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Human Pathology (2011) xx, xxx–xxx

Original contribution

Folliculocentric B-cell–rich follicular dendritic cellssarcoma: a hitherto unreported morphological variantmimicking lymphoproliferative disordersLuisa Lorenzi MDa, Silvia Lonardi BS a, Giulia Petrilli MDa, Francesco Tanda MDb,Michelangelo Bella MDb, Licia Laurino MD c, Rossi Giuseppe MDd,Fabio Facchetti MD, PhDa,⁎

aDepartment of Pathology I, University of Brescia, 25123 Brescia, ItalybInstitute of Pathology, University of Sassari, 07100 Sassari, ItalycDepartment of Pathology, Hospital of Treviso, 31100 Treviso, ItalydDepartment of Hematology, Spedali Civili di Brescia, 25123 Brescia, Italy

Received 28 January 2011; revised 18 February 2011; accepted 24 February 2011

0d

Keywords:Follicular dendritic cellsarcoma;

Lymph node;Immunohistochemistry

Summary We report three cases of follicular dendritic cell sarcoma (FDCS) showing a hithertoundescribed histological pattern consisting of nodular tumor growth associated with small Blymphocytes. FDCS tumor cells consistently showed large epithelioid features and were intermingledwith small lymphocytes in the nodules in two cases, whereas they formed cohesive aggregatessurrounded by lymphocyte mantle in the other. These features were easily confused withlymphomatous proliferations and, in particular, subtypes of Hodgkin lymphoma, high-grade follicularlymphoma, and germinotropic large B-cell lymphomas. The diagnosis was established by the use of abroad panel of antibodies that showed a variable expression of the FDC markers CD21, CD23, CD35,clusterin, podoplanin, claudin 4, epidermal growth factor receptor, and CXCL13. The associated Blymphocytes revealed a mantle zone B phenotype, with expression of CD20 and PAX5, together withTCL1 and IgD. Of notice, in all cases, morphological features suggesting hyaline-vascular Castlemandisease were recognized in the interfollicular areas, containing scattered epithelioid cells similar tothose found in the nodules, thus providing a useful clue for FDCS diagnosis. Of the 3 cases, 1presented multiple recurrences unresponsive to chemotherapy and radiotherapy and finally died ofdisease 14 years after diagnosis. This study further emphasizes the extreme variability ofmorphological presentation of FDCS and expands the spectrum of lesions showing a nodular growthpattern occurring in human lymph nodes.© 2011 Elsevier Inc. All rights reserved.

1. Introduction

⁎ Corresponding author.E-mail address: facchett@med.unibs.it (F. Facchetti).

046-8177/$ – see front matter © 2011 Elsevier Inc. All rights reserved.oi:10.1016/j.humpath.2011.02.029

Follicular dendritic cell sarcoma (FDCS) is a rareneoplasm showing a broad spectrum of clinical presentationsas well as of cytological and histological features. About twothirds of cases occur in lymph nodes, and the remaining cases

2 L. Lorenzi et al.

have been reported in various sites, such as tonsils, oralcavity, soft tissues, mediastinum, and abdominal organs.Because of the extreme variability of the cell compositionand patterns of growth that characterizes FDCS, the diagnosisrequires a high index of suspicion, and it is necessarily basedon immunohistochemistry, aimed at confirming the expres-sion of follicular dendritic cells (FDC) markers (such asCD21, CD23, CD35, clusterin) and to exclude thoseassociated with the main mimickers (such as epithelialneoplasms, sarcomas, and melanoma) [1-3].

In this study, we describe three cases of nodal FDCSpresenting hitherto unreported histological features, consist-ing of an intrafollicular growth of atypical epithelioid FDC,that markedly resembled lymphomatous proliferations, par-ticularly subtypes of Hodgkin lymphoma, high-grade follic-ular lymphoma, and germinotropic large B-cell lymphomas.

2. Patients, materials, and methods

The main clinical features of the three cases aresummarized in Table 1.

Formalin-fixed, paraffin-embedded tissue was availablefrom all cases and glutaraldehyde-fixed material from case 1.

