Type A thymoma in a pet rabbit

Abstract

A 4-y-old, spayed female, mixed-breed domesticated rabbit (Oryctolagus cuniculus domesticus) was presented because of progressive bilateral exophthalmos, with a large mediastinal mass in the cranial thorax. Palliative radiation therapy was elected, and 4 fractions of 5 Gy were delivered twice weekly under general anesthesia using 3-dimensional conformal radiation therapy for a total dose of 20 Gy, guided by an on-board cone beam CT scan. Quality-of-life and respiratory rate improved before sudden death that followed an episode of dyspnea. The overall survival time following initial diagnosis was 93 d, with 68 d after the first dose of radiation. An autopsy was performed, and the mass was diagnosed as a type A thymoma. The diagnosis was confirmed with positive immunohistochemical labeling of the neoplastic cells for cytokeratin 5/6 and cytokeratin 7.

Thymomas are cranial mediastinal neoplasms that arise from the thymic epithelium. According to the 2004 World Health Organization classification scheme, thymomas are classified based on the histologic appearance of the neoplastic epithelial cells and the proportion of non-neoplastic lymphocytes. ⁶ Thymomas are the most common mediastinal neoplasm in rabbits; however, they have rarely been classified according to the WHO classification.^1,3–5 We report here a type A thymoma in a rabbit.

A 4-y-old, spayed female, mixed-breed domesticated rabbit (Oryctolagus cuniculus domesticus) was presented because of progressive bilateral exophthalmos over a period of ~1 mo. The rabbit lived comfortably in a home with another rabbit and several cats, with access to an enclosed garden area. She had no medical history prior to presentation, other than routine dental care, and had never been hospitalized. The owners reported a slight decrease in energy, but appetite remained normal. She was prescribed enrofloxacin (Baytril; Elanco) initially, and a dental procedure was scheduled ~5 wk later. Thoracic radiographs obtained during the dental procedure revealed a cranial thoracic mass, dorsal tracheal displacement, and an interstitial pattern within the caudal lung lobes (Fig. 1). The presumptive clinical diagnosis was thymoma. The rabbit was referred to the Michigan State University (MSU) Veterinary Medical Center’s oncology clinic (East Lansing, MI, USA) for further care.

Figures 1–6.

Type A thymoma in a pet rabbit. Figure 1. Lateral chest radiograph. The cranial thorax is expanded by a soft tissue opacity. Figure 2. A white-to-tan, firm, nodular thymic mass expands the mediastinum. Figure 3. Dense spindloid neoplastic cells compress the remaining thymus. H&E. 4× magnification. Figure 4. Neoplastic cells are arranged in interlacing streams and bundles. H&E. 20× magnification. Figure 5. Large numbers of neoplastic cells label for cytokeratin 7. Immunohistochemistry for cytokeratin 7, DAB (brown), hematoxylin counterstaining. 10× magnification. Figure 6. Occasional neoplastic cells label strongly for cytokeratin 5/6. Immunohistochemistry for cytokeratin 5/6, DAB (brown), hematoxylin counterstaining. 10× magnification.

Upon examination at MSU, ~3 wk after the initial diagnosis, the rabbit appeared healthy, hydrated, properly conditioned, and weighed 2.8 kg. Positional exophthalmos was observed and found to be most pronounced upon ventroflexion of the neck. Thoracic auscultation revealed normal lung and heart sounds, although audibility was poor over the left hemithorax.

Palliative radiation therapy was elected; additional testing (including bloodwork and fine-needle aspiration) was declined given financial concerns. A total of 20 Gy was prescribed for the rabbit’s mass, in 4 fractions of 5 Gy, to be administered twice weekly under general anesthesia using 3-dimensional conformal radiation therapy guided by an on-board cone beam CT scan. It was initially very difficult to anesthetize the rabbit because of positional airway occlusion precluding the use of a V-gel supraglottic airway device. Following completion of the 2-wk-long (10-d duration) radiation regimen, physical examination revealed minimal positional exophthalmos and a respiratory rate reduced from 50 brpm to 36 brpm. Her owners noted that her energy levels had returned to normal at home.

The rabbit died at home after sudden onset of respiratory distress 131 d after initial presentation to her primary care veterinarian, which was 93 d after the diagnosis, and 68 d after her initial fraction of radiation therapy. Her owners observed a change in respiration the night before she died, despite a continued appetite; an obstructive respiratory pattern was observed with a decreased respiratory rate and increased respiratory effort. She decompensated quickly over the course of 12 h and was noted to begin using accessory muscles for respiration and posture with a strained, extended neck. Death occurred shortly after cyanosis of the mucous membranes was observed. Cardiopulmonary resuscitation was not attempted. Permission was granted for an autopsy to be performed, and the rabbit’s body was transported to MSU shortly after death.

An autopsy was performed; the thoracic cavity contained 18 mL of yellow-to-pink fibrinous exudate. A white-to-tan, firm, nodular 7 × 5 × 1-cm mass in the mediastinum extended from the thoracic inlet to the diaphragm and adhered to the dorsal and left lateral surfaces of the pericardium (Fig. 2). There were several adhesions to the thoracic wall. Sections of the mass were collected and fixed in 10% neutral-buffered formalin, processed routinely, and 5-μm sections stained with H&E. The mediastinal mass was composed of a densely cellular population of neoplastic spindle cells arranged in interlacing streams and bundles, supported by dense fibrovascular stroma (Figs. 3, 4). Neoplastic cells had scant eosinophilic cytoplasm, indistinct borders, and finely stippled ovoid-to-fusiform nuclei. Anisokaryosis and anisocytosis were moderate, and no mitoses were observed in 10 hpfs (400×, 2.37 mm²). Rare karyomegalic neoplastic cells were present. Regions of the neoplasm were arranged in loose, fine streams, with small aggregates of lymphocytes and macrophages. Rarely, these foci were mineralized.

