Cytologic and immunohistochemical characterization of a primitive neuroectodermal tumor in the brain of a dog

A 6-year-old intact female Pointer dog was presented for evaluation of acute onset of ataxia, circling, and head tilt. Neurologic assessment revealed overall decreased postural reaction, left-sided hemiparesis with incoordination, rigidity of fore- and hindlimbs, strabismus of the right eye, and bilateral horizontal nystagmus. Using magnetic resonance imaging, a mass lesion was identified in the cerebrum adjacent to the left side of the cerebellum compressing the brain stem ventrally. The mass was incompletely resected, and during surgery fine-needle aspiration and biopsy of the mass were performed. Cytologically, smears were highly cellular and contained predominantly small to medium-sized discrete round cells with high nuclear to cytoplasmic ratios and round nuclei with rare deep clefts or indentation, smooth chromatin, and indistinct nucleoli. Numerous cytoplasmic fragments were noted in the background. The primary diagnosis was lymphoma; other differential diagnoses included neuroendocrine tumor and poorly differentiated tumor of neural origin. The histologic diagnosis was lymphoma, and the lesion was presumed to be metastatic. On immunohistochemical analysis, the cells expressed neither CD3 nor CD79a. Re-examination of the histologic section revealed disorganized sheets of cells with multifocal palisading and perivascular arrangements of rosette-like structures. An expanded panel of antibodies to vimentin, cytokeratin, glial fibrillary acid protein (GFAP), neuron-specific enolase (NSE), synaptophysin (SYN), S-100, and CD45 was applied to histologic sections. Neoplastic cells were immunoreactive for vimentin, NSE, and S-100. Based on the histologic appearance and immunophenotype of the tumor, a diagnosis of primitive neuroectodermal tumor (PNET) was made. PNET, although rare in dogs, should be considered as a differential diagnosis for round cell tumors in the brain.     

Immunohistochemical and histochemical stains for differentiating canine cutaneous round cell tumors

Immunohistochemical and histochemical stains are useful adjunct techniques in the diagnosis of canine cutaneous round cell tumors, which can appear histologically similar. We applied a panel of monoclonal antibodies (recognizing tryptase, chymase, serotonin for mast cells; CD1a, CD18, MHC class II for histiocytes; CD3 for T lymphocytes; CD79a for B lymphocytes and plasma cells) and one histochemical stain (naphthol AS-D chloroacetate for chymase activity) to formalin-fixed, paraffin-embedded sections of canine cutaneous mast cell tumors, histiocytomas, lymphosarcomas, plasmacytomas, and unidentified round cell tumors. Of 21 tumors with a histologic diagnosis of mast cell tumor, 7/7 (100%) grade I, 6/7 (85.7%) grade II, and 3/7 (42.9%) grade III tumors were diagnosed as mast cell tumors based on positive staining for tryptase antigen and chymase activity. Mast cells were positive for both tryptase antigen and chymase activity, indicating equal efficacy of tryptase immunohistochemistry and chymase histochemistry. Chymase was detected immunohistochemically in both tumor and nontumor cells, while serotonin was not detected in most mast cell tumors, and thus, neither was useful in the diagnosis of mast cell tumors. Immunohistochemistry to detect CD18 and MHC class II was equally effective in staining histiocytomas, although lymphosarcoma must be ruled out through the use of CD3 and CD79a immunohistochemistry. Immunohistochemistry using three different monoclonal antibodies to human CD1a showed no cross-reactivity in canine histiocytomas and was not useful. A final diagnosis was obtained for 4/5 (80%) of the unidentified tumors, indicating the usefulness of multiple stains in poorly differentiated round cell tumors.     

