Where did the white cells go?

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Where did the white cells go?

FIGURE 1: Fine needle aspirate of mediastinal mass. Rafts of epithelial cells with pale cytoplasm and indistinct cell borders. Occasional small lymphocytes are present in the background. Mast cells were also present (not shown here).

Genevieve D’Amours, from SVS Laboratories, Ian Millward, from Veterinary Referral Services and Sara Bruce, from Cambridge Pet Vets consider the case of a cat with a severe and persistent neutropenia.

STARSKY, A 13-YEAR-OLD male domestic short-hair cat, was taken to his veterinarian because of weight loss in the previous two or three months. He was tachycardic and hyperthyroidism was suspected.

On serum biochemistry there was a mild increase in globulins at 53g/L (reference range 27–49) and the alkaline phosphatase was mildly elevated at 114U/L (reference range 0–85), but the T4 was normal at 22nmol/L (reference range 20–40). Haematology showed a severe neutropenia, with neutrophils at 0.2 x109/L (reference range 1.5–10.8) with no left shift. There was no anaemia or thrombocytopenia. A week later, the neutropenia was still present at 0.4 x109/L.

Chest radiographs indicated a 3cm x 6cm ovoid soft tissue mass in the cranial mediastinum, extending from the second to the fifth rib, with contact on the dorsal aspect of sternebrae 2–5 and causing border effacement of the cranial aspect of the cardiac silhouette. On ultrasonography the mass had soft tissue echogenicity with some fluid pockets. An ultrasound-guided fine needle aspirate of the mass was taken.

On cytology there was a mixed population of cells consisting mainly of large rafts of poorly preserved epithelial cells. The cells had pale cytoplasm and indistinct cytoplasmic borders and ovoid nuclei with fine chromatin and single small nucleolus (Figure 1). Scattered throughout the rafts was a moderate number of heavily granulated mast cells. Around the rafts of epithelial cells was a moderate number of small to medium lymphocytes with occasional large lymphocytes. There were also occasional large, foamy macrophages containing dark pigment and rare, plump, fusiform mesenchymal cells. Occasional cholesterol crystals were present in the background.

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This mixed cell population was highly suggestive of a mixed cell thymoma.

A median sternotomy was performed through sternebrae 2–5 and the soft tissue mass was identified in the mediastinum immediately cranial to the heart. The mass was packed with moist swabs then carefully dissected from the adjacent tissues through blunt dissection. The mass was in contact with the cranial vena cava over approximately 2cm, but appeared to dissect free from the blood vessel wall. A number of small blood vessels were ligated and transected near the cranial end of the mass.

The mass was removed en bloc, and the thoracic cavity was thoroughly lavaged with warm saline solution. A fenestrated closed suction drain was placed through the sixth intercostal space and the mediastinum was closed routinely using three alternating figure-of-eight sutures using 22-gauge orthopaedic wires, 3-0 polydioxanone (PDS II, Ethicon) and 4-0 Nylon (Ethilon, Ethicon).

The patient was placed on morphine 0.2mg/kg every four hours, and a 12.5mcg transdermal fentanyl patch (Fentanyl,

FIGURES 2 AND 3:Mediastinal mass. Large islands of epithelial cells admixed with moderate numbers of lymphocytes with minimal supporting stroma.

Sandoz) was applied. Meloxicam 0.05mg/ kg orally (Meloxicam, Apex) was started post operatively and continued for 14 days.

The chest drain was removed 12 hours later, when the drainage volume was noted to reduce significantly to less than 5ml/kg/hour. Starsky was placed on soft foods only for seven days and cage confinement for six to eight weeks. He recovered well from the surgery and was discharged 36 hours afterwards.

On histology the mass consisted of a well-demarcated, partially encapsulated neoplasm composed of large islands of polygonal epithelial cells admixed with a variable number of small lymphocytes,

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with the lymphocytic population representing approximately 25% of the mass (Figures 2 and 3). The epithelial cells had abundant pale eosinophilic cytoplasm and round-to-indented nuclei with prominent single central nucleoli. There was mild anisocytosis and anisokaryosis, and mitotic figures were rare. A moderate number of mast cells were scattered throughout the neoplasm (Figure 4).

There were several small to large cystic areas of necrosis containing numerous clear acicular cholesterol clefts, a large number of foamy macrophages and multinucleated giant cells. Occasional large lymphoid follicles were also present. This confirmed a diagnosis of a benign thymoma. In all sections there were areas where the capsule was absent, with neoplastic cells extending to the tissue margins, so there is concern for a potential recurrence of the lesion.

It was suspected that the thymoma was the cause of Starsky’s neutropenia. Thymomas have been recognised as a cause of many immune-mediated conditions, most notably myasthenia gravis in dogs and exfoliative dermatitis in cats (Atwater et al., 1994; Robat et al., 2013). In humans thymomas are the neoplasms most commonly associated with paraneoplastic autoimmune diseases such as myasthenia gravis, systemic lupus erythematosus, bullous pemphigoid, dermatomyositis, rheumatoid arthritis, ulcerative colitis, acquired red cell aplasia and other cytopenias (Shelly et al., 2011).

Autoimmune diseases develop because of a loss of self-tolerance. Precursor T cells are generated in the bone marrow, but they mature in the thymus through a process of maturation, differentiation and selection. Positive and negative selection allows the survival of cells that recognise major histocompatibility complex molecules but eliminates autoreactive cells. It is assumed that thymic neoplasms allow for the maturation of autoreactive T cells, but the mechanisms are poorly understood. With epithelial thymomas this could occur due to the genetic mutation of the epithelial cells with subsequent changes in antigenic expression, while with lymphocytic thymomas it could occur through the escape of immature lymphocytes into circulation by avoiding passage through the medulla where maturation occurs. Another theory is that abnormal T helper cells may activate B cells to produce autoantibodies (Shelly et al., 2011).

FIGURE 4:Neoplastic population of epithelial cells admixed with moderate numbers of small lymphocytes and mast cells.

While these autoimmune diseases can sometimes undergo remission following thymectomy, others can develop even after the tumour has been resected. Starsky is doing quite well clinically but had a persistent neutropenia with neutrophils of 0.5 x109/L three weeks after surgery. We are hopeful that his neutrophils remain at a level sufficient to prevent illness and that the tumour will not recur.

REFERENCES: Atwater SW, Powers BE, Park RD, Straw RC,

Ogilvie GK, Withrow SJ. Thymoma in dogs: 23 cases (1980–1991). Journal of the American Veterinary Medical Association 205(7), 1007–13, 1994

Robat CS, Cesario L, Gaeta R, Miller M,

Schrempp D, Chun R. Clinical features, treatment options, and outcome in dogs with thymoma: 116 cases (1999–2010). Journal of the American Veterinary Medical Association 243(10), 1448–54, 2013

Shelly S, Agmon-Levin N, Altman A, Shoenfeld

Y. Thymoma and autoimmunity. Cellular & Molecular Immunology 8(3), 199–202, 2011. doi:10.1038/ cmi.2010.74

AUSTRALIAN AND NEW ZEALAND COLLEGE OF VETERINARY SCIENTISTS