Aquaporin-1 expression in fluoro-edenite-induced mesothelioma effusions: An approach by cell-block procedure.

OBJECTIVE: Aquaporin 1 (AQP-1) is a water channel protein found in cell membranes, whose expression has been considered an independent favourable prognostic factor in pleural malignant mesothelioma (MM). The aim of this study was to evaluate the expression of AQP-1 and its prognostic value in a series of pleural MM effusions, from a geographical area with high concentrations of fluoro-edenite (FE). METHODS: We selected 25 MM cases from Biancavilla (Italy), an area with high environmental concentrations of FE. Cytological samples, cell-blocks (CB), clinical and follow-up data were available for all cases. Immunohistochemistry for calretinin, CK5/6, WT1, CK7 and TTF1 was used on CB sections to confirm the cytological diagnosis of MM. Immunohistochemistry for AQP-1 was performed and high expression was defined when ≥50% of tumour cells showed linear and circumferential membranous staining. RESULTS: The cohort included 16 men and nine women (median age: 67.5 years; range: 49-88 years). The median survival was 14 months (range 1.5-60 months), with a significant value (P = 0.006). All cases have been histologically confirmed and classified as epithelioid (16 cases), biphasic (seven cases) and sarcomatoid (two cases). AQP-1 high expression has been observed in 16 cases. Comparing AQP-1 high expression to the survival of corresponding patients, a significant association with a slight increased overall survival of 12 months has been demonstrated. Nine patients with a AQP-1 score less than 50% showed a shorter median overall survival (7 months). CONCLUSIONS: AQP-1 high expression is detectable on cytological samples of FE-induced MM with a prognostic value.

The post-surgical era of GBM: How molecular biology has impacted on our clinical management. A review.

Glioblastoma (GBM) is the most common glioma in adults, with incidence increasing by 3% per year. According to the World Health Organization Classification of Central Nervous System Tumors, GBM is considered a grade IV tumor due to its malignant behavior. The aim of this review is to summarize the main biological aspects of GBM. In particular, we focused our attention on those alterations which have been proven to have an impact on patients' outcome, mainly in terms of overall survival (OS), or on the tumor response to therapies. We have also analyzed the cellular biology and the interactions between GBM and the surrounding environment.

Cytological preparations for molecular analysis: A review of technical procedures, advantages and limitations for referring samples for testing.

Minimally invasive procedures such as endobronchial ultrasound-guided transbronchial needle aspiration (EBUS-TBNA) must yield not only good quality and quantity of material for morphological assessment, but also an adequate sample for analysis of molecular markers to guide patients to appropriate targeted therapies. In this context, cytopathologists worldwide should be familiar with minimum requirements for refereeing cytological samples for testing. The present manuscript is a review with comprehensive description of the content of the workshop entitled Cytological preparations for molecular analysis: pre-analytical issues for EBUS TBNA, presented at the 40th European Congress of Cytopathology in Liverpool, UK. The present review emphasises the advantages and limitations of different types of cytology substrates used for molecular analysis such as archival smears, liquid-based preparations, archival cytospin preparations and FTA (Flinders Technology Associates) cards, as well as their technical requirements/features. These various types of cytological specimens can be successfully used for an extensive array of molecular studies, but the quality and quantity of extracted nucleic acids rely directly on adequate pre-analytical assessment of those samples. In this setting, cytopathologists must not only be familiar with the different types of specimens and associated technical procedures, but also correctly handle the material provided by minimally invasive procedures, ensuring that there is sufficient amount of material for a precise diagnosis and correct management of the patient through personalised care.

Fine needle cytology pre-surgical differentiation of parathyroid neoplasms: Is it reliable?