Cytologic,histologic, and immunohistochemical features of maxillofacial alveolar rhabdomyosarcoma in a juvenile dog

Abstract: A 15‐month‐old castrated male dog with a history of intermittent epistaxis and sneezing was admitted for the examination of a maxillofacial mass. An impression smear of a biopsy sample from the cauliflower‐shaped gingival mass contained numerous round cells, 5–25 μm in diameter, which contained a moderate amount of clear to pale blue cytoplasm and resembled lymphoid cells. Mitotic figures were frequently observed. The mass was diagnosed as malignant round cell neoplasia. On histologic examination the tumor was composed of diffusely arranged, small, atypical round cells with a small amount of fibrovascular stroma. Immunohistochemically, the cells were negative for CD3, CD18, CD20, CD79α, cytokeratin, melan‐A, chromogranin A, α‐smooth muscle actin, and myoglobin but positive for vimentin and desmin. The cells also had strong positive nuclear staining for myogenin and MyoD1. A diagnosis of solid‐pattern alveolar rhabdomyosarcoma was made on the basis of morphologic and immunohistochemical results. Alveolar rhabdomyosarcoma should be considered in the differential diagnosis of tumors in juvenile dogs, especially when cytologic findings reveal round, undifferentiated cells.     

Diagnosis of canine renal lymphoma by cytology and flow cytometry of the urine

Lymphoma is a common hematopoietic neoplasm of dogs. A definitive diagnosis typically requires the collection of samples via fine-needle aspirate or biopsy. A unique case of canine renal T-cell lymphoma diagnosed using urine sediment microscopy with flow cytometry and PCR for Antigen Receptor Rearrangement (PARR) is presented. A fresh urine sample was collected via a urinary catheter and immediately prepared for cytologic examination, flow cytometry, and PARR. The flow cytometric study revealed that 83% of the cells were large CD3⁺CD8⁺ T cells, while PARR identified a clonally rearranged T-cell receptor gene, supporting the flow cytometry findings. Despite supportive care, the patient progressed to anuric renal failure and was humanely euthanized. A necropsy was performed, and tissues from the upper and lower urinary tracts were collected. Histologically, the right and left kidneys were infiltrated by a neoplastic round cell population effacing the cortex and medulla. Immunohistochemistry for the T- and B-cell antigens CD3 and CD20, respectively, revealed that the neoplastic population within the kidney demonstrated diffuse, strong, membranous to intracytoplasmic CD3 expression while lacking CD20 expression. These results confirmed the diagnosis of renal T-cell lymphoma. This is the first known report of canine lymphoma diagnosed using either urine flow cytometry or clonality testing. Therefore, in select cases, urine flow cytometry and/or PARR are feasible to perform on urine-derived cells as a quick and cost-effective means to aid in the diagnosis of urinary tract lymphoma.     

Histiocytic chorioretinitis in a dog

A 3‐year‐old castrated male mixed‐breed dog presented with an acute bullous retinal detachment and thickened choroid of the right eye. Subretinal cytology revealed an atypical cell proliferation suggestive of neoplasia. The eye was enucleated, and the original diagnosis was a histologically benign choroidal melanocytic tumor. Further diagnostics revealed no other systemic abnormalities other than a nonhealing shoulder wound. Six months later, the left eye developed a bullous retinal detachment. This eye responded well to systemic steroids and the dog regained vision within a few weeks of initiating therapy. Results of immunohistochemistry with Melan‐A and CD204 of the previously enucleated right eye caused a revision of the histologic diagnosis from melanocytic tumor to histiocytic chorioretinitis. This case highlights the subtle and sometimes confusing distinction between neoplastic and inflammatory processes on both cytology and histopathology.     

Expression of the KIT protein (CD117) in primary cutaneous mast cell tumors of the dog.

Thirty-one canine cutaneous masses, diagnosed as mast cell tumors (MCT) by histopathologic analysis, were used to evaluate the immunohistochemical pattern of expression of KIT protein (CD117), a type III tyrosine kinase protein involved in mast cell growth and differentiation. Lesions were graded as I (well differentiated), II (intermediate differentiation), or III (poorly differentiated) according to the following morphologic features: invasiveness, cellularity and cellular morphology, mitotic index, and stromal reaction. Immunohistochemical KIT expression was compared with histologic grade and some histomorphologic features (cell differentiation and nuclear grade) evaluated separately. A possible predictive role of biologic behavior in MCTs for KIT expression was also investigated. Immunohistochemical analysis revealed three different patterns of KIT expression: a cytoplasmic diffuse pattern, a membranous pattern with immunostaining located on the cell surface, and a cytoplasmic perinuclear pattern, where KIT expression was detected in the cytoplasm of the neoplastic mast cells, close to the nucleus. Statistical analysis showed a close relationship between different KIT immunohistochemical patterns and histologic grade (P < 0.00000), cell differentiation (P < 0.00000), and nuclear grade (P < 0.0024). According to Kaplan-Meier-estimated survival curves compared by survival analysis, KIT expression was significantly associated with survival time (P = 0.037) but not cancer-free interval (P = 0.50). Similar to other well-known histomorphological features, KIT expression is a useful parameter of malignancy in cutaneous MCTs. KIT expression also predicted the biological behavior of the tumors in this study.     

