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Cytology and the

Diagnosis of Neoplasia
Diplomate ACVP (Clinical Pathology) SPECIMEN PREPARATION
Associate Professor Choosing an area from which to collect a sample for
Department of Veterinary Biosciences cytology and the method of collection depend on what
College of Veterinary Medicine abnormality is detected clinically. Neoplastic process-
The Ohio State University es can result in discrete tumor masses, invade normal
Columbus, Ohio 43210 parenchyma and cause organ enlargement, induce in-
flammation or hemorrhage, or result in abnormal se-
KEY WORDS cretions or effusions. There are numerous references
cytology describing sample collection techniques from a wide
neoplasia variety of tissues, and specific techniques are not dis-
fine-needle aspiration biopsy cussed in this article.
impression smear In general, samples for cytologic evaluation are col-
malignancy lected by fine-needle aspiration, touch impression, or
gentle tissue scraping. Superficial cutaneous and sub-
Cytology refers to the microscopic evaluation of cutaneous nodules and readily accessible tissues such
cells. Cytologic evaluation can be very useful in the as peripheral lymph nodes are easily sampled by fine-
clinical diagnosis of neoplasia. Samples for cytology needle aspiration. Ulcerated masses can be sampled
can be collected from a wide variety of sites and by impression smears, fine-needle aspiration, or gen-
many different tissues. Sample collection is relatively tle tissue scraping. Ultrasound is useful for guiding
noninvasive, and most samples can be collected on an needle aspiration biopsies of focal or diffuse neoplas-
outpatient basis. Both sample collection and specimen tic infiltrations involving internal organs; using ultra-
preparation can be performed using inexpensive sound increases the likelihood of obtaining a diagnos-
equipment that is readily available in most veterinary tic sample and decreases the risk of complications.
practices. In-house interpretations can be made the Specimen preparation and submission are addressed
same day, and interpretations from reference laborato- elsewhere in these Proceedings (see pp. 79).
ries frequently are available within 24 hours. Compli-
cations of sample collection for cytologic evaluation CYTOLOGIC INTERPRETATION
are uncommon and usually are limited to minor hem- Interpretation of cytologic specimens requires
orrhage. Infection, injury to adjacent structures, and knowledge of normal cellular and tissue morphology,
dissemination of neoplastic cells are extremely un- recognition of the limitations of cytology, and experi-
common. ence. To avoid unrealistic expectations and overinter-
Although cytology should be viewed as a screening pretation, cytologic findings should be correlated with
tool, many reactions can be classified as inflammato- other clinical and laboratory findings and with infor-
ry, hyperplastic, or neoplastic. For neoplastic pro- mation about tumor incidence, site predilection, and
cesses, an experienced cytologist can definitively di- gross morphology. For specimens submitted to a ref-
agnose several specific neoplasms, make a tentative erence laboratory for interpretation, it is useful for the
diagnosis of neoplasia for many types of tumors, cytologist to know signalment, a brief history, rele-
identify sites of tumor metastasis, and monitor tumor vant physical examination findings, previous therapy,
regrowth following anticancer therapy. Information a summary of results of pertinent diagnostic tests, the
gained from cytology may be useful in establishing a site from which the sample was collected, and the
diagnosis, determining a prognosis, and formulating a clinicians tentative diagnosis.
diagnostic or therapeutic plan. The most significant Evaluation of stained cytologic preparations re-
disadvantage of cytology is the absence of tissue ar- quires a routine pattern that includes gross and micro-
chitecture. The arrangement of neoplastic cells within scopic examination. The cytologic smear or imprint
tissues is critical in determining the diagnosis of should be observed grossly to evaluate the quality of
many types of tumors, in evaluating surgical margins, the preparation and to locate cellular areas on the slide
and in establishing whether a tumor is benign or ma- to be examined microscopically. The slide is then
lignant.1 evaluated with the 10 objective to estimate cellulari-