Immunohistochemistry for pancytokeratin, CD204, E-cadherin, calretinin, cytokeratin 5/6, and cytokeratin 7 (Table 1) was performed on additional sections of the mediastinal neoplasm. Large regions of neoplastic spindle cells had positive cytoplasmic labeling for cytokeratin 7 (Fig. 5) and were multifocally positive for cytokeratin 5/6 (Fig. 6). Neoplastic cells were negative for CD204, E-cadherin, and calretinin. The aggregates of lymphocytes were admixed with small numbers of CD204- or E-cadherin–positive histiocytic cells. Small numbers of spindle cells near these round cells were positive for pancytokeratin, but most neoplastic cells were negative for pancytokeratin. Based on the location, histologic features, and immunohistochemistry findings, this neoplasm is consistent with type A thymoma.^6,7

Table 1.

Antibodies with source, clone and manufacturer, concentration used, retrieval information, and detection method.

Antibody	Host	Source	Clone	Antigen retrieval	Dilution	Chromogen	Autostainer	Positive canine control
CD204	Mouse	Medicinal Chemistry Pharmaceutical, KT022	SRA-E5	PT low*	1:1,000	DAB	Bond-Max; Leica	Histiocytic proliferation
E-cadherin	Mouse	BD Bioscience, 610181	36/E-cadherin	ER1†	1:300	DAB	Bond-Max; Leica	Skin
Calretinin	Mouse	Dako, M7245	Dak-Calret-1	ER1	1:100	DAB	Bond-Max; Leica	Eye
Pancytokeratin plus	Mouse	BioCare, CM162C	AE1/AE3 + 5D3	ENZ3‡	1:200	DAB	Bond-Max; Leica	Skin
Cytokeratin 5/6	Mouse	Dako, M7237	D5/16 B4	PT high§	1:50	DAB	Dako Link 48; Agilent	Skin
Cytokeratin 7	Mouse	Dako, M7018	OV-TL 12/30	ER2¦	1:75	DAB	Bond-Max; Leica	Lung

^* Low pH retrieval for 20 min (Dako PT Link; Agilent).

^† Heat retrieval for 20 min (ER1, Bond epitope retrieval solution 1; Leica).

^‡ Enzymatic retrieval for 15 min (Dako proteinase K; Agilent).

^§High pH retrieval for 20 min (Dako PT Link; Agilent).

^¦Heat retrieval for 10 min (ER2, Bond epitope retrieval solution 2; Leica).

Type A thymoma is an uncommon neoplasm in domestic animals and is composed primarily of spindloid thymic epithelial cells and rare-to-absent lymphocytes. In dogs, 70% of cases are type B thymomas, and are composed of epithelioid cells and small lymphocytes. The ratio of the epithelial cell population to non-neoplastic lymphocytes determines the subclassification of type B thymomas. The most common subtypes in dogs are type B1 (26%) and B2 (36%). ⁶ Thymomas have been reported in domestic rabbits, although detailed histologic descriptions and accurate classification are lacking.^1,3–5 In a review of 2,970 neoplasms in domestic rabbits, thymomas were reported in 48 cases, with a prevalence of 2.1%; the thymomas were predominantly type B1 based on their microscopic features (pers. comm., C.A. Bertram, 2021 Sept 1). ¹ Additionally, thymomas were more common in rabbits > 6-y-old, with a prevalence of 9.4%, and prevalence was higher in neutered male and female rabbits than in intact rabbits. ¹

Given how uncommonly type A thymomas are reported, we used immunohistochemistry (PAN Plus; Biocare Medical) to prove that the spindle cells were consistent with neoplastic thymocytes and to rule out other differential diagnoses. After observing no immunoreactivity with PAN Plus in most neoplastic cells, individual cytokeratins (CK5/6 and CK7) were chosen based on their reported reactivity in thymomas. ⁷ Based on the location and spindloid nature of the neoplasm, a sarcomatoid mesothelioma was a potential differential, but the neoplasm was calretinin negative. Given the rare karyomegalic neoplastic cells, a histiocytic sarcoma was also considered, but the neoplasm was CD204 negative. Regions of this mass had CD204-negative and E-cadherin–positive histiocytic cells, lymphocytes, and a few pancytokeratin-positive spindle cells. These regions were suspected to represent the remaining compressed thymus.

Although the Masaoka–Koga staging system is more accurate in prognosticating thymomas in humans than the WHO classification scheme, morphologic subtyping based on the WHO histologic criteria of thymomas is performed routinely. ² Specifically, type B3 thymomas have been associated with an intermediate survival time, whereas type C thymomas consistently had the worst prognosis. ² Although significant prognostic differences have not yet been determined for the morphologic subtypes of thymomas in rabbits, adherence to the WHO classification system is necessary to accurately differentiate the various subtypes from sarcomas and lymphomas. Consistently accurate classification of thymoma subtypes will also allow future clinical outcome studies. In our case, biopsies could not be performed prior to palliative radiation. Therefore, the possibility that irradiation may have altered the histomorphologic appearance of this thymoma should also be considered.