osseous metaplasia within a canine insulinoma

C A S E R E P O R T

Osseousmetaplasia within a canine insulinomaEmily M. Pieczarka, Duncan S. Russell, Kelly S. Santangelo, Famke Aeffner, Mary Jo Burkhard

Department of Veterinary Biosciences, College of Veterinary Medicine, The Ohio State University, Columbus, OH, USA

KeyWords

Bone, dog, islet cell carcinoma, pancreas

Correspondence

Mary Jo Burkhard, Veterinary Biosciences,

1925 Coffey Rd., Columbus, OH 43210, USA

E-mail: [email protected]

DOI:10.1111/vcp.12117

Abstract: An 11-year-old male castrated mixed-breed dog was presented

for exercise intolerance, tetraparesis, and persistent hypoglycemia. Abdom-

inal ultrasound examination revealed 2 nodules within the right limb of

the pancreas. Cytology from one nodule was consistent with a carcinoma

of neuroendocrine origin, with a primary differential diagnosis of insulino-

ma. Histologic evaluation and immunohistochemistry for synaptophysin

and insulin confirmed the diagnosis of insulinoma. Additionally, there was

a solitary nodule of mineralized compact bone composing approximately

60% of the mass. To the authors’ knowledge, this is the first report of osse-

ousmetaplasia within an insulinoma (islet cell carcinoma).

Case Presentation

An 11-year-old male castrated mixed-breed dog was

presented to the Ohio State University Veterinary

Medical Center (OSUVMC) for evaluation of exercise

intolerance and tetraparesis for several weeks’ dura-

tion, as well as persistent hypoglycemia. Serum chem-

istry performed at OSUVMC showed sustained marked

hypoglycemia (36 mg/dL; reference interval [RI]

77–126 mg/dL), mild hypokalemia (3.92 mEq/L; RI

4.2–5.4 mEq/L), and mildly elevated total serum ALP

activity (148 IU/L; RI 15–120 IU/L]) with significantly

increased corticosteroid-induced ALP isoenzyme activ-

ity (138 IU/L; RI 0–6 IU/L). An abdominal ultrasound

examination identified 2 hypoechoic nodular struc-

tures measuring 2.9 9 1.9 cm (cranial nodule) and

3.3 9 1.4 cm (caudal nodule) in the right limb of the

pancreas. The echogenicity of these nodules was heter-

ogenous with focal hyperechoic shadowing regions. A

fine-needle aspirate of one pancreatic nodule revealed

many variably sized clusters of large, round to polygo-

nal shaped cells within a background of blood and

numerous bare nuclei (Figure 1). Neoplastic cells dis-

played mild-to-moderate anisocytosis and anisokaryo-

sis. Nuclei were round to oval, centrally placed, and

had a fine chromatin structure with 1–3 prominent

round to ovoid nucleoli. These cells had moderate

amounts of basophilic cytoplasm that frequently con-

tained multiple small punctate vacuoles. Occasional

cell clusters contained distinct intercellular junctions

and/or piling of cells, as well as multiple ruptured cells.

Rare binucleated cells, nuclear molding, and mitotic

figures were observed. These findings were compatible

with a carcinoma of neuroendocrine origin. The

primary differential diagnosis was an insulinoma.

Thoracic radiographs were acquired prior to sur-

gery and revealed no evidence of metastasis. A partial

pancreatectomy was performed of the right limb along

with surgical excision of 2 regional lymph nodes, and

Figure 1. Fine-needle aspirate of a pancreatic mass in a dog. Note a

cluster of malignant epithelial cells with moderate amounts of pale baso-

philic cytoplasm with varying numbers of small punctuate vacuoles, and

round nuclei with a fine chromatin structure and prominent nucleoli.

There are also a few bare nuclei and ruptured cells, suggestive of neuro-

endocrine origin. ModifiedWright–Giemsa. Bar = 10 lm.

Vet Clin Pathol 43/1 (2014) 89–93©2014 American Society for Veterinary Clinical Pathology and European Society for Veterinary Clinical Pathology 89