ty of the sample, observe cell-to-cell associations, ten-

tatively identify cell types, and locate areas to be ex- CYTOLOGIC FEATURES
amined with higher magnification. Scanning the entire OF MALIGNANT CELLS
slide with the 10 objective is important as many cy-
Cellular Features
tologic samples have cells that are distributed on only Pleomorphism
a small part of the slide.2 Anisocytosis
The 100 (oil immersion) objective is used to ex- Macrocytosis
amine cellular detail. Changes in nuclear and cyto- Variation in nuclear to cytoplasmic ratio
plasmic morphologic characteristics of neoplastic Increased nuclear to cytoplasmic ratio (except
cells are best evaluated at this higher magnification. lymphoid cells)
Only intact cells should be examined and interpreted. Variation in stages of differentiation
Disrupted cells reveal artifactually enlarged nuclei,
pale-staining diffuse chromatin, and nucleolar promi- Nuclear Features
nence, all of which may be misinterpreted as cytolog- Anisokaryosis
ic characteristics of neoplastic cells.2 Macronuclei
Multinuclearity with abnormal nuclei
Abnormal mitotic figures
Inflammation and Hyperplasia Finely dispersed or coarsely clumped chromatin
Many tumors elicit an inflammatory response that Thickened, angular, or indented nuclear
may mask the presence of neoplastic cells. Inflamma- membrane
tion is characterized by a mixed population of cells in- Nuclear molding
cluding neutrophils, lymphocytes, plasma cells, Macronucleoli
eosinophils, monocytes, and macrophages.3 The pres- Multiple nucleoli
ence of inflammation may be problematic in the cyto- Irregularly shaped nucleoli
logic diagnosis of neoplasia because of the small Anisonucleoliosis
amount of tissue evaluated and the lack of architec-
ture to define the demarcation between inflammation Cytoplasmic Features
and neoplasia. Increased basophilia
Abnormal vacuoles or granules
Inflammatory reactions often result in hyperplasia of
Phagocytosis of other cells
surrounding tissues. Cells from hyperplastic tissue re-
semble normal cells except they appear more imma-
ture. Cytologic characteristics of hyperplastic cells in-
clude large nuclei with poorly condensed chromatin
and prominent nucleoli. Cytoplasm often is basophilic. large and pleomorphic in size and shape when com-
Hyperplastic cells have a relatively constant nuclear- pared with normal cells of the same type (see box
to-cytoplasmic (N:C) ratio (nuclear size compared above).
with the amount of cytoplasm present), an important Benign neoplastic lesions yield cells that are rela-
feature in distinguishing hyperplastic from neoplastic tively uniform in size and appearance. Nuclei have a
cells.3 The distinction between hyperplasia and benign similar chromatin pattern, and nucleoli usually are
neoplasia often must be made histologically. small and regular in outline or inconspicuous.1 Rarely
do nuclei of benign cells exceed two to three times the
Cytologic Features of Neoplasia size of homologous red blood cells in diameter. There
Neoplasia usually is recognized cytologically by the is minimal variation in N:C ratio in cells from most
presence of cells that are neither inflammatory nor benign tumors. The cytoplasm from benign neoplastic
normally expected from the site of collection.4 For ex- cells appears to be at a similar stage of differenti-
ample, the presence of squamous epithelial cells in a ation.1
lymph node aspirate is highly suggestive of metastatic In contrast to benign neoplastic lesions, cells from
neoplasia. Because neoplasms are clonal expansions most malignant tumors are very pleomorphic in ap-
of a particular type of cell, cells from a specific tumor pearance. Although the degree of pleomorphism may
often appear similar cytologically. This is described as directly correlate with clinical behavior for some tu-
a uniform or monomorphic population of cells, even mors, there are exceptions. For example, cells from
though the cells may be morphologically dissimilar. some tumors that behave in a relatively benign man-
The cytologic features of neoplastic cells vary consid- ner may appear very pleomorphic (e.g., histiocytoma)
erably with cell type. In general, neoplastic cells are and cells from some clinically aggressive malignant