On paraffin sections, immunohistochemistry for the follow-ing antigens was performed: CD21 (mouse IgG2a, clone 2G9;Thermo Scientific, Fremont, CA), CD23 (mouse IgG1, clone1B12; Thermo Scientific), CD35 (mouse IgG1, clone Ber-MAC-DRC; Dako, Glostrup, Denmark ), clusterin (mouseIgG1, clone 7D1; Novocastra Laboratories, Newcastle uponTyne, UK), podoplanin (mouse IgG1, clone D2-40; SignetLaboratories, Dedham, MA), epidermal growth factor receptor(EGFR,EGFRpharmDx IHCkit system, clone 2-18C9;Dako),claudin 4 (mouse IgG1, clone 3E2C1; Invitrogen, Carlsbad,CA), CXCL13 (goat polyclonal; R&D Systems Minneapolis,MN), CD20 (mouse IgG2a, clone L26; Dako), PAX5 (mouseIgG1, clone 24/PAX-5; BD Biosciences, San Jose, CA), TCL1(mouse IgG1, clone 27D6/20; Medical & Biological Labora-

Table 1 Clinical features of the three cases of folliculocentric B-cell

Case Age (y)/sex Presentation and clinical history

1 57/ F Asymptomatic submandibular swelling(maximum diameter 4.0 cm)

Subsequent diagnosis of noninvasivebladder urothelial carcinoma

2 14/ F Asymptomatic laterocervical and occipitallymphoadenopathy (maximum diameter 5.0 cm)

3 66/ M Retroperitoneal mass and loco regionallymphoadenopathy (maximum diameter 5.5 cm)Incidental finding during workup for metastaticmelanoma

Abbreviations: AWD, alive with disease; DOD, dead of disease.

tories, Naka-ku,Nagaya, Japan), IgD (rabbit polyclonal;Dako),CD3 (rabbit, clone SP7; Thermo Scientific), CD45 (mouseIgG1, clone RP2/18-RP2/22; Novocastra Laboratories),CD117 (rabbit polyclonal; Dako), CD1a (mouse IgG1, clone010; Dako), S100 protein (rabbit polyclonal; Dako), EBV-LMP1 (mouse IgG1, clone CS1, CS2, CS3, CS4; NovocastraLaboratories), BDCA2/CD303 (mouse IgG1, clone 124B3.13;Dendritics, Lyon, France), CD30 (mouse IgG1, clone BerH2;Dako), CD15 (mouse IgM, clone Ab3; Thermo Scientific),HMB45 (mouse IgG1, clone HMB45; Dako), EMA (mouseIgG1, clone GP1.4; Novocastra), AE1+AE3 (mouse monoclo-nal cocktail; Biogenex, San Ramon, CA). Reactivity wasrevealed using Envision Mouse/Rabbit-HRP (Dako) orbiotinylated anti-goat IgG (Vector Laboratories, Burlingame,CA), followed by diaminobenzidine.

Double immunostain for PAX5 (brown) and CD23 (blue)was performed on case 3, as previously described [4] andusing Mach4 MR-alkaline phosphatase (Biocare Medical,Concord, CA) followed by Ferangi Blue as chromogen(Biocare Medical) to visualize the second immune reaction.

Detection of Epstein-Barr virus (EBV) by in situhybridization of EBV-encoded RNA (EBV/EBER) wasperformed using the PNA ISH Detection Kit (Dako).

Electron microscopic analysis was performed using themicroscope Philips EM 301 (Philips, Eindhoven, TheNetherlands).

3. Results

Morphologically, the lymph nodes from all three patientsshowed altered architecture, with an obvious nodular growthpattern. In cases 1 and 2, the nodules were large (3-4 timeslarger than a reactive follicle) and composed of small maturelymphocytes intermingled with atypical epithelioid cells.The latter were either scant (thus resulting in “dark” nodulesdue to the marked predominance of small lymphocytes) orabundant (resulting into “pale” nodules) (Fig. 1A-D). In case

–rich FDCS

Treatment Outcome (follow-up duration)

Multiple surgicalresections, chemotherapy,and radiotherapy

Multiple local relapses involvingmuscles, soft tissues, and skin.

Unresponsive to chemotherapyand radiotherapy.DOD after 14 years

Surgical resection AWD (18 mo)

Chemotherapy No response to chemotherapy

AWD (48 mo)

Fig. 1 Histological patterns of folliculocentric FDCS, showing either macronodules with a predominant lymphocytic composition (A–B,cases 1 and 2) or small nodules surrounded by mantle lymphocytes (E, case 3). The morphological features of the atypical cells are similar in allcases and show large epithelioid cells with prominent eosinophilic nucleoli and abundant cytoplasm; note in D some binucleated cells. Tumorcells are scattered among numerous lymphocytes in cases 1 and 2 (C and D), whereas they form a cohesive aggregate in the center of the nodulein case 3 (F). A-F, hematoxylin and eosin.

