Ebook Atlas of non-gynecologic cytology: Part 2

Continued part 1, part 2 of ebook Atlas of non-gynecologic cytology provide readers with content about: pancreaticobiliary tract cytology; liver cytology; kidney and adrenal gland cytology; urine cytology; serous effusion cytology; lymph node cytology;... Please refer to the ebook for details!

© Springer International Publishing AG, part of Springer Nature 2018

X Jing et al (eds.), Atlas of Non-Gynecologic Cytology, Atlas of Anatomic Pathology,

https://doi.org/10.1007/978-3-319-89674-8_6

Pancreaticobiliary Tract Cytology

Judy Pang and Andrew Sciallis

Introduction

The major indications for cytologic evaluation of the

pancre-aticobiliary tract are a pancreatic mass and/or a bile duct

stricture Endoscopic ultrasound-guided fine-needle

aspira-tion (EUS-FNA) is the primary modality for obtaining tissue

diagnosis; it has largely replaced percutaneous FNA because

it allows for real-time visualization of the needle, provides

better visualization of small lesions than CT guidance, and

enables the sampling of regional lymph nodes and

assess-ment of invasion of local structures, thus providing

simulta-neous diagnosis and staging [1 3] The sensitivity of

EUS-FNA for solid masses has been reported to range from

54 to 95%, with specificity approaching 100% [4] The role

of EUS-FNA is less clear for cystic lesions, as the sensitivity

is generally lower and more variable in detecting a neoplastic

mucinous cyst than a solid neoplasm, ranging from 23 to

100% [5] Furthermore, the sensitivity of detecting a

malig-nancy in a neoplastic mucinous cyst is reported to be 29%,

with 100% specificity [6 7]

Endoscopic retrograde cholangiopancreatography

(ERCP) with bile duct brushings for cytology is an

addi-tional minimally invasive modality to obtain material for

tis-sue diagnosis, which can be helpful in the assessment of

pancreatic neoplasms, particularly ductal adenocarcinomas

The sensitivity of brushings is reported to be lower than that

of EUS-FNA, 44–72% [8 11], but the specificity approaches

100%, similar to EUS-FNA

The Papanicolaou Society of Cytopathology has

pro-posed a terminology scheme for the reporting of

pancreati-cobiliary cytology, utilizing a six-tiered system, as shown

on Table 6.1 [12]

Sampling of pancreatic head masses is performed using a transduodenal approach, whereas a transgastric approach is used for the body and tail masses It is important for the pathologist to be aware of the approach so that contaminat-ing normal duodenal mucosa (Fig. 6.1) and gastric mucosa (Fig. 6.2) is not misinterpreted as lesional tissue

J Pang, M.D ( * ) · A Sciallis, M.D

Department of Pathology, The University of Michigan,

Ann Arbor, MI, USA

e-mail: jcpang@med.umich.edu ; sciallis@med.umich.edu

• Others – Well-differentiated neuroendocrine tumor – Solid pseudopapillary tumor

– Intraductal papillary mucinous neoplasm, all grades of

dysplasia – Mucinous cystic neoplasm, all grades of dysplasia

V Suspicious (for malignancy)

VI Positive or malignant

• Pancreatic ductal adenocarcinoma

• Cholangiocarcinoma

• Acinar cell carcinoma

• Poorly differentiated (small-cell and large-cell) neuroendocrine carcinoma

a b

Fig 6.1 Duodenal epithelium is often seen in aspirates of masses from the pancreatic head or proximal body Typically seen are flat sheets of

epithelial cells with uniform small nuclei studded with occasional goblet cells (a) Diff-Quik stain; (b) Papanicolaou stain

Fig 6.2 Gastric epithelium is often seen in aspirates of masses from

the pancreatic distal body or tail It typically appears as flat sheets of

mucinous epithelial cells with uniform small nuclei In this image,

adja-cent to the gastric epithelium is a small cluster of disordered malignant

cells (Papanicolaou stain)

Fig 6.3 Benign pancreatic ductal epithelial cells have uniform small

round nuclei and are arranged in cohesive, evenly spaced honeycomb sheets (Diff-Quik stain)

Fig 6.4 Benign acinar cells are reminiscent of “grapelike” clusters

when associated with fibrovascular tissue (Papanicolaou stain)

Normal Pancreas

Figures 6.3, 6.4, and 6.5 show examples of normal

pancre-atic elements

Solid Pancreatic Masses

EUS-FNA of solid pancreatic masses is not always

neces-sary when a solid mass detected on imaging is considered

to be resectable, as a benign cytology does not entirely

exclude a malignancy It is most useful in patients who have

unresectable disease or are poor surgical candidates, in

whom tissue diagnosis is necessary prior to the initiation of

chemotherapy or radiation [13] It is also helpful when it is

not clear from clinical and radiologic findings whether a

mass is attributable to a benign process such as pancreatitis

(Figs. 6.7, 6.8, and 6.9), when the patient has a prior history

of another malignancy, or when a lymphoma is suspected

In these scenarios, surgical resection may not be indicated

Pancreatitis

Figures 6.6 and 6.7 show features of pancreatitis

Pancreatic Ductal Adenocarcinoma

Pancreatic ductal adenocarcinoma can be identified from a

number of characteristics of its cytomorphology, as

illus-trated in Figs. 6.8, 6.9, 6.10, 6.11, 6.12, and 6.13:

• Disordered, crowded epithelial sheets (“drunken

honeycomb”)

• Single malignant cells

• Irregular nuclear contours (grooves, convolutions)

• Irregular chromatin (clearing and clumping)

• Nuclear enlargement

• Prominent nucleoli

• Nuclear pleomorphism (4:1 nuclear diameter size

differ-ence within a single cluster/sheet)

• Prominent mucinous vacuolization

Fig 6.5 Benign acinar cells are polygonal with abundant granular

cytoplasm (Papanicolaou stain)

a

b

Fig 6.6 Stromal fragments in chronic pancreatitis (a) Diff-Quik

stain; (b) Papanicolaou stain

Fig 6.7 Pancreatitis Cohesive cluster of slightly crowded ductal cells

with slightly enlarged round to oval nuclei, smooth nuclear membranes, and small nucleoli There is little variation in nuclear diameter within the same sheet (<4:1 difference) There is also a background of neutro- phils (Papanicolaou stain)

Fig 6.8 Crowded clusters of epithelial cells and loss of honeycomb

arrangement in pancreatic ductal adenocarcinoma (Diff-Quik stain)

Fig 6.9 Single malignant cells with enlarged nuclei and prominent

nucleoli in pancreatic ductal adenocarcinoma (Diff-Quik stain)

Fig 6.10 Loosely cohesive clusters of ductal adenocarcinoma with

nuclear enlargement, admixed with benign acinar cells (Diff-Quik stain)

a

b

c

Fig 6.11 Prominent mucinous vacuolization resulting in low nuclear

to cytoplasmic (N:C) ratio, is commonly encountered in noma Irregular nuclear contours, nuclear enlargement, and 4:1 nuclear

adenocarci-diameter size difference can also be appreciated in these images (a) Diff-Quik stain; (b) Papanicolaou stain; (c) Cell block section with

hematoxylin and eosin (H&E) stain

Pancreatic Neuroendocrine Tumor

Pancreatic neuroendocrine tumors (PanNETs) can be fied by several cytomorphologic features:

identi-• Highly cellular aspirate

• Single cells often with bare nuclei

• Pseudorosettes and small clusters

• Uniform, round or oval, eccentrically placed nuclei (plasmacytoid)

• Moderate to abundant cytoplasm

• Salt-and-pepper chromatin and distinct nucleoliThese tumors are histologically separated into well- differentiated (low-grade and intermediate-grade) tumors (Figs. 6.14, 6.15, 6.16, 6.17, and 6.18) and poorly differentiated (high-grade) neuroendocrine carcinomas (Figs. 6.19 and 6.20)

Fig 6.12 Mitotic figures, although not diagnostic of malignancy, are

more frequently seen in adenocarcinoma and would rarely be seen in

benign processes such as pancreatitis In this cluster of malignant cells,

other features of adenocarcinoma are seen, including nuclear

enlarge-ment, irregular nuclear contours, and 4:1 nuclear diameter size

differ-ence (Diff-Quik stain)

a

b

Fig 6.13 (a and b) Nuclear clearing and irregular nuclear contours

and grooves are often seen in adenocarcinomas Prominent mucinous

vacuolization and 4:1 nuclear diameter size difference are also seen in

these images (Papanicolaou stain)

Fig 6.14 Highly cellular aspirate with loose clusters in a low-grade

pancreatic neuroendocrine tumor (PanNET) (Diff-Quik stain)

Fig 6.15 Pseudorosettes and small clusters in a low-grade PanNET

(Diff-Quik stain)

Fig 6.16 Aspirates of low-grade PanNETs often consist of

predomi-nantly dyscohesive cells with eccentrically placed nuclei (Diff-Quik stain)

Fig 6.17 Salt-and-pepper chromatin and distinct nucleoli in low-

grade pancreatic neuroendocrine tumors are better visualized with

Papanicolaou stain

Fig 6.18 Low-grade PanNETs can appear very bland on ThinPrep

On closer inspection, the cells have eccentrically placed nuclei and salt-

and- pepper chromatin

a

b

Fig 6.19 (a and b) High-grade PanNET with nuclear molding and

scant cytoplasm, similar in cytomorphology to small-cell carcinoma (Diff- Quik stain)

Fig 6.20 High-grade PanNET with nuclear molding and scant

cyto-plasm (Papanicolaou stain)

Immunostains are utilized to confirm the diagnosis, as

acinar cell carcinoma (ACC) and solid pseudopapillary

tumors (SPT) can have similar cytomorphology, as discussed

below PanNETs are typically positive for neuroendocrine

markers such as synaptophysin (Fig. 6.21) and chromogranin

(Fig. 6.22) A panel of immunostains is recommended, as

both ACC and SPT can be positive for neuroendocrine

mark-ers (Table 6.2) [14–16]

The proliferative rate is used to grade PanNETs because

it provides prognostic information that may influence

clini-cal management Staining for Ki-67 has been found to be

useful in FNA specimens in this regard [17] The Ki-67

index is less than 3% for low-grade tumors, 3–20% for

intermediate- grade tumors, and greater than 20% for

high-grade tumors [18] Caution should be noted, as a

higher-grade focus may not have been sampled in FNA

specimens

Acinar Cell Carcinoma

Several cytomorphologic features will point to acinar cell carcinoma (ACC):

• Highly cellular aspirate

• Single cells, loose aggregates, naked nuclei

• Prominent nucleoli

• Granular cytoplasmThe typical “grapelike cluster” arrangement of benign acinar cells is usually absent Figures 6.23 and 6.24 illustrate the appearance of aspirates from these tumors

Solid Pseudopapillary Tumor

Solid pseudopapillary tumor (SPT) is also recognized by its characteristic cytomorphologic features:

• Highly cellular aspirate

• Vascular stalks lined by neoplastic cells

Fig 6.21 PanNETs stain positive for synaptophysin

Table 6.2 Immunohistochemical profiles of pancreatic

neuroendo-crine tumor (PanNET), acinar cell carcinoma (ACC), and solid papillary tumor (SPT)

Fig 6.22 PanNETs stain positive for chromogranin

Fig 6.23 Aspirates of acinar cell carcinoma (ACC) are typically

highly cellular, consisting of numerous isolated cells, loose aggregates, and naked nuclei (Diff-Quik stain)

• Round to oval or bean-shaped nuclei

a

b

c

Fig 6.24 The nuclei of ACC are round to oval, with smooth nuclear

contours Delicate, granular cytoplasm is also seen In contrast to

nor-mal, benign acinar cells, prominent nucleoli are usually encountered (a

and b) Diff-Quik stain; (c) Papanicolaou stain

Fig 6.25 Vascular stalks lined by neoplastic cells is a helpful feature

in the diagnosis of solid pseudopapillary tumor (SPT) (Diff-Quik stain)

Fig 6.26 Vascular stalks lined by neoplastic cells in SPT (Papanicolaou

stain)

Fig 6.27 There is frequently a predominance of dyscohesive single

cells with round to oval, sometimes bean-shaped nuclei that are

eccen-trically placed As such, low-grade PanNETs and ACC are considered

in the differential diagnosis (Diff-Quik stain)

Fig 6.28 Hyaline globules can sometimes be seen in SPTs and can be

a helpful diagnostic clue (Diff-Quik stain)

Fig 6.29 Cell block sections often demonstrate the vascular stalks

lined by neoplastic cells (H&E stain)