tumors are not very pleomorphic (e.g., canine thyroid determining the degree of differentiation but are not
carcinomas). as useful as nuclear criteria for distinguishing between
Malignant cells often exhibit moderate to marked benign and malignant cells. The cytoplasm of neo-
variation in cell size, which is referred to as anisocy- plastic cells may be more basophilic than the cyto-
tosis. Cells from malignant tumors usually are macro- plasm of the normal cell counterpart. Some neoplastic
cytic compared to nonneoplastic cells of the same cells may have abnormal cytoplasmic vacuolization
type. The N:C ratio often varies markedly from cell to and/or granulation.4,9
cell. For most tumors, the N:C ratio increases with
malignant transformation. Lymphoid cells, however, NEOPLASTIC EFFUSIONS
normally have a high N:C ratio. The N:C ratio ap- Recognition of neoplastic cells in body cavity effu-
pears decreased in many lymphoid neoplasms, com- sions depends on the cytologists ability to identify an
pared to normal lymphocytes.5 Malignant cells from abnormal cell type and to recognize the criteria of ma-
some tumors appear to be at different stages of differ- lignancy. This frequently is difficult because neo-
entiation and may exhibit asynchrony between nucle- plasms involving the thoracic or abdominal cavity can
ar and cytoplasmic maturation. For example, some cause inflammation and mesothelial cell hyperplasia.
cells from a squamous cell carcinoma may appear to Reactive mesothelial cells may closely resemble cells
be very immature, some may appear relatively ma- from malignant neoplasms and are characterized by
ture, some may appear to have a mature nucleus with increased basophilia, variation in cell size, binucle-
immature cytoplasm, and some may appear to have an ation or multinucleation, and frequent mitoses.1012 Re-
immature nucleus with mature cytoplasm. active mesothelial cells may occur in large clusters,
Nuclear changes are of primary importance in deter- which also is characteristic of some malignant neo-
mining malignancy of a neoplasm. In the absence of plasms. Unfortunately, there are no morphologic crite-
inflammation or other causes of dysplasia, the pres- ria that clearly distinguish between reactive mesothe-
ence of more than three nuclear criteria of malignancy lial cells and cells from malignant neoplasms.
in the majority of the cells is considered highly sug- Mediastinal lymphoma is a common cause of neo-
gestive of malignant neoplasia.6,7 Malignant cells may plastic effusion in small animals, especially in cats.4
have nuclei as large as 10 to 12 microns in diameter Effusions from animals with mediastinal lymphoma
(macronuclei), which is about the size of a neutrophil. often contain large numbers of neoplastic lympho-
There may be variation in nuclear size (anisokaryosis) cytes, which most frequently are larger than normal,
among malignant cells. Multinuclearity is not a crite- mature lymphocytes. The nucleus usually is round but
rion of malignancy unless the nuclei within the cell may be indented or irregular in shape. There is a fine
exhibit nuclear criteria of malignancy.8 Nuclei that chromatin pattern, and one to several nucleoli may be
vary in size within the same cell and cells that have present. The cytoplasm is variable in amount and
odd numbers of nuclei are indications of malignancy. staining intensity.12
Frequent normal mitotic figures do not necessarily in- In addition to mediastinal lymphoma, the most
dicate neoplasia.4 However, abnormal mitotic figures commonly diagnosed tumors in small animals involv-
usually are associated with malignant neoplasms.3,4 ing pleural and peritoneal cavities are adenocarcino-
Nuclear chromatin patterns vary, depending on the ma (mammary, bronchiolar, thyroid, pancreatic,
type of tumor. Chromatin may appear less clumped, sweat gland), transitional cell carcinoma, and squa-
as in neoplastic lymphocytes, or may be coarsely mous cell carcinoma. Neoplastic cells from mast cell
clumped and unevenly distributed, as in some carci- tumors and mesotheliomas also can be seen in body
nomas.6 The nuclear membrane may appear irregular- cavity effusions.4,10,13 These tumors often exfoliate
ly thickened, angular, or indented. Some malignant readily, and numerous neoplastic cells are evident cy-
cells show nuclear molding around adjacent nuclei or tologically. However, in many tumor-induced effu-
cytoplasmic vacuoles. 1,8 Nucleoli may be large sions, neoplastic cells are not detected by cytologic
(macronucleoli) in malignant cells. Nucleoli greater examination.10,14 Tumors that may cause neoplastic
than 5 microns in diameter (red blood cells are 5 to 7 effusions but usually exfoliate poorly include heman-
microns in diameter) are highly suggestive of malig- giosarcoma, melanoma, chemodectoma, and pheo-
nancy. Nucleoli also may be irregularly shaped, and chromocytoma.10
more than one nucleolus per cell may be present. Pericardial effusions also can be evaluated cytologi-
Anisonucleoliosis (variation in nucleolar size) occurs cally. Although neoplasia is a relatively common
relatively frequently in malignant cells. cause of pericardial effusion in dogs, cytologic evalu-
Cytoplasmic changes in neoplastic cells may aid in ation of pericardial fluid has been shown to be unreli-