Veterinary Clinical Pathology ISSN 0275-6382

all tissues were submitted for routine histopathologic

examination. Gross examination of the surgical speci-

men revealed a solitary, hard, roughly round, white to

tanmass about 9 mm in diameter within the right limb

of the pancreas. The second nodule identified via ultra-

sound was not found on gross examination of the sur-

gical specimen despite complete sectioning of the

tissue. Microscopically, approximately 50% of the

mass was composed of densely arranged packets of

monomorphic polygonal cells interspersed by a deli-

cate fibrovascular network. The mass was partially

delineated by a thick fibrous capsule, with nodular

aggregates of neoplastic cells that infiltrated the cap-

sule and the surrounding parenchyma. The architec-

ture of the lymph node had been entirely effaced by

similar cells. They had ample, finely granular eosino-

philic cytoplasm, with basilar to centrally located

round nuclei, which had clumped chromatin, and a

single centrally located nucleolus. There was mild-

to-moderate anisocytosis and anisokaryosis with

occasional binucleated cells. Mitotic figures were rare

(< 1 per 10 high-power fields). Embedded within the

neoplasm and composing the remainder of the mass,

there was a single nodule of broad and irregularly

arranged, anastomosing trabeculae of mineralized

compact bone. Bony trabeculae were lined by a single

layer of osteoblasts, and the matrix contained loosely

arranged osteoblasts within lacunae (Figures 2 and 3).

Numerous neoplastic cells and pigment-laden macro-

phages (hemosiderophages) were scattered between

the trabeculae.

Immunohistochemistry of the neoplasm was per-

formed by the avidin-biotin-peroxidase method (ABC

elite reagent; Vector Laboratories, Burlingame, CA,

USA). Primary antibodies included synaptophysin

(1:100; Dako, Glostrup, Denmark), chromogranin A

(1:400; Dako), and insulin (1:50; Invitrogen, Grand

Island, NY, USA). Diaminobenzidine was used as the

chromogen; slides were counterstained with Harris’

hematoxylin. Neoplastic cells showed strong intracel-

lular staining with synaptophysin immunohistochem-

istry (not shown), supportive of the diagnosis of

neuroendocrine carcinoma. Subsequent staining of

the neoplastic cells with insulin was also strongly posi-

tive, compatible with a diagnosis of an insulinoma

(Figure 4). The chromogranin A immunohistochemis-

try was not interpretable due to excessive background

staining.

The dog recovered well following surgery. At the

time of discharge, the tetraparesis had significantly

improved and blood glucose was stable. Within one

month of discharge postoperatively, the dog was re-

presented for increased liver enzymes, tetraparesis,

and respiratory distress. Fluid was noted in the abdo-

men, and cytology of an abdominal fluid sample

showed neutrophilic inflammation with clusters of

malignant neuroendocrine epithelial cells consistent

with an exudate accompanied by a neoplastic effusion.

Based on cell morphology, the neuroendocrine cells

were most consistent with the previously diagnosed

Figure 2. Histologic section of the right limb of the pancreas in a dog.

The parenchyma is disrupted by an expansile, partially encapsulated and

mildly infiltrative, multilobulated mass that consists of densely arranged,

monomorphic polygonal cells. Approximately 50% of the mass is com-

posed of metaplastic, mature trabecular bone. H&E. Bar = 5 mm.

Figure 3. Histologic section of bone within an islet cell carcinoma in

the pancreas of a dog. Monomorphic polygonal cells are interspersed

by delicate fibrovascular stroma, typical of neuroendocrine packeting.

Cells have moderate amounts of finely granular eosinophilic cyto-

plasm, basilar or centrally located round nuclei, clumped chromatin,

and a single prominent nucleolus. Interspersed amongst the neoplas-

tic cells are broad and irregularly arranged, anastomosing trabeculae

of mineralized compact bone containing loosely set osteoblasts within

lacunae. H&E. Bar = 100 lm.

Vet Clin Pathol 43/1 (2014) 89–93©2014 American Society for Veterinary Clinical Pathology and European Society for Veterinary Clinical Pathology90

Osseous metaplasia within a canine insulinoma Pieczarka et al

islet cell carcinoma. However, per owner request, no

further testing was performed to evaluate the extent of

metastasis. Due to the grave prognosis, the dog was

humanely euthanized. A necropsy was declined.

Discussion

Insulin-secreting beta-cell neoplasia (insulinoma) is

the most common tumor arising from pancreatic islets

in the dog.1 The majority of canine insulinomas are

malignant solitary masses, predominantly identified in

the duodenal (right) limb of the pancreas.2–4 Beta-cell

adenomas occur less frequently than carcinomas, but

both tumors may cause significant endocrine distur-

bances and clinical signs through oversecretion of

insulin.5–7 Insulinomas have been found in dogs from

3–15 years of age, with a mean of 9 years.8 There is no

apparent sex predilection and no breed predisposition

has been reported; however, most cases involve

medium- to large-breed dogs.8 Islet cell adenomas

typically occur as small (1–3 cm), single or less often

multiple nodules that are sharply delineated and sur-

rounded by a partial or complete thin, fibrous capsule.

