Pathology in Practice

Kristina O. Goncarovs-Gran Department of Veterinary Clinical Sciences, School of Veterinary Medicine, Purdue University, West Lafayette, IN 47907.

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 DVM, MS, DACVIM
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Chad B. Frank Department of Comparative Pathobiology, School of Veterinary Medicine, Purdue University, West Lafayette, IN 47907.

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Aubrey N. Baird Department of Veterinary Clinical Sciences, School of Veterinary Medicine, Purdue University, West Lafayette, IN 47907.

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Laurent L. Couetil Department of Veterinary Clinical Sciences, School of Veterinary Medicine, Purdue University, West Lafayette, IN 47907.

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Jose A. Ramos-Vara Department of Comparative Pathobiology, School of Veterinary Medicine, Purdue University, West Lafayette, IN 47907.

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History

A 2.5-year-old 62.6-kg (137.7-lb) pregnant female Huacaya alpaca was evaluated because of a mass associated with its left shoulder region. The mass was first detected 1 month prior and had doubled in size in the intervening period. The owners had not noted it to cause signs of pain, cause lameness, or affect the alpaca's eating or locomotion. There was no known traumatic event preceding the appearance of the mass. No other animals on the farm had similar lesions currently or in the past. The alpaca was reported to be in its 10th month of gestation, as determined on the basis of its last breeding date.

Diagnostic test results were unremarkable except for mild hyperfibrinogenemia consistent with mild systemic inflammation. Diagnostic testing was performed again 3 months later, following an uneventful parturition and weaning of the cria. At that time, the alpaca had lost 27% of its previous body weight and weighed 45.8 kg (100.8 lb), and the increasing size of the mass was interfering with its ability to cush and ambulate. The alpaca was euthanized and underwent necropsy.

Clinical and Gross Findings

At the final evaluation prior to euthanasia, a large irregularly shaped, firm mass was present medial to the left scapula (Figure 1). The mass measured 37 × 37 cm and displaced the scapula laterally approximately 7 cm. Ultrasonography of the mass (performed at the initial evaluation) was suggestive of fluid with regions of unaffected soft tissue interspersed throughout. Normal borders of the left scapula, thoracic vertebrae, and ribs were visualized. Gram staining and cytologic evaluation of a fine-needle aspirate revealed many round, spindle-shaped, or stellate neoplastic cells with moderate to marked anisocytosis and anisokaryosis. These cells had discrete borders with a large amount of basophilic cytoplasm and few to many intracytoplasmic punctate vacuoles. The nuclei were round to ovoid with variably numbered and shaped nucleoli. Normal and atypical mitotic figures were infrequently observed.

Figure 1—
Figure 1—

Photographs of the left side of a 2.75-year-old female Huacaya alpaca evaluated because of a mass associated with its left shoulder region prior to euthanasia (A) and the subcutaneous mass removed from the shoulder region at necropsy (B). The mass had been detected initially 4 months earlier but had expanded to 37 × 37 × 7 cm and inhibited locomotion. In panel A, the spine and dorsocaudal edge of the scapula are marked with black lines. At necropsy, the shoulder mass was cut to reveal firm, tan tissue with areas of cavitation, hemorrhage, and necrosis.

Citation: Journal of the American Veterinary Medical Association 243, 8; 10.2460/javma.243.8.1113

At necropsy, the subcutaneous mass measured 38.5 × 37 cm and displaced the left scapula laterally by 7.6 cm. It extended from the neck of the scapula caudally to the ninth rib and dorsally to the spinous processes of the thoracic vertebrae. The mass did not infiltrate any osseous structures. It was pale tan to white, varied in consistency from firm to soft and fluctuant, and was well demarcated and easily removed from adjacent tissues. On cut section (Figure 1), the mass was composed of slightly firm tan to white tissue that surrounded coalescing foci of soft and friable dark red to brown necrotic parenchyma.

Additionally, the cortex of a cranial mediastinal lymph node contained a 1.5-cm-diameter, well-demarcated, pale tan to white, slightly firm, smooth mass. There were 3 spherical to irregularly shaped, tan to white, slightly firm nodules that ranged in size from 1.7 × 2.2 cm to 3.0 × 2.7 cm along the dorsal aspect of the fourth, seventh, and eighth intercostal spaces.