Figure 1. Aspirate from a mast cell tumor. Three mast cells and one Figure 2. Neoplastic lymphocytes with convoluted nuclei typical of some
eosinophil are present. (Wrights-Giemsa stain, 1000). cutaneous lymphomas. (Wrights-Giemsa stain, 1000).

able in distinguishing neoplastic from nonneoplastic diagnosed cytologically than histologically.18 Aspi-
pericardial effusions.15 rates from mast cell tumors usually are very cellular.
Mast cells predominate and are usually round to oval
SKIN TUMORS in shape; numbers of eosinophils and/or fibroblasts
Tumors involving the skin and subcutaneous tissue are variable.3,4,18 The round to oval nucleus has aggre-
occur frequently in dogs and cats. Samples from cuta- gated chromatin but may stain poorly or may be ob-
neous and subcutaneous masses are obtained easily scured by the presence of distinct cytoplasmic gran-
for cytologic evaluation and can be used to differenti- ules (Figure 1).18 These granules are round or oval in
ate between neoplastic and nonneoplastic processes. shape and fine to coarse in size and are the striking
Cytologic evaluation resulted in the correct diagnosis feature that allows the specific identification of mast
for 74% of skin and subcutaneous tumors.16 The cyto- cells. With most Wrights-Giemsa stains, the granules
logic appearance of some of the more common skin are blue-black to purple-red. Diff-Quika may stain
tumors in dogs and cats is presented below. the granules poorly or not at all.
Canine mast cell tumors can be graded according to
Round Cell Tumors their degree of granulation and cellular anaplasia.
Several skin tumors can be definitively diagnosed Well-differentiated mast cells contain numerous gran-
with confidence using cytology. Most of these are re- ules, whereas poorly differentiated ones contain few
ferred to as round cell tumors or discrete cell tumors granules. Although dogs with anaplastic mast cell tu-
because of their characteristic appearance both cyto- mors have significantly shorter survival times than
logically and histologically.2 Cells from round cell tu- those with more well-differentiated tumors, these tu-
mors microscopically are round and have well-de- mors are highly unpredictable. Even well-differentiat-
fined cytoplasmic margins. These cells do not have ed mast cell tumors can be widely metastatic.6 The
cell-to-cell attachments and therefore appear as sepa- primary skin mass may have well-differentiated mast
rate cells. Most exfoliate well when sampled by fine- cells, whereas metastatic sites in the same animal may
needle aspiration or impression.6,9 Round cell tumors contain poorly differentiated tumor cells.19 Mast cell
include mast cell tumors, lymphosarcoma, plasmacy- tumors must be differentiated from inflammatory pro-
tomas, histiocytomas, and transmissible venereal tu- cesses containing mast cells. The presence of other in-
mors. Cells from basal cell tumors and melanomas flammatory cells such as neutrophils and macro-
also may appear as discrete round cells cytologically, phages usually makes this distinction relatively easy,
but histologically these tumors are not considered to but histologic evaluation may be required in some
be round cell tumors.7 cases.

Mast Cell Tumor Lymphoma

Mast cell tumors are common skin tumors in dogs17; Primary cutaneous lymphoma in dogs and cats is
in cats these tumors more commonly involve internal rare. Most cutaneous lymphomas typically exfoliate
organs but also can occur as cutaneous masses. Cyto- cells well and consist of a uniform population of poor-
logic evaluation is an accurate method for the diagno- ly differentiated lymphoid cells. 7,20 Cytologically,
sis of mast cell tumors.4 In fact, some are more readily a
American Scientific.

Figure 3. Aspirate from a plasma cell tumor. Note the perinuclear clear Figure 4. Aspirate from a histiocytoma. (Wrights-Giemsa stain, 1000).
area in some cells. (Wrights-Giemsa stain, 1000).

