High-throughput RNA sequencing of the T cell receptor alpha and beta chains for simultaneous clonality and biological analyses in Sezary syndrome.

BACKGROUND: Previous investigations pointed out a role for antigen stimulation in Sezary syndrome (SS). High-throughput sequencing of the T cell receptor (TR) offers several applications beyond diagnostic purposes, including the study of T cell pathogenesis. METHODS: We performed high-throughput RNA sequencing of the TR alpha (TRA) and beta (TRB) genes focusing on the complementarity-determining region 3 (CDR3) in 11 SS and one erythrodermic mycosis fungoides (MF) patients. Five psoriasis patients were employed as controls. Peripheral blood CD4+ cells were isolated and RNA sequenced (HiSeq2500). High-resolution HLA typing was performed in neoplastic patients. RESULTS: Highly expanded predominant TRA and TRB CDR3 were only found in SS patients (median frequency: 94.4% and 93.7%). No remarkable CDR3 expansions were observed in psoriasis patients (median frequency of predominant TRA and TRB CDR3: 0.87% and 0.69%, p < 0.001 compared to SS). CDR3 almost identical to the predominant were identified within each SS patient and were exponentially correlated with frequencies of the predominant CDR3 (R2 = 0.918, p < 0.001). Forty-six different CDR3 were shared between SS patients displaying HLA similarities, including predominant TRA and TRB CDR3 in one patient that were found in other three patients. Additionally, 351 antigen matches were detected (Cytomegalovirus, Epstein-Barr, Influenza virus, and self-antigens), and the predominant CDR3 of two different SS patients matched CDR3 with specificity for Influenza and Epstein-Barr viruses. CONCLUSIONS: Besides detecting clonality, these findings shed light on the nature of SS-related antigens, pointing to RNA sequencing as a useful tool for simultaneous clonality and biological analysis in SS.

Diagnostic Value of Genotypic Analysis in Primary Cutaneous Lymphomas using Standardized BIOMED-2 Polymerase Chain Reaction Protocols: Experience in Daily Clinical Practice.

BIOMED-2 Concerted Action BMH4-CT98-3936 (BIOMED-2) PCR protocols are an important diagnostic tool in the evaluation of cutaneous lymphomas. The aim of this study was to assess the diagnostic value of the genotyping results obtained by these techniques in daily clinical practice. A total of 360 paraffin-embedded skin samples were retrospectively reviewed from 114 cutaneous T-cell lymphomas and 35 cutaneous B-cell lym-phomas. A total of 249 biopsies from 180 patients with benign lymphoid infiltrates served as controls. T-cell receptor and immunoglobulin gene rearrangements were assessed using the BIOMED-2 method. A combined T-cell receptor gamma and beta assay approach reliably distinguished cutaneous T-cell lymphomas from benign skin T-cell infiltrates (sensitivity 89.4%; specificity 81.5%). Analysis of complete immunoglobulin heavy chain rearrangements also differentiated cutaneous B-cell lymphomas from benign B-cell infiltrates (sensitivity 85.7%; specificity 82.4%). In conclusion, the full BIOMED-2 protocol is a useful aid combined with clinical, histological and immunophenotypical findings for assessment of lymphoid clonality in skin lymphoid proliferations.

Heinz-Lippmann disease as an underrecognized cause of chronic venous insufficiency-associated cutaneous ulcers: Clinical and imaging findings.

Venous insufficiency is a frequent cause of consultation in primary care settings. Heterotopic ossification, consisting of an abnormal formation of true bone in extraskeletal soft tissues, is an underrecognized complication of chronic venous insufficiency that may cause torpid ulcers. We report a case of 78-year-old woman, with a long-standing history of venous insufficiency and tibial fracture, showing a non-healing ulcer associated with subcutaneous calcifications of the left lower extremity. Gold standard of imaging diagnosis are both plain radiographs and computed tomography but also magnetic resonance imaging could be useful for assessing the characteristics of the pathology. We describe a case of Heinz-Lippmann disease, diagnosed by using both computed tomography and magnetic resonance imaging.

Bullous pemphigoid induced by dipeptidyl peptidase-4 inhibitors. Eight cases with clinical and immunological characterization.

