-2 using the Focus assay compared to 5×103copies/mL and 2×104copies/mL, respectively for the EraGen assay. Both assays showed equal performance for VZV with a LOD of 5×103copies/mL. Analytical specificity testing showed no cross reactivity with other selected DNA viruses. Both assays showed similar performance when clinical samples were tested using both extracted and unextracted specimens. ConclusionCommercially available ASRs can be successfully multiplexed for the PCR detection of HSV-1, HSV-2, and VZV using dermal specimens. Either direct testing or nucleic acid extracted specimens can be used with similar performance in these assays.

Dermatopathology Practice in Ethiopia

Dermatologic diseases are extremely common among the Ethiopian population and are a significant cause of morbidity. However, few studies exist in the literature that describe the incidence and clinical and histologic features of biopsied cutaneous lesions. To categorize the cutaneous diseases observed in skin biopsies at the All African Leprosy Rehabilitation and Training Center (ALERT) in Addis Ababa, Ethiopia, and to describe the clinical and histologic features of dermatopathologic diagnoses most frequently encountered in this practice setting. Pathology reports of 2342 cutaneous specimens received at ALERT in Addis Ababa, Ethiopia, were reviewed to determine the range and frequency of cutaneous diseases and dermatoses diagnosed from January 2007 through December 2010. The range of cutaneous diseases observed in skin biopsies at ALERT was varied and included inflammatory dermatoses (27%), infectious diseases (24%), and malignant and benign cutaneous neoplasms (22% and 20%, respectively). The most common conditions observed in this study were squamous cell carcinoma (8% of total cases), eczema (6% of total cases), leishmaniasis (6% of total cases), tuberculosis (6% of total cases), and benign nevi (4% of total cases).

Diagnostic utility of direct immunofluorescence findings around hair follicles and sweat glands in immunobullous disease

Inherent to some immunobullous disorders is potential for intraepidermal or dermal-epidermal junction fragility, a phenomenon that may compromise biopsy specimen integrity and direct immunofluorescence (DIF) interpretation. In these situations, cutaneous adnexal structures (e.g. hair follicles, sweat apparatus) usually remain intact. Whether periadnexal DIF findings are reliable in diagnosing immunobullous conditions is unknown. We evaluated 56 cutaneous specimens with diagnostic immunoglobulin (Ig) deposition patterns that contained adnexal structures. In a corollary study, we examined 145 hematoxylin-eosin-stained frozen specimens to determine biopsy factors associated with the presence of adnexal structures. Periadnexal DIF findings offered diagnostic sensitivity in conditions with linear or cell-surface Ig deposition or lupus band. Periadnexal DIF findings were unreliable in dermatitis herpetiformis. Biopsy specimens from scalp and genitalia were most likely to contain folliculosebaceous units and sweat duct apparatus, respectively. Relative depth of biopsy correlated directly with the likelihood of identifying sweat duct apparatus but not folliculosebaceous units. Periadnexal DIF findings may add diagnostic sensitivity in DIF evaluation of pemphigoid, pemphigus and lupus erythematosus. Pathologists can guide clinicians to biopsy certain anatomic sites and to obtain sufficient biopsy depth to increase the probability of capturing adnexal structures and, therefore, diagnostic yield from DIF specimens.

Expression of Fas and Fas ligand in primary cutaneous squamous cell carcinoma in association with grade of tumor differentiation

The aim of this study was to evaluate whether Fas (CD95) and Fas ligand (FasL) immunoreactivity in cutaneous squamous cell carcinoma (SCC) are correlated with grade of tumor differentiation. A total of 31 cutaneous SCC specimens excised during 1997-2004 were collected from the four branch hospitals of the Catholic University of Korea. A tissue microarray technique was used for immunohistochemical staining. Expression of Fas and FasL was evaluated in correlation with the grade of tumor differentiation. Statistical analysis was performed using Fisher's exact test and exact trend test. Six of 31 SCCs exhibited strong (+++) intensity for Fas immunostaining, and all of these were well differentiated. Five of 31 SCCs exhibited strong (+++) intensity for FasL staining; all of these were moderately or poorly differentiated. Fas was expressed more strongly in well-differentiated SCCs than in poorly differentiated tumors. FasL was expressed more strongly in moderately and poorly differentiated tumors. These findings indicate that Fas and FasL play important roles in immune surveillance and grade of differentiation in SCC.

Molecular Biologic Assessment of Cutaneous Specimens of Ocular Rosacea

To characterize the molecular biologic environment of pathological skin in ocular rosacea and to differentiate the levels of inflammatory molecules in ocular rosacea from those of normal skin. The concentrations of 48 molecules were assayed in cutaneous biopsies taken from patients with ocular rosacea and from normal controls. There were very few molecular differences between the 2 groups, and 43 of the 48 molecules that were measured in this study were not significantly different between the 2 groups. The concentrations of 5 molecules (interleukin-1β, interleukin-16, stem cell factor, monocyte chemotactic protein-1, and monokine induced by γ-interferon) were significantly enriched in ocular rosacea. Ocular rosacea is a highly ordered molecular process and involves elevations in the concentrations of specific molecules. The particular pattern of enrichment supports the notion that ocular rosacea represents a disorder of innate immunity. Furthermore, these molecules may represent novel therapeutic targets in the future management of this disorder.