these cells are larger than neutrophils and have a mod-

erate amount of blue-staining cytoplasm. The nucleus
is round or irregularly shaped and has a finely stippled
chromatin pattern. One to several nucleoli may be
present. Nuclei may be strikingly convoluted in some
cutaneous lymphomas (Figure 2). Cutaneous lym-
phoma composed of small lymphocytes occasionally
may be present. These lymphocytes are smaller than
neutrophils and cannot be distinguished cytologically
from hyperplastic lymphoid tissue. Although cytology
is useful in tentatively diagnosing cutaneous lym-
phoma, histologic confirmation is recommended.
Figure 5. Aspirate from a transmissible venereal tumor. (Wrights-Giem-
sa stain, 1000).
Cutaneous Plasmacytoma
Cutaneous plasmacytomas are benign neoplasms
that should be included in the differential diagnosis of
cutaneous round cell tumors. These tumors occur at with distinct borders that may appear scalloped.9 His-
solitary sites and most commonly involve the skin of tiocytomas may be infiltrated with lymphocytes and
digits, lips, and ears in dogs. Cells from cutaneous plasma cells. Histiocytomas also may be ulcerated
plasmacytomas are round to oval and have predomi- and secondarily inflamed; in these cases the cytology
nantly round nuclei with coarsely clumped chromatin resembles chronic inflammation and the definitive di-
and a single, small nucleolus. There is a variable agnosis may require histopathology.3 Differentiation
amount of amphophilic to pale basophilic cytoplasm. among histiocytoma and large cell lymphoma, histio-
A perinuclear clear area representing the Golgi zone cytic lymphoma, transmissible venereal tumor, basal
is typical of plasma cells (Figure 3). Mitotic activity is cell tumor, and anaplastic, agranular mast cell tumor
low to moderate, and there may be binucleate and also may be difficult without histologic evaluation.3,6,18
multinucleate cells.21
Transmissible Venereal Tumor
Histiocytoma Transmissible venereal tumor (TVT) is a discrete
Benign histiocytomas are very common tumors of cell tumor of dogs. Such tumors most commonly in-
the dermis and subcutis of young dogs.17 Cytologic volve mucous membranes of the external genitalia,
samples from histiocytomas contain a uniform popu- but extragenital tumors also occur.17 Cytologic sam-
lation of round, oval, or irregularly shaped cells mor- ples from TVTs usually are very cellular and contain
phologically resembling monocytes or epithelioid round to oval cells with round or oval nuclei.6 The nu-
cells.3,18 The eccentric nuclei of these cells are variable cleus often is eccentric and varies in size. The chro-
in size and shape.12 The chromatin is dispersed and matin is coarsely granular, and there is usually a sin-
nucleoli usually are not prominent (Figure 4).3 There gle prominent nucleolus.4,9 TVT cells have abundant
is a moderate amount of pale bluestaining cytoplasm cytoplasm with distinct cytoplasmic borders.3 The cy-

melanomas may be difficult to recognize cytological-

ly. In most cases, careful observation reveals a few
pigmented melanocytes. If no melanin granules are
found, melanomas are difficult to distinguish from
other mesenchymal tumors cytologically.6,9
Cytologically, cells from melanomas usually occur
singly or in small clusters.4,9,19 Both round and spin-
dle-shaped cells often are found in the same neo-
plasm. Animals with melanomas in which the pre-
dominant cell type is round have shorter survival
times than those with melanomas of predominantly
spindle-shaped cells. 23 Nuclei in melanocytes are
Figure 6. Aspirate from a malignant melanoma. A melanocyte with fine round to oval. There is a moderate amount of cyto-
pigment granules and a melanophage with coarse pigment granules are
present. (Wrights-Giemsa stain, 1000).
plasm with ill-defined cytoplasmic borders. 9
Melanocytes from a melanoma must be differentiated
from pigmented macrophages. Macrophages may
contain large coarse granules of hemosiderin or
toplasm may appear pale blue or moderately ba- melanin and, when compared with melanocytes, usu-
sophilic and often contains distinct vacuoles (Figure ally are larger and have vacuolated cytoplasm. The
5). Cytologic samples from TVTs also may contain hemosiderin and melanin pigment granules in
plasma cells, lymphocytes, and macrophages.9,18 Al- macrophages are more coarse and variably sized than
though the unique location of most TVTs is helpful in those in melanocytes (Figure 6).6
cytologic interpretation, definitive differentiation of
TVT from lymphoma and histiocytoma may require In contrast to the discrete cell tumors and mela-
histologic evaluation. TVT cells usually have more nomas described above, most other neoplasms require
abundant cytoplasm than most neoplastic lymphoid histopathology for definitive diagnosis. However, cy-
cells and are more uniformly round in nuclear and cel- tologic evaluation often can distinguish neoplasia
lular shape than cells from a histiocytoma.3 from inflammation and may be useful in differentiat-
ing neoplasia from hyperplasia. If the cytologic diag-
Melanoma nosis is neoplasia and the cells do not resemble those
Melanomas are relatively common in dogs and rare from a discrete cell neoplasm or melanoma, there are
in cats. Melanomas often are not classified as discrete some general criteria (discussed below) to tentatively
cell tumors as described above nor do they fit neatly diagnose the tumor as epithelial or mesenchymal in
with the epithelial and mesenchymal tumors described origin.
below. Melanomas are of neuroectodermal origin, and
the cells may have characteristics of both mesenchy- Epithelial Tumors of the Skin
mal and epithelial cells in the same tumor.6 Most cuta- There are several characteristics of epithelial tumors
neous melanomas are benign whereas oral and distal in general that are useful cytologically in tentatively
extremity melanomas most often are malignant.22 diagnosing the cell of origin. Epithelial tumors exfoli-
Melanocytes produce melanin pigment, which cyto- ate relatively easily. There may be organized groups
logically appears as brown to greenish black cytoplas- of cells and evidence of cell-to-cell adherence. Indi-
mic granules that are very fine and of uniform size. vidual epithelial cells are round or polyhedral and
Melanin granules often are more obvious cytological- have abundant cytoplasm with well-defined cytoplas-
ly than histologically, and their presence is the most mic borders. The cytoplasm of epithelial cells differs
important criterion used to identify melanocytes. greatly among tumor types with respect to amount,
Melanomas may be very sparsely or heavily pigment- color, granularity, and vacuolation.9 Many epithelial
ed.3 If the tumor is heavily pigmented, the mass ap- tumors involving the skin are of low grade cytologic
pears dark grossly and is easily recognized cytologi- malignancy, making the cytologic distinction between
cally. The sample may appear brown even on benign and malignant neoplasms difficult. Tumors
unstained slides. In general the less pigment, the more with many characteristics of malignancy can be called
anaplastic the cells appear cytologically.6 However, malignant; however, if few malignant characteristics
the amount of melanin present does not necessarily are present, the confirmation that the tumor is benign
indicate the degree of malignancy.17 Poorly pigmented must be made histopathologically.6