BACKGROUND: Dipeptidyl peptidase-4 (DPP-4) inhibitors have increasingly been identified as causative agents of bullous pemphigoid. The clinical and immunological characteristics of this pemphigoid variant are still unclear. The objective of our study was to analyze the clinical and immunological features of patients with pemphigoid induced by DPP-4 inhibitors. METHODS: All patients diagnosed with DPP-4 inhibitor-associated bullous pemphigoid at dermatology departments in three Spanish centers during the period 2013 to 2015 were included. ELISA assays for the NC16A domain of BP180 and BP230 were performed. Immunoblot studies using epidermal/dermal extracts and the C-terminal, NC16A and LAD-1 regions of BP180 were also carried out. RESULTS: A total of eight patients were identified (5 treated with vildagliptin, 2 with linagliptin, and one with sitagliptin). Of these, four presented the classical inflammatory phenotype of bullous pemphigoid and four a noninflammatory phenotype. The ELISA for BP180 (NC16A domain) was positive in six patients at diagnosis. Most patients reacted to more than one BP180 antigenic site (LAD-1 and/or C-terminal domain) on the immunoblot. Two patients showed no reaction against the NC16A domain of BP180 on either the ELISA or immunoblot but recognized either LAD-1 or both LAD-1 and the C-terminal domain. Only one of the NC16A-negative patients had a noninflammatory subtype of bullous pemphigoid. CONCLUSIONS: Patients with DPP-4 inhibitor-induced BP may present either an inflammatory or a noninflammatory phenotype of BP. IgG response against other BP180 regions different from the NC16A domain, such as LAD-1 and the C-terminal domain, could be pathogenically relevant to the onset of DPP-4 inhibitor-induced BP.

Inefficient differentiation response to cell cycle stress leads to genomic instability and malignant progression of squamous carcinoma cells.

Squamous cell carcinoma (SCC) or epidermoid cancer is a frequent and aggressive malignancy. However in apparent paradox it retains the squamous differentiation phenotype except for very dysplastic lesions. We have shown that cell cycle stress in normal epidermal keratinocytes triggers a squamous differentiation response involving irreversible mitosis block and polyploidisation. Here we show that cutaneous SCC cells conserve a partial squamous DNA damage-induced differentiation response that allows them to overcome the cell division block. The capacity to divide in spite of drug-induced mitotic stress and DNA damage made well-differentiated SCC cells more genomically instable and more malignant in vivo. Consistently, in a series of human biopsies, non-metastatic SCCs displayed a higher degree of chromosomal alterations and higher expression of the S phase regulator Cyclin E and the DNA damage signal γH2AX than the less aggressive, non-squamous, basal cell carcinomas. However, metastatic SCCs lost the γH2AX signal and Cyclin E, or accumulated cytoplasmic Cyclin E. Conversely, inhibition of endogenous Cyclin E in well-differentiated SCC cells interfered with the squamous phenotype. The results suggest a dual role of cell cycle stress-induced differentiation in squamous cancer: the resulting mitotic blocks would impose, when irreversible, a proliferative barrier, when reversible, a source of genomic instability, thus contributing to malignancy.

Chronic scalp ulceration as a late complication of fluoroscopically guided cerebral aneurysm embolization.

Fluoroscopy-induced chronic radiation dermatitis resulting from prolonged exposure to ionizing radiation during interventional procedures has been documented in the medical literature. However, this condition often requires a high clinical suspicion in order to establish a correct diagnosis. In this report, the development of deep scalp ulceration with bone exposure following the endovascular coiling of an anterior communicating artery aneurysm 8 years before is described. A skin biopsy specimen demonstrated changes consistent with chronic radiation dermatitis and ruled out malignancy. This case report expands the clinical manifestation spectrum of fluoroscopy-induced chronic radiation skin injury and highlights the importance of recognizing these lesions early to prevent morbidity related to radiation-induced skin damage.

Cutis laxa-like mycosis fungoides.

Mycosis fungoides is the most common form of primary cutaneous T-cell lymphoma. Several clinical and clinicopathological variants of mycosis fungoides have been reported. A 75-year-old woman presenting with multiple ill-defined areas of marked cutaneous wrinkling on the trunk and extremities is reported. Histopathological examination showed characteristic features of mycosis fungoides along with an interstitial dermal infiltrate composed predominantly of atypical lymphocytes with histiocytes intermingled within the collagen bundles. A focal reduction and fragmentation of elastic fibers was demonstrated. This observation illustrates a peculiar and previously unreported clinicopathological presentation of mycosis fungoides: cutis laxa-like mycosis fungoides, expanding the spectrum of mycosis fungoides variants associated with abnormalities of the dermal elastic fibers.