History

A captive 12.5-year-old 35.7-kg (78.5-lb) hemicastrated Mexican gray wolf (Canis lupus baileyi) had a 2.5-month history of right carpal swelling associated with mild lameness. The swollen area was nonulcerated, firm, and nonmobile and abutted the bone. Radiography revealed a locally extensive soft tissue opacity subtended by periosteal reaction and slight radio-lucency of the distal portion of the radius. Thoracic radiography revealed widespread pulmonary nodular radiopacities. Abdominal ultrasonography detected an 8-cm-diameter, cavitated mass with soft tissue opacity in the right caudal portion of the abdomen. The wolf was euthanized via IV administration of pentobarbital, and the carcass was submitted for postmortem examination. Nine months earlier, the wolf had undergone surgery to correct a 360° gastric torsion, during which a splenectomy was performed. Eighteen months earlier, the wolf underwent unilateral orchiectomy because of a right testicular mass that was diagnosed as a Sertoli cell tumor. Results of hematologic assessments prior to the hemicastration and euthanasia were unremarkable. Serum samples had been collected at various time points and archived.

Gross Findings

At the proximal dorsomedial aspect of the right carpus, there was a locally extensive, firm, spherical, 7.5 × 6.5 × 5.5-cm mass that abutted the joint, expanded the subcutis, and raised the nonulcerated skin (Figure 1). On section, the mass was tan to light brown to red and composed of cavitated areas filled with tan soft material separated by coalescing firm tan to light brown tissue. The mass did not extend from bone, but areas adjacent to the radial cortex contained hard spicules (periosteal reaction). Forty percent of the lung lobes had widespread, randomly distributed, multifocal to coalescing, raised, well-demarcated, firm, white to tan masses with a diameter of up to 3 cm. Adjacent parenchyma was variably atelectatic and congested. In the caudodorsal portion of the abdomen, there was a smooth, nodular, soft to firm, 6.5- to 7-cm-diameter mass adhered to the parietal peritoneum of the sublumbar region. The mass did not involve the surrounding viscera, although sublumbar lymph nodes were obliterated. On section, the mass was friable and had multiple white to tan firm hemorrhagic nodules among frequent cavities filled with opaque, pink to red to brown, thick material. The right ureter was focally adhered to the outer aspect of the sublumbar mass without obstruction. One pole of the left adrenal gland was disrupted and enlarged by a firm, 2 × 2 × 1-cm, well-demarcated mass. The wolf had atrophy of the remaining left testicle, a small symmetric prostate gland, a ventral lingual mass, a urinary bladder mural mass, and mild pancreatic hemorrhage.

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Photographs of a Mexican gray wolf (Canis lupus baileyi) evaluated for a swelling of the proximal portion of the right carpus causing mild lameness of 2.5 months’ duration. A—The right carpal mass is firm, prominent, and raises intact skin. B—The right carpal mass occupies the subcutis and is well-demarcated, tan, and firm. C—In the right side of the thorax, the lungs have widespread, multifocal to coalescing, tan, firm masses (asterisks) with fibrous adhesions and adjacent variable atelectasis and congestion. Inset—View of a cut section of a lung mass.

Citation: Journal of the American Veterinary Medical Association 257, 6; 10.2460/javma.257.6.599

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

Histopathologic and Immunohistochemical Findings

Samples of the masses from the right carpus, the lungs, the left adrenal gland, and the sublumbar region were histologically similar. Tissue architecture was replaced by a poorly demarcated, unencapsulated, moderately cellular, and infiltrative neuroendocrine-like tumor. The cuboidal to pseudostratified columnar tumor cells were arranged in trabeculae and clusters of palisading serpentine bands supported by a thin extensive fibrovascular network or in some areas supported by thick bands of fibrous connective tissue. Occasionally, tumor cells were arranged as tubules surrounding a lumen, which did or did not contain blood and sloughed cells (Figure 2). For each cell, the borders were variably distinct with moderate granular eosinophilic cytoplasm and a round to oblong nucleus with coarse chromatin and multiple magenta nucleoli. There was moderate anisokaryosis, moderate anisocytosis, mild nuclear pleomorphism, and bizarre mitoses. There were > 50 mitotic figures in 10 hpf (400X). In sections of the sublumbar mass, ≤ 50% of the tissue was replaced by necrosis with fibrin and acute and chronic hemorrhage. There was frequent vascular invasion. Neoplastic cells in the lung tissue were strongly vimentin positive, strongly inhibin (INH-α) positive, multifocally neuron-specific enolase (NSE) positive, cytokeratin negative, and melan-A negative. The remaining testis had diffuse atrophy of the seminiferous tubules with fibrosis of the tunica albuginea but no tumor was present. The prostate had a preserved architecture with glandular atrophy and no squamous metaplasia. The lingual mass was a fibrolipoma. The urinary bladder mass was a leiomyoma.