Figure 7. Aspirate from a squamous cell carcinoma. Two large neoplastic Figure 9. Aspirate from a perianal gland adenoma. (Wrights-Giemsa
squamous epithelial cells are present. The neutrophils are typical of the in- stain, 1000).
flammatory response that often accompanies squamous cell carcinomas.
(Wrights-Giemsa stain, 1000).

to predict either cytologically or histologically. Aspi-

rates from these tumors often show cords or ribbons of
small, uniform epithelial cells. In some samples there
may be only individual cells. The cells have a high N:C
ratio with scant basophilic cytoplasm (Figure 8). The
cells are not distinctive and appear benign. Histologic
confirmation is recommended because similar-appear-
ing basilar reserve cells can be seen cytologically with
other cutaneous tumors.9 Pigmented basal cell tumors
contain melanocytes and may be difficult to distinguish
cytologically from melanomas.6

Figure 8. Aspirate from a basal cell tumor. (Wrights-Giemsa stain, Sebaceous Gland Tumor
Sebaceous gland tumors occur commonly in old
dogs,17 and most are benign. Cells from sebaceous
Squamous Cell Carcinoma gland tumors often exfoliate in clumps, so it may be
Squamous cell carcinomas usually exfoliate well. difficult to distinguish individual cells cytologically.
Those that are poorly differentiated are characterized The cells have small, centrally located nuclei and
by large, anaplastic cells with giant, polymorphous abundant foamy cytoplasm. There are occasional re-
nuclei. The cytoplasmic borders may appear angular serve cells that appear smaller and have more ba-
due to keratin production (Figure 7). Small, kera- sophilic cytoplasm and a higher N:C ratio.9 Sebaceous
tinized squamous cells and keratin debris usually are gland adenocarcinomas cytologically appear as clus-
present and aid in the diagnosis of squamous cell car- ters of highly basophilic reserve cells with malignant
cinoma.6 Keratin may induce a chronic inflammatory characteristics. There are occasional cells with cyto-
reaction. Chronic inflammation often is associated plasmic secretory material, which sometimes may be
with epithelial cell dysplasia, which may be difficult quite abundant and displace the nucleus to the periph-
to differentiate cytologically from a well-differentiat- ery of the cell (signet ring cell).6
ed squamous cell carcinoma.2 The definitive diagnosis
of squamous cell carcinoma should be made histologi- Perianal Gland Tumor
cally. Perianal gland tumors, most of which are adenomas,
are common in intact, aged male dogs.16 The differen-
Basal Cell Tumor tiation of nodular hyperplasia of the perianal glands
Basal cell tumors arise from multipotential germinal from adenoma is difficult cytologically or histologi-
epidermal cells and are common in the skin of cats and cally. 6 Cells from perianal gland tumors exfoliate
dogs.6,17 The tumors may be pigmented, especially in readily and appear as variably sized clusters or indi-
cats, and often contain cystic spaces.6 Basal cell tumors vidual cells that are polygonal in shape. The cells
usually are benign, but the biologic behavior is difficult have round, central nuclei that frequently contain nu-