Fig 6.30 Nuclear staining for beta-catenin is the key diagnostic

marker of SPTs

Fig 6.31 Nuclear staining for beta-catenin in the neoplastic cells with

adjacent benign glandular epithelium showing only membranous ing without nuclear staining is useful as an internal negative control

stain-Fig 6.32 A negative E-cadherin immunostain is helpful in

differenti-ating SPT from PanNET, which is positive for E-cadherin This image shows negative staining in the neoplastic cells, with positive staining in the benign glandular epithelium as an internal positive control

Fig 6.33 Aspirates of pseudocysts are paucicellular, consisting of

mixed inflammatory cells and histiocytes Yellow, hematoidin-like ment can sometimes be seen

Pancreatic Cysts

Cystic lesions in the pancreas are being identified with

increas-ing frequency because of increased utilization of imagincreas-ing

stud-ies to evaluate abdominal complaints As such, many of these

are incidental findings A CT scan is often the initial test by

which a cystic lesion is detected, and certain features can be

suggestive of a particular diagnosis [19] The finding of a

soli-tary, septated cystic lesion in the tail of the pancreas in a woman

is highly suggestive of mucinous cystic neoplasm (MCN)

Grapelike clusters of involved side branch ducts are the most

common finding in an intraductal papillary mucinous

neo-plasm (IPMN) Septa and excrescent nodules along the dilated

main pancreatic duct can be seen in main duct IPMN EUS has

been reported to be the ideal tool for the evaluation of cystic

lesions [19] In addition to enabling the performance of FNA,

EUS images can suggest a diagnosis EUS of pseudocysts may

show a thick wall with floating debris EUS of serous

cystade-nomas typically demonstrates a multiloculated, microcystic

lesion with little free fluid in the locules MCNs are often

uni-locular and contain highly viscous, clear fluid that may be

dif-ficult to aspirate The appearance of IPMN on EUS ranges

from simple, unilocular lesions to complex cystic masses

Classifying pancreatic cysts as non-mucinous (i.e.,

pseu-docyst, serous cystadenoma) versus mucinous can be

chal-lenging on cytology [5 13] Differentiating neoplastic mucin

from contaminating mucin from the gastrointestinal tract

with absolute certainty is difficult on cytology preparations,

as is differentiating normal gastrointestinal and pancreatic

epithelium from the epithelial lining of mucinous cysts with

minimal atypia [5 13] Cyst fluid CEA level is the most

accurate test for the diagnosis of a neoplastic mucinous cyst,

with a diagnostic accuracy of 79% when levels are greater

than 192 ng/mL [5 20] However, CEA levels do not predict

the presence or absence of malignant transformation in

neo-plastic mucinous cysts [20]

Pseudocyst

Aspirates of pseudocysts are paucicellular, consisting of

mixed inflammatory cells and histiocytes Yellow,

hematoidin- like pigment can sometimes be seen (Fig. 6.33)

Other than gastrointestinal contamination, no extracellular

mucin or epithelial cells are present

Serous Cystadenoma

Aspirates of serous cystadenoma are sparse in cellularity and

consist of small, bland-appearing cuboidal cells in flat sheets

and loose clusters (Fig. 6.34) These cuboidal cells are rich in

glycogen (Figs. 6.35 and 6.36)

a

b

Fig 6.34 (a and b) Aspirates of serous cystadenoma are sparse in

cel-lularity and consist of small, bland-appearing cuboidal cells in flat sheets and loose clusters, as seen in these images from a ThinPrep preparation (Papanicolaou stain)

Neoplastic Mucinous Cyst

Cytology cannot differentiate between mucinous cystic

neo-plasm (MCN) and intraductal papillary mucinous neoneo-plasm

(IPMN), nor can it reliably differentiate high-grade dysplasia

from invasive adenocarcinoma (Figs. 6.37, 6.38, 6.39, and

6.40) Features of malignancy should be identified using the

same criteria as for ductal adenocarcinomas, however, as

clin-ical management would be affected (Fig. 6.41) The

manage-ment of neoplastic mucinous cysts is multifactorial and

includes assessment of the patient’s surgical risk, symptoms,

and the likelihood of a benign diagnosis on cytology Findings

on imaging, including size, presence of mural nodule, and

dilatation of the main pancreatic duct, are also considered

When the clinical and imaging findings do not show clear

indications for surgery, an abnormal cytology could result in

Fig 6.35 The cuboidal cells of serous cystadenomas are glycogen-

rich, as highlighted with a PAS stain

Fig 6.36 Predigestion with diastase abolishes the glycogen in the

cuboidal cells of a serous cystadenoma

a

b

Fig 6.37 Thick, colloid-like extracellular mucin is seen in aspirations

of neoplastic mucinous cysts (a) Diff-Quik stain; (b) Papanicolaou

stain

Fig 6.38 The neoplastic mucinous epithelium (right) demonstrates

disordered nuclei and apical mucin in contrast to normal intestinal

epi-thelium (left) (Papanicolaou stain)

b

Fig 6.39 (a and b) Although the nuclei of neoplastic mucinous cysts

are disordered, enlarged, and have irregular nuclear contours,

signifi-cant nuclear pleomorphism (4:1 nuclear diameter size difference)

should be absent in mucinous cysts with low or moderate dysplasia

(Papanicolaou stain)

a

b

c

Fig 6.40 (a–c) Fibrovascular cores and papillary-like projections can

sometimes be seen in aspirates of neoplastic mucinous cysts, suggestive

of an intraductal papillary mucinous neoplasm However, this is not absolute, and correlation with imaging to determine communication with or without the pancreatic duct is necessary (Diff-Quik stain)

a b

Fig 6.41 (a and b) Malignant features in this neoplastic mucinous cyst including mitotic figures and a 4:1 nuclear diameter size difference are

easily identified in these images (Papanicolaou stain)

Fig 6.42 ThinPrep preparation of a bile duct brushing with both

benign ductal epithelium (left) and adenocarcinoma (right) Nuclear

enlargement and membrane irregularity, prominent nucleoli, and 4:1

nuclear diameter size difference are evident in the cluster of malignant cells, in contrast to the flat honeycomb sheet consisting of uniform, small nuclei of benign ductal epithelium (Papanicolaou stain)

triaging the patient to surgery In the international consensus

guidelines for the management of branch-duct IPMN, a

cytol-ogy interpretation of suspicious or positive for malignancy

would result in the patient being triaged to surgery [21]

Bile Duct Brushings

The same criteria used in the diagnosis of pancreatic ductal

adenocarcinoma in EUS-FNA are used to evaluate for

malig-nant cells in a bile duct brushing (Fig. 6.42) Given the

limi-tations of cytology, indeterminate diagnoses such as

“atypical” or “suspicious” are not infrequent, especially in

patients with primary sclerosing cholangitis (PSC), where reactive changes can mimic carcinoma As such, adjunct fluorescence in situ hybridization (FISH) testing using the UroVysion™ probe set has been advocated to improve the sensitivity of detecting malignancies, compared with routine cytology alone [22] The sensitivity of positive FISH testing ranges from 34 to 53% in the literature, compared with 8–38% for positive routine cytology in head-to-head com-parisons [23–30] The specificity of FISH is slightly lower than that of routine cytology, but it remains high (from 89 to 100%) [23–30] These probes are directed to chromosome 3 (CEP3), chromosome 7 (CEP7), chromosome 17 (CEP17), and the 9p21 locus The most common FISH abnormalities

are polysomy, trisomy, and tetrasomy A cell with polysomy

(more than two copies) of multiple probes is defined as

“polysomy,” but if each probe displays exactly four copies,

the cell is defined as “tetrasomy.” A cell with three copies of

a single probe is defined as “trisomy.” The cutoff for

consid-ering a case positive for these abnormalities is at least five

cells with polysomy, ten or more cells with trisomy, and ten

or more cells with tetrasomy Once the threshold for

poly-somy is reached, a positive interpretation is rendered If

thresholds for polysomy and an additional abnormality, such

as trisomy, are met, the case is classified as polysomy, as this

abnormality is very specific for malignancy Polysomy is

interpreted as a positive FISH result, whereas trisomy and

tetrasomy are considered equivocal results Table 6.3

sum-marizes the FISH interpretation of pancreaticobiliary

brush-ings [22]

References

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2 Erickson RA, Garza AA.  Impact of endoscopic ultrasound on

the management and outcome of pancreatic carcinoma Am J

Gastroenterol 2000;95:2248–54.

3 Volmar KE, Vollmer RT, Routbort MJ, Creager AJ. Pancreatic and

bile duct brushing cytology in 1000 cases: review of findings and

comparison of preparation methods Cancer 2006;108:231–8.

4 Hewitt MJ, McPhail MJ, Possamai L, Dhar A, Vlavianos P,

Monahan KJ.  EUS-guided FNA for diagnosis of solid

pancre-atic neoplasms: a meta-analysis Gastrointest Endosc 2012;

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5 Thosani N, Thosani S, Qiao W, Fleming JB, Bhutani MS, Guha

S. Role of EUS-FNA-based cytology in the diagnosis of mucinous

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Dig Dis Sci 2010;55:2756–66.

6 Pitman MB, Genevay M, Yaeger K, Chebib I, Turner BG,

Mino- Kenudson M, Brugge WR.  High-grade atypical epithelial

cells in pancreatic mucinous cysts are a more accurate

predic-tor of malignancy than “positive” cytology Cancer Cytopathol

2010;118:434–40.

7 Pitman MB, Lewandrowski K, Shen J, Sahani D, Brugge W,

Fernandez-del Castillo C. Pancreatic cysts: preoperative diagnosis

and clinical management Cancer Cytopathol 2010;118:1–13.

8 Layfield LJ, Wax TD, Lee JG, Cotton PB. Accuracy and

morpho-logic aspects of pancreatic and biliary duct brushings Acta Cytol

1995;39:11–8.

9 McGuire DE, Venu RP, Brown RD, Etzhorn KP, Glaws WR, Abu- Hammour A. Brush cytology for pancreatic carcinoma: an analysis

of factors influencing results Gastrointest Endosc 1996;44:300–4.

10 Ryan ME.  Cytologic brushings of ductal lesions during ERCP. Gastrointest Endosc 1991;37:139–42.

11 Jing X, Wamsteker EJ, Li H, Pu RT.  Combining fine needle aspiration with brushing cytology has improved yields in diag- nosing pancreatic ductal adenocarcinoma Diagn Cytopathol 2009;37(8):574.

12 Pitman MB.  Overview of diagnostic terminology and reporting In: Pitman MB, Layfield LJ, editors The Papanicolaou Society of Cytopathology system for reporting pancreaticobiliary cytology Cham: Springer; 2015 p. 1–3.

13 Pang JC, Minter RM, Kwon RS, Simeone DM, Roh MH.  The role of cytology in the preoperative assessment and management

of patients with pancreaticobiliary tract neoplasms J Gastrointest Surg 2013;17:501–10.

14 Pitman MB. Pancreas and biliary tree In: Cibas ES, Ducatman BS, editors Cytology: diagnostic principles and clinical correlates 4th

ed Philadelphia: Elsevier Saunders; 2014 p. 399–422.

15 Ohara Y, Oda T, Hashimoto S, Akashi Y, Miyamoto R, Enomoto T,

et al Pancreatic neuroendocrine tumor and solid-pseudopapillary neoplasm: key immunohistochemical profiles for differential diag- nosis World J Gastroenterol 2016;22:8596–604.

16 Tajima S, Waki M, Azuma M, Koda K, Ohata A.  E-cadherin- negative acinar cell carcinoma of the pancreas: report of a case showing a solid pseudopapillary growth pattern Med Mol Morphol 2016;49:177–81.

17 Farrell JM, Pang JC, Kim GE, Tabatabai ZL. Pancreatic docrine tumors: accurate grading with Ki-67 index on fine-needle aspiration specimens using the WHO 2010/ENETS criteria Cancer Cytopathol 2014;122:770–8.

18 Klimstra DS, Modlin IR, Coppola D, Lloyd RV, Suster S.  The pathologic classification of neuroendocrine tumors: a review of nomenclature, grading, and staging systems Pancreas 2010;39: 707–12.

19 Brugge WR. The use of EUS to diagnose cystic neoplasms of the pancreas Gastrointest Endosc 2009;69(2 Suppl):S203–9.

20 Petrone MC, Arcidiacono PG. Role of endosocopic ultrasound in the diagnosis of cystic tumours of the pancreas Dig Liver Dis 2008;40:847–53.

21 Tanaka M.  International consensus on the management of ductal papillary mucinous neoplasm of the pancreas Ann Transl Med 2015;3:286.