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Photomicrographs of sections of affected lung tissue from the wolf in Figure 1. A—Tissue architecture has been replaced by a poorly demarcated, unencapsulated, moderately cellular, and infiltrative neuroendocrine-like tumor. The tumor has tubules, clusters, and trabeculae composed of cuboidal to columnar pseudostratified cells that form palisading serpentine bands supported by a thin extensive fibrovascular network. H&E stain; bar = 40 μm. Inset B—Neoplastic cells have strong cytoplasmic immunoreactivity to inhibin-α. Immunohistochemical stain for inhibin-α; bar = 80 μm. Inset C—Following immunohistochemical staining for neuron-specific enolase, moderate cytoplasmic immunoreactivity of the neoplastic cells is evident. Immunohistochemical stain for neuron-specific enolase; bar = 80 μm.

Citation: Journal of the American Veterinary Medical Association 257, 6; 10.2460/javma.257.6.599

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Results of Serum Sample Analysis

Serum samples obtained from the wolf when it was healthy (8 years old), at the time of unilateral orchiectomy, and at 8, 8.5, and 18 months after unilateral orchiectomy had been opportunistically collected and archived. Following the euthanasia and postmortem examination of the wolf, these samples underwent hormone analysis for anti-Müllerian hormone (AMH), testosterone, estradiol, and progesterone concentrations (Table 1). The notable finding was persistently high serum AMH concentration at and since the time of the unilateral orchiectomy.

Morphologic Diagnosis and Case Summary

Morphologic diagnosis and case summary: malignant Sertoli cell tumor in the subcutis of the proximal region of the right carpus, lungs, left adrenal gland, and sublumbar region in a 12.5-year-old hemicastrated Mexican gray wolf.

Comments

Eighteen months prior to euthanasia of the wolf, a primary Sertoli cell tumor had been identified in the animal's right testicle. At the time of unilateral orchiectomy, the tumor had likely already metastasized, at least to the sublumbar lymph nodes; thereafter, it most likely disseminated to the lungs, the left adrenal gland, and the proximal region of the right carpal subcutis. Sertoli cell tumor metastases to the liver, lungs, kidneys, spleen, and adrenal glands in a dog have been described¹; however, metastasis to the subcutis of the proximal region of the right carpus is unusual. The wolf's carpal mass and limping alerted clinicians to the possibility of a more systemic problem, and subsequent thoracic radiographic and abdominal ultrasonographic examinations revealed probable disseminated neoplasia that warranted euthanasia. For the wolf of the present report, a diagnosis of metastatic Sertoli cell tumor was supported by the histopathologic and immunohistochemical findings and results of the postmortem hormone analysis of banked serum samples, which revealed persistently high AMH concentration.