Figure 10. Aspirate from a lipoma. Two adipocytes are present. Figure 11. Aspirate from a hemangiopericytoma. (Wrights-Giemsa
(Wrights-Giemsa stain, 1000). stain, 1000).

cleoli. There is abundant pinkish-blue cytoplasm that adipocytes cytologically. Adipocytes are very large
is distinctly granular (Figure 9). The cells resemble cells that are distended with fat (Figure 10). Slides
hepatocytes, and these tumors often are referred to as from lipomatous tumors should always be examined
hepatoid gland tumors. Perianal adenocarcinomas are microscopically because many malignant tumors in-
rare. Cells from perianal adenocarcinomas usually ex- vade subcutaneous fat and neoplastic cells can be de-
hibit variation in nuclear size and number and size of tected cytologically.6
nucleoli; well-differentiated perianal adenocarcino- Liposarcomas occur rarely.17 Aspirates from liposar-
mas are difficult to differentiate from adenomas cyto- comas usually contain numerous cells. These cells
logically or histologically.6 have abundant cytoplasm histologically, but the cell
membrane may be difficult to appreciate cytological-
Mesenchymal Tumors of the Skin ly. Usually there are some cells that contain cytoplas-
Cytologic characteristics of mesenchymal tumors mic vacuoles, which vary in size and number. Often
are different from those described for epithelial neo- there are fat vacuoles in the background.6 The diagno-
plasms. In general, mesenchymal tumors exfoliate sis of lipomas frequently is made cytologically.
poorly because the cells are embedded in extracellular
matrix such as fibrous connective tissue, cartilage, or Hemangiopericytoma
bone. Cytologically, the cells present are individual Hemangiopericytoma is a tumor of questionable ori-
cells or unorganized clusters. Cells from most mes- gin that may arise from cells around blood vessels.17
enchymal tumors are spindle shaped, with cytoplas- The tumor most commonly occurs on the extremities
mic extensions and ill-defined cytoplasmic borders. of older dogs. Cells exfoliate relatively well compared
Nuclei may be fusiform or oval.1 Even if the cell of with other mesenchymal tumors. Usually there are in-
origin cannot be determined cytologically, a tentative dividual cells with round, uniform nuclei. The cyto-
diagnosis of mesenchymal tumor usually can be plasm varies in amount and cytoplasmic membranes
made. Benign or well-differentiated malignant mes- are indistinct (Figure 11). In thick areas of the smear,
enchymal tumors may resemble actively proliferating there may be whorls of cells, which represent the
granulation tissue.6 Definitive diagnosis and determi- characteristic swirling pattern seen histologically.
nation of biologic behavior of most mesenchymal tu- There may be occasional multinucleate cells.6 Cyto-
mors require histologic confirmation. logically, it may be difficult to distinguish heman-
giopericytomas from fibrosarcomas, and the diagnosis
Lipomas should be confirmed histologically.
Lipomas occur commonly in the subcutaneous tis-
sues of dogs and cats. Grossly, aspirates from lipomas LYMPHOID NEOPLASIA
appear oily and smears do not dry on microscope Lymphadenopathy may be associated with lym-
slides.1 The cells often rupture, and the alcohol fixa- phoid hyperplasia, inflammation, or hematopoietic or
tive used in some stains dissolves the fat; thus the metastatic neoplasia. Fine-needle aspiration cytology
slide often appears acellular cytologically. Intact cells of enlarged lymph nodes often can provide a quick di-
from lipomas cannot be distinguished from normal agnosis with minimum inconvenience and expense.