BACKGROUND: Fine needle cytology (FNC) of a parathyroid neoplasia (PN) is reliable, but needs to be confirmed by Parathormone (PTH) and Thyroglobulin (TG) immunoassay on needle washing or by immunocytochemistry (ICC) evaluation. The differentiation between parathyroid adenoma (PA), atypical adenoma (PAA) and carcinoma (PC) is difficult on histology or even impossible on FNC. The aim of this study was to evaluate possible cytological criteria to classify FNC-PN further. METHODS: Twenty-three FNC samples of PN and parathyroid cysts were rather then have been reviewed. The series includes 18 PNs, 4 cysts and 1 Thyr3B (histologically diagnosed as PA). Cytological features were: cellularity, patterns (follicular, solid or papillary), clear, oncocytic, isolated cells, nuclear atypia, cytoplasmic inclusions, nucleoli and mitoses. Data were compared with the histological controls. RESULTS: Seventeen PNs, 2 cysts and 1 Thyr3B FNC samples were histologically diagnosed as PA (16), PAA (2) and PC (2). Two cysts and 1 PN were not confirmed histologically. Cytological features and incidences were: high cellularity (1 PA, 1 PAA, 2 PCs), follicular (8 PAs, 1 PAA), solid (5 PAs, 1 PC), papillary pattern (1PA, 1 PAA, 1 PC), clear cells (4 PAs, 1 PAA, 2 PCs), oncocytic cells (6 PAs, 1 PAA, 2 PCs), isolated cells (5 PAs, 2 PAAs, 2 PCs), nuclear atypia (2 PAs, 1 PAA, 2 PCs), cytoplasmic inclusions (4 PAs, 2 PCs), nucleoli (2 PCs) and mitoses (2 PCs). CONCLUSION: Evident nucleoli and mitoses may suggest the differentiation between PA and PC. However, further investigations are required to confirm these preliminary observations.

Innate immunity in cutaneous melanoma.

The skin immune system is composed of a vast network of immune cells, including lymphocytes, macrophages, neutrophils, dendritic cells and Langerhans cells, which not only are involved in inflammatory responses but also contribute to homeostatic function and may participate in the various steps of carcinogenesis. Many studies support the notion that innate immunity has a key role in the development, growth and prognosis of cutaneous malignant melanoma (MM), through the release of pro- and/or anti-inflammatory cytokines and tumour growth factors. The tumour environment in a major subset of cutaneous MM shows evidence of a T cell-infiltrated phenotype, but there is less known about the presence and the phenotype of other immune system cells. Response to immunotherapy is largely correlated with the presence of T cells in the tumour microenvironment, while the regulation exerted by stromal components such as macrophages and mast cells has been less investigated. In the current report, we review the recent literature, focusing our attention on the role of macrophages, dendritic cells, mast cells and natural killer cells in orchestrating MM progression, to better understand tumour immunobiology. The identification of new therapeutic targets and the application of approaches aimed at modulating crosstalk between immune and tumour cells, could have a crucial impact on immunotherapy and result in better clinical outcome. We hope this review will be helpful in cutaneous MM research.

Randomized comparison of power Doppler ultrasonography-guided core-needle biopsy with open surgical biopsy for the characterization of lymphadenopathies in patients with suspected lymphoma.

The sensitivity of lymph node core-needle biopsy under imaging guidance requires validation. We employed power Doppler ultrasonography (PDUS) to select the lymph node most suspected of malignancy and to histologically characterize it through the use of large cutting needle. Institutional review board approval and informed consent were obtained for this randomized clinical trial. In a single center between 1 January 2009 and 31 December 2015, patients with lymph node enlargement suspected for lymphoma were randomly assigned (1:1) to biopsy with either standard surgery or PDUS-guided 16-gauge modified Menghini needle. The primary endpoint was the superiority of sensitivity for the diagnosis of malignancy for core-needle cutting biopsy (CNCB). Secondary endpoints were times to biopsy, complications, and costs. A total of 376 patients were randomized into the two arms and received allocated biopsy. However, four patients undergoing CNCB were excluded for inadequate samples; thus, 372 patients were analyzed. Sensitivity for the detection of malignancy was significantly better for PDUS-guided CNCB [98.8%; 95% confidence interval (CI), 95.9-99.9] than standard biopsy (88.7%; 95% CI, 82.9-93; P < 0.001). For all secondary endpoints, the comparison was significantly disadvantageous for conventional approach. In particular, estimated cost per biopsy performed with standard surgery was 24-fold higher compared with that performed with CNCB. The presence of satellite enlarged reactive and/or necrotic lymph nodes may impair the success of an open surgical biopsy (OSB). PDUS and CNCB with adequate gauge are diagnostic tools that enable effective, safe, fast, and low-cost routine biopsy for patients with suspected lymphoma, avoiding psychological and physical pain of an unnecessary surgical intervention.