22 Fritcher EGB, Kipp BR, Halling KC, Clayton AC. FISHing for creatobiliary tract malignancy in endoscopic brushings enhances the sensitivity of routine cytology Cytopathology 2014;25:288–301.

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RF, et al A multivariable model using advanced cytologic

meth-Table 6.3 UroVysion™ fluorescence in situ hybridization (FISH) interpretation for pancreaticobiliary brushing specimens

FISH cell

type Definition

Cutoff value

Test interpretation Clinical significance Disomy 2 copies of each probe NA Negative Expected signal pattern in benign epithelium

Trisomy 3 copies of a single probe

ods for the evaluation of indeterminate pancreatobiliary strictures

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© Springer International Publishing AG, part of Springer Nature 2018

X Jing et al (eds.), Atlas of Non-Gynecologic Cytology, Atlas of Anatomic Pathology,

https://doi.org/10.1007/978-3-319-89674-8_7

Liver Cytology

Derek B. Allison, David Borzik, and Qing Kay Li

Introduction

Advanced imaging techniques can detect many benign and

malignant liver lesions, including simple liver cysts, focal

nodular hyperplasia, hepatic adenoma, hepatic hemangioma,

hepatocellular carcinoma, cholangiocarcinoma, and the

majority of metastatic carcinomas [1 5] Despite these

increasingly precise techniques, it is still critical to obtain

lesional tissue for morphological diagnosis and molecular

characterization of lesions that require medical or surgical

management [1, 6 7] Furthermore, small lesions less than

1 cm in size may not be well characterized by a radiological

image study [8 10] Currently, percutaneous fine needle

aspi-ration (FNA) and needle core biopsy are the most commonly

used procedures for the morphological evaluation of liver

lesions These procedures are usually performed under

ultra-sound guidance or CT guidance in an outpatient setting

Studies have shown that these procedures have a high

diag-nostic accuracy, with minimal risks for patients The

sensitiv-ity of these procedures for diagnosing malignant tumors has

been reported to be 90% (range, 67–100%), with 100%

speci-ficity [7 10, 11] Percutaneous FNA biopsy has been reported

to have a 100% positive predictive value for liver malignancy,

59.1% negative predictive value, and 92.4% overall accuracy

[7 10, 11] The main challenge of diagnosing liver masses is

to differentiate a primary from a metastatic tumor [12–14]

Several factors may affect the diagnostic sensitivity and

accu-racy, such as the operator’s skill and experience, the size and location of the lesion, the quality of cytological smears, and the cytopathologist’s expertise [7 15–18]

Endoscopic ultrasound-guided FNA (EUS-FNA) is the latest diagnostic and staging tool, with a sensitivity

of 82–94% and a specificity of 90–100% [2 4 5 10, 11] This procedure is safe and accurate but highly operator- dependent EUS-FNA can access the left lobe of the liver, hilum, proximal right lobe, gallbladder, extrahe-patic biliary system, and perihilar lymph nodes It is especially useful for small and deep-seated left lobe lesions, which cannot be easily accessed by percutane-ous FNA [2 4 5 10, 11]

During FNA and/or biopsy procedures, several types of cytological specimens can be prepared, including direct smears, cell block preparations, and core biopsies Direct smears play a critical role in the on-site assessment of lesions Unlike core biopsy and cell block preparations, direct smear preparations do not require formalin fixation and can be per-formed during rapid on-site evaluation for specimen ade-quacy [7 16, 17] Core needle biopsy and cell block preparations are still the preferred techniques for obtaining tumor samples, particularly in patients whose lesion requires ancillary studies or molecular characterization

This chapter summarizes the key findings for the accurate morphological diagnosis of benign, malignant, and meta-static lesions of the liver

Department of Pathology and Oncology, The Johns Hopkins

Medical Institutions, Baltimore, MD, USA

e-mail: qli23@jhmi.edu

7

Benign Bile Duct Epithelium, Normal Liver

Cells, and Benign Lesions

The cytological features of benign bile duct epithelial cells,

benign liver cells, and the most commonly seen benign liver

lesions are summarized in Table 7.1

Table 7.1 Summary of main cytological features of benign conditions of the liver

Conditions Main findings Key features of differential diagnosis

Benign liver

cells

Sheets, trabeculae or tissue fragments, and/or dispersed individual cells Centrally located round to oval-shaped nuclei, small nucleoli, granular chromatin, and intranuclear pseudoinclusions Binucleation Dense cytoplasm with bile and lipofuscin pigments

Polygonal cells with centrally located nuclei and prominent small nucleoli and dense cytoplasm Smooth nuclear membrane Normal N:C ratio

Differential diagnosis: hepatic adenoma, focal nodular hyperplasia, well-differentiated hepatocellular carcinoma, melanoma, and metastatic carcinomas

Bile duct

epithelium (bile

ductules)

Small clusters or sheets of cuboidal cells

Epithelial appearance Ovoid nuclei with granular chromatin, inconspicuous nucleoli, scant cytoplasm Nuclear disarray may be present

Cells are smaller than hepatocytes Ovoid, darkly stained, overlapping nuclei Scant cytoplasm Normal N:C ratio

Differential diagnosis: cholangiocarcinoma, metastatic adenocarcinomas

Kupffer cells Intermediate-sized cells Elongated nuclei,

vacuolated cytoplasm, and cytoplasmic hemosiderin pigment

Resemble macrophages

Differential diagnosis: endothelial cells, cholangiocarcinoma, metastatic adenocarcinoma

Histiocytes Loosely formed two-dimensional clusters or

dispersed individual cells, with coffee shaped nuclei, fine chromatin, inconspicuous nucleoli, and foamy cytoplasm

bean-No nuclear atypia Abundant cytoplasm with vacuoles and pigment Normal N:C ratio

Differential diagnosis: cholangiocarcinoma, hepatocellular carcinoma Focal nodular

hyperplasia

Benign-appearing hepatocytes No nuclear atypia Presence of benign bile duct epithelial cells

No nuclear atypia Normal N:C ratio Benign bile duct epithelial cells

Differential diagnosis: hepatic adenoma, regenerating nodule in cirrhosis Hepatic

adenoma

Benign-appearing hepatocytes Absence of bile duct epithelial cells

Variable cellularity on slides No nuclear atypia Normal N:C ratio

Differential diagnosis: well-differentiated hepatocellular carcinoma Bile duct

hamartoma

Benign ductal epithelium Benign- appearing hepatocytes Scattered stromal cells

Scant specimen, no malignant features

Differential diagnosis: cholangiocarcinoma, metastatic adenocarcinoma Hemangiomas Scant tissue fragments with closely packed,

thin-walled capillaries Benign-appearing endothelial cells Blood in the background

Bloody specimen Thin-walled capillaries Benign-appearing endothelial cells Hemosiderin-laden macrophages

Differential diagnosis: spindle cell lesions such as granulomatous hepatitis, leiomyosarcoma, melanoma, spindle cell carcinoma

Extramedullary

hematopoiesis

Variable amount of megakaryocytes, nucleated red blood cells, and various stages of white blood cell maturation

Multinucleated megakaryocytes

Differential diagnosis: Reed-Sternberg cells seen in Hodgkin lymphoma Mallory body Cytoplasmic eosinophilic inclusion with

characteristic “twisted-rope” appearance Red color by Papanicolaou stain and blue color by Diff-Quik stain

Commonly seen in alcohol-related liver disease and hepatocellular carcinoma

Differential diagnosis: eosinophilic body in urothelial cell carcinoma Bile pigment Cytoplasmic pigment Dark-green color by

Papanicolaou and Diff-Quik stains

Seen in both benign hepatocytes and hepatocellular carcinoma

Differential diagnosis: melanin pigments, lipofuscin, and hemosiderin pigments

Lipofuscin Cytoplasmic pigment Golden-brown color by

Papanicolaou stain; green-brown color by Diff-Quik stain

Normal findings due to cellular aging or degenerative changes

Differential diagnosis: melanin pigments, bile pigments, and hemosiderin pigments

Hemosiderin Cytoplasmic pigment Yellow-brown color by

Papanicolaou stain; blue-brown color with

Diff-Quik stain

Seen in normal liver and abnormal iron metabolism

N:C ratio nuclear-cytoplasmic ratio

Benign Bile Duct Epithelial Cells

Benign bile duct epithelium appears as small clusters and/or

sheets of cohesive, uniform cells Of note, the size of benign

bile duct epithelial cells is smaller than that of normal

hepa-tocytes These cells have round to ovoid nuclei, dark and

granular chromatin, inconspicuous or occasionally small

nucleoli, and scant cytoplasm Focal nuclear overlap and

nuclear disarray are not uncommon findings and may mimic

an adenocarcinoma, but in adenocarcinoma (including

pri-mary and metastatic lesions), tumor cells will have a

mark-edly increased nuclear-cytoplasmic ratio (N:C ratio), coarse

or hyperchromatic chromatin, prominent nucleoli, and, in

many cases, vacuolated cytoplasm Therefore, making

par-ticular note of the nuclear features as well as imaging and

clinical characteristics will be very useful for the accurate

assessment of a cluster of epithelial cells (Fig. 7.1)

Benign Liver Cells

Benign liver cells (Fig. 7.2) may be present in several terns, including scattered clusters, flat sheets, or dispersed individual cells The specimen also may reveal trabecular arrangements and may contain many tissue fragments Most commonly, benign hepatocytes are found as sheets or as dis-persed, individual, large, polygonal cells with centrally placed round to oval nuclei, granular chromatin, small and prominent nucleoli, and, most importantly, a normal N:C ratio Binucleation is often encountered and should not be interpreted as cytologic atypia Bile and lipofuscin pigments are often identified in the cytoplasm Bile stains dark green

pat-by Papanicolaou and Diff-Quik methods, whereas lipofuscin stains golden brown by Papanicolaou stain and green-brown

by Diff-Quik Benign liver cells may be seen in cirrhosis, hepatic adenomas, focal nodular hyperplasia, nodular regen-erative hyperplasia, and other conditions During the cyto-logical evaluation of slides, it is important to know the location of the lesion, the type of procedure performed, and how the sample was obtained (such as the path of the needle) because benign hepatocytes may be the predominant cell type on the slide and can obscure rare lesional cells in certain circumstances

Fig 7.1 Benign bile duct epithelial cells (Papanicolaou, 40×)

Fig 7.2 Benign hepatocytes (Diff-Quik, 20×)

Bile Duct Hamartoma

The bile duct hamartoma (Fig. 7.3), also known as von

Meyenburg complex, Meyenburg complex, or biliary

hamar-toma, is a benign tumor-like developmental malformation of

the liver [19–21] The incidence of this lesion is estimated to

be between 1% and 3%, based on autopsy series [19–21] It

is usually an incidental finding, but it may be associated with

autosomal dominant polycystic kidney disease and Caroli’s

disease The bile duct hamartoma may be singular (located

throughout the liver parenchyma) or multifocal (located

mainly in the subcapsular area) These lesions are usually

smaller than 1 cm FNA reveals bland-appearing columnar

cells haphazardly arranged in tubules, sheets, and two-

dimensional clusters The cells can be admixed with

scat-tered stromal cells, which constitute the dense, fibrous

stroma that surrounds the bile ducts in the liver, but this

material is not always easily aspirated Surrounding benign-

appearing hepatocytes, which are more readily aspirated,

may comprise a majority of the cellularity

The main differential diagnosis for a bile duct hamartoma

includes cholangiocarcinoma and metastatic

adenocarci-noma In cholangiocarcinomas and adenocarcinomas, tumor

cells form three-dimensional clusters with large,

hyperchro-matic nuclei, coarse chromatin, irregular nuclear membranes,

prominent nucleoli, and vacuolated cytoplasm Appropriate

immunohistochemistry (IHC), based on the patient’s history

of malignancy, will be useful when a metastasis is suspected

In combination with cytology, clinical imaging will be most

helpful in differentiating between a bile duct hamartoma and

cholangiocarcinoma [19–21]

Cirrhosis

Although not routinely assessed on FNA, cirrhotic livers may be sampled because of the presence of an ill-defined cirrhotic nodule, which is indeterminate for a mass on an image study On FNA, the specimen can be variably cellular with hepatocytes, bile duct epithelium, and stromal cells/fragments (Fig. 7.4) Hepatocytes may appear both normal

or mildly atypical—showing some degree of anisocytosis, binucleation, slightly increased N:C ratios, prominent nucle-oli, or intranuclear inclusions

