Luigi Rossiello

Three-point checklist of dermoscopy

Background: Dermoscopy used by experts has been demonstrated to improve the diagnostic accuracy for melanoma. However, little is known about the diagnostic validity of dermoscopy when used by nonexperts. Objective: To evaluate the diagnostic performance of nonexperts using a new 3-point checklist based on a simplified dermoscopic pattern analysis. Methods: Clinical and dermoscopic images of 231 clinically equivocal and histopathologically proven pigmented skin lesions were examined by 6 nonexperts and 1 expert in dermoscopy. For each lesion the nonexperts assessed 3 dermoscopic criteria (asymmetry, atypical network and blue-white structures) constituting the 3-point method. In addition, all examiners made an overall diagnosis by using standard pattern analysis of dermoscopy. Results: Asymmetry, atypical network and blue-white structures were shown to be reproducible dermoscopic criteria, with a kappa value ranging from 0.52 to 0.55. When making the overall diagnosis, the expert had 89.6% sensitivity for malignant lesions (tested on 68 melanomas and 9 pigmented basal cell carcinomas), compared to 69.7% sensitivity achieved by the nonexperts. Remarkably, the sensitivity of the nonexperts using the 3-point checklist reached 96.3%. The specificity of the expert using overall diagnosis was 94.2% compared to 82.8 and 32.8% achieved by the nonexperts using overall diagnosis and 3-point checklist, respectively. Conclusion: The 3-point checklist is a valid and reproducible dermoscopic algorithm with high sensitivity for the diagnosis of melanoma in the hands of non-experts. Thus it may be applied as a screening procedure for the early detection of melanoma.

Identification of genes down-regulated during melanoma progression: a cDNA array study.

Abstract: In order to identify genes relevant for melanoma development, we carried out cDNA array experiments employing an in vitro model of human melanoma progression, consisting of two cell lines: one, LP, derived from a primary melanoma and the other, LM, from its metastatic supraclavicular lymph node. Basic cDNA array data identified 26 genes as down‐regulated in the LM cell line. Northern blot analysis confirmed an effective transcriptional down‐regulation for five out of 13 genes analyzed. The products of these five genes belong to different functional protein types, such as transcription and translation regulators (Edg‐2, eIF‐3 p110, and RNPL/RBM3), extracellular communicators (PRSS11) and members of the major histocompatibility complex (β2‐microglobulin). Some previously described differences in expression patterns, such as loss of HLA I, were confirmed by our array data. In addition, we identified and validated for the first time the reduced expression level of several genes during melanoma progression. In particular, reduced Edg‐2 gene product expression was also confirmed in a group of 50 primary melanomas and unrelated metastases. In conclusion, comparative hybridization by means of cDNA arrays assisted in identifying a series of novel progression‐associated changes in gene expression, confirming, at the same time, a number of previously described results.

Ferritin Contributes to Melanoma Progression by Modulating Cell Growth and Sensitivity to Oxidative Stress

Purpose: Employing an in vitro model system of human melanoma progression, we previously reported ferritin light chain (L-ferritin) gene overexpression in the metastatic phenotype. Here, we attempted to characterize the role of ferritin in the biology of human melanoma and in the progression of this disease. Experimental Design: Starting from the LM human metastatic melanoma cell line, we engineered cell clones in which L-ferritin gene expression was down-regulated by the stable expression of a specific antisense construct. These cells were then assayed for their growth capabilities, chemoinvasive properties, and sensitivity to oxidative stress. Additionally, ferritin protein content in primary and metastatic human melanomas was determined by immunohistochemistry. Results: Artificial L-ferritin down-regulation in the LM cells strongly inhibited proliferation and chemoinvasion in vitro and cell growth in vivo. In addition, L-ferritin down-regulated cells displayed enhanced sensitivity to oxidative stress and to apoptosis. Concurrently, immunohistochemical analysis of a human melanoma tissue array revealed that ferritin expression level in metastatic lesions was significantly higher (P < 0.0001) than in primary melanomas. Furthermore, ferritin expression was constantly up-regulated in autologous lymph node melanoma metastases when compared with the respective primary tumors in a cohort of 11 patients. Conclusions: These data suggest that high ferritin expression can enhance cell growth and improve resistance to oxidative stress in metastatic melanoma cells by interfering with their cellular antioxidant system. The potential significance of these findings deserves to be validated in a clinical setting.