Common canine testicular tumors include seminoma, interstitial or Leydig cell tumors, and Sertoli cell tumors. These tumors are often benign and can be distinguished on the basis of their macroscopic appearance, microscopic features, and immunohistochemical profile.² Malignant forms may develop, albeit infrequently, and malignancy is best supported by documented metastasis rather than histologic appearance alone.² Macroscopic differentiation of testicular tumors relies on color and texture of the neoplasms. Seminomas are tan or pink-gray throughout, interstitial cell tumors are typically soft and tan to orange with multifocal hemorrhage, and Sertoli cell tumors are firm, tan, and lobulated by fibrosis.³ Microscopically, these tumors have histologic features that usually allow a definitive diagnosis to be made. Seminomas form intratubular or diffuse sheets of large round cells, each with scant cytoplasm and a large nucleus; the sheets of large round cells are often infiltrated by lymphocytes. Interstitial cell tumors have cells arranged in sheets with microvesiculated, lipid-filled cytoplasm that often contains lipofuscin. Sertoli cell tumors are composed of either normal Sertoli cells or pleomorphic pseudostratified cell populations that tend to palisade along fibrous stroma or form tubular structures.^3,4 The histologic appearance of a pseudostratified tubular arrangement described as palisading neoplastic cells along a thick basement membrane is sufficient to classify a testicular tumor as a Sertoli cell tumor.⁵ When the diagnosis of a testicular tumor is equivocal or when there is a suspicion of a testicular tumor metastasis, immunohistochemical analyses can help determine the testicular cell of origin, such as in the case described in the present report. Several immunohistochemical markers for testicular tumors are described, with variable labeling patterns.² A suggested marker panel to distinguish among the 3 most common canine testicular tumors usually consists of immunohistochemical stains for INH-α (positive results for Sertoli cell tumors and interstitial cell tumors), KIT (positive results for germ cell tumors), E-cadherin (positive results for Sertoli cell tumors), GATA-4 (positive results for Sertoli cell tumors), NSE (positive results for Sertoli cell tumors), and PGP 9.5 (positive results for germ cell tumors).⁶ Vimentin is usually expressed in all testicular tumor cell types. Seminomas are negative for cytokeratin, whereas normal Sertoli cells are negative for cytokeratin with reports of focal positivity in Sertoli cell tumors.⁶ Labeling for melan-A in Sertoli cell tumors is inconsistent with preferential labeling of interstitial cells.^2,6 Immunohistochemical analysis for AMH is another useful option that may allow diagnosis of a Sertoli cell tumor,^2,7 although this was not pursued in the case described in the present report. However, on the basis of combined assessment of the patient's clinical signs and the tumor's macroscopic and microscopic features, undertaking such an extensive immunohistochemical profile may not be required. For the wolf of the present report, the immunohistochemical profile used on the lung mass was guided by assessment of published data and the availability and cost of the analytic components; the findings indicated that the mass was vimentin positive, INH-α positive, NSE positive, cytokeratin negative, and melan-A negative, which supported the histologic diagnosis of Sertoli cell tumor. Sertoli cell tumors are common in older dogs and are usually unilateral, and their incidence increases in cryptorchid individuals.² Sertoli cell tumor metastasis typically develops in cryptorchid dogs, but abdominal infiltration may also occur with intrascrotal tumors.¹ Rarely, Sertoli cell tumors develop in stallions, rams, tomcats, and bulls.²

Sertoli cell tumors can also be diagnosed on the basis of serologic detection of AMH. Anti-Müllerian hormone has a role in the early stages of fetal male genital tract organogenesis, whereby it induces the regression of the Müllerian ducts.⁷ Anti-Müllerian hormone is expressed in the Sertoli cells of canine fetuses, neonates, and puppies < 45 days old and in Sertoli cell tumors, but is not evident in other testicular cells, including Leydig cells and spermatogonia.⁷ In normal human and dog testes, the expression of AMH is typically highest during the period of testicular differentiation until puberty and decreases to low or undetectable amounts in adulthood.⁷ However, the re-expression of AMH by Sertoli cells occurs in dogs and humans that develop Sertoli cell tumors.^7,8 In humans, the level of AMH expression in Sertoli cell tumors is less than the level detected in fetal testes during testicular differentiation but greater than the level detected in testes in adults.⁸ For the wolf of the present report, the serum AMH concentration at the time of unilateral orchiectomy was approximately 177 times that of when it was a healthy adult, which was consistent with the diagnosis of a testicular Sertoli cell tumor. Following the presumably curative removal of the primary testicular Sertoli cell tumor, a progressive decrease in serum AMH would have been expected. In calves, the decrease in circulating AMH concentration should follow first-order kinetics, with an approximate half-life of AMH of 48 hours.⁹ On the basis of this rate of elimination, the wolf's serum AMH concentration should have approached 0 ng/mL within 30 days after tumor removal. However, the wolf's serum AMH concentration remained high. At 18 months after surgery, the serum AMH concentration was > 6,000 ng/mL, suggesting persistence of tumor tissue likely as a result of metastasis prior to tumor removal.

Sertoli cell tumors may precipitate a feminization syndrome as a result of overproduction of estrogen. In affected males, hyperestrogenemia may cause a combination of symmetric alopecia, hyperpigmentation, gynecomastia, and galactorrhea and result in attraction of other males.^10,11 High estrogen concentration may induce structural effects such as prostatic squamous metaplasia and myelotoxicosis.^11,12 For the wolf of the present report, the clinical signs, results of hematologic assessments prior to euthanasia, and findings of histologic examination of the prostate did not support the presence of an estrogen-producing Sertoli cell tumor. This was corroborated by the results of the serum estradiol analysis at the time of euthanasia, which was below the upper limit of the reference interval established for male domestic dogs. Meanwhile, serum testosterone concentration was < 0.01 ng/mL at the time of euthanasia, consistent with left-sided testicular atrophy.