Bilateral thyroid follicular compact-cellular carcinoma in a llama

Abstract

An 18-y-old female llama (Lama glama) in Saskatoon, Saskatchewan was examined during a routine herd check, and a mass was detected in the ventral cervical area just below the angle of the jaw. No clinical signs were evident except for the mass and chronic loss of body condition. Postmortem examination revealed bilateral enlargement of the thyroid gland with multiple cysts. Histopathology of the thyroid gland revealed follicular compact-cellular carcinoma lesions, with infiltration of neoplastic thyroid follicular cells in regional lymph nodes.

Thyroid function is essential for maintenance of metabolism and homeostasis of the internal milieu. Decreased thyroid function produces a range of clinical signs, including reproductive, neuromuscular, dermatologic, cardiac, ocular, and gastrointestinal disruption, and alterations of cellular metabolism.⁵ A robust body of work has contributed to our understanding of thyroid diseases in small animals, but this understanding has not been translated to other species, such as farm or wild animals. During a routine herd check in May, an 18-y-old female llama (Lama glama) was found with a swelling just below the angle of the jaw. The llama belonged to a herd of 25 llamas and alpacas and had been born and maintained on the farm. Llamas in the herd had free access to water and good quality hay during the winter months and pasture during the grazing season. Annual management included vaccination with a 3-way clostridial vaccine, deworming by parenteral administration of ivermectin, and supplementation with vitamins A, D, and E, and selenium in the spring. The llama in question had a history of infectious keratoconjunctivitis the previous summer; however, this condition resolved after antibiotic therapy. Additionally, the llama had a history of being treated for a tooth root abscess 2 y ago. The llama had normal appetite, interacted with other members of the herd, and was attentive and responsive to its environment. Body condition score was 5 of 10 (thin = 1, fat = 10) at the time of examination in May, and no further actions were taken.

The llama continued to lose body condition, reaching a score of 3 of 10 later in the fall. The llama was thin and weighed 120 kg. Clinical parameters such as heart rate (62 beats/min; herd average: 70 ± 8 beats/min), respiration rate (30 breaths/min; herd average: 33 ± 9 breaths/min), and rectal temperature (37.5°C; herd average: 37.4 ± 0.7°C) were not altered compared to other members of the herd. No abnormalities were detected in gastric compartment contraction frequency or intestinal borborygmi, and rectal palpation revealed no abnormalities. Manual palpation of the ventral neck revealed a moderately firm, fluctuant, and painless bilateral mass ventral to the larynx.

Ultrasonographic examination of the neck mass was performed using a 5-MHz linear-array probe (Mylab 5; Esaote North America, Indianapolis, IN) in transverse and longitudinal planes. The affected area had multiple cystic cavities of various sizes, in proximity to the carotid artery and trachea. Cysts were 3–10 mm diameter with variable degrees of echogenicity. Color Doppler imaging was used to distinguish between vascular structures and the nonvascular cysts. As part of the annual cull of older animals in the herd, the affected llama was euthanized with another female of similar age (18 y) by intravenous administration of sodium pentobarbital (Euthanyl; Bimeda-MTC Animal Health, Cambridge, Ontario, Canada).

Blood samples were collected from the affected llama and an age- and sex-matched control llama from the same herd just prior to euthanasia and submitted to Prairie Diagnostic Services (PDS; University of Saskatchewan, Saskatoon, Canada) for a complete blood count, serum biochemistry, and total tetraiodothyronine (T4). Normal reference intervals for blood count and chemistry were obtained from the International Species Information System (ISIS) for llamas > 15 y old.¹⁵ A mild left shift without neutrophilia and without evidence of toxic change was reported in both llama samples. Mild hyperglycemia was noted in both the affected and control llama at 10.4 mmol/L and 12.9 mmol/L, respectively (reference interval [RI]: 4.8–10.0 mmol/L), likely associated with handling stress. Total T4 was 126 nmol/L in the affected llama and 91 nmol/L in the age- and sex-matched herdmate (RI: 50–262 nmol/L).¹⁵