Liver cirrhosis is a well-known risk factor for the opment of hepatocellular carcinoma (HCC), which is the most important entity in the differential diagnosis In fact, the presence of abundant bile duct epithelium is a helpful feature to distinguish cirrhosis from hepatic adenomas and well-differentiated HCC. Hepatic adenomas do not contain intralesional bile duct epithelium In HCC, tumor cells show

devel-a wide rdevel-ange of cytomorphologicdevel-al devel-atypidevel-a In well- differentiated forms, tumor cells resemble normal liver cells and may form trabeculae, cords, or nests Typically, however, they have an increased N:C ratio, large round nuclei, naked nuclei, and bile pigment in the cytoplasm In poorly differen-tiated forms, malignant cells are markedly polygonal and dyscohesive, with pleomorphic nuclei and giant tumor cells The most notable feature of HCC, particularly in well- differentiated tumors, is the identification of sinusoidal cap-illaries surrounding the markedly thickened trabeculae of neoplastic cells

Fig 7.4 Cirrhosis (Papanicolaou, 10×) Fig 7.3 Bile duct hamartoma (Diff-Quik, 20×)

Fatty Liver Changes

Fatty change, known as hepatic steatosis, can be the result of

a number of diverse physiologic changes and can occur in

the background of benign and malignant lesions In fact, it

has been reported that steatosis can be identified in up to

40% of HCCs [22, 23] It is also interesting to note that the

prevalence of fatty change decreases as HCC tumor size

increases [22, 23] Steatosis is best appreciated by a Diff-

Quik stain as intracytoplasmic vacuoles or as dispersed

bub-bles in the background of the slide (Fig.  7.5) In a

well-differentiated HCC, the finding of tumor cells with

prominent fatty change should not be confused with a benign

lesion or a histiocytic process

Extramedullary Hematopoiesis

Extramedullary hematopoiesis (EMH) is always an mal finding in adults and can be caused by bone marrow fail-ure disorders such as myelofibrosis [20] In such conditions, EMH commonly involves the liver and spleen FNA cytol-ogy reveals a variable amount of megakaryocytes, nucleated red blood cells, and various stages of white blood cell matu-ration The most easily identifiable feature of EMH is the presence of megakaryocytes [24] Sometimes, however, megakaryocytes can be confused with Reed-Sternberg cells, which are seen in Hodgkin lymphoma Megakaryocytes are much larger than Reed-Sternberg cells and typically have three to five lobes (Fig. 7.6) In addition, other features of Hodgkin lymphoma should be absent in the specimen Finally, the mixed cell types and lack of clonal expansion should favor a benign process over a neoplastic process

present (Diff-Quik, 20×)

Focal Nodular Hyperplasia

Focal nodular hyperplasia (FNH) is typically an incidental

finding most commonly found in women of reproductive age

[20, 25, 26] Recent studies have reported that the lesion

involves both females and males equally [25, 26] On

imag-ing, FNH can form a tumor-like mass that may warrant tissue

sampling, though most suspected FNH lesions can be

moni-tored with serial imaging and do not require tissue sampling

[20, 25, 26] When sampling is required because of

indeter-minate radiographic features, FNA specimens are often

bloody and contain clusters or dispersed individual, benign-

appearing liver cells with either no atypia or, at most, mild

nuclear atypia Cells have abundant granular cytoplasm and

normal N:C ratios Bile duct epithelial cells, often forming

long tubular structures, are also identified in most cases

because bile ducts may proliferate at the periphery of the

lesions (Fig. 7.7) Like cirrhotic nodules, FNH nodules are

surrounded by bands of fibrosis, which may or may not be

present as stromal cells or fragments of fibrous tissue on

aspirates

The main differential diagnosis includes hepatic

adeno-mas and well-differentiated HCCs In hepatic adenoadeno-mas,

the bile duct epithelial cells should be absent In HCC

(particularly well-differentiated carcinomas), tumor cells

may recapitulate the normal architecture of hepatocytes,

forming trabeculae, cords, and nests Cytologically,

how-ever, HCC cells have increased N:C ratios, large round

nuclei, prominent nucleoli, and naked nuclei The most

notable feature of HCC is the identification of sinusoidal

capillaries surrounding markedly thickened trabeculae of

neoplastic cells, which will be absent in both FNH and

hepatic adenomas

Hepatic Adenoma

Hepatic adenoma, also referred to as a liver cell adenoma, is an uncommon benign tumor most often occurring in women of child-bearing age who have a prolonged history of using oral contraceptives [27–29] In the general population, this lesion is rare and has been associated with androgen therapy and under-lying metabolic diseases such as diabetes mellitus and glycogen storage disease Hepatic adenomas can become large, outgrow their vascular supply, and develop intratumoral hemorrhage or undergo spontaneous rupture into the peritoneal cavity [27–29] Therefore, accurate diagnosis is important because of the poten-tial life-threatening risk of hemorrhage Cytologically, FNA specimens are hypercellular and contain clusters or dispersed individual, benign- appearing, monotonous hepatocytes with little to no nuclear atypia and normal N:C ratios (Fig. 7.8) No bile duct epithelial cells are present within the lesion, but a few bile duct epithelial cells may be present from aspiration of sur-rounding normal liver parenchyma

It is important to distinguish hepatic adenomas from other benign liver tumors such as hemangiomas (spindle cell lesion, mixed with benign liver cells) and focal nodular hyperplasia (presence of bile duct epithelial cells), because malignant trans-formation to HCC may occur in up to 10% of patients [27–29] Unlike HCC, hepatic adenomas should have preserved reticulin scaffolding and lack significant nuclear atypia Finally, several molecular subtypes have been identified in hepatic adenomas, such as inactivating mutations in hepatocyte nuclear factor 1A, activating mutations in β-catenin, activation of inflammatory signaling pathways, and several other genetic alterations [29] It has been reported that activating mutations of the β-catenin gene are associated with a higher risk of malignant transformation and the development of tumor bleeding [29]

Fig 7.8 Hepatic adenoma with an absence of bile duct epithelium

(Papanicolaou, 20×)

Fig 7.7 Focal nodular hyperplasia with benign hepatocytes and bile

duct epithelium (Diff-Quik, 10×)

Hemangioma

Hepatic hemangiomas are the most common mesenchymal

tumors of the liver; they are reported to be present in up to

20% of the general population, based on several autopsy and

imaging studies [20, 30, 31] Many of these tumors are

pres-ent at the time of birth [30, 31] The causes of this lesion are

not clear, but it has been postulated that sex hormone

imbal-ances may play a role, as it has been found that the size of

tumors increases markedly during pregnancy [30, 31] Most

tumors are less than 4 cm in size, are typically solitary and

subcapsular in location, and are discovered incidentally on

imaging performed for unrelated reasons Although both

ultrasound and CT can be used for the detection of tumors,

magnetic resonance imaging (MRI) is the best imaging

tech-nique for the characterization of the tumor, particularly with

the advanced MRI techniques and usage of hepatocyte-

specific contrast agents On MRI, tumors show a

heteroge-neous appearance Extensive scarring can mimic malignant

neoplasms of the liver such as cholangiocarcinoma,

requir-ing tissue samplrequir-ing for accurate classification [30, 31]

These tumors comprise dilated vascular spaces lined by

flat, bland-appearing endothelial cells without atypia, with

various amounts of intervening fibrous septa On FNA, these

characteristics are often displayed as scant tissue fragments

with embedded and closely packed, thin-walled capillaries in

a background of abundant blood The endothelial cells of

these capillaries are notably bland The background of slides

may reveal benign liver cells, stromal cells, and hemosiderin-

laden macrophages (Fig. 7.9)

The main differential diagnosis includes spindle cell

lesions such as granulomatous hepatitis, leiomyosarcoma,

melanoma, spindle cell carcinoma, and others GLUT-1 is an

immunohistochemical marker that is highly specific for

hemangioma and can be used to differentiate hemangioma

from vascular malformations [32]

Hydatid Cyst

Hydatid cysts (echinococcal cysts) are caused by tion of the larval stage of the cestode (or tapeworm) of the

accidental swallowing of eggs found in feces of dogs Larvae develop over the course of years to form fluid-filled cysts in various organs, particularly the liver Clinically, these present as a large, cystic lesion; though rare in the United States, this is the most common cause

of liver cysts worldwide [33, 34] Cysts can grow to siderable size and contain a large amount of fluid contain-ing infectious scolices On imaging, cysts are typically solitary; about a quarter are lined by a wall of calcifica-tion If suspected clinically, cysts should not be aspirated, out of concern for cyst fluid leakage and subsequent ana-phylactic shock The cysts have an acellular wall made from both host tissue (pericyst) and larval tissue (endo-cyst) FNA specimens contain fragments of the laminated cystic wall, scolices, hooklets, and fluid (Fig.  7.10) Morphologically, the hooklets have the distinctive appear-ance of shark teeth and are acid-fast positive The differ-ential diagnosis includes other cysts of parasitic origin, and clinical and microbiological correlation is often required for the accurate diagnosis

con-Fig 7.9 Hemangioma with large, intact vascular structures

( Diff- Quik, 20×)

Fig 7.10 Hydatid cyst of the liver with fibrous cystic wall

(hematoxy-lin and eosin [H&E], 20×)

Primary Malignant Tumors

The most common primary malignant tumors of the liver are

hepatocellular carcinoma (HCC), accounting for 80–85% of

liver tumors [35, 36] The incidence of HCC has more than

doubled over the past 30 years in the United States [35, 36],

largely owing to the increased incidence of hepatitis B and

hepatitis C infections (particularly in the so-called “baby

boomer” population), alcohol abuse, and obesity and type II

diabetes [35, 36] It is well known that the development of

HCC is also associated with many other insults that damage

the liver, such as alcohol- and non-alcohol-related cirrhosis,

aflatoxin B1 exposure, and parasite infections [37, 38] HCC

is most commonly seen in the sixth and seventh decades of

life and is two to three times more common in men than in

women [35, 36] Recent molecular studies have also found

that mutations of TP53 and β-catenin (CTNNB1) are

fre-quently associated with HCC [39]

Clinically, tumors may present as a single mass or

multi-ple masses throughout the liver [37, 38] Because of advances

in imaging technology, 80% of HCC cases can be diagnosed

in liver nodules larger than 2 cm Therefore, hepatic nodules

that are sampled by FNA are more likely to be smaller than

2 cm, lacking diagnostic imaging features of HCC [40] The cytological diagnosis of a well-differentiated HCC, as well

as the grading of the tumor, can be challenging, particularly

in cirrhotic patients [15, 41, 42] Other primary tumors, including cholangiocarcinoma, hepatoblastoma, neuroendo-crine tumors, and many others, are much less common and are most difficult to distinguish from metastatic carcinomas [5 43]

Importantly, metastases are much more common than mary hepatic malignancies in non-cirrhotic livers [5 43, 44] Most metastases can be diagnosed on the basis of cytomor-phology, based on the similarity of the sample with features

pri-of the primary malignancy, but it is usually necessary to firm the diagnosis by IHC study Furthermore, in difficult cases, IHC markers may be used to aid in the differentiating

con-a primcon-ary liver tumor from con-a metcon-astcon-asis A pcon-anel of mcon-arkers

is usually performed, including liver markers of arginase 1, glypican-3, hepatocyte paraffin 1 (Hep Par 1), CD10, and other IHC markers of various organ origins [45–48]

The cytological features of major types of malignant tumors are summarized in Table 7.2

Table 7.2 Summary of main cytological features of malignant tumors

Conditions Main findings Key features of differential diagnosis

Resemble normal hepatocytes, but loss of the reticulin scaffolding Numerous naked nuclei on slides Endothelial cell surrounding thickened cords of tumor cells

Differential diagnosis: regenerating nodule of cirrhosis, liver cell adenoma, focal nodular hyperplasia

pleomorphism Numerous naked nuclei on slides

Numerous pleomorphic individual tumor cells with hyperchromatic nuclei, irregular nuclear shape, large prominent nucleoli

Differential diagnosis: metastatic carcinoma, cholangiocarcinoma

Large polygonal tumor cells separated by dense lamellar material

Differential diagnosis: other types of hepatocellular carcinoma Cholangiocarcinoma Clusters, crowded sheets, disorganized

honeycomb groups, and dispersed individual cuboidal cells Large nuclei, prominent nuclei, scant cytoplasm High N:C ratio

No cytoplasmic bile pigment, no naked nuclei on slides

Similar to other type of adenocarcinoma

Differential diagnosis: hepatocellular carcinoma, metastatic adenocarcinoma

Hepatoblastoma In fetal cell variant, tumor cells resemble normal

liver cells In the anaplastic variant, tumor cells reveal a pleomorphic appearance with high mitotic activity and abundant tumor cell necrosis

In the embryonal and small-cell undifferentiated variant, the tumor reveals a primitive and undifferentiated appearance with hyperchromatic nuclei and scant cytoplasm