Of all the nerve! A subcutaneous forelimb mass on a cat

A 16-year-old, male, neutered cat had a 2.5 X 1.5 cm mass on the medial aspect of the right carpus. Cytologic examination of a fine-needle aspirate of the mass indicated a markedly pleomorphic population of plasmacytoid to histiocytic-appearing cells. The cytologic diagnosis was malignant neoplasia of probable mesenchymal or round cell origin. The right forelimb was surgically removed and the scapular, axillary, and prescapular lymph nodes were excised. Malignant fibrous histiocytoma was tentatively diagnosed histologically; however, the tumor cells subsequently were found to be negative for histiocytic (MAC 387, antitrypsin), T-cell (CD3), and B-lymphocyte (immunoglobulin light chains, Ly 5/CD45R) markers, and positive for glial fibrillary acidic protein, vimentin, and S-100. Based on the immunohistochemical results, the diagnosis was modified to malignant peripheral nerve sheath tumor (PNST). Six months after surgery, the cat was reported to be well and had no evidence of metastasis. PNSTs are rare tumors in cats, and are considered as synonymous with schwannomas, neurofibrosarcomas, and hemangiopericytomas. In this cat, the plasmacytoid and pleomorphic appearance of the PNSTcells in cytologic and histologic specimens was unusual, and made it difficult to reach an accurate diagnosis without immunocytochemistry.     

Large anaplastic spinal B‐cell lymphoma in a cat

Abstract: A 5‐year‐old female spayed domestic shorthair cat was presented for evaluation of tetraparesis. The neurologic lesion was localized to the cervical spinal segment (C1–C6). A left axillary mass was identified, and the results of fine needle aspiration cytology indicated malignant round cell neoplasia of possible histiocytic origin. The cells were large, had marked anisocytosis and anisokaryosis, occasional bi‐ and multinucleation, and cytoplasmic vacuolation. Euthanasia was performed due to the poor prognosis associated with severe, progressive neurologic signs and a malignant neoplasm. Postmortem examination revealed spinal cord compression and an extradural mass at the C1–C2 spinal segment, with neoplastic cells in the adjacent vertebral bodies, surrounding skeletal muscle, left axillary lymph node, and bone marrow from the right femur. The initial histologic diagnosis was anaplastic sarcoma, but immunohistochemical results indicated the cells were CD20+ and CD45R+ and CD3?, compatible with a diagnosis of B‐cell lymphoma. CD79a staining was nonspecific and uninterpretable. Weak to moderate CD18 positivity and E‐cadherin positivity were also observed. Clonality of the B‐cell population could not be demonstrated using PCR testing for antigen receptor gene rearrangement. To the authors' knowledge, this is the first reported case of a feline spinal anaplastic B‐cell lymphoma exhibiting bi‐ and multinucleated cells. The prognostic significance of this cell morphology and immunophenotype is unknown.     