Immunoglobulin heavy and light chains and T-cell receptor beta and gamma chains PCR assessment on cytological samples. A study comparing FTA cards and cryopreserved lymph node fine-needle cytology.

OBJECTIVES: To evaluate and compare the DNA yield and quality extracted from lymph node fine needle cytology (FNC) samples stored on FTA cards to those cryopreserved, and to assess the immunoglobulin heavy and light chains (IGHK) and T-Cell receptor beta and gamma chains (TCRBG) PCR tests. METHODS: DNA extractions were performed on FNC of 80 non-Hodgkin lymphomas (NHL), four myelomas and 56 benign reactive hyperplasias (BRH) cryopreserved and stored on FTA cards. The JAK2 gene was amplified to assess the DNA integrity and the IGHK/TCRBG clonality status was tested. RESULTS: IGHK monoclonality was found in 99% of B-cell NHL and 100% of myeloma. TCRBG monoclonality was found in 100% of T-cell NHL. TCRBG polyclonality was detected in 97% of B-cell NHL, 100% of myeloma and 96% of BRH. IGHK/TCRBG PCR data were confirmed by histological and/or follow-up controls. No differences were found in the DNA quality between cryopreservation and FTA cards storage methods. CONCLUSIONS: IGHK/TCRBG PCR of the lymphoproliferative process on FTA cards is comparable to those cryopreserved. FTA cards can be used to store lymph node FNC for further molecular investigations.

Surgical management of cervico-mediastinal goiters: Our experience and review of the literature.

AIM: We analyze and discuss the clinical presentation, the diagnostic procedures and the surgical technique in relation to post-operative complications and results in cervico-mediastinal thyroid masses admitted in Thoracic Surgery Unit of AOU Second University of Naples from 1991 to 2006 and in Thoracic Surgery Unit of AOU "S. Giovanni di Dio & Ruggi D'Aragona" of Salerno over a period of 3 years (2011-2014). METHODS: We reviewed 97 patients who underwent surgical treatment for cervico-mediastinal goiters. 47 patients (49.2%) had cervico-mediastinal goiter, 40 patients (40%) had mediastino-cervical goiter and 10 patients (10.8%) had mediastinal goiter. 73 cases were prevascular goiters and 24 were retrovascular goiters. We performed total thyroidectomy in 40 patients, subtotal thyroidectomy in 46 patients and in 11 cases the resection of residual goiter. In 75 patients we used only a cervical approach, in 21 patients the cervical incision was combined with median sternotomy and in 1 patient with transverse sternotomy. RESULTS: Three patients (3.1%) died in the postoperative period (2 cardio-respiratory failure and 1 pulmonary embolism). The histologic study revelead 8 (7.7%) carcinomas. Postoperative complications were: dyspnea in 9 cases (10.7%), transient vocal cord paralysis in 6 patients (9.2%), temporary hypoparathyroidism in 9 patients (9.2%) and kidney failure in 1 case (0.9%). CONCLUSIONS: The presence of a cervico-mediastinal thyroid mass with or without respiratory distress requires a surgical excision as the only treatment option. Thyroid masses extending to the mediastinum can be excised successfully by cervical incision. Bipolar approach (cervical incision and sternotomy) has an excellent outcome, achieving a safe resection, especially in large thyroid masses extending to the mediastinum with close relations to mediastinal structures and in some limited cases (carcinoma, thyroiditis, retrovascular goiter, ectopic goiter). Postoperative mortality and morbidity is very low, independent of surgical techniques. Other surgical approaches for excision of a Posterior Mediastinal Thyroid Goiter reported in literature are: VATS techniques to remove an ectopic intrathoracic goiter, robot-assisted technique for the removal of a substernal thyroid goiter, with extension into the posterior mediastinum.