Heterogeneous group of neoplasms with wide variation in their morphology

Differential diagnosis: hepatocellular carcinoma, small-cell carcinoma, metastatic carcinomas

Metastatic

adenocarcinoma

Three-dimensional clusters, papillary and acinar arrangements of columnar cells Hyperchromatic nuclei with prominent nucleoli, coarse chromatin,

“lacy” cytoplasm, and cytoplasmic vacuolization (cytoplasmic mucin)

Malignant columnar cells Irregular nuclear membrane, coarse chromatin, prominent nucleoli, and cytoplasmic vacuole (mucin production) High N:C ratio

Differential diagnosis: cholangiocarcinoma, hepatocellular carcinoma

Metastatic squamous

cell carcinoma

Dyscohesive clusters, loosely formed dimensional cellular sheets, and scattered individual polymorphic cells Hyperchromatic nuclei Smudgy chromatin With or without cytokeratin formation

two-Polygonal, rounded, elongated, or tadpole-shaped cells Large, dark nuclei, smudgy chromatin, and dense cytoplasm (cytokeratin formation) or without cytokeratin formation

Differential diagnosis: poorly differentiated hepatocellular carcinoma

Fine chromatin (salt-and-pepper appearance), paranuclear blue bodies, mitosis, necrosis, and apoptotic bodies

Differential diagnosis: lymphoma, basaloid squamous cell carcinoma, poorly differentiated adenocarcinoma Carcinoid Loosely cohesive clusters and scattered individual

cells Rosette-like arrangement Relatively uniform tumor cells, with fine (salt-and- pepper) chromatin and moderate cytoplasm No mitosis or necrosis

Monomorphic appearance of tumor cells, with fine chromatin, inconspicuous nucleoli Branching capillaries in the

background No mitosis or necrosis

Differential diagnosis: atypical carcinoid, small-cell carcinoma Metastatic melanoma Scattered individual large cells with prominent

nucleoli Cytoplasmic melanin pigment

Binucleation with “mirror” arrangement

Hodgkin lymphoma)

Dispersed individual atypical lymphoid cells with coarse chromatin and irregular nuclear membrane, prominent nucleoli High N:C ratio and scant cytoplasm Increased mitotic activity and the presence of background lymphoglandular bodies

Tumor cells range from small to large in size, depending on the type of lymphoma Monomorphic population of lymphocytes

in SLL/CLL. Polymorphic population in other types

Differential diagnosis: reactive lymphocytes, small-cell carcinoma, poorly differentiated carcinoma

Sarcomas Four major cytomorphologic categories

depending on predominant tumor type:

• Epithelioid and clear cell morphology

• Spindle cell morphology

• Biphasic spindle and epithelioid morphology

• Small round cell morphology

Heterogeneous group of mesenchymal neoplasms with wide variation in their morphology, genetics, immunoprofile, and clinical behavior

Differential diagnosis:

CLL chronic lymphocytic leukemia, N:C ratio nuclear-cytoplasmic ratio, SLL small B-cell lymphocytic lymphoma

Hepatocellular Carcinoma

In HCC, tumor cells show a wide range of

cytomorpho-logical variability [15, 41, 42] Generally, FNA of HCC

reveals a hypercellular specimen composed of dispersed,

single cells and tissue fragments with various

architec-tural and cytological features indicative of the

differentia-tion (grade) of the tumor Further, there should be a

notable absence of bile ducts in lesional tissue; but bile

ducts located adjacent to the carcinoma in the patient may

be aspirated, smeared, and displaced next to tumor cells

on the slide In a well- differentiated HCC, tumor cells

resemble relatively normal liver cells and form wide

tra-beculae, cords, and nests with only a mildly to moderately

increased N:C ratio Large round nuclei, prominent

nucle-oli, naked nuclei, and bile pigment in the cytoplasm are

typical findings In poorly differentiated forms, malignant

cells are polygonal and dyscohesive, with pleomorphic

nuclei, naked nuclei, and giant tumor cells The most

notable feature of HCC, particularly in well- differentiated

tumors, is the presence of sinusoidal capillaries

surround-ing markedly thickened trabeculae of neoplastic cells—a

phenomenon referred to as “endothelial wrapping.” This

feature, not seen in benign liver cells and/or

cholangiocar-cinoma, is essentially pathognomonic Another common

feature is the presence of small vessels traversing tissue

fragments

In difficult cases, IHC markers may be used to aid in the differential diagnosis A panel of markers is usually performed, including arginase 1, glypican-3, Hep Par 1, CD10, and CAM5.2 for proving hepatocyte tissue origin [45–47] For the differential diagnosis of cholangiocarci-noma, the panel should include polyclonal or monoclonal carcinoembryonic antigen (CEA), CK7, and maspin [47] Cholangiocarcinoma is positive for CEA, CK7, and maspin Polyclonal CEA and CD10 demonstrate a cana-licular staining pattern in HCC but a cytoplasmic pattern

in cholangiocarcinoma The demonstration of α-fetoprotein (AFP) positivity points toward a malignant tumor of hepatocellular origin, but it is only positive in 30% of HCC cases [46] It is also positive in nonsemino-matous germ cell tumors and extrahepatic AFP-producing carcinomas [46] Hep Par 1 antibody (clone OCH1E5.2.10), developed in 1993 by Wennerberg et al., stains normal and neoplastic hepatocytes [48] Several studies have reported that this antibody is a sensitive marker for HCC (80–90%) [46–48] However, recent studies have found that it also frequently stains gastric carcinomas (30–47%) [47] Several other tumors can also stain positively for Hep Par

1, including yolk sac tumors and carcinomas of the nal cortex, lung, colon, and ovary It also stains cholan-giocarcinoma in 50% of cases, and unfortunately, poorly differentiated HCCs are more likely to be negative for Hep Par 1 [47, 48]

Well-Differentiated Hepatocellular Carcinoma

Tumor cells of well-differentiated HCC resemble normal

liver cells and form wide trabeculae, cords, and nests with an

increased N:C ratio Tumor cells have large, round nuclei

with prominent nucleoli The cytoplasm may contain

promi-nent bile pigments and/or lipofuscin pigments, which are

golden and brown in color with Papanicolaou staining—a

feature that is very helpful in distinguishing HCC from a

metastatic carcinoma Naked nuclei are also commonly

pres-ent on the slide The most notable feature in HCC,

particu-larly in well-differentiated tumors, is the loss of the reticulin

scaffolding pattern and the presence of endothelial

wrap-ping, as described above (Fig. 7.11) This feature is best seen

on core biopsy specimens but can also be visualized on

smears Unfortunately, the mild nuclear atypia seen in well-

differentiated HCCs can be confused with many entities,

including nodular regenerative hyperplasia in cirrhosis As a

result, clinical and radiographic correlation, such as the

pres-ence of “rapidly enlarged liver mass” is incredibly helpful in

the diagnosis

Poorly Differentiated Hepatocellular Carcinoma

In a poorly differentiated tumor, numerous pleomorphic individual tumor cells with hyperchromatic nuclei, irregular nuclear shape, prominent nucleoli, and dense granular cyto-plasm are often appreciated Tumor cells may be arranged in cords, clusters, or dispersed as individual cells Numerous naked nuclei with large, prominent cherry-red nucleoli and intranuclear pseudoinclusions are also seen on smears (Fig. 7.12) Dense cytoplasm with cytoplasmic bile/lipofus-cin pigments are less likely to be present in a poorly differ-entiated case, but when they are present, they are particularly helpful in distinguishing HCC from a metastatic carcinoma

Fig 7.12 Poorly differentiated hepatocellular carcinoma (Diff-Quik,

20×)

Fig 7.11 Well-differentiated hepatocellular carcinoma with

endothe-lial wrapping

Fibrolamellar Hepatocellular Carcinoma

The fibrolamellar variant of HCC is usually seen in patients

under the age of 30 years; it represents approximately 5% of

HCCs This subtype has a favorable prognosis and,

impor-tantly, does not arise in a background of a cirrhotic liver The

cytomorphological features of fibrolamellar HCC reveal

large, polygonal tumor cells separated by fibrous tissue on a

core biopsy specimen The tumor cells are much larger than

the cells of a well-differentiated HCC.  On smears, tumor

cells reveal large nuclei and abundant oxyphilic cytoplasm;

the N:C ratio of tumor cells may be in a normal range or

slightly increased, but nuclear membranes should be

irregu-lar (Fig. 7.13) The tumor cells also contain prominent

nucle-oli and intracytoplasmic hyaline globules and pale bodies

The dense lamellar material is also seen in slides, separating

loosely cohesive tumor cells

Recent studies have shown that the fibrolamellar variant

is frequently positive for CK7 and CD68, less frequently

positive for glypican-3, and negative for AFP [47]

Hepatoblastoma

The hepatoblastoma is a rare tumor, but it is the most

com-mon primary malignant liver tumor in infancy and childhood

[49, 50] Patients present with a large, solitary hepatic mass,

most often in the right lobe; most patients have an elevated

serum AFP level [49, 50] Hepatoblastoma can be syndromic

(i.e., Beckwith-Wiedemann syndrome) or sporadic

Hepatoblastoma is a heterogenous group of neoplasms,

including two subtypes, with either entirely epithelial

com-ponents or mixed epithelial and mesenchymal comcom-ponents

Based on the differentiation of the epithelial component, tumors are divided into several subtypes, including well- differentiated fetal, poorly differentiated embryonal, macro-trabecular, and small-cell undifferentiated subtypes [49, 50]

In FNA specimens, findings vary depending on the cell type of the tumor but are generally hypercellular (Fig. 7.14) [50] In the fetal cell variant, tumor cells resemble normal liver cells but are smaller in size In the anaplastic variant, tumor cells reveal a more pleomorphic appearance, with high mitotic activity and abundant tumor cell necrosis The fetal and embryonal patterns are often present in a single tumor; the pure fetal type is associated with a more favor-able prognosis In the embryonal and small-cell undifferen-tiated variant, the tumor reveals a primitive and undifferentiated appearance with hyperchromatic nuclei and scant cytoplasm, with cells forming sheets, rosettes, tubules, and trabeculae Finally, mixed mesenchymal com-ponents may be present and are characterized by fibrous tissue containing spindle cells with or without heterologous elements

The main differential diagnosis of hepatoblastoma is HCC and metastatic small round blue cell tumors, such as Wilms’ tumor, embryonal sarcoma, and neuroblastoma Positive staining with Hep Par 1 and glypican-3 in the fetal and embryonal types supports the diagnosis of a liver primary Hepatoblastoma is also positive for high-molecular- weight cytokeratin, whereas HCC is negative for high-molecular-weight cytokeratin [40, 51] Finally, β-catenin is positive in as many as 70% of cases Clinical and radio-graphic correlation is extremely helpful in determining pri-mary versus metastatic tumors, as occult small round blue cell tumors are rare in this population

Cholangiocarcinoma

Cholangiocarcinoma is a group of neoplasms that arise from

cholangiocytes located within various regions of the biliary

tree [52] Based on the anatomic location, the tumors can be

classified as intrahepatic, perihilar, or distal

cholangiocarcino-mas Of the three types, the intrahepatic cholangiocarcinoma

is the most common Most cholangiocarcinomas are

recog-nized as well-differentiated, moderately differentiated, or

poorly differentiated adenocarcinomas Recently, a distinct

subtype known as a combined hepatocellular- cholangiocellular

carcinoma was recognized by the World Health Organization

(WHO) classification; it represents less than 1% of all liver

cancers and has a poor prognosis [53] In the combined HCC/

cholangiocarcinoma variant, the tumor consists of both HCC

and cholangiocarcinoma morphology Because it is difficult to

definitely diagnose it on FNA, it is not further discussed here

Although most patients with cholangiocarcinoma have no

known risk factors, cirrhosis and viral hepatitis B and C

recently have been recognized as risk factors, particularly in

patients with intrahepatic cholangiocarcinomas [52, 53]

Other well-known risk factors for intrahepatic

cholangiocar-cinoma include primary sclerosing cholangitis, congenital

liver malformations, infection with the parasitic liver flukes

to Thorotrast (thorium dioxide), a chemical formerly used in

medical imaging [52, 53] Recent molecular studies have

shown that several genetic alterations have been identified,

including RAS-RAF-MEK-ERK, PI3K-AKT-mTOR, EGFR

and ERBB2, and KRAS signaling pathways [52] Clinically,

most intrahepatic cholangiocarcinomas can be diagnosed by

image studies in non-cirrhotic livers, but when a malignant

mass is present in a cirrhotic liver, the next differential

diag-nostic step is to distinguish cholangiocarcinoma from HCC,

which requires a morphological assessment of the mass

The cytomorphological features of cholangiocarcinoma

(particularly intrahepatic cholangiocarcinoma) include

hypercellular aspirates composed of tissue fragments with

overlapping cells in abnormal architectural patterns such as

sheets of cells with a disorganized honeycomb appearance,

three-dimensional clusters, disorganized acini, or dispersed

atypical individual cells Similar to other types of

adenocarcinoma, tumor cells have an increased N:C ratio

with large oval nuclei, irregular nuclear membranes, coarse chromatin, prominent nucleoli, and either scant cytoplasm or more abundant mucin-containing cytoplasm Mitotic activity

is typically present, and tumor necrosis may be appreciated (Fig. 7.15)

