Fabio Pagni

Comparison of 5-tiered and 6-tiered diagnostic systems for the reporting of thyroid cytopathology: a multi-institutional study.

At present, thyroid fine‐needle aspiration (FNA) specimens are diagnosed using a tiered classification scheme, with the most popular of these being the 5‐tiered and 6‐tiered systems. In this study, the authors present their institutional experiences using these 2 different systems and evaluate their efficacy based on the surgical follow‐up.

Combined hepatocellular‐cholangiocarcinoma: A population‐level analysis of an uncommon primary liver tumor

Combined hepatocellular‐cholangiocarcinoma (cHCC‐CC) is a rare primary liver cancer. Our aims were to analyze the demographic, clinical, and pathological characteristics of cHCC‐CC at a population level and to investigate the effects of these features as well as different management strategies on the prognosis. The Surveillance, Epidemiology, and End Results (SEER) database was analyzed for 1988‐2009. Data analyses were performed with chi‐square tests, analyses of variance, Kaplan‐Meier curves, and Cox proportional hazards regression. Four hundred sixty‐five patients with cHCC‐CC, 52,825 patients with hepatocellular carcinoma (HCC), and 7181 patients with cholangiocarcinoma (CC) were identified. cHCC‐CC was more common in patients who were white, male, and older than 65 years. Treatment was more frequently nonsurgical/interventional. Patients with cHCC‐CC, HCC, and CC had 5‐year overall survival (OS) and disease‐specific survival rates of 10.5%, 11.7%, and 5.7% (P < 0.001) and 17.8%, 21.0%, and 11.9% (P < 0.001), respectively. For cHCC‐CC patients, an increasing invasiveness of the therapeutic approach was significantly associated with prolonged survival (P < 0.001). In a multivariate model, black race, a distant SEER stage, and a tumor size of 5.0 to 10.0 cm were independently associated with lower survival for cHCC‐CC patients; a year of diagnosis after 1995 and surgical treatment with minor hepatectomy, major hepatectomy (MJH), or liver transplantation (LT) were independently associated with better survival for cHCC‐CC patients. Patients diagnosed with cHCC‐CC, HCC, and CC and treated with LT had 5‐year OS rates of 41.1%, 67.0%, and 29.0%, respectively (P < 0.001). In conclusion, cHCC‐CC patients appear to have intermediate demographic, clinical, and survival characteristics in comparison with HCC and CC patients. cHCC‐CC patients undergoing LT showed inferior survival in comparison with HCC patients, and the role and indications for LT in cHCC‐CC have yet to be defined. At this time, MJH may be considered the best therapeutic approach for such patients. Liver Transpl 20:952–959, 2014.

Virtual Surgical Pathology in Underdeveloped Countries: The Zambia Project

Only 1 surgical pathology laboratory is available in Zambia, a country with a population of 12 million people. Since 2004 the Italian association of pathologists Patologi Oltre Frontiera has been working to create a virtual laboratory through the use of telemedicine. The project has involved staining histologic preparations on site, with the interpretation of imaged slides performed abroad through telepathology. Starting in April 2007, all surgical specimens obtained in Mtendere Mission Hospital, Chirundu, Zambia, were submitted for microscopic examination through whole-slide scans. Two independent Italian pathologists evaluated the cases by means of satellite connection and the final diagnoses were sent to Zambian clinicians via the internet. This article describes the spectrum of diagnoses made via telepathology for the Zambian population. Also, we analyze the concordant and discordant data between this telepathology method and traditional microscopy in a developing country. Moreover, we provide possible solutions for providing pathology services in other underdeveloped countries.

'Indeterminate for malignancy' (Tir3/Thy3 in the Italian and British systems for classification) thyroid fine needle aspiration (FNA) cytology reporting: morphological criteria and clinical impact

The British system (Thy1–5), the Bethesda system for reporting thyroid cytopathology (BSRTC) and the Italian Society of Anatomic Pathology and Cytology (SIAPEC) classification represent the most important international classifications for thyroid cytopathology. Irrespective of the system used, the ‘indeterminate’ categories are still debated among cytopathologists, particularly with regard to diagnostic criteria, clinical impact of subclassification and role of molecular techniques.