Evidence of Key Role of Cdk2 Overexpression in Pemphigus Vulgaris

The pathogenesis of pemphigus vulgaris (PV) is still poorly understood. Autoantibodies present in PV patients can promote detrimental effects by triggering altered transduction of signals, which results in a final acantholysis. To investigate mechanisms involved in PV, cultured keratinocytes were treated with PV serum. PV sera were able to promote the cell cycle progression, inducing the accumulation of cyclin-dependent kinase 2 (Cdk2). Microarray analysis on keratinocytes detected that PV serum induced important changes in genes coding for one and the same proteins with known biological functions involved in PV disease (560 differentially expressed genes were identified). Then, we used two different approaches to investigate the role of Cdk2. First, small interfering RNA depletion of Cdk2 prevented cell-cell detachment induced by PV sera. Second, pharmacological inhibition of Cdk2 activity through roscovitine prevented blister formation and acantholysis in the mouse model of the disease. In vivo PV serum was found to alter multiple different pathways by microarray analysis (1463 differentially expressed genes were identified). Major changes in gene expression induced by roscovitine were studied through comparison of effects of PV serum alone and in association with roscovitine. The most significantly enriched pathways were cell communication, gap junction, focal adhesion, adherens junction, and tight junction. Our data indicate that major Cdk2-dependent multiple gene regulatory events are present in PV. This alteration may influence the evolution of PV and its therapy.

Primary cardiac T-cell lymphoma

Primary cardiac lymphoma (PCL), defined as a lymphoma clinically mimicking cardiac disease, with the bulk of the tumor located intrapericardially, is extremely rare in immunocompetent patients. Clinical manifestations vary depending on sites of involvement in the heart and include chest pain, arrhythmias, pericardial effusion, and heart failure. Diagnosis is often difficult and may require invasive procedures; in some cases, diagnosis is not made until autopsy. Histologically, nearly all cases of PCL reported thus far have been of B-cell origin. In this report, we describe a case of PCL of T-cell origin in an adult immunocompetent patient, the second reported in the literature to the best of our knowledge, and provide a brief overview of the features of previously published PCL cases.

Autologous bone marrow cell therapy and metabolic intervention in ischemia-induced angiogenesis in the diabetic mouse hindlimb.

Peripheral arterial disease (PAD) is a major health problem especially when associated to diabetes. Administration of autologous bone marrow cells (BMC) is emerging as a novel intervention to induce therapeutic angiogenesis in experimental ischemic limb models and in patients with PAD. Since tissue ischemia and diabetes are associated with an overwhelming generation of oxygen radicals and detrimental effects due to formation of glycosylation end-products, metabolic intervention with antioxidants and L-arginine can confer beneficial effects beyond those achieved by BMC alone. The effects of cotreatment with intravenous BMCs and metabolic vascular protection (1.0% vitamin E, 0.05% vitamin C, and 6% L-arginine) were examined in the ischemic hindlimb of diabetic and non diabetic mice. BMC therapy increased blood flow and capillary densities and Ki67 proliferative marker, and decreased interstitial fibrosis. This effect was amplified by metabolic cotreatment, an intervention inducing vascular protection, at least in part, through the nitric oxide pathway, reduction of systemic oxidative stress, and macrophage activation.

Lingual traumatic ulceration (Riga–Fede disease)

An 11‐month‐old male infant was referred to our clinic because of a painful ulcer of approximately 5 months’ duration on the ventral surface of the tongue ( Fig. 1 ). On physical examination, the lesion was circular (3 cm × 2 cm) with erythematous, raised, and indurated borders.