The main differential diagnosis of cholangiocarcinoma includes HCC (particularly in tumors with gland-like archi-tectures) and metastatic adenocarcinomas In HCC, tumor cells typically do not form true glandular structures, and mucin should be absent by staining with mucicarmine For the differential diagnosis of cholangiocarcinoma, the panel should include polyclonal or monoclonal carcinoembryonic antigen (CEA), CK7, and maspin [47] Cholangiocarcinoma

is positive for CEA (cytoplasmic staining), CK7, and maspin In metastatic adenocarcinoma, the tumor may resemble features seen in the primary tumor, and IHC mark-ers and a thorough clinical and radiographic history will be imperative to the workup It has been suggested that a four-marker panel of S100P, pVHL, CK17, and MUC5AC can be used to differentiate intrahepatic cholangiocarci-noma from a metastatic pancreatic adenocarcinoma Intrahepatic cholangiocarcinoma has a staining pattern of S100P−/pVHL+/MUC5AC−/CK17−, whereas a metastatic pancreatic adenocarcinoma has a staining pattern of S100P+/pVHL−/MUC5AC−/CK17+ [47]

Fig 7.15 Cholangiocarcinoma (Papanicolaou, 40×)

Lymphoma

The liver is often involved in patients with non-Hodgkin

lym-phoma (NHL) In fact, the prevalence in lymlym-phoma patients

has been reported to range from 15% to 50% [54] The most

common subtypes identified are diffuse large B-cell

lym-phoma (DLBCL), T-cell lymlym-phomas, Burkitt’s lymlym-phoma,

follicular lymphoma, marginal zone B-cell lymphoma

(MZL), and small B-cell lymphocytic lymphoma (SLL) [54]

Primary hepatic lymphoma (PHL) is defined as lymphoma

confined to the liver without evidence of involvement of any

other organ system, including the spleen, lymph nodes, bone

marrow, or other lymphoid structures [54, 55] PHL is

extremely rare, accounting for less than 0.01% of all cases of

NHL; it typically occurs in middle-aged men The hepatic

involvement by NHL typically presents as multiple nodular

lesions in the liver, with a propensity to infiltrate around the

portal tracts The most frequent clinical finding is

hepatomeg-aly Jaundice is rare, occurring in less than 5% of patients

FNAs of lymphomas are typically cellular, with

dyscohe-sive neoplastic cells Depending on the type of lymphoma,

the neoplastic cells may have a monomorphic or pleomorphic

appearance In general, the cytological features of lymphoma

include monomorphic cells with hyperchromatic nuclei with

clumped (soccer ball-like) chromatin, irregular nuclear

mem-brane, and scant basophilic cytoplasm The background of the

smear may reveal karyorrhexis and lymphoglandular bodies,

which are fragments of cytoplasm A thorough history, flow

cytometry, and IHC staining with lymphocytic markers can

be very helpful in the subclassification of lymphomas

Diffuse Large B-Cell Lymphoma

Diffuse large B-cell lymphoma (DLBCL) is a biologically

and clinically diverse group of diseases The FNA reveals a

cellular specimen with dyscohesive, large, atypical lymphoid

cells (Fig. 7.16) Three morphological variants are most

commonly seen: centroblastic, immunoblastic, and

anaplas-tic Most cases are polymorphic, with a mixture of

centro-blastic and immunocentro-blastic cells The centrocentro-blastic variant,

the most common subtype, reveals tumor cells of medium to

large size with high N:C ratios, oval or round nuclei, fine

chromatin, and single or multiple prominent nucleoli

Immunoblasts have a basophilic cytoplasm and a central

nucleolus The anaplastic variant consists of large tumor

cells with pleomorphic nuclei; the cells may resemble Hodgkin cells or Reed-Sternberg cells

DLBCL can also mimic several carcinomas The main ferential diagnosis includes HCC, poorly differentiated ade-nocarcinoma, and small-cell carcinoma In HCC, tumor cells reveal hyperchromatic nuclei, coarse chromatin, large promi-nent nucleoli, intranuclear pseudoinclusions, and cytoplasmic bile pigment Numerous naked nuclei and tumor necrosis are also seen in HCC In poorly differentiated adenocarcinomas, tumor cells form three-dimensional clusters with large, hyper-chromatic nuclei, coarse chromatin, irregular nuclear mem-brane, prominent nucleoli, and vacuolated cytoplasm In small-cell carcinomas, tumor cells reveal fine (salt-and-pep-per) chromatin with nuclear crowding and molding

Immunochemical markers can be very helpful in the ferential diagnosis For example, DLBCL should be reactive with CD45 and at least one B-cell marker, such as CD20, PAX5, or CD79a It should be noted, however, that a subset

dif-of cases can be positive with P63 and other cytokeratin ers Additionally, based on the staining patterns of CD10, BCL-6, and MUM1, DLBCL is divided into two types: ger-minal center and non-germinal center, with the former hav-ing a better prognosis Finally, BCL2, BCL6, and MYC rearrangements, as well as Epstein-Barr virus (EBV) status, have important clinical indications regarding therapeutic options and disease prognosis

mark-Fig 7.16 Diffuse large B-cell lymphoma, germinal center type (H&E, 40×)

Hodgkin Lymphoma

Hodgkin lymphoma (HL) is a rare malignant neoplasm

that can involve a variety of anatomic sites, including the

liver The Hodgkin and Reed-Sternberg (HRS) tumor cells

of classic HL (cHL) and nodular lymphocyte-predominant

HL (NLPHL) are derived from mature B cells, but these

neoplastic cells are often obscured by a reactive

inflamma-tory infiltrate that makes up the bulk of the lesion Recently,

Epstein-Barr virus has been considered to be a risk factor

in a subset of cHL [56] The HL is composed of

mononu-clear Hodgkin and binucleated or multinucleated

Reed-Sternberg cells, which have prominent nucleoli, and a

variable mixture of nonneoplastic small lymphocytes,

eosinophils, neutrophils, histiocytes, and plasma cells

(Fig. 7.17) [57]

The characteristic finding is HRS cells [56, 57] HRS

cells show expression of CD30 and MUM1, variable

expres-sion of CD15, and weak expresexpres-sion of PAX5 However,

HRS cells may lose their B-cell phenotype and show an

unusual expression of other hematopoietic cell lineage

markers, leading to confusion with other hematopoietic

malignancies The binucleated or multinucleated

Reed-Sternberg cells most likely derive from the mononuclear

Hodgkin cells through a process of tumorigenesis involving

genetic alterations and numerous intracellular signaling

pathways [56] The process is also mediated by cellular

interactions within the lymphoma microenvironment The

genetic alterations involve members of the nuclear factor-κB

(NF-κB) and JAK/STAT pathways, as well as major

histo-compatibility complex expression

The main differential diagnosis of cHL is small B-cell lymphocytic lymphoma (SLL) and poorly differentiated car-cinoma In SLL, the FNA consists of monomorphic, small lymphocytes with a notable absence of HRS cells The lym-phoma cells express B-cell markers but are negative for CD30 and CD15 expression In poorly differentiated carci-noma, tumor cells are arranged in tight clusters, sheets, or syncytial groups Tumor cells reveal hyperchromatic nuclei with coarse, granular chromatin, prominent nucleoli, high N:C ratio, and positive staining by cytokeratin

(Papanicolaou, 20×)

Epithelioid Hemangioendothelioma

Epithelioid hemangioendothelioma is a rare vascular tumor

that is most often seen in adults over 30 years of age; it can

occur in the bone, soft tissue, lung, and liver [57] The tumor

can arise in medium- to large-sized vessels and can cause

painful thrombophlebitis Overall, the tumor has an

interme-diate prognosis, between a benign hemangioma and a

malig-nant angiosarcoma

On FNA specimens, the cytological features include a

bloody smear with variable amounts of sheets, clusters, and

single, polygonal cells in a mixed background, which may

contain myxohyaline stroma, osteoclast-like giant cells,

hemosiderin-laden macrophages, or areas of calcifications

Tumor cells can have abundant, vacuolated cytoplasm with

intracytoplasmic lumina referred to as “blister cells.” These

cells may be large and atypical, spindled, or plasmacytoid in

shape and may contain large nuclei with vesicular chromatin,

prominent nucleoli, and pseudoinclusions (Fig. 7.18) Tumor

cells are strongly positive for CD31 and CD34 and focally

positive for cytokeratin and SMA [57]

The differential diagnosis includes spindle cell lesions of

the liver, such as granulomatous hepatitis, leiomyosarcoma,

melanoma, spindle cell carcinoma, and other vascular

lesions Glut1 is an immunohistochemical marker, highly

specific for hemangioma, and can be used to differentiate

hemangioma from epithelioid hemangioendothelioma In

addition, the tumor may mimic an epithelioid mesothelioma

Immunomarkers of mesothelial cells (calretinin, D2–40, and

others) may help with the differential diagnosis, though

D2–40 staining may be seen in a subset of epithelioid

hemangioendotheliomas

Metastasis to the Liver

The liver is a common site of metastasis, given the blood supply it receives from the portal vein and the hepatic artery [5 10] In fact, the liver is the most common site to be involved by hematogenous metastatic spread by gastrointes-tinal primaries In addition to the colon, pancreas, and stom-ach, common primary sites also include the lung, breast, kidney, ovary, and prostate [5 10] Metastases typically present as multiple nodules that involve both lobes of the liver and are much more common than HCC, particularly in

a non-cirrhotic liver In the majority of liver metastasis cases, patients have a known malignant history of a primary tumor elsewhere, but some patients do not have a known primary tumor When there is no known primary or malignant history, certain cytological features may suggest a specific primary site Tumors from different primary sites can have quite a lot

of overlapping cytomorphology, however For example, both metastatic colonic and pancreatic adenocarcinomas reveal malignant columnar cells with tumor necrosis Therefore, IHC markers may be necessary for the differential diagnosis

As a result, every effort should be made to prepare a cell block and obtain a core needle biopsy for ancillary studies The cytological features of some commonly seen metastatic tumors are summarized in Table 7.2

Metastatic Colonic Adenocarcinoma

Metastatic colonic adenocarcinoma has several unique logical features Tumor cells reveal a tall, columnar “picket fence” appearance with hyperchromatic, pencil-shaped nuclei, coarse chromatin, and prominent nucleoli The pres-ence of “dirty” necrosis is also seen (Fig. 7.19) The finding

cyto-of tall columnar cells favors a colonic primary over other adenocarcinomas Metastases of other tumors may have dif-ferent cytological features For example, breast carcinoma tumor cells form tight, three-dimensional “cannon ball” clus-ters and, overall, are smaller than colonic adenocarcinomas and may have intranuclear inclusions Renal cell carcinomas usually reveal centrally located nuclei and clear cytoplasm, but a variable amount of oncocytic cytoplasm can be seen and can be difficult to distinguish from a primary HCC Lung adenocarcinoma may have a variety of morphological fea-tures, so it must be worked up with IHC IHC staining with TTF-1, napsin A, CK7, CK20, and CDX2 may aid the dif-ferential diagnosis in difficult cases TTF-1, napsin A, and CK7 are positive in lung adenocarcinoma, whereas CK20 and CDX2 are positive in colonic adenocarcinoma Finally, expression of GATA3 may indicate a breast or urothelial pri-mary carcinoma

Fig 7.18 Epithelioid hemangioendothelioma (Diff-Quik, 20×)

Metastatic Lung Adenocarcinoma

In a metastatic adenocarcinoma of the lung, tumor cells may reveal several cytological appearances In well-differentiated adenocarcinomas, tumor cells show acinar and/or papillary arrangements or form three-dimensional clusters Tumor cells are intermediate or large in size, with hyperchromatic nuclei, coarse chromatin, and prominent nucleoli Tumor cells usually have vacuolated or clear cytoplasm (Fig. 7.20)