Islet cell carcinomas are usually larger than adenomas,

are multilobular, invade adjacent pancreatic paren-

chyma, and may metastasize. Histologic evidence of

capsular invasion is considered a critical diagnostic

feature of malignancy.9

The clinical signs of tetraparesis and clinicopatho-

logic findings of persistent hypoglycemia, mild hypo-

kalemia, and mildly increased serum ALP activity in

the current case are consistent with previous reports of

dogs with insulin-secreting islet cell tumors.2–4 In

healthy dogs, hypoglycemia of less than 80 mg/dL is

corrected through complete inhibition of insulin

secretion, while counterregulatory hormones such as

glucagon, catecholamines, growth hormone, and

glucocorticoids are increased.8 However, neoplastic

beta cells secrete insulin independent of blood glucose

concentration, resulting in persistent and unregulated

hypoglycemia accompanied by normal or increased

blood insulin levels in most cases. As a consequence,

hypoglycemia of the central nervous system or neuro-

glyocopenia leads to the clinical signs of tetraparesis.8

It is well-recognized that hyperinsulinism induces

hypokalemia via intracellular potassium shifting, inde-

pendent of glucose transport.10 Although serum insu-

lin was not measured in the current case, the mild

hypokalemia is likely the result of hyperinsulinemia

due to unregulated insulin secretion from tumor cells.

The mechanism of increased ALP in dogs with insuli-

nomas has not been clearly defined, but may be associ-

ated with increased secretion of glucocorticoids in

response to the hypoglycemia, and/or chronic stress.

This explanation is supported in the current case

because the majority of increased serum ALP activity

was attributable to the corticosteroid-induced ALP iso-

enzyme. Although bone ALP has been linked to

increased osteoblastic activity in tumors such as osteo-

sarcoma,11 the amount of bone formation in the cur-

rent case was likely insufficient to appreciably

contribute to total ALP activity. In a study comparing

dogs with mammary tumors with and without the

presence of osseous transformation, there was no

significant difference in bone ALP activity between the

2 groups.12 Furthermore, when the 2 groups were

independently compared to a control population, bone

ALP activity was significantly increased only in the

group with mammary tumors lacking bone, suggesting

that small amounts of bone formation in tumors may

not significantly affect measured serum activity of

bone ALP.

Intratumoral bone can be found in a variety of epi-

thelial and mesenchymal neoplasms other than those

primarily derived from osteoblasts. In people, mature

bone has been reported in a number of extra-osseous

neoplasms. In veterinary species, bone is commonly

identified in fibromas derived from the periodontal lig-

ament, maxillary fibromas of young horses, canine

mixed mammary tumors, and in some chondrosarco-

mas.13–16 Less frequently, bone has been described

within canine mixed thyroid tumors, mixed apocrine

sweat gland tumors, gastrointestinal adenocarcinomas,

oral malignant melanomas, and a pancreatic exocrine

adenocarcinoma in a ferret.17–22 The current case

Figure 4. Histologic section of an islet cell carcinoma in a dog. Strong

positive staining for insulin within neoplastic cells, consistent with a diag-

nosis of insulinoma. Diaminobenzidine chromogen, Harris’ Hematoxylin

counterstain. Bar = 25 lm.


Pieczarka et al Osseous metaplasia within a canine insulinoma

represents the first report of bone formation within an

insulinoma (islet cell carcinoma) in animals or people.

Intratumoral bone formation can originate from

2 basic mechanisms, either osseous metaplasia or

endochondral ossification. In osseous metaplasia,

nonosteoblastic cells are directly transformed into os-

teoblasts. However, identifying the cell of origin

responsible for metaplastic bone can be challenging.

In canine mammary tumors, for example, it is still

unclear whether bone forms from metaplasia of

epithelial cells, stromal connective tissue, or basal/

myoepithelial cells.23,24 In endochondral ossification,

ground substance is secreted and converted into

hyaline cartilage, which subsequently undergoes

endochondral ossification. For instance, myoepitheli-

al cells in benign or malignant mammary tumors can

secrete a mucinous ground substance that converts

to cartilage, and then undergoes endochondral ossi-

fication.16 As there was no evidence of ground

substance or cartilage in the current case, it was con-

cluded that the bone formed via osseous metaplasia.

Acknowledgments

Special thanks to Drs. Bridget Urie and Andrew Mercurio

for their clinical management and follow-up with the case.

The authors also thankMarc Hardman for providing techni-

cal assistance with the images.

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osseous metaplasia within a canine insulinoma

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