Postmortem examination of the neck mass was completed in the field. The mass was partially attached to the sternocephalicus muscle and firmly attached to the trachea (Fig. 1). Given the location and appearance of the mass compared to the unaffected llama, it was concluded that the mass corresponded to the thyroid gland. The thyroid glands were bilaterally enlarged in the affected llama compared to the normal llama (affected llama: left lobe 4 × 3 × 10 cm, right lobe 3.5 × 2.5 × 8.5 cm; normal llama: left lobe 1.5 × 1 × 4 cm, right lobe 1.5 × 1 × 4 cm; Fig. 1).Variably sized cysts were apparent on the surface of the mass. Contents of the cystic structures varied from gelatinous to firm consistency; other cysts contained bloody or viscous fluid. Abundant blood vessels were observed crossing the surface of the mass. Postmortem evaluation of the remainder of the carcasses was not performed.

Figures 1–3.

Macroscopic and microscopic appearance of the thyroid gland and deep cervical lymph node of the affected llama. Figure 1. Ventral view of the thyroid gland of the affected llama at the junction of the trachea (left) and larynx (right). Figure 2. Relationship between neoplastic areas (compact-cellular, left arrow; follicular, right arrow) and thyroid follicles (asterisks). H&E. Figure 3. Thyroglobulin (brown; arrows) in cytoplasmic vacuoles within (compact-cellular) neoplastic cells. Immunohistochemistry. Figure 4. Cranial deep cervical lymph node with thyroid follicular tissue (arrows) interspersed among lymphoid follicles (asterisks). H&E.

Tissue samples from the thyroid gland and one of the deep cervical lymph nodes (craniomedial to the gland) from the affected and unaffected llamas were fixed in 10% neutral-buffered formalin, washed in phosphate-buffered saline, and submitted to PDS. Paraffin-embedded sections (5-µm thick) were mounted on glass slides, stained with hematoxylin and eosin, as well as Masson trichrome for collagen or fibrous tissue identification. Immunohistochemistry for thyroglobulin was also performed.

Histologic examination revealed that the thyroid was composed of variably sized, well-differentiated follicles filled with colloid, as well as areas of solid tumor growth (Fig. 2). The ratio of follicular type to compact-cellular type was approximately 1 to 3. Within solid areas, the neoplastic cell population was composed of islands and trabeculae of epithelial cells separated by a fine fibrovascular stroma. The mass was expansive and compressed the normal thyroid architecture (Fig. 2); no evidence of capsular or vascular invasion was noted. Cells were cuboidal-to-polygonal, with distinct cell borders, had abundant granular cytoplasm and often a large vacuole containing amorphous eosinophilic material. The oval nucleus was often eccentrically placed and molded by the large vacuole (Fig. 3). The nuclei contained finely clumped chromatin, a single basophilic nucleolus, varied in size up to 6-fold above that of the normal range for the thyroid gland, and frequently contained round, eosinophilic inclusions similar in color to the large vacuoles (pseudoinclusions). No mitotic figures were observed, but frequent binucleate cells were present. In addition to hemorrhages, a central area of necrosis and mineralization was also present. Immunohistochemistry confirmed the presence of thyroglobulin within follicle lumens (not pictured) and within intracytoplasmic vacuoles of neoplastic cells (Fig. 3).¹⁰ No bacteria were observed in the examined tissues.

Two deep cervical lymph nodes were sampled for histopathology. Up to 50% of the lymph nodes were replaced by neoplastic follicular cells within the cortex and subcapsular sinuses composed of foci of thyroid follicles of variable size surrounded by small amounts of fibrous connective tissue (desmoplasia), interspersed with small areas of compact-cellular growth (Fig. 4). Foci of hemorrhage and aggregates of hemosiderin-laden macrophages (pigment granulomas) were also seen throughout the lymph node.