In a poorly differentiated adenocarcinoma, the size of tumor cells may be smaller than for well-differentiated tumors; one should not mistake this tumor with smaller tumor cells for a small-cell carcinoma, which is a distinct entity with a very poor prognosis Small-cell carcinoma has fine (salt-and- pepper) chromatin, nuclear crowding, and molding with a high mitotic index and a background of necrosis In HCC, tumor cells are arranged in cords or clusters or are dispersed

as individual cells with centrally located nuclei, large nent nucleoli, and dense cytoplasm containing cytoplasmic bile and/or lipofuscin pigments Numerous naked nuclei are also seen in HCC Cholangiocarcinomas have many cyto-logical features that overlap with those of metastatic adeno-carcinomas, and it is difficult to separate these tumors from metastatic adenocarcinoma based on morphology alone Positive immunostains of TTF1 and napsin A may help in the differentiating lung adenocarcinoma from other carcinomas

promi-In difficult cases, a mucicarmine stain is often helpful in proving that a tumor is in fact an adenocarcinoma

Fig 7.20 Metastatic well-differentiated lung adenocarcinoma

( Diff- Quik, 20×)

Fig 7.19 Metastatic poorly differentiated colonic adenocarcinoma

(Papanicolaou, 40×)

Metastatic Squamous Cell Carcinoma

Metastatic squamous cell carcinomas usually form loose

clus-ters and are dispersed as individual tumor cells The

cytologi-cal features of tumor cells include large nuclei with smudgy

chromatin, nuclei with variation in size and shape, and dense

cytoplasm (indicative of keratin formation) (Fig. 7.21) The

dense cytoplasm is blue in color with a Diff- Quik stain and

red-pink (eosinophilic) in color with Papanicolaou stain

Prominent nucleoli can be seen in poorly differentiated

squa-mous cell carcinomas and should not be confused with poorly

differentiated adenocarcinomas Metastatic squamous cell

carcinomas are immunoreactive with p40 and CK5/6 Tumor

cells may also express p16, depending on the primary site

(such as a head and neck squamous cell carcinoma) The

expression of p63, another marker for squamous

differentia-tion, can be found in approximately 50% of adenocarcinomas

Therefore, the interpretation of p63 should be cautious,

par-ticularly in a poorly differentiated carcinoma

Metastatic Breast Carcinoma

In metastatic breast carcinoma, tumor cells form tight, three- dimensional clusters and are generally smaller than adeno-carcinoma cells of the lung or GI tract The presence of intracytoplasmic lumina, called “magenta bodies,” is also a characteristic feature of metastatic breast carcinomas Magenta bodies are variably sized red-to-purple perinuclear inclusions seen by Diff-Quik stains (Fig. 7.22) Breast carci-noma cells typically have prominent nucleoli, but this feature does not distinguish them from adenocarcinomas of other primary locations Naked nuclei can be seen in metastatic melanoma and HCC, but they should be absent in breast car-cinomas The immunomarker GATA3 may aid in the differ-ential diagnosis of the breast primary Other immunomarkers such as estrogen receptor (ER), progesterone receptor (PR), mammaglobin, and GCDFP-15 (gross cystic disease fluid protein-15) may also be useful in the identification of a breast primary

Fig 7.21 Metastatic keratinizing squamous cell carcinoma

(Papanicolaou, 20×) Fig 7.22 Metastatic breast carcinoma with rare cytoplasmic lumen

(Diff-Quik, 40×)

Metastatic Renal Cell Carcinoma

The most common sites of metastatic renal cell carcinomas

(RCC) include the lung, liver, lymph nodes, and bones

Aspirates from a clear cell RCC are usually cellular and

often quite bloody In a metastatic clear cell RCC, tumor

cells are arranged in clusters or dispersed as individual cells

with round to ovoid, hyperchromatic nuclei, irregular nuclear

membrane, variably prominent nucleoli (depending on the

ISUP grade), and clear cytoplasm The tumor cells have a

relatively low N:C ratio, due to the abundant cytoplasm

Focally, tumor cells may also show large, hyperchromatic

nuclei with multinucleated giant cells, eosinophilic

cyto-plasm, and associated areas of necrosis, particularly in ISUP

grade III and IV tumors Finally, these tumors often have a

prominent small vessel, which should not be confused with

the endothelial wrapping or dissecting vascular structures

present in HCC (Fig. 7.23)

Metastatic Carcinoid

Carcinoids, the most common gastrointestinal endocrine neoplasms, frequently involve the liver, whereas primary hepatic carcinoids are extremely rare Metastatic carcinoids

to the liver most often come from a primary tumor in the intestine, pancreas, or lung Carcinoids, despite differing

embryological origin, have common phenotypic tics The cytological features of carcinoids are characterized

characteris-by acini, rosette arrangements, and dispersed individual tumor cells Tumor cells reveal a fine chromatin pattern (salt- and- pepper chromatin), inconspicuous or small nucleoli, and scant cytoplasm (Fig. 7.24) Nuclear grooves are not seen in carcinoids, and mitoses are extremely rare

The cellular proliferative rate (Ki67 labeling) should be evaluated in tumors because it is a useful marker for the accu-rate grading of neuroendocrine neoplasms In contrast to car-cinoids, high-grade neuroendocrine tumors reveal nuclear crowding and molding, a high mitotic index, and tumor necro-sis In HCC, tumor cells reveal hyperchromatic nuclei, coarse chromatin, large prominent nucleoli, and intranuclear pseu-doinclusions Numerous naked nuclei and tumor necrosis are also seen in HCC.  In adenocarcinomas, tumor cells form three-dimensional clusters with large, hyperchromatic nuclei, coarse chromatin, irregular nuclear membranes, prominent nucleoli, and vacuolated cytoplasm Finally, cholangiocarci-nomas form two-dimensional or three- dimensional clusters, crowded sheets, or dispersed large typical cells with coarse granular rather than fine chromatin and lacy or vacuolated cytoplasm indicative of mucin production In difficult cases, the differential diagnosis of a carcinoid from other carcinomas depends on IHC staining with neuroendocrine markers Carcinoid tumors are positive for synaptophysin, chromo-granin, and CD56 Recently, INSM1 has been considered a better marker for the identification of neuroendocrine tumors

Fig 7.24 Metastatic carcinoid (Papanicolaou, 20×) Fig 7.23 Metastatic renal cell carcinoma with prominent nucleoli

(Papanicolaou, 40×)

Metastatic Endocrine Tumor of the Pancreas

Pancreatic neuroendocrine neoplasm (PEN), usually located

in the body or tail of the pancreas, can be of two types, cystic

or solid [58] The liver is often involved, even if the primary

tumor is only 1–2  cm in size The majority of tumors are

functional and secrete a variety of hormones, such as insulin,

glucagon, somatostatin, vasoactive intestinal polypeptide

(VIP), pancreatic polypeptide, serotonin,

adrenocortico-tropic hormone (ACTH), or calcitonin Cytological aspirates

of the solid type are composed of sheets and/or loosely

cohe-sive cell clusters Similar to other neuroendocrine tumors,

PENs often form pseudorosettes and acinar arrangements

Tumor cells are relatively uniform and intermediate in size,

with scant to moderate amounts of cytoplasm The chromatin

of tumor cells is finely granular (salt-and-pepper) in texture,

and the nucleoli are small and inconspicuous (Fig. 7.25)

Sometimes the tumor cells can have eccentrically located

nuclei and a plasmacytoid appearance Finally, capillaries

may be attached to or surround tumor cells

On immunostudies, tumors are usually positive for

cyto-keratin, synaptophysin, chromogranin, INSM1, and CD56

Additionally, amyloid may be seen in a subset of cases,

particu-larly insulinomas A mitotic count and Ki67 proliferation index

should be performed to properly assess the tumor grade The

differential diagnosis of a metastatic small-cell carcinoma may

be considered when nuclear crowding and molding are present

in the setting of a high mitotic index and tumor necrosis

Metastatic Small-Cell Carcinoma

On FNA, small-cell carcinomas are typically lar and contain a dirty, necrotic debris with apoptotic bodies and crush artifact Tumor cells are relatively small (two to three times the size of mature lymphocytes) and display nuclear overlap and nuclear molding The N:C ratio is extremely high, owing to scant cytoplasm The nuclei contain fine chromatin, indistinct nucleoli, and frequent mitosis On slides, crush artifact seen as “blue strips” of nuclear material is also characteristic (Fig. 7.26)

hypercellu-In poorly differentiated adenocarcinomas, tumor cells are larger, with hyperchromatic nuclei, coarse chromatin, prominent nucleoli, and vacuolated cytoplasm The tumors also reveal acinar and three-dimensional arrangements that are typically absent in small-cell carcinomas In lym-phoma, tumor cells have a high N:C ratio with coarse chro-matin and nucleoli, without nuclear molding and crowding

In HCC, tumor cells form cords, tubules, sheets, or lated individual cells with hyperchromatic nuclei, coarse chromatin, large prominent nucleoli, and intranuclear pseudoinclusions Additionally, cytoplasmic bile/lipofus-cin pigments are characteristically seen in HCC IHC stains with neuroendocrine markers for synaptophysin, chromogranin, CD56, and INSM1, as well as Ki67, are helpful in the differential diagnosis of small-cell carcinoma

(Papanicolaou, 40×)

Metastatic Urothelial Cell Carcinoma

In metastatic urothelial cell carcinoma, the tumor cells form

clusters, sheets, or individual large, pleomorphic cells with

large, irregular nuclei and dense cytoplasm Tumor cells

show an increased N:C ratio, hyperchromatic nuclei with

coarse (chunk of charcoal) chromatin, irregular nuclear

membranes, and round eosinophilic cytoplasmic inclusions

(Melamed-Wolinska bodies) (Fig. 7.27) Melamed-Wolinska

bodies are also commonly seen in degenerated urothelial

cells, particularly in voided urine The pathogenesis of these

bodies is still not fully understood, and they can be found in

both benign and malignant conditions When they are seen in

an extraurinary cytological specimen, this finding may

sug-gest that the lesion is of urothelial origin

The main differential diagnosis of metastatic urothelial

cell carcinoma is metastatic squamous cell carcinoma and

HCC, as both tumors display dense cytoplasm In HCC,

tumor cells have large, prominent nucleoli and cytoplasmic

bile and/or lipofuscin pigments, which stain golden brown

color with the Papanicolaou method Numerous naked nuclei

are also seen in HCC.  In addition, the presence of the so-

called cercariform cells, a clue for squamous differentiation,

may be seen in urothelial cell carcinoma, which can make

distinction from a pure squamous cell carcinoma difficult

The “cercariform cells” are large tumor cells with long,

cytoplasmic tails Immunostains of the

high-molecular-weight cytokeratin, CK903 (34BE12), and GATA3 are

posi-tive in urothelial cell carcinomas and can be helpful in

indicating urothelial origin

Metastatic Adenocarcinoma of the Prostate

The most predominant cytological features of metastatic adenocarcinoma of the prostate include a cellular speci-men with three-dimensional clusters and microacinar and cribriform arrangements of malignant cells Tumor cells are small to intermediate in size with hyperchromatic nuclei, coarse granular chromatin, prominent nucleoli, and vacuolated or clear cytoplasm There is usually an absence

of significant inflammation or cellular necrosis in the background (Fig. 7.28) Higher-grade metastatic tumors, especially Gleason grade V, may pose a diagnostic chal-lenge on the basis of cytology alone Tumor cells may be more dyscohesive, with a complete loss of acinar or cribri-form architecture The individual tumor cells of a poorly differentiated prostate carcinoma may have marked nuclear atypia and hyperchromasia not seen in well-differentiated tumors

Immunostains with prostate markers such as PSA (prostate- specific antigen), PSAP (prostate-specific acid phosphatase), prostate-specific membrane antigen, and NKX3.1 are usually positive in metastatic tumor cells However, a small percentage of prostatic adenocarcinomas, particularly those treated with androgen deprivation therapy, may lose the expression of prostate markers Therefore, cor-relation with the patient’s malignant history is critical for an accurate diagnosis

Fig 7.28 Metastatic adenocarcinoma of the prostate (Papanicolaou, 40×) Fig 7.27 Metastatic urothelial cell carcinoma with rare eosinophilic

cytoplasmic inclusions (Papanicolaou, 40×)

Metastatic Gastrointestinal Stromal Tumor

Gastrointestinal stromal tumor (GIST) metastatic to the liver

often displays clusters of spindle cells with focal palisading

nuclei on smears, and the cytoplasm is generally pale and

fibrillar, with perinuclear vacuoles indenting the nucleus

Other cytological features of GIST include sheets or loosely

formed clusters of spindle cells with oval or bipolar nuclei,

bland chromatin, inconspicuous nucleoli, and vesicular

cyto-plasm (Fig. 7.29)