Malignant pilomatricoma in a soft‐coated Wheaten Terrier

Abstract: A 3‐year‐old, castrated male, soft‐coated Wheaten Terrier was presented for evaluation of mild lameness, fecal incontinence, lumbosacral pain, and lack of anal tone. Magnetic resonance imaging scan showed a large (8 × 6 × 5 cm) mass invading and expanding the pelvic bones, sacrum, and associated structures. A fine‐needle aspirate of the mass contained many neoplastic cells with high nuclear to cytoplasmic ratios and rare spindle and inflammatory cells. The neoplastic cells were 12–16 μm in diameter, round to cuboidal, basaloid in appearance, and arranged both individually and in loosely cohesive clusters with variably distinct cell borders. Given the location, signalment, and cytologic findings, differential interpretations included a primitive embryonal tumor (eg, neuroblastoma or nephroblastoma in an atypical location) or poorly differentiated carcinoma. The owner elected euthanasia due to the poor prognosis. Abnormal gross findings on necropsy included the pelvic mass and multiple firm, pale, pink‐tan nodules in the lung, which proved to be metastases. On histologic examination, the mass and nodules were composed of irregular islands, lobules, and nests of basaloid cells, which transitioned abruptly into large lakes of “ghost” cells with areas of ossification and calcification, consistent with a diagnosis of malignant pilomatricoma. This unusual presentation of a pilomatricoma adds to our knowledge of expected cytologic findings for this tumor.     

Multifocal cutaneous histiocytic sarcoma in a young dog and review of histiocytic cell immunophenotyping

A 9‐month‐old male Great Dane had progressive generalized nodular dermatopathy for several months. There were > 100 raised, alopecic, firm, painful nodules throughout the skin. Aspirates from several lesions yielded moderate numbers of irregularly round or polygonal to spindle‐shaped cells with mild to moderate anisocytosis and few inflammatory cells, and the cytologic interpretation was proliferation of mesenchymal or histiocytic cells. On histopathologic examination, nodules were composed of densely packed sheets of round to spindle‐shaped cells with mild anisokaryosis and low mitotic activity. Multifocal histiocytic sarcoma with a spindle‐cell pattern was diagnosed based on morphologic features and intense expression of CD18. Additional immunophenotypic analysis on frozen sections of tissue confirmed the diagnosis of histiocytic sarcoma; expression of CD18, CD45, CD1a, CD11b, and CD11c, limited expression of Thy‐1 (CD90) and CD80, and lack of expression of CD4, CD11d, and CD86 indicated that the cells were likely interstitial dendritic cells; a review of reactive and neoplastic dendritic cells is provided. Based on staging, internal organs were not affected. Sequential treatment with lomustine and doxorubicin failed to prevent progression of the cutaneous lesions, and the dog died 3 months after initial diagnosis. At necropsy, a focus of neoplastic cells was present in one lymph node, but except for skin other organs were not involved. The clinical presentation of histiocytic sarcoma may be unusual, and neoplastic cells may lack overt features of malignancy on cytologic and histopathologic examination. In some instances, immunophenotyping is required to differentiate histiocytic sarcoma from other histiocytic disorders.     

Disseminated histiocytic sarcoma with peripheral blood involvement in a Bernese Mountain dog

Abstract: A 6‐year‐old Bernese Mountain dog was presented with a history of lethargy and weight loss of 2 weeks duration. On physical examination the dog had pale mucous membranes and tachypnea. Ultrasound examination revealed hepatomegaly, splenomegaly, and mesenteric lymphadenomegaly. Results of a CBC included marked normocytic normochromic nonregenerative anemia, marked thrombocytopenia, and moderate leukocytosis with mild neutrophilia and a large population of unclassified round cells (6.2 × 10³/μL). The unclassified cells occasionally were bi‐ or multinucleated and had variably abundant pale basophilic cytoplasm that contained multiple irregular clear vacuoles and occasionally erythrocytes. Fine needle aspirate specimens of the mesenteric lymph nodes and spleen were composed of a population of round pleomorphic cells with the same features as the circulating cells. On flow cytometric analysis of peripheral blood, the unclassified cells expressed CD18, CD45, CD11c, CD1c, and CD14; immunocytochemical analysis of blood smears also indicated the cells were positive for CD1c, CD1a, and CD11c. The dog died a few hours after referral. The histologic interpretation of samples collected from spleen, liver, and lymph nodes was malignant neoplasia of histiocytic origin. Immunohistochemical staining yielded negative results for CD11d, a marker of red‐pulp macrophages, ruling out hemophagocytic histiocytic sarcoma. Based on clinical and pathologic findings, the final diagnosis was disseminated histiocytic sarcoma (DHS) with peripheral blood involvement. To our knowledge, DHS in a dog with evidence and immunophenotyping of neoplastic cells in peripheral blood has been reported only rarely.     