3Folliculocentric B-cell-rich FDCS

3, the nodules were of the size of reactive B follicles; theywere surrounded by a rim of small mature lymphocytes andcontained in the center cohesive aggregates of atypicalepithelioid cells (Fig. 1E-F).

Cytologically, the atypical epithelioid cells were similarin all cases and showed large nuclei with prominenteosinophilic nucleoli and abundant cytoplasm; occasionalbinucleated cells were detected; mitoses were rare. Scattered

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atypical epithelioid cells were found in the internodular areasin case 1 and, to a lesser extent, in case 2.

Of notice, morphological features suggesting hyaline-vascular Castleman disease (focal hyalinosis, increase ofvessels with high endothelial features) were recognized inthe interfollicular areas of all cases; in addition, in case 2,some abnormal hypervascular B follicles containing dys-plastic FDC were found, together with large clusters ofBDCA2/CD303+ plasmacytoid dendritic cells/plasmacytoidmonocytes in the interfollicular areas, whereas in case 3,marked interfollicular plasmacytosis was obvious.

On immunohistochemistry, the atypical cells variablyexpressed follicular dendritic cell antigens (Table 2, Fig. 2).In addition, they were negative for all leukocyte antigens(CD3, CD15, CD20, CD30, CD45, CD117, CD303, andPAX5), as well as for melanocytic (S100 and HMB45) andepithelial antigens (EMA; low– and high–molecular weightcytokeratins). EBV was negative in tumor cells; in case 3,scattered EBV+ lymphocytes were recognizable on EBER.

In all cases, small lymphocytes within the nodules andsurrounding the clusters of atypical cells showed a mantlezone B-cell phenotype, with expression of CD20, PAX5,TCL1, and IgD (Fig. 3). The close contact between FDCScells and surrounding B lymphocytes was highlighted by thedouble staining for PAX5 and CD23 shown in Fig. 2D.

Remarkably, in recurrent tumors from case 1, this nodulargrowth pattern was still recognizable, although the majorityof tumor cells formed large and confluent aggregates, withevident increase of cytological atypia and number of mitoses.Furthermore, in some tumor recurrences (nodal andcutaneous), tumor cells were intermingled with largenumbers of CD1a+ Langerin+ S100+ Langerhans cells andeosinophils (Fig. 2F).

Electron microscopy performed in case 1 showed thepresence in atypical cells of numerous interdigitatingcytoplasmic processes with intercellular desmosome-likejunctions (Fig. 2E), further confirming the folliculardendritic nature.

Table 2 Immunophenotype of tumor cells in the three cases offolliculocentric B-cell–rich FDCS

Antigen Case

1 2 3

CD21 ++ + +++CD23 − +++ −CD35 ++ ++ −Clusterin +++ +++ ++ wPodoplanin +++ ++ ++ wEGFR + a +++ +Claudin 4 ++ ++ ++ wCXCL13 ++ a ++ w ++ w

FDCS-positive cells are indicated as percentage of the tumor population:− b1%; + 1%-25%; ++ 25%-50%, and +++ N50%; w, weak staining.

a Evaluated on recurrent tumors.

4. Discussion

We report three cases of FDCS showing a hithertoundescribed histological pattern, consisting of tumorgrowth within B follicles resulting in a nodular architec-ture. Tumor cells were consistently large and withepithelioid features; in two cases, they were intermingledwith small mature lymphocytes within the nodules,whereas in the other, they formed cohesive aggregatessurrounded by mantle lymphocytes.

These unusual features add to the great morphologicalvariability of FDCS, a neoplasm characterized by severalgrowth patterns, with fascicles, coarse, or delicate stori-form arrays, trabeculae, and whorls [1,5]. Cytologicalheterogeneity is also remarkable, and tumor cells aregenerally spindle but can be oval, polygonal, or epitheli-oid, and different cell types can be present in the sametumor [1,2]. Consequently, the diagnosis of FDCS ischallenging, and pathologists often must exclude sarcomas,carcinomas, and other neoplasms [2], and extensiveimmunohistochemical analysis is therefore mandatory forFDCS diagnosis.

In the present three cases, the nodular growth patterntogether with the association of tumor cells with smallmantle B-cell lymphocytes first prompted consideration ofthe diagnosis of a lymphoid neoplasm. In particular, cases1 and 2 evoked nodular lymphocyte-predominant Hodgkinlymphoma or lymphocyte-rich classical Hodgkin lympho-ma, whereas case 3 strongly suggested the diagnosis ofuncommon large cell lymphomas with intrafolliculargrowth, such as high-grade follicular lymphoma and the“germinotropic” variants of large B-cell lymphoma asso-ciated or not with the expression of human herpesvirus8 and EBV antigens by tumor cells [6-8].