The value of repeat biopsy in the management of lupus nephritis: an international multicentre study in a large cohort of patients

BACKGROUND The International Society of Nephrology/Renal Pathology Society (ISN/RPS) classification represents the gold standard for the histological evaluation of Systemic Lupus Erythematosus (SLE) nephritis. A repeat biopsy (RB) might be an important tool to provide information on long-term renal outcomes and optimal therapy. Aims of this study were to evaluate the use of the ISN/RPS classification and the role of RB in routine clinical practice. METHODS A total number of 142 patients with SLE nephritis and with adequate reference and RB samples were included in this multicentre retrospective study. A meticulous histological examination was centrally performed on first and RB and compared with clinical variables and follow-up data. RESULTS Morphological features of the ISN/RPS classification: at first and RB, significant differences were observed between segmental classes (III, IV-S) and Class IV-G in mesangial proliferation, wire loops and tuft necrosis. Clinical features and ISN/RPS classification: the correlation between serum creatinine, proteinuria, blood pressure levels and histological classes at first and RB demonstrated more severe renal disease in Class IV-G, both at first and RB. Agreement between ISN/RPS classification at first and RB: 40.8% of patients changed the histological class. Fifty per cent of Class II (mild mesangial form) were reclassified as Class IV-G at RB, whereas 18.9% of Class IV-G were reclassified as Class II. The transition among segmental (III/IV-S) and mesangial forms (II/IV-G) was extremely rare. The comparison between the clinical parameters at the final follow-up and the ISN/RPS classification confirmed that the trend of serum creatinine and proteinuria between the different classes was better described at the RB (higher in Class IV-G) than on the first biopsy. CONCLUSIONS The histopathological data suggest that morphological differences between segmental and global forms do exist, possibly due to different pathogenetic mechanisms. An RB strategy could provide additional information on long-term renal outcomes. A strategy of protocol biopsies could be useful in perspective future trials to better understand the therapeutic response and the natural history of this disease.

An alternative approach in endocrine pathology research: MALDI-IMS in papillary thyroid carcinoma

To the Editor, Many different molecular techniques (polymerase chain reaction (PCR), DNA sequencing, fluorescence in situ hybridization (FISH)) have been introduced in thyroid pathology [1]. Fewer studies evaluated the role of proteomic analysis in the research of new useful targets [2, 3]. Matrixassisted laser desorption/ionization (MALDI) imaging mass spectrometry (IMS) is a unique technology that explores the spatial distribution of biomolecules directly in situ, thus integrating molecular and morphological information. Therefore, we are investigating the potential role of MALDIIMS in detecting new diagnostic targets in papillary thyroid carcinoma (PTC). We have addressed the issue of molecular stratification of PTC in a small cohort of samples, evaluating both the genomic profile of the main genes of interest (BRAF, N-H-K RAS point mutations, and PAX8/PPARγ or RET/PTC rearrangements) and the proteomic profile shown byMALDIIMS analysis. We have collected ex vivo cytological specimens (see “Technical Note”), in order to analyse a sample perfectly mimicking the in vivo fine-needle aspiration (FNA) biopsy, while respecting the ethical requirements. Unsupervised proteomic analysis of the samples was followed by comparison with histopathology and genetic classification of the patients (Table 1). Data generated by MALDI-IMS were submitted to hierarchical cluster analysis (HCA), in order to evaluate the different proteomic expressions in the cases under study. HCA allowed to cluster proteomic spectra based on their similarity on a dendrogram: spectra showing comparable features were grouped under the same node of the dendrogram; then, it was possible to select nodes and assign them a specific colour (Fig. 1) [4]. Node A groups together three cytological specimens (two collected from patient 1 and one from patient 2), all histologically diagnosed as hyperplastic benign nodules. Node B groups together specimens collected from patients 3, 4 and 5, who were affected by PTC comparable for stage and histotype (Table 1). Node C, instead, groups a microcarcinoma (follicular variant (fv) of PTC), from patient 6, and a nodule histologically classified as uncertain malignant potential (UMP) tumour (patient 7). The last one, originated in a multinodular goitre environment and showed ambiguous morphological features, defined as borderline between a hyperplastic nodule and an “incipient” fv,PTC. The three distinct nodes (A, B, C) generated by HCA analysis confirmed that MALDI-IMS can potentially discriminate between benign and malignant thyroid lesions. Moreover, the homologies between cases 6 and 7 highlight that MALDI-IMS is able to identify a PTC even when the classic diagnostic morphological aspects are still unclear and ambiguous (mild nuclear clearing, rare grooves, no pseudoinclusions). This finding is in agreement Veronica Mainini and Fabio Pagni equally contributed to this paper.

α‐1‐antitrypsin detected by MALDI‐Imaging in the study of glomerulonephritis: its relevance in chronic kidney disease progression

Idiopathic glomerulonephritis (GN), such as membranous glomerulonephritis, focal segmental glomerulosclerosis (FSGS), and IgA nephropathy (IgAN), represent the most frequent primary glomerular kidney diseases (GKDs) worldwide. Although the renal biopsy currently remains the gold standard for the routine diagnosis of idiopathic GN, the invasiveness and diagnostic difficulty related with this procedure highlight the strong need for new diagnostic and prognostic biomarkers to be translated into less invasive diagnostic tools. MALDI‐MS imaging MALDI‐MSI was applied to fresh‐frozen bioptic renal tissue from patients with a histological diagnosis of FSGS (n = 6), IgAN, (n = 6) and membranous glomerulonephritis (n = 7), and from controls (n = 4) in order to detect specific molecular signatures of primary glomerulonephritis. MALDI‐MSI was able to generate molecular signatures capable to distinguish between normal kidney and pathological GN, with specific signals (m/z 4025, 4048, and 4963) representing potential indicators of chronic kidney disease development. Moreover, specific disease‐related signatures (m/z 4025 and 4048 for FSGS, m/z 4963 and 5072 for IgAN) were detected. Of these signals, m/z 4048 was identified as α‐1‐antitrypsin and was shown to be localized to the podocytes within sclerotic glomeruli by immunohistochemistry. α‐1‐Antitrypsin could be one of the markers of podocyte stress that is correlated with the development of FSGS due to both an excessive loss and a hypertrophy of podocytes.

Analytical validity of a microRNA-based assay for diagnosing indeterminate thyroid FNA smears from routinely prepared cytology slides

The majority of thyroid nodules are diagnosed using fine‐needle aspiration (FNA) biopsies. The authors recently described the clinical validation of a molecular microRNA‐based assay, RosettaGX Reveal, which can diagnose thyroid nodules as benign or suspicious using a single stained FNA smear. This paper describes the analytical validation of the assay.

Proteomics in thyroid cytopathology: Relevance of MALDI-imaging in distinguishing malignant from benign lesions.

Several proteomic strategies are used extensively for the purpose of biomarker discovery and in order to obtain insights into the molecular aspects of cancers, using either body fluids or tissue as samples. Among them, MALDI‐imaging can be applied to cytological thyroid specimens to investigate the molecular signatures of different pathological conditions and highlight differences in the proteome that are of relevance for diagnostic and pathogenetic research. In this study, 26 ex‐vivo fine needle aspirations from benign thyroid nodules (n = 13) and papillary thyroid carcinomas (n = 13) were analyzed by MALDI‐imaging. Based on the specific protein signatures capable of distinguishing the aforementioned patients, MALDI‐imaging was able to correctly assign, in blind, the specimens from ten additional FNABs to a malignant or benign class, as later confirmed by the morphological classification. Moreover, some proteins presented a progressive overexpression in malignant phenotypes when compared with Hashimotos thyroiditis and hyperplastic/follicular adenoma. This data not only suggests that a MALDI‐imaging based approach can be a valuable tool in the diagnosis of thyroid lesions but also in the detection of proteins that have a possible role in the promotion of tumorigenic activity.

MALDI imaging mass spectrometry in glomerulonephritis: feasibility study

The in‐situ proteomics technology known as matrix‐assisted laser desorption/ionization imaging mass spectrometry (MALDI‐IMS) is a powerful technique that combines traditional histology and proteomics.