Formulate differential diagnoses from the history, clinical findings, and Figure 1—then turn the page

Histopathologic Findings

Sections of the left shoulder mass and cranial mediastinal and intercostal lymph nodes were examined microscopically. Histologically, the mass was highly cellular, locally invasive, and nonencapsulated. It was composed of sheets of round to polygonal or spindloid cells admixed with scant collagenous stroma. Neoplastic cells had moderate to abundant pale eosinophilic cytoplasm with distinct cell margins and ovoid to irregularly shaped eccentric nuclei with several prominent amphophilic nucleoli. Cellular pleomorphism was marked. Many cells contained hyaline cytoplasmic inclusions. The mitotic index was 61 figures/ten 400×fields. Neoplastic cells focally infiltrated and partially effaced the surrounding fascia. Coalescing foci of necrosis were scattered throughout the mass tissue. Cross-striations were not evident in sections stained with H&E (Figure 2) or phosphotungstic acid–hematoxylin stain. The lymphoid parenchyma of the cranial mediastinal and intercostal lymph nodes was completely or partially effaced by sheets of pleomorphic neoplastic round to spindloid cells similar to the ones associated with the left shoulder mass, confirming metastasis.

Figure 2—
Figure 2—

Photomicrograph of a section of the mass in the left shoulder region of the alpaca in Figure 1. Notice the marked nuclear atypia and mitotic figures (arrows). H&E stain; bar = 50 μm.

Citation: Journal of the American Veterinary Medical Association 243, 8; 10.2460/javma.243.8.1113

Immunohistochemical analysis of mass tissue sections was performed to further classify the neoplasm. Immunohistochemical staining for cytokeratins,a vimentin,b desmin,c actin (a broad-range marker for all types of muscle or a marker for smooth muscle),d myoglobin (rabbit polyclonale), MyoD1,f CD3,g CD79a,h lysozymes,i and CD68j was performed. Tissue control samples (tissue samples collected from dogs during biopsy procedures or obtained from canine cadavers during necropsies) were similarly processed; known positive internal controls present in the alpaca tissues examined were also used, except for staining for MyoD1 for which only a canine control was used. Neoplastic cells were positive for vimentin (broad-range mesenchymal marker), desmin, actin (the broad-range muscle marker), and myoglobin and MyoD1 (striated muscle markers). Labeling was cytoplasmic, except for MyoD1 labeling, which had a nuclear expression. No uptake was observed with epithelial markers (cytokeratin), leukocytic markers (CD3, CD79a, lysozyme, and CD68), or the smooth muscle actin marker (a smooth muscle, myoepithelium, and myofibroblast marker). Rhabdomyosarcomas are derived from striated muscle, and tumor tissues are typically not positive for smooth muscle actin; on the basis of the cellular phenotype of this tumor and immunohistochemical results, a diagnosis of embryonal rhabdomyosarcoma was reached.

Morphologic Diagnosis and Case Summary

Morphologic diagnosis: embryonal rhabdomyosarcoma with metastases to the cranial mediastinal and intercostal lymph nodes.

Case summary: embryonal rhabdomyosarcoma in the shoulder region of an alpaca.

Comments

Rhabdomyosarcoma is a type of soft tissue sarcoma rarely encountered in veterinary medicine. This case is the first report of this tumor type in a camelid species, to our knowledge. Soft tissue sarcomas are neoplasms that develop from mesenchymal tissue and are named according to the cell type of origin. Rhabdomyosarcomas are soft tissue sarcomas originating from skeletal muscle cell progenitors.1 Immunohistochemical staining is usually necessary for accurate diagnosis of these tumors. Specifically, most rhabdomyosarcomas express vimentin, skeletal muscle actin, myoglobin, and MyoD1.2,3 Rhabdomyosarcomas can be classified further histologically as embryonal, alveolar, pleomorphic, or botryoid, all of which have been described in veterinary medicine.4 Embryonal rhabdomyosarcoma and the botryoid variant are histologically characterized by sheets of round (myoblast-like) to spindle (myotube-like) cells, which are often embedded within a mucinous stroma. In contrast, alveolar rhabdomyosarcomas are composed of small round cells arranged in clusters or aggregates separated by fibrous tissue. Pleomorphic rhabdomyosarcomas lack the features of the other subtypes and are composed of haphazardly arranged pleomorphic spindle cells.