Overexpression of RNA helicase p68 protein in cutaneous squamous cell carcinoma

RNA helicase p68 is a prototypic DEAD-box RNA helicase. Recent studies indicate that p68 plays important role in cancer development and progression. However, the role of p68 protein expression in cutaneous squamous cell carcinoma (SCC) remains unknown. To elucidate the expression of p68 protein in cutaneous SCC. The level of p68 protein was examined by double immunofluorescent staining in 24 samples of human cutaneous SCC tissue specimens and their adjacent tissues and in 6 normal foreskin samples to compare the expression of p68 with that of Ki-67. Overexpression of p68 protein was seen in all 24 SCC cases (100%), whereas very low expression of p68 was detected in normal foreskin. Moreover, p68 protein expression was higher in cases of cutaneous SCC with metastasis than in cases without metastasis. Additionally, p68 had a similar expression pattern to that of Ki-67. The high frequency of p68 expression in cutaneous SCC indicates that p68 might be involved in the development and progression of cutaneous SCC.

Effects of epidermal growth factor receptor and insulin‐like growth factor 1 receptor inhibition on proliferation and intracellular signaling in cutaneous SCCHN: Potential for dual inhibition as a therapeutic modality

Combined inhibition of epidermal growth factor receptor (EGFR) and insulin-like growth factor-1 receptor (IGF-1R) has been proposed as a therapy for cutaneous squamous cell carcinoma of the head and neck (SCCHN). Receptor expression and downstream signaling were assessed in cutaneous squamous cell carcinoma (SCC) cell lines and patient samples. EGFR and IGF-1R signaling was inhibited in cutaneous SCC cell lines using erlotinib and/or picropodophyllin. EGFR and IGF-1R were overexpressed in cutaneous SCCHN specimens relative to normal skin. Dual inhibition of both receptors prevented cell growth and decreased activation of Akt and p42/44 mitogen-activated protein kinase (MAPK) more effectively than either inhibitor alone. Dual inhibition of EGFR and IGF-1R is effective at blocking cell growth, and is correlated with inhibition of Akt and p42/44 MAPK, suggesting that this may be a promising treatment for cutaneous SCCHN.

Expression and significance of CXCR7 chemokine receptor in cutaneous malignant tumors and their cell lines

Objective To investigate the expression of CXCR7 in several cutaneous malignant tumors including cutaneous squamous cell carcinoma (SCC),basal cell carcinoma (BCC) and invasive cutaneous malignant melanoma and their cell lines,as well as its significance.Methods Tissue specimens were obtained from the lesions of 30 patients with cutaneous squamous cell carcinoma,25 patients with basal cell carcinoma and 30 patients with cutaneous malignant melanoma.Immunohistochemistry was performed to detect the expression of CXCR7 protein in these tissue specimens and several cell lines (A375 human melanoma cells,M14 human melanoma cells,A431 human epidermoid carcinoma cells,HaCaT human keratinocytes).The mRNA expression of CXCR7 in these cell lines was measured by reverse transcription PCR.Results CXCR7 protein was apparently expressed in invasive cutaneous malignant melanoma.The high expression rate of CXCR7 protein was significantly elevated in cutaneous malignant melanoma tissue specimens compared with SCC and BCC tissue specimens [80％ (24/30) vs.26.67％ (8/30) and 8％ (2/25),x2 =17.16,28.36,both P ＜ 0.05].CXCR7 mRNA was expressed in A375,M14 and A431 cells,but not in HaCaT cells,with the strongest expression observed in A375 cells.Immunohistochemistry revealed the expression of CXCR7 protein only in A375 cells.Conclusions CXCR7 is highly expressed in cutaneous malignant melanoma and A375 cells,which may be involved in the malignant invasion and metastasis of melanoma. Key words: Melanoma; Carcinoma, squamous cell; Carcinoma, basal cell; Receptors, chemokine; Cell line, tumor

P16 protein expression in primary cutaneous melanoma with positive and negative lymph node biopsies: Particular aspects of a study performed at the Hospital de Clinicas de Porto Alegre, Brazil.

Cutaneous melanoma dermal invasion, identified through measurement of maximum tumour thickness and sentinel lymph node (SLN) biopsy, is important to establish melanoma prognosis and progression. P16 protein expression has been shown to be a predictive factor for melanoma evolution and prognosis. To investigate p16 protein expression in cutaneous melanomas with and without SLN metastasis. Sixty-seven paraffin-embedded cutaneous melanoma specimens of patients who had undergone SLN investigation were evaluated from 1995 to 2007. SLN biopsy was negative for metastasis in 34 of these patients (controls); in the remaining 33 patients, SLN biopsy was positive (cases). The expression of p16 protein in the primary tumour was measured using an immunohistochemical assay. The samples were classified according to their nuclear expression. P16 nuclear expression was absent in 14 cases and in 15 controls; P=0.812. There was no statistically significant difference in p16 nuclear expression between cases and controls. The present study does not support the findings of other studies that suggest p16 protein expression is important in the prognosis of cutaneous melanoma.

Collision of two rare adnexal tumors with folliculosebaceous differentiation

To the Editor: Cutaneous collision tumor, the coexistence of more than one neoplasm in a single cutaneous specimen, is unusual. Most of the collision tumors are accidental, such as a combination of basal cell carcinoma and melanocytic nevus; however, some of them may arise from the involvement of related cell types in cases of adnexal tumors. However, to our knowledge the combination of desmoplastic trichoepithelioma (DTE) and chondroid syringoma (CS) has never been reported.

Identification of Rickettsial Infections by Using Cutaneous Swab Specimens and PCR

To determine the usefulness of noninvasive cutaneous swab specimens for detecting rickettsiae, we tested skin eschars from 6 guinea pigs and from 9 humans. Specimens from eschars in guinea pigs were positive for rickettsiae as long as lesions were present. Optimal storage temperature for specimens was 4°C for 3 days.