With experience, the cytologic diagnosis of lymph-

adenopathy, including that caused by lymphoid neo-
plasia, can be highly accurate.16,24,25 Knowledge of his-
tory and physical examination findings is useful in the
cytologic evaluation of lymphoid tissue, and it is im-
portant for the clinician to communicate this informa-
tion to the cytologist. For example, immunization may
cause diffuse paracortical hyperplasia to the extent
that the lymphocytes may appear neoplastic.26 A dog
with multiple peripheral lymph node enlargements is
more likely to have lymphoma than a dog with a sin-
gle enlarged lymph node. Young cats with enlarged
peripheral lymph nodes may have nonneoplastic pro- Figure 12. Aspirate from a lymph node from a dog with lymphoma.
(Wrights-Giemsa stain, 1000).
liferative disease instead of lymphoma.27

Cytology of Normal Lymph Nodes monomorphic population of cells at the same stage of
The cytologist must be familiar with normal lymph development.3 The mixed lymphoid population in hy-
node morphology. Normal lymph nodes consist of a perplastic lymph nodes may appear similar to normal
heterogenous population of cells. The predominant nodes but with increased numbers of prolymphocytes
cells are mature lymphocytes, which cytologically ap- and lymphoblasts. There may be increased numbers
pear smaller than neutrophils. The nucleus is similar of plasma cells and macrophages, and an occasional
to red blood cells in size. 28,29 Mature lymphocytes neutrophil may be present. Numbers of mitotic figures
have a very high N:C ratio, coarsely aggregated chro- may appear increased, but the figures are normal mor-
matin, and scant cytoplasm. Nucleoli usually are ab- phologically.3,26
sent, but there may be one small nucleolus.28,29 Pro-
lymphocytes, less mature lymphoid cells, are slightly Cytology of Neoplastic Lymph Nodes
larger than neutrophils. The nuclear chromatin in pro- Lymphoid neoplasms are relatively common in dogs
lymphocytes is moderately clumped, and there is a and cats and can involve multiple tissues including
moderate amount of cytoplasm. Mature lymphocytes lymph nodes, intestine, thymus, and hematopoietic or-
and prolymphocytes constitute 90% to 95% of the gans. In lymph nodes, the normal heterogeneous pop-
normal lymph node population.4 Lymphoblasts are ulation of mature lymphocytes is replaced by a mono-
large cells (two to four times the size of mature lym- morphic (or occasionally dimorphic) population of
phocytes) with fine nuclear chromatin and one to two lymphoid cells.26 Plasma cells and inflammatory cells
nucleoli. Cytoplasm may be scant or abundant. Lym- are rare. The morphology of the cells varies with the
phoblasts represent less than 15% of the cells in nor- degree of differentiation and type of lymphocyte.
mal lymph nodes.28,29 Most lymphoid neoplasms involve immature lym-
Occasional plasma cells may be present in aspirates phoid cells. Lymphoma involving immature cell types
from normal lymph nodes. Plasma cells have eccen- usually can be diagnosed with confidence cytological-
tric nuclei, abundant basophilic cytoplasm, and a peri- ly. Lymphomas involving morphologically mature
nuclear clear area.28 Inflammatory cells such as neu- lymphocytes occasionally occur and are particularly
trophils, macrophages, and mast cells are rare in most difficult to diagnose cytologically.3,18,26
normal lymph nodes. However, the cell population in The degree of differentiation and the histologic clas-
lymphoid tissue may vary according to location. Lym- sification of lymphoma may affect response to treat-
phoid tissue associated with the gastrointestinal tract ment and prognosis. A classification system of canine
may appear to have slightly increased numbers of lymphomas based on the National Cancer Institute
plasma cells and inflammatory cells because of regu- (NCI) Working Formulation of Non-Hodgkins Lym-
lar antigenic exposure.29 phomas in humans recently has been described.30,31
The histologic classification is based on tissue archi-
Cytology of Hyperplastic Lymph Nodes tecture, cell morphology, and mitotic rate; the cyto-
Lymphoid neoplasms must be differentiated from logic classification is based almost entirely on nuclear
lymphoid hyperplasia. Hyperplastic lymph nodes con- characteristics including nuclear size, shape, and
tain lymphoid cells at all stages of differentiation chromatin pattern and nucleolar number and size.
whereas lymphoid neoplasms usually contain a While cytologic classification is not as reliable as his-

tologic classification, it is sufficient to recognize the etic tumors, and evaluation of oncogenes and chromo-
common cell types.30,31 No classification of feline lym- somal abnormalities to determine prognosis.
phomas has been published using this system.
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