Rhabdomyosarcomas in humans are relatively well described; however, these neoplasms are sufficiently rare in veterinary medicine that the literature is limited to individual case reports and case series. Thus, it is difficult to draw scientific conclusions regarding the prevalence of rhabdomyosarcomas in domestic animals. In a recent retrospective analysis5 of neoplasia in camelids, rhabdomyosarcoma was not identified in any of 551 biopsy and necropsy samples examined over a 5-year period. Likewise, in a 10-year survey of equine tumors published in 1982, no rhabdomyosarcomas were reported.6 In a study7 in 1975 in which 244 neoplasms involving the skin in horses were evaluated, there was only 1 case of rhabdomyosarcoma. Descriptions of rhabdomyosarcomas in horses consist of 6 individual case reports8–13 and 1 case series of 8 animals.14 Six rhabdomyosarcomas in cattle have been reported.4,15–19

Rhabdomyosarcomas have been identified in the abdominal wall,14 tongue,10,14 shoulder region,9,14 masseter muscle,14 buccal muscles,14 heart,14 urinary bladder,11 uterus,12 hind limbs,8 and deep digital flexor tendon13 in horses. In cattle, rhabdomyosarcomas involving the abdominal wall,15 brisket,16 pleura,4 thoracic wall,18 perifemoral soft tissue,17 and heart19 have been reported. The location of the mass between the thorax and scapula in the case described in this report is therefore not particularly unusual.

In humans, soft tissue sarcomas, including rhabdomyosarcomas, are generally highly metastatic with a predilection for the lymph nodes, lungs, and bone marrow. Masses located on the extremities have a higher rate of metastasis than masses located elsewhere.20 Of the 14 equine cases of rhabdomyosarcoma, 4 involved metastases; the primary tumors involved the hind limbs,8 deep digital flexor tendon,13 and urinary bladder.11 Only 2 of the 6 bovine cases involved metastases; the primary tumors involved the abdominal wall15 and brisket.16 In both species, metastases were detected in the liver, lungs, kidneys, adrenal glands, uterus, and lymph nodes. Lymph node metastases were also evident in the case described in the present report.

Histologically, the rhabdomyosarcoma in the alpaca of the present report was similar to those previously described. Immunohistochemical analysis is necessary for a definitive diagnosis because features such as nonuniform texture and color of surfaces, marked cellular pleomorphism, high mitotic index, and local invasion of surrounding tissues are not specific to this soft tissue tumor. Previous reports4,7–19 of rhabdomyosarcomas in horses and cattle consistently used MyoD1 or myogenin labeling for confirmation of muscle differentiation and myoglobin and MyoD1 for confirmation of striated muscle differentiation, as was done in this case.

Although it is difficult to draw any conclusions regarding the biological behavior of rhabdomyosarcoma in domestic animals from the available literature, several observations can be made. Most of the described cases, including that of the alpaca of this report, were evaluated because of a mass that had expanded dramatically over a period of just a few months. Most affected animals either died or were euthanized because of poor prognosis, but several patients survived long term following complete excision.14 However, this approach may not be feasible in many cases, including the alpaca of this report, because of the location and size of the mass or anatomic structures involved. Additionally, one must take into consideration the likelihood of metastases and economic value of the animal when choosing such a radical treatment.

a.

Dako M0821, monoclonal mouse anti–human cytokeratin antibody, DAKO Denmark A/S, Glostrup, Denmark.

b.

Dako M7020, monoclonal mouse anti–human vimentin antibody, DAKO Denmark A/S, Glostrup, Denmark.

c.

Dako M0760, monoclonal mouse anti–human desmin antibody, DAKO Denmark A/S, Glostrup, Denmark.

d.

Dako M0635, monoclonal mouse anti–human actin antibody, DAKO Denmark A/S, Glostrup, Denmark.

e.

Dako A0324, polyclonal rabbit anti–human myoglobin antibody, DAKO Denmark A/S, Glostrup, Denmark.

f.

Dako M3512, monoclonal mouse anti–MyoD1 antibody, DAKO Denmark A/S, Glostrup, Denmark.

g.

Dako A0452, polyclonal rabbit anti–human CD3 antibody, DAKO Denmark A/S, Glostrup, Denmark.

h.

Dako M7051, monoclonal mouse anti–CD79α antibody, DAKO Denmark A/S, Glostrup, Denmark.

i.

Dako A0099, polyclonal rabbit anti–human lysozymes antibody, DAKO Denmark A/S, Glostrup, Denmark.

j.

Dako M0718, monoclonal mouse anti–human CD68 antibody, DAKO Denmark A/S, Glostrup, Denmark.

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