Defining the involvement of proteinases in pemphigus vulgaris : Evidence of matrix metalloproteinase-9 overexpression in experimental models of disease

Pemphigus vulgaris (PV) acantholysis represents a complex phenomenon wherein a number of factors cooperates. PV serum is known to modulate important cellular events, including kinase activity, transcriptional regulation, and proteinase expression. Indeed, transduction of signals to the cell triggered by PV serum may induce proteinase up‐regulation potentially responsible for disruption of epidermal adhesion and, ultimately, blister formation. Here, we sought to investigate this hypothesis by using both in vivo and in vitro models of PV. Microarray analysis on mouse skin tissues suggested that the equilibrium between extracellular proteinases and their inhibitors moved towards enhanced proteolytic activity in PV neonatal mouse model, at least on the transcriptional level. Conversely, genes codifying cell adhesion proteins were dramatically down‐regulated. The effects of PV serum on the protein level were then studied in vitro both in keratinocyte monolayers and skin organ cultures focusing on matrix metalloproteinase (MMP) 9 expression and activity. By means of Western blotting, zymography, and living cell immunofluorescence studies, we showed that MMP‐9 was early overexpressed in keratinocytes exposed to PV serum, and subsequently secreted in the culture medium. However, we failed to demonstrate extracellular activation of MMP‐9, since it was found in its 92 kDa inactive form in serum‐free culture supernatants. Taken together, our data demonstrated that proteinase expression, particularly of MMP‐9, is modulated by PV serum and associated with PV acantholysis. J. Cell. Physiol. 212: 36–41, 2007.

Differential expression of cyclooxygenases in hypertrophic scar and keloid tissues

Hypertrophic scar (HS) and keloid (KL) are two forms of an abnormal cutaneous scarring process, mainly characterized by excessive extracellular matrix deposition and fibroblast proliferation. Despite the increased understanding of the molecular and cellular events leading to HS and KL, the pathogenesis of these lesions remains poorly understood. A pivotal role in the formation of abnormal scars has been ascribed to transforming growth factor‐β, whose activity appears to be mediated through a link with pathways acting via cyclooxygenases (COX‐1 and COX‐2). To date, there is no report on the in vivo expression of COX‐1 and COX‐2 in human HS and KL tissues. Therefore, using immunohistochemistry and Western blot analysis, we investigated 36 cases of KL, 32 cases of HS, and 25 cases of normal skin in order to define the localization and distribution of COX‐1 and COX‐2 in the tissues of these scar lesions and the overlying epidermis. The results mainly show the following: (a) a significant overexpression of COX‐1 in HS tissues and the overlying epidermis as compared with normal skin and KL tissues and (b) a significant overexpression of COX‐2 in KL tissue and the overlying epidermis in contrast to normal skin and HS tissues. Our data support the hypothesis that both COXs are involved in the pathogenesis of scar lesions in different ways and, particularly, COX‐1 in the formation of HS and COX‐2 in the formation of KL. In addition, the overexpression of COX‐1 and COX‐2 in the epidermis overlying HS and KL tissues, respectively, underlines the importance of epithelial–mesenchymal interactions in the pathogenesis of scar lesions.

Wagner-Meissner neurilemmoma of the vulva

A 24‐year‐old woman was seen in consultation for a painless tumor of 6 cm in diameter of the vulva. The excised oval tumor was situated in the lower part of the dermis and encapsulated ( Fig. 1A ). It was completely composed of multiple adjacent cell complexes, each containing 5–20 laminar cells and resembling Wagner–Meissner touch corpuscles ( Fig 1B and 1C ). The nuclei of the laminar cells were located at the periphery of the corpuscles or were arranged transversely to the long axis of these structures along the lamellae. The nuclei were oval or showed one or several nuclear invaginations. Mitoses were not observed. These corpuscles showed a positive immunohistochemical reaction for S‐100 ( Fig. 1D ), vimentin, and neuron‐specific endase (NSE) (not shown), but were negative for CD‐68, desmin, and α‐actin. The gross and microscopic morphology and the immunohistochemical findings were compatible with a diagnosis of Wagner–Meissner neurilemmoma of the vulva. The patient remained disease free for 2 years after surgery.