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Unusual presentation of a T-cell-rich large B-cell lymphoma in the nictitating membrane in a seven-year-old Bavarian warmblood gelding
  1. James Oliver Rushton1,
  2. Andrea Fuchs-Baumgartinger2 and
  3. Barbara Nell1
  1. 1Department of Companion Animals and Horses, Veterinary University Vienna, Austria
  2. 2Department of Pathobiology, Veterinary University Vienna, Austria
  1. Correspondence to Dr James Oliver Rushton, james.rushton{at}vetmeduni.ac.at

Abstract

A seven-year-old, 550-kg, Bavarian warmblood gelding was referred to the Ophthalmology Service of the Veterinary University Vienna, due to a mass on the bulbar aspect of the nictitating membrane of the left eye. Thorough ophthalmic examination of the left eye revealed a red-pink mass, about 1 cm in diameter and 4 mm thickness, on the bulbar aspect of the nictitating membrane. Histopathological and immunohistological examination of the excised mass identified a T-cell-rich large B-cell lymphoma. Follow-up examination revealed no recurrence of the mass 18 months after initial presentation. To the authors' knowledge this is the first report of a histopathologically confirmed T-cell-rich large B-cell lymphoma of the ocular adnexa in the horse, with no recurrence or metastasis 18 months after initial presentation.

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Background

Lymphomas are the most common neoplasms in horses, which may occur at any age, with the highest incidence between 4 years and 10 years of age (Taintor and Schleis 2011). The overall reported prevalence in the general equine population is between 0.002 per cent and 0.5 per cent (Rendle and others 2012). No established risk factors for equine lymphomas have been identified as yet (Schnoke and others 2013). Lymphomas are traditionally classified as multicentric, alimentary, mediastinal, cutaneous and solitary (Taintor and Schleis 2011). However the classification by anatomical location has been rendered obsolete, due to a more recent classification based on the present cell type (Rendle and others 2012). This system is more likely to predict the behaviour of the underlying equine lymphoma (Rendle and others 2012). Lymphomas of the diffuse, mixed type are commonly seen in horses with prevalences of 33 per cent (Kelley and Mahaffey 1998) to 43 per cent (Durham and others 2012). This type of lymphoma has been classified in the WHO classification system as T-cell-rich large B-cell lymphoma (TCRLBCL) (Valli and others 2002). TCRLBCL is a slowly progressive lymphoma with a mixed population of small T lymphocytes and larger neoplastic B-cells (Valli and others 2002). This subtype of lymphoma may be encountered in horses in the cutaneous, multicentric and gastrointestinal form (Durham and others 2012). The skin type occasionally presents as a single subcutaneous mass or more commonly as multiple round, flattened disc-like lesions arising along subcutaneous lymphatics (Valli and others 2002). This type of lymphoma typically affects the skin of predominantly female horses (Valli 2007). A case of TCRLBCL in the tongue of a horse has also been described (Rhind and Dixon 1999). To the authors’ knowledge there is no report of the occurrence of lymphoma confirmed as TCRLBCL of the ocular adnexa in horses, which was successfully treated with excisional biopsy alone. This report describes the unusual location and successful surgical treatment of TCRLBCL in the third eyelid of a seven-year-old Bavarian warmblood gelding.

Case presentation

A seven-year-old, 550 kg, Bavarian warmblood gelding was referred to the Ophthalmology Service of the Veterinary University Vienna, due to a mass on the bulbar aspect of the nictitating membrane of the left eye (OS). The mass had increased in size for three to four weeks prior to presentation. At the time of examination the horse had been with the current owner for a year, a prepurchase examination had been performed, with no ocular abnormalities found. The horse was intended to be used for cross-country performance.

Investigations

Upon presentation the horse was in an excellent physical state of health. A complete ophthalmic exam of both eyes (OU) was performed using slit lamp biomicroscopy (Kowa portable slit-lamp SL-14, CR Medical, Austria) as well as direct and indirect ophthalmoscopy (Heine Omega 2C of Heine Optotechnik GmbH & Co KG, Germany). Thorough examination revealed that both eyes were visual and comfortable, with a normal menace response and dazzle reflex; direct and consensual pupillary light reflexes were normal OU. Detailed examination of OS revealed mildly hyperaemic conjunctivae. Upon eversion of the third eyelid under topical anaesthesia (Novain 0.4 per cent ‘Agepha’, Austria), a red-pink mass, about 1 cm in diameter and 4 mm thickness, on the bulbar aspect of the nictitating membrane was identified (Fig 1). Digital palpation of the enlargement revealed a dense mass with a slightly rough surface attached to the nictitating membrane by a stalk of 6–7 mm in diameter. The cornea of the left eye was unaffected by the mass and negative on fluorescein stain (Fluorescein Papier; Haag-Streit International, Switzerland). The remaining findings of the ocular examination included two bullet hole retinal lesions interpreted as inactive chorioretinitis in the right eye (OD) and a peripapillary coloboma OS.

FIG 1:

Photograph of the mass on the bulbar aspect of the third eyelid (courtesy of Dr Claudia Siedler)

In order to determine the aetiological origin of the mass a cytobrush sample and fine needle aspiration biopsy were taken and submitted to cytology. Analysis of both samples revealed infiltration of lymphatic cells and mild epithelial cell dysplasia. Due to the inconclusive results of cytology three incisional biopsy samples of the mass were obtained under standing sedation (0.6 mg/kg xylazine [Xylasol; Dr E Graub AG, Switzerland] and 0.01 mg/kg butorphanol [Butomidor; Richter Pharma, Austria]) as well as topical anaesthesia, and submitted for histopathological examination. The predominating cell types in all samples included lymphocytes, plasma cells and neutrophils, hence the cytological diagnosis chronic inflammation was concluded.

Differential diagnosis

Based on the results of cytology and histology, prolapse of retrobulbar fat tissue was eliminated from the list of differential diagnoses, rendering chronic inflammation the most likely cause for the mass. However a neoplastