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Aggressive B-cell lymphomas present as a heterogeneous spectrum of disease. A primary diagnosis in the bone marrow (BM) may be challenging in terms of diagnostic classification and clinical handling, owing to limited architectural information. Aggressive B-cell lymphomas can be subdivided into entities that typically present primarily in the BM, and cases with BM involvement in which the bulk of disease is present in other organs. One main topic at the 2018 BM workshop of the European Association of Haematopathology/Society of Hematopathology was therefore aggressive B-cell lymphomas with a primary BM presentation. The spectrum of cases submitted to this topic gave a good overview of commonly encountered problems, as well as unusual manifestations, and highlighted areas of imprecise disease definitions and diagnostic grey zones. The categories submitted to the workshop included cases of Burkitt lymphoma (BL) with unusual features, high-grade B-cell lymphomas (HG-BCLs) with and without so-called double/triple-hit, and diffuse large B-cell lymphomas (DLBCLs) with a primary BM presentation. Areas of difficulties included the morphological boundaries of HG-BCL not otherwise specified, cases with MYC and bcl-2 or bcl-6 translocations and terminal deoxynucleotidyl transferase (TdT) expression, which were categorised as B-cell lymphoblastic leukaemia/lymphoma if most cells showed TdT positivity, and the clinicopathological overlap between intravascular large B-cell lymphoma, CD5-positive DLBCL, and DLBCL with primary presentations in the BM, spleen, and liver. This review summarises our understanding of the main aggressive B-cell lymphoma categories with a common primary BM presentation and potential problem areas, and makes suggestions for the immunophenotypic and genetic work-up, illustrated by the interesting and challenging cases submitted to the workshop., (© 2020 The Authors. Histopathology published by John Wiley & Sons Ltd.)

Calreticulin (CALR) mutations are detected in the majority of JAK2 wild type patients with essential thrombocythemia (ET). Unlike JAK2 V617F and MPL point mutations, CALR mutations are highly heterogeneous, with several types of indels being reported so far. CAL2 is a monoclonal antibody specifically recognizing the C-neoterminal peptide derived from all the frameshift mutations of CALR. We retrospectively analysed 172 ET patients diagnosed at our Institution from 1980 to 2015. In JAK2 V617F - and MPL W515K/L -wild type patients CALR mutations were searched on peripheral blood and CAL2 immunostaining was performed on bone marrow. In addition, bone marrow biopsies were histologically reviewed for megakaryocytic features. Thirty-one patients (18%) were CALR-mutated. Concordance between molecular and immunohistological detection of CALR mutations was near complete, albeit a single patient was found to be positive by molecular tests only. Two patterns were defined in CAL2-positive bone marrow samples, characterized by staining of almost only megakaryocytes (pattern A: 41%) or staining of megakaryocytes and ≥ 2% small non megakaryocytic elements (pattern B: 59%), at least partially being myeloid precursors. Pattern B biopsies had higher cellularity and number of megakaryocytes compared to pattern A samples. In this series, CAL2 allowed rapid and cost-efficient identification of CALR-mutated ET patients. The biological significance of different staining pattern should be confirmed in wider and independent series.

Patients with Systemic Mastocytosis (SM) need a highly sensitive diagnostic test for D816V detection of the KIT receptor gene. Along with histology/cytology and flow cytometry evaluation, bone marrow (BM) from 110 consecutive adult patients referred with a suspicion of SM to Multidisciplinary Outpatient Clinic for Mastocytosis in Verona were tested both by Amplification Refractory Mutation System Reverse Transcriptase quantitative real time Polymerase Chain Reaction (ARMS-RT-qPCR) and RT-PCR+Restriction Fragment Length Polymorphism (RFLP) followed by Denaturing-High Performance Liquid Chromatography (D-HPLC) and Sanger sequencing. ARMS-RT-qPCR identified D816V mutation in 77 patients, corresponding to 100% of cases showing CD25(+) mast cells (MCs) whereas RT-PCR+RFLP/D-HPLC+sequencing revealed D816V mutations in 47 patients. According to the 2008 WHO criteria 75 SM, 1 Cutaneous Mastocytosis (CM), 1 monoclonal MC activation syndrome (MMAS), and 1 SM Associated with Haematologic Non-Mast Cell Disorder (SM-AHNMD) were diagnosed. Seventeen out 75 SM patients (23%) would have not satisfied sufficient WHO criteria on the basis of the sole RT-PCR+RFLP: these patients had significantly lower serum tryptase levels and amount of CD25(+) MCs. Therefore, ARMS-RT-qPCR might result particularly useful, in patients that do not fulfil major BM histological criterion, for the recognition of indolent SM with a very low MC burden., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

CD49d, the α4 chain of the VLA‐4 integrin, is a negative prognosticator in chronic lymphocytic leukemia (CLL) with a key role in CLL cell‐microenvironment interactions mainly occurring via its ligands VCAM‐1 and fibronectin. In the present study, we focused on EMILIN‐1 (Elastin‐MIcrofibriL‐INterfacer‐1), an alternative VLA‐4 ligand whose role has been so far reported only in non‐hematological settings, by investigating: i) the distribution of EMILIN‐1 in CLL‐involved tissues; ii) the capability of EMILIN‐1 to operate, via its globular C1q (gC1q) domain, as additional adhesion ligand in CLL; iii) the functional meaning of EMILIN‐1 gC1q/VLA‐4 interactions in CLL. EMILIN‐1 is widely present in the CLL‐involved areas of bone marrow biopsies (BMBs) without difference between CD49d negative and positive cases, displaying at least three different expression patterns: "fibrillar", "dot‐like" and "mixed". The lack in CLL‐BMB of neutrophil elastase, whose proteolytic activity degrades EMILIN‐1 and impairs EMILIN‐1 function, suggests full functional EMILIN‐1 in CLL independently of its expression pattern. Functionally, EMILIN‐1 gC1q domain promotes adhesion of CLL cells through specific interaction with VLA‐4, and releases pro‐survival signals for CLL cells, as demonstrated by enhanced ERK and AKT phosphorylation and impairment of in‐vitro‐induced apoptosis. EMILIN‐1/VLA‐4 interaction can efficiently contribute to the maintenance of the neoplastic clone in CLL. [ABSTRACT FROM AUTHOR]

Aims CD30 expression by bone marrow ( BM) mast cells ( MC) has been reported recently in systemic mastocytosis ( SM) patients. The aim of this study was to investigate the potential diagnostic and prognostic value of CD30 expression in SM as assessed by multiparameter flow cytometry. Methods and results A total of 163 consecutive BM samples corresponding to 142 SM patients and 21 non-mastocytosis cases were studied. CD30 was positive in most SM patients (80%), but in only one non-mastocytosis case (4.8%). When combined with CD25, CD30 contributed to an improved accuracy over that of CD25 alone (98% versus 93%) mainly because most (eight of nine) of the well-differentiated SM ( WDSM), who lacked CD25, were CD30+. Similar levels of expression of CD30 were observed among all different subgroups of SM except mast cell leukaemia; among indolent SM ( ISM) patients, no significant association was observed between the levels of CD30 expression and other clinical and biological features of the disease. Conclusions The increased expression of CD30 associated with absence of CD25 contributes to the diagnosis of WDSM and its distinction from other subtypes of SM. By contrast, CD30 expression did not contribute either to prognostic stratification of ISM or to the differential diagnosis between ISM and aggressive SM cases. [ABSTRACT FROM AUTHOR]