On morphology, the likelihood that atypical cells wererepresented by FDC could be inferred by their cytologicalfeatures and especially by the large nuclei with a delicatenuclear membrane, the distinct eosinophilic nucleoli, and theabundant eosinophilic cytoplasm [1].

Of note, in all cases, morphological features of hyaline-vascular Castleman disease were recognized in the inter-follicular areas, where, in addition, scattered epithelioid cellssimilar to those found in the nodules were found in cases 1and 2, thus providing a useful clue for FDCS diagnosis. It isknown that 10% to 20% of cases of FDCS are associatedwith hyaline-vascular Castleman disease [9,10], a lesioncharacterized by clonal proliferation of FDC [11-13], whichshows dysplastic features [14] and phenotypical changes[3,15] comparable to those observed in FDCS, thussupporting the hypothesis that it represents a precursor ofthis neoplasm [2,9,16].

In the three cases reported here, tumor cells showedvariable expression of FDC antigens, with the notablenegativity of CD23 in two cases (1 and 3); this reinforces therequirement of using a broad panel of markers [1-3] forFDCS diagnosis, especially taking into consideration that

Fig. 2 Immunohistochemical reactivity of tumor cells in FDCS, with strong expression of clusterin (A, case 1), podoplanin (B, case 2), andCD21 (C, case 3). In D, a double stain for CD23 (blue) and Pax5 (brown) shows CD23+ tumor cells surrounded by numerous Pax5+ Blymphocytes (case 2). Electron microscopy (E, case 1) illustrates the elongated cytoplasmic processes of FDC, intermingled with smalllymphocytes. Note a desmosome between two adjacent FDC (inset). (F) Recurrent nodal lesion in case 1: numerous Langerhans cells, stainedby anti-CD1a (inset and left) are associated with FDCS cells. Immunohistochemistry for clusterin (A), podoplanin (B), CD21 (C), and CD1a(F, inset and left), hematoxylin counterstain; double immunohistochemistry for CD23 and Pax5 (D); hematoxylin and eosin (F, right).

5Folliculocentric B-cell-rich FDCS

none of them is absolutely specific. Furthermore, this studyconfirms the usefulness in FDCS diagnosis of CXCL13, andclaudin 4, two recently reported antigens expressed by bothnormal and neoplastic FDC [3,17].

This variant of B-cell–rich FDCS differs from the“inflammatory pseudotumor-like” FDCS described byCheuk et al [18], in which patients typically present systemicsymptoms, their tumors occurring in abdominal sites and

Fig. 3 The B-cell rich composition of the FDCS nodules, as shown by anti-CD20 immunostain (A, B, case 1; E, case 3); these Blymphocytes express a mantle zone phenotype, with positivity for IgD (C, case 2) and TCL1 (D, case 2; F, case 3). Immunohistochemistry forCD20 (A, B and E), IgD (C), and TCL1 (D and F); hematoxylin counterstain.

6 L. Lorenzi et al.

showing a diffuse rather than a nodular growth pattern, withtumor cells infected by EBV and dispersed amongheterogeneous inflammatory cell population.

Despite the occurrence of neoplastic FDC within Bfollicles seeming to indicate an early stage of tumordevelopment or even in situ FDCS, the clinical evolution

of the single case (1) with a long follow-up was dismal,characterized by several local recurrences unresponsive toboth chemotherapy and radiotherapy and eventual death 14years after the diagnosis. FDCS was traditionally consid-ered an indolent, low-grade neoplasm [19], but recentstudies that included patients with a long follow-up showed

7Folliculocentric B-cell-rich FDCS

that FDCS has features of an intermediate malignancytumor, associated with a substantial risk of recurrences andmetastasis [2,20,21].

An intriguing feature observed in case 1 was theprominent infiltration of Langerhans cells found in recurrenttumors and resembling a process described in varioushematological and solid neoplasms [22]. The biologicalsignificance of this phenomenon is still controversial, and itis argued that it may represent an exaggerated reactiveprocess rather than a second neoplasm [23].

In conclusion, in this study, we describe three cases ofnodal FDCS presenting with hitherto unreported histolog-ical features that further emphasize the extreme variabilityof presentation of FDCS and expands the spectrum oflesions showing a nodular growth pattern occurring inhuman lymph nodes.

Acknowledgments

The authors thank Cristina Rossini, Roberto Paracchini,and Franco Alpi for their valuable technical support. Thisstudy was partially supported by Fondazione Beretta andFondazione Berlucchi (to F.F.).

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