Training and practice of cytotechnologists: a discussion forum focused on Europe.

OBJECTIVES: To discuss the role and training of cytotechnologists (CTs) in Europe, to identify areas of good practice and to provide an informed opinion to those providing guidelines for training and practice in Europe. METHODS: All members of the Editorial Advisory Board of Cytopathology were invited to take part in a 'discussion forum' for which six topics were circulated in advance concerning the roles of CTs with regard to: (1) pre-screening slides; (2) 'signing out' reports; (3) carrying out ancillary techniques; (4) supervising laboratory staff; (5) taking part in rapid on-site evaluation (ROSE) of fine needle aspirates (FNAs); and (6) whether CTs were trained specifically in cytopathology or in general histopathology. Notes of the meeting were circulated by email and a final report was agreed by 22 participants from 17 predominantly European countries. RESULTS: Training for CTs throughout Europe was variable, especially for non-gynaecological cytology, which was inconsistent with the range of activities required. The participants recommended graduate entry, preliminary training in general laboratory technology, and subsequent training to take account of the probability and, in some centres, the reality of primary cervical cancer screening changing from cytology to human papillomavirus (HPV) testing. They further recommended that CTs should perform HPV tests and take part in ROSE for FNAs, and they supported the European Federation of Cytology Societies developing guidelines for training and practice. CONCLUSION: With CT training added to a university-based education in laboratory or biomedical science, a career in cytotechnology should be an attractive option involving a diverse range of laboratory and clinically based activities.

Fine needle aspiration cytology of lymphoproliferative lesions of the oral cavity.

OBJECTIVE: Oral cavity non-Hodgkin lymphoma (OCL) is a rare condition that may be clinically and radiologically indistinguishable from other pathologies of the mouth. A complete excision or adequate biopsy of the OCL may be difficult. Fine needle aspiration (FNA) cytology has been successfully utilized in the pre-operative diagnosis of oral masses and in lymphoma involving other anatomical areas. Our experience with FNA pre-operative cytological diagnosis of 16 OCLs is reported herein. METHODS: The results of FNA cytology on 16 consecutive lymphoproliferative lesions of the oral cavity collected over an 8-year period in three institutions were retrieved. Sampled lesions were submucosal masses of different sizes bulging into the oral cavity. Rapid on-site evaluation (ROSE) and routine cytological staining were performed. Immunocytochemistry (ICC), flow cytometry (FC) and polymerase chain reaction (PCR) of the IGH (immunoglobulin heavy) locus were performed on additional passes according to ROSE. RESULTS: Fourteen OCLs, one myeloma and one florid reactive lymphoid hyperplasia (FRLH) were diagnosed by FNA. OCLs were diagnosed as large B-cell (eight cases) and small B-cell (six cases) lymphomas. Histology revealed eight diffuse large B-cell lymphomas (DLBCL), four lymphomas of mucosa-associated lymphoid tissue (MALT), two follicular lymphomas and one FRLH; no false-negative or false-positive results were diagnosed, but accurate subclassification was obtained in four cases only. CONCLUSIONS: FNA diagnosis of OCLs may be hampered by the rare incidence, anatomical context and difficulties in obtaining a sufficient amount of cells. Ancillary techniques should be used according to ROSE; a pre-operative FNA cytology diagnosis can avoid unnecessary extensive surgery and speed up the institution of therapeutic procedures.

Non lymphomatous clonal B-Cell populations in enlarged lymph nodes in acquired immunodeficiency syndrome.