The macroscopic location, large size and bilateral involvement, histopathologic features, and positive immunoreactivity for thyroglobulin is consistent with the diagnosis of thyroid follicular compact-cellular carcinoma, and, to our knowledge, has not been reported previously in llamas. The diagnosis of carcinoma is supported by invasion of at least 2 subjacent deep cervical lymph nodes by neoplastic thyroid follicles of markedly variable size with interspersed compact-cellular growth, consistent with what was observed in the main thyroid tumor. These findings were key in differentiating between adenocarcinoma and adenoma, given that adenomas are often present as nodular lesions that are well demarcated from surrounding tissue and do not invade other organs.²

The etiology of thyroid neoplasia in llamas is unknown, and there is a scarcity of information available regarding neoplasia in camelids in general. In a retrospective study of postmortem examinations of llamas over 3 y in the northwestern USA, 15 of 77 llamas had gross thyroid lesions, and 14 of 77 were devoid of gross lesions but had microscopic lesions, such as ruptured follicular epithelia and focal areas of hyperplasia.⁷ Based on the same study, a clear relationship with advanced age was established, which is consistent with our case and with studies in horses⁴ and dogs.^1,5

Hyperthyroidism occurs frequently in cats with thyroid hyperplasia or adenoma. Hyperthyroidism is uncommon in dogs, with nonfunctional thyroid carcinoma being the most common of canine thyroid tumors. In horses, thyroid tumors are typically nonfunctional, although hyperthyroidism has been reported.¹³ Common laboratory abnormalities that may be found in hyperthyroid cats include increased serum alkaline phosphatase, alanine aminotransferase, and aspartate aminotransferase.² Hyperthyroidism may also lead to a decrease in serum creatinine from muscle mass loss and because of concomitant chronic kidney disease, which itself can be masked by the hyperthyroid state.¹⁶ However, none of these parameters were altered in our llama. We estimated the blood concentration of total T4 in the affected and normal llama; however, we lacked reference values for camelids in our geographic area. Based on reference values from ISIS and other sources,^6,14,15 we conclude that the thyroid carcinoma in our llama was not functional. Given that dentition was normal, we attribute the low body condition score of the llama to the age of the llama.

Several causes of thyroid carcinoma have been described in the literature. In humans, mutations of the thyroid receptor have been linked to thyroid adenomas and hyperplasia.¹² As well, mice expressing the oncogene ret/PTC1 develop papillary thyroid carcinomas.⁹ Nutritional imbalances, such as iodine deficiency, have not been associated with thyroid tumors; however, they are associated with goiter, as has ingestion of goitrogenic substances. Goiter is an enlargement of the thyroid gland that is not associated with inflammation or neoplasia, and is characterized by low levels of T3 and T4.² We discounted the diagnosis of goiter in our case given that a marked proliferation of follicular cells in multiple areas was detected, with areas of necrosis and metastasis, which is not consistent with goiter.

A variety of treatments are available for thyroid tumors. In cats, surgical removal, radioactive iodine administration, dietary change, and pharmacologic approaches are used.^3,11 In horses, surgical removal and immunohistochemical confirmation of the excised adenoma are recommended given that poor correlation has been found between macroscopic and microscopic characteristics (Troillet A, et al. Thyroid gland tumours in the horse: clinical diagnostic, treatment and histological characterization. Proc Brit Equine Vet Assoc Congress; Sept 2014; Birmingham, UK). We euthanized the llama in our case based on the low likelihood of survival (based on age and low body condition score) in harsh winter conditions in Saskatchewan. Although we did not perform an autopsy of the whole llama, distant metastasis, especially to the lung, is a consequence of thyroid tumors in other species.⁸ The extent of regional lymph node invasion suggests that metastasis may also have occurred to other organs, although this is speculation as information is not available on the behavior of thyroid tumors in llamas.