IHC of tumor cells is positive for CD34, c-kit, and DOG1

The differential diagnosis of a spindle cell lesion in liver

FNA cytology is broad, including virtually all tumors with

spindle cell morphology, such as spindle cell carcinoma,

leiomyoma, sarcomatoid mesothelioma, melanoma,

angiomyolipoma, and others Therefore, the correlation of

cytological features with clinical and imaging findings is

extremely important and may help to narrow the differential

diagnosis Primary GISTs are more frequently seen in the

bowel or stomach wall and are less commonly found within

the soft tissue In a poorly differentiated carcinoma,

epitheli-oid tumor cells form three-dimensional clusters with large,

hyperchromatic nuclei, coarse chromatin, irregular nuclear

membranes, and prominent nucleoli In sarcomatoid

meso-thelioma, tumor cells have an epithelioid appearance with

hyperchromatic, oval nuclei, prominent nucleoli, and dense

cytoplasm Mesothelial cell markers (calretinin, D2–40, and

others) may aid in the differential diagnosis Finally, CK5/6

and P40 are typically positive for spindle cell squamous cell

carcinoma

Metastatic Melanoma

In metastatic melanoma, tumor cells are usually arranged in loose clusters or dispersed as individual cells Nuclei of tumor cells are eccentrically located (plasmacytoid appear-ance) and highly variable in size, with fine to coarse, granu-lar chromatin and a single prominent, cherry-red nucleolus The cytoplasm tends to be abundant and may or may not contain melanin pigment The N:C ratio may vary signifi-cantly and is not always increased Melanin pigment appears coarsely granular and dark brown on Papanicolaou stain Other characteristic features include multinucleation, binu-cleation, and intranuclear pseudoinclusions (Fig. 7.30).Some cytological features of metastatic melanoma may overlap with features of HCC, such as individual cells with prominent large nucleoli, intranuclear inclusions, cytoplasmic pigment, binucleation, and the presence of a plasmacytoid appearance The cytological differential diagnosis of a meta-static melanoma and HCC can be difficult In HCC, tumor cells have large, prominent nucleoli and dense cytoplasm that contains cytoplasmic bile and/or lipofuscin pigments, staining golden and brown in color with the Papanicolaou method Numerous naked nuclei are also seen in HCC.  In difficult cases, IHC stains of SOX10, S100, HMB45, and Melan-A may be helpful for the diagnosis of melanoma Among these markers, S100 may stain both nuclei and cytoplasm with 90% sensitivity, but it is not very specific HMB45 stains cytoplasm and is more specific but less sensitive than S100 (only 0–20% positivity in desmoplastic/spindle cell melanoma) Melan-A stains cytoplasm but it is negative in desmoplastic/spindle cell melanoma SOX10 has higher sensitivity and specificity for the melanoma In summary, a combination of melanoma IHC markers is preferred if sufficient material is available

Fig 7.29 Metastatic gastrointestinal stromal tumor (GIST) (Diff-

Metastatic Wilms’ Tumor

Nephroblastoma (Wilms’ tumor) is the most common

malig-nant primary renal tumor in children before the age of

5 years The tumor is usually multiphasic and shows

blaste-mal, epithelial, and mesenchymal components Generally, in

a FNA specimen, numerous dyscohesive, small round blue

cells are found, representing the blastemal component, with

scant cytoplasm, nuclear overlap, coarse chromatin, and

small nucleoli Further, rare ganglion-like, large, pale cells

may be present Tumor epithelial cells form characteristic

tubular-like structures and tight clusters, representing

primi-tive tubular and glomerular-like structures The

mesenchy-mal cells reveal an elongated spindle cell appearance, which

may show varying degrees of differentiation In fact, the

presence of skeletal muscle differentiation is highly

indicative of a nephroblastoma in a child The background of

smears may reveal fibrillary material (neuropil) (Fig. 7.31)

The tumor cells can be positive for AE1/AE3, WT1,

vimentin, and desmin and variable for CD99/MIC2 The

main differential diagnosis is rhabdomyosarcoma and PNET

(primitive neuroectodermal tumor)/Ewing’s sarcoma In

rhabdomyosarcoma, tumor cells have more eosinophilic

cytoplasm In PNET/Ewing’s sarcoma, tumor cells reveal a

“small blue cell” appearance, with hyperchromatic nuclei,

scant cytoplasm, and a high N:C ratio; they should not

con-tain heterologous differentiation The background of the

smear may reveal a tigroid appearance The tumor may also

contain “light” cells with pale nuclei and fine granular

chro-matin and “dark,” dying cells with hyperchromatic nuclei

and smudged chromatin Similarly to Wilms’ tumor, PNET/Ewing’s sarcoma coexpresses CD99/MIC2 (the glycoprotein

of MIC2) and vimentin Other nonspecific markers can be

found in PNET/Ewing’s, including S-100, neuron-specific enolase, CD75, and synaptophysin However, the PNET/Ewing’s tumor usually expresses FLI1 (Friend leukemia

integration-1), which is the product of the fusion gene EWS

PNET/Ewing’s is known to carry a characteristic

transloca-tion of t(11;22)(q24;q12) and/or fusion gene EWS/FLI1 The

detection of these abnormalities may aid in the diagnosis

Fig 7.31 Metastatic Wilms’ tumor with blastemal, mesenchymal, and

tubal epithelial components (H&E, 20×)

Metastatic Leiomyosarcoma

Smooth muscle tumors, including leiomyoma and

leiomyo-sarcoma, may metastasize to the liver Aspiration of a

leio-myosarcoma typically reveals a cellular specimen comprising

clusters of spindle cells with cigar-shaped, hyperchromatic

nuclei, coarse chromatin, inconspicuous nucleoli, and

fibril-lary cytoplasm Mitosis and tumor necrosis are also

fre-quently present (Fig. 7.32)

The differential diagnosis of a leiomyosarcoma includes a

variety of spindle cell lesions For example, poorly

differen-tiated carcinoma and sarcomatoid carcinomas may share

spindle cell morphology with cells having variably shaped

ovoid nuclei and prominent nucleoli In addition, these

enti-ties often express cytokeratins, making the differential

diag-nosis even more difficult Findings of marked cytological

atypia, atypical mitotic figures, and markedly increased

cel-lularity are usually features favoring a diagnosis of

carci-noma In spindle cell carcinoma, tumor cells reveal elongated

or bizarre nuclei, prominent nucleoli, and scant cytoplasm

In desmoplastic/spindle melanoma, tumor cells reveal oval

or bizarre nuclei and prominent nucleoli; they do not contain

cytoplasmic melanin pigment In contrast, the characteristic

cytological features of smooth muscle tumors include

eosin-ophilic cytoplasm with a fibrillary appearance, classic cigar-

shaped nuclei with finely dispersed chromatin, and no

prominent nucleoli Immunostains with muscle markers

(smooth muscle actin and desmin) are helpful for the

differ-ential diagnosis of the tumor

Metastatic Adrenal Cortical Carcinoma

FNA of a metastatic adrenal cortical carcinoma typically reveals a cellular smear containing cohesive clusters and contains cohesive clusters of small to intermediate-sized tumor cells Tumor cells have slightly hyperchromatic nuclei, fine and evenly distributed chromatin, and inconspicuous nucleoli Focal anisonucleosis, nuclear atypia, and naked nuclei dispersed in a bubbly and granular background may

be found Finally, mitotic activity and necrosis are monly identified (Fig. 7.33)

com-Malignant cortical carcinomas may show cytological tures that overlap those of HCC. In HCC, tumor cells have large, prominent nucleoli and dense cytoplasm that contains cytoplasmic bile and/or lipofuscin pigments, staining golden and brown in color with the Papanicolaou method In diffi-cult cases, IHC stains of cytokeratin and inhibin are positive

fea-in metastatic adrenal cortical carcfea-inoma and can help with the correct diagnosis

Fig 7.32 Metastatic leiomyosarcoma (Papanicolaou, 20×)

Fig 7.33 Metastatic adrenal cortical carcinoma (H&E, 40×)

Metastatic PNET/Ewing’s Sarcoma

PNET/Ewing’s sarcoma is a primitive neoplasm of neural

crest origin It accounts for approximately 5% of malignant

tumors in children and young adults The tumor may consist

of two distinct cell types, small “dark” cells and large “light”

cells The “light” cells reveal pale nuclei and fine granular

chromatin; the “dark” cells represent dying cells with

hyper-chromatic nuclei and smudged chromatin The background

of smears reveals a tigroid appearance The tumor cells may

lose specific differentiation and display small, round nuclei

with finely granular chromatin and scant cytoplasm, giving a

“small round blue cell” appearance (Fig. 7.34) Tumor cells

may be positive for CD99, neurofilament, and chromogranin

and are typically negative for cytokeratin and muscle

mark-ers Similar to a subset of Wilms’ tumors, PNET/Ewing’s

sarcoma tumors co-express CD99/MIC2 (the glycoprotein of

MIC2) and vimentin Other nonspecific markers may be

pos-itive in PNET/Ewing’s sarcoma, including S-100, neuron-

specific enolase, CD75, and synaptophysin PNET/Ewing’s

sarcoma usually expresses FLI1, which is the product of the

fusion gene EWS/FLI1—a characteristic translocation of

t(11;22)(q24;q12) The detection of these abnormalities may

aid in the diagnosis

Metastatic Papillary Thyroid Carcinoma

Metastasis of papillary thyroid carcinoma (PTC) to the liver

is a rare event, typically occurring in patients with a long clinical latent period Low clinical suspicion or remote clini-cal history of the PTC may complicate an accurate diagnosis The features that are typically present in primary PTC are usually present in metastatic PTC [14] These characteristic findings on FNA include sheets of tumor cells with or with-out overt papillary-like architecture; pale oval nuclei; evenly distributed, fine, powdery chromatin; prominent nuclear grooves; and intranuclear inclusions (Fig. 7.35) Certain cytological features may pose a diagnostic challenge in this location, particularly when focal acinar features and three- dimensional clusters of tumor cells are present

In general, the differential diagnosis of metastatic PTC in the liver is broad and includes commonly seen carcinomas such as metastatic adenocarcinoma from the lung, pancreato-biliary tract, colon, breast, or prostate In addition, the acinar arrangement, relatively small tumor cell size, and finely granular chromatin may lead to the consideration of a well- differentiated neuroendocrine neoplasm, such as carcinoid or pancreatic neuroendocrine tumors Nuclear grooves, crowd-ing and molding, intranuclear pseudoinclusions, and periph-erally placed micronucleoli are features of metastatic PTC. The presence of cohesive fragments of tumor cells and punched-out nuclear inclusions, with a lack of pale, powdery chromatin, should raise concern for a metastatic carcinoma IHC can be very helpful in the workup, as thyroglobulin and TTF-1 are positive for a metastatic PTC, whereas other immunomarkers (TTF, CDX2, NKX3, and others) may be positive for metastatic adenocarcinomas of different origins

Fig 7.35 Metastatic papillary thyroid carcinoma with classic

intranu-clear pseudoinclusions (Papanicolaou, 40×)

Fig 7.34 Metastatic PNET/Ewing’s sarcoma (Diff-Quik, 10×)

Metastatic Endometrioid Adenocarcinoma

FNA of a metastatic endometrioid adenocarcinoma reveals

columnar tumor cells with hyperchromatic, pencil-shaped

nuclei, coarse chromatin, and prominent nucleoli The

pres-ence of “dirty” necrosis is also commonly seen (Fig. 7.36)

The finding of columnar cells and tumor necrosis may be

confused with metastatic colonic adenocarcinoma, but in

colonic adenocarcinoma, the tumor cells reveal a tall,

colum-nar “picket fence” appearance In metastatic breast

carcino-mas, tumor cells form tight, three-dimensional “cannon ball”

clusters, are usually smaller than those of endometrioid

ade-nocarcinoma, and may contain intranuclear inclusions

Metastatic renal cell carcinomas usually reveal centrally

located nuclei and clear cytoplasm Metastatic lung

adeno-carcinomas may have a variety of morphological features,

making it difficult to determine the tissue of origin

IHC staining with PAX8, EMA, TTF-1, napsin A, CK7,

CK20, and CDX2 may aid the differential diagnosis in

diffi-cult cases Metastatic endometrioid carcinomas are positive

for PAX8 and EMA. TTF-1, napsin A, and CK7 are positive

in lung adenocarcinoma, whereas CK20 and CDX2 are

posi-tive in colon adenocarcinoma Expression of GATA3 may

indicate a breast or urothelial primary

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Fig 7.36 Metastatic endometrioid carcinoma (Diff-Quik, 20×)