Clonal B-cell populations in non-lymphomatous processes have been sporadically reported in enlarged reactive lymph nodes and mucosa-associated lymphoid cell populations. These generally small clones are considered non-malignant proliferations of B-lymphocytes determined by an abnormal response to bacterial or viral antigen stimulation. In cases reported in literature, clonality was detected by light chain assessment and or by polymerase chain reaction (PCR) analysis of immunoglobulin heavy chain (IgH) gene in histologically and clinically proven non lymphomatous processes. In this study the clinical, cytological, phenotypical and pathological features of three HIV patients in which non-lymphomatous clonal B-cell populations detected in enlarged lymph nodes are reported. All the patients complained for later cervical lymph nodes enlargement, positive at the FDG-positron emission tomography scan. Fine needle cytology, coupled with flow cytometry showed atypical lymphoid cell proliferations and kappa (2 cases) or lambda (1 case) light chain restriction. Reactive, non lymphomatous nature of these processes were then proven by histological control in two cases and by clinical follow-up in the last one; corresponding clinical and pathological aspects are discussed. Clonal B-cell populations in non-lymphomatous processes can sporadically occur in enlarged reactive lymph nodes in immunodeficiency as well as in autoimmune processes. Awareness of the phenomenon and attention should be paid in the evaluation of corresponding pathological features and in the clinical management of corresponding patients.

Low-dose valgancyclovir as cytomegalovirus reactivation prophylaxis in allogeneic haematopoietic stem cell transplantation.

The efficacy and safety of low dose oral valgancyclovir (VGCV) as cytomegalovirus (CMV) reactivation prophylaxis was retrospectively evaluated in 32 consecutive patients which underwent allogeneic HLA-matched related and unrelated hematopoietic stem cell transplantation (HSCT). Thirty HSCT recipients showed pretransplant CMV seropositivity. Fifteen received a myeloablative conditioning regimen, while seventeen patients received a reduced-intensity conditioning regimen. Twenty-one patients received graft-versus-host disease (GVHD) prophylaxis with cyclosporin A (CsA) and methotrexate (MTX), and the others CsA with MTX and anti-thymocyte globulin. CMV infection was monitored weekly using polymerase chain reaction (PCR). VGCV was administered orally at a dose of 450 mg daily for six months. Six patients developed a positive CMV-PCR on average 56 days after HSCT successfully treated with VGCV at 1800 mg/day, except one who developed fatal gastrointestinal CMV disease. At the time of CMV reactivation, four patients had been affected by grade II-IV acute GVHD and two by an extensive chronic GVHD. No significant specific VGCV-related toxicity was encountered. Seven patients presented hematological toxicity which did not require drug discontinuation. Our data suggest that low dose VGCV is safe and effective as CMV reactivation prophylaxis in allogeneic HSCT recipients. These results require further validation in prospective randomized studies.

Successful management of pulmonary mucormycosis with liposomal amphotericin B and surgery treatment: a case report.

Mucormycosis is an increasingly recognized invasive fungal infection (IFI) in patients with acute myeloid leukemia (AML) and after allogeneic (allo) stem cell transplantation (HSCT); it is mainly due to the severe and prolonged neutropenia related to high-dose chemotherapy. In such patients, the lung is the most frequently involved site in mucormycosis. Since rapidly progressive dissemination may occur after pulmonary mucormycosis in hematologic malignancies, early diagnosis and prompt initiation of an effective antifungal therapy is mandatory for a successful outcome. We report the case of a young AML patient who developed, early after the onset of neutropenia in the first induction phase of chemotherapy, a rapidly progressive pulmonary IFI, successfully treated with liposomal Amphotericin-B (LAmB) and then with a limited open toracothomy biopsy, clearly establishing diagnosis of mucormycosis and removing lung infiltrate. Secondary prophylaxis with LamB, applied during both consolidation therapy and myeloablative sibling allogeneic HSCT, was effective to prevent IFI recurrence despite the development of grade I acute graft-versus-host disease (GVHD) and limited chronic GVHD requiring immunosuppressive treatment. Our case report further provide evidence that the combined surgical and LAmB therapy is an effective and safe choice for the management of pulmonary mucormycosis in hematological immunocompromised patients.