A peripheral primitive neuroectodermal tumor with generalized bone metastases in a puppy

A peripheral primitive neuroectodermal tumor (pPNET), most consistent with a human Ewing's sarcoma, is described in a 5-month-old male Australian Shepherd puppy. The first tumor site detected was in the left frontal bone of the skull with apparent subsequent rapid metastases to multiple sites in the axial and appendicular skeleton and bone marrow, kidneys, and perihyphophyseal meninges. Radiographically, all bone lesions were lytic and there was also a humeral bone fracture. Histologically, the tumor was diagnosed as a small round blue cell tumor. At this stage, the differential diagnosis included a lymphoma, rhabdomyosarcoma, and a PNET of the peripheral nervous system. However, the cells had positive expression of triple neurofilament antigens as detected immunocytochemically. The cells were negative for a broad panel of canine-specific leucocyte cell marker antigens for desmin, smooth muscle actin, synaptophysin, and CD99. Ultrastructurally, the cells contained occasional dense core neurosecretory granules and intermediate filaments with intercellular desmosomal-like junctions and abundant glycogen clusters. Based on the age of the dog, the clinical history, the distribution of gross lesions, histologic characteristics of a small round blue cell tumor, and immunocytochemical and ultrastructural evidence of neuroectodermal differentiation, a diagnosis of a pPNET similar to a human Ewing's sarcoma was made.     

Keratitis and periocular lesions associated with equine herpesvirus‐3 in a 3‐month‐old filly

A 3‐month‐old Quarter Horse filly presented with corneal ulceration in the right eye with extensive coalescing periocular ulcerations, erosions, and cutaneous crusts. Similar periocular lesions were present around the left eye, on the gingival mucosa, and on the cutaneous and mucosal surfaces of the lips. Based on the severity of the filly's corneal lesions, expense and duration of treatment, euthanasia was elected. Histological post mortem examination revealed numerous hyperplastic and/or dysplastic epithelial cells adjacent to areas of ulceration and erosion with intranuclear viral inclusion bodies. Equine herpesvirus‐3 (EHV‐3) was identified by polymerase chain reaction from the right cornea and lip. The virus was isolated from the right cornea, right eyelid and lip. The dam presented with multifocal to coalescing perineal vesicles. EHV‐3 was confirmed by polymerase chain reaction from the vulvar lesions and the mare recovered spontaneously. This is the first case of EHV‐3 corneal infection reported in horses and emphasises that EHV‐3 should be included as a differential diagnosis for vesicular lesions involving the equine periocular and oronasal epithelium.     

Olfactory neuroblastoma in a cat

CASE SUMMARY: A 12.5-year-old castrated male domestic shorthair cat was presented for evaluation of dyspnea. Physical examination suggested upper airway disease. Computed tomography revealed the presence of a nasopharyngeal mass. Biopsies obtained via rhinoscopy led to a diagnosis of olfactory neuroblastoma. CLINICAL CHALLENGES: Olfactory neuroblastoma (or esthesioneuroblastoma) is a rare malignant neoplasm that arises from olfactory neuroepithelium. It was first described in human medicine in 1924. Clinical signs are variable, and diagnosis may prove challenging as this tumor, which is a differential for spontaneous intranasal tumors in cats, may be mistaken for poorly differentiated carcinomas or round cell tumors. Given its rarity, treatment options for this neoplasm have not been thoroughly evaluated and there is no standard treatment protocol. LITERATURE REVIEW: A review of veterinary and human literature is provided, encompassing clinical signs, staging and grading systems, treatment options and prognostic factors.     

Laryngeal rhabdomyoma in a dog

A 3‐year‐old, female spayed Australian Shepherd dog was presented to Veterinary Healthcare Associates in Winter Haven, FL with a history of respiratory stridor, difficulty swallowing, and a change in bark for approximately 3 months. Radiographs revealed a soft tissue mass caudal to the epiglottis. Oral and computerized tomographic (CT) examinations were performed under general anesthesia and revealed a small firm mass in the caudal larynx on the right side. Cytologic evaluation of a fine‐needle aspirate of the mass revealed cells consistent with a rhabdomyoma, oncocytoma, or granular cell tumor. Histopathologic interpretation was rhabdomyoma or oncocytoma. Histochemical staining with periodic acid‐Schiff was mostly negative with a low number of weakly positive cells and with phosphotungstic acid‐hematoxylin was strongly positive with visible cross striations. Tumor cells did not express pancytokeratin AE1/AE3, were strongly immunoreactive for myoglobin and muscle‐specific actin, and did not express vimentin except for a low number of weakly immunoreactive cells. These findings confirmed a diagnosis of rhabdomyoma. Primary neoplasia of the larynx is rare, and widely varying types of neoplasms have been documented in this location. Due to the cytologic and histologic similarities between rhabdomyoma, oncocytoma, and granular cell tumor, these neoplasms should be differentiated using histochemical staining and immunohistochemical analysis.     

A metastatic secretory gastric plasmacytoma with aberrant CD3 expression in a dog

A 10‐year‐old crossbred dog was presented with a 6‐week history of hematemesis, melena, anorexia, and lethargy. Clinical evaluation revealed a gastric mass with a regional lymphadenomegaly as well as a monoclonal gammopathy manifesting as hyperglobulinemia. Cytologic and histopathologic analyses were consistent with a round cell neoplasm; neoplastic cells showed nuclear immunoreactivity for MUM1 and diffuse cytoplasmic reactivity for CD3. Polymerase chain reactions performed on fixed and fresh tissue identified a clonal rearrangement with an IgH primer set. An extramedullary plasmacytoma (EMP) was confirmed by cellular morphology and molecular diagnostics. Following an objective response to chemotherapy, the dog was euthanized 8 months after diagnosis, and a postmortem examination confirmed the clinical findings. This is the first reported case of a monoclonal gammopathy secondary to a gastric EMP coupled with aberrant expression of CD3 in an aggressive plasmacytic tumor, and highlights the utility of molecular diagnostics for classifying atypical hemolymphoid neoplasms.     

Immunohistochemical expression of E-cadherin does not distinguish canine cutaneous histiocytoma from other canine round cell tumors

Immunohistochemistry for E-cadherin (ECAD) has been used to distinguish canine cutaneous histiocytoma from other leukocytic neoplasms ("round cell tumors"). To determine the specificity of this test, 5 types of canine cutaneous round cell tumors were evaluated for immunohistochemical expression of ECAD. Tumors of all 5 types had variable cytoplasmic, plasma membrane, and/or paranuclear ECAD expression: All 13 cutaneous histiocytomas were ECAD+; all but 1 of 14 mast cell tumors expressed ECAD; 10 of 12 epitheliotropic lymphomas reacted with E-cadherin antibody; of 72 plasmacytomas, 54 were ECAD+; and 5 of 5 histiocytic sarcomas were positive. Conclusions based on these results include the following: First, immunoreactivity for ECAD is not limited to leukocytes of cutaneous histiocytoma; second, antibody to ECAD also labels neoplastic cells in most mast cell tumors, plasmacytomas, cutaneous histiocytic sarcomas, and epitheliotropic lymphomas; third, although most histiocytomas have membranous ECAD expression, the immunoreactivity varies among round cell tumors and is frequently concurrent in different cellular compartments; fourth, the distinctively paranuclear ECAD expression pattern in epitheliotropic lymphomas might distinguish them from other round cell tumors; and, fifth, ECAD should be used with other markers (eg, MUM1 for plasmacytomas, KIT for mast cell tumors, CD3 and CD79a for lymphomas) to distinguish among canine round cell tumors.     

Cytology of canine cutaneous round cell tumors. Mast cell tumor, histiocytoma, lymphosarcoma and transmissible venereal tumor.

Sixty-four canine cutaneous round cell tumors were divided into 25 mast cell tumors, 15 histiocytomas, nine cutaneous lymphosarcomas and 15 transmissible venereal tumors. The final diagnosis was made from cytologic, clinical and histologic findings. Cytologic features were significantly distinctive in mast cell tumor, transmissible venereal tumor, and most cases of histiocytoma and lymphosarcoma to allow a diagnostic opinion. This opinion was supported by subsequent histologic examination. In some instances cytology was considered essential in rendering a diagnostic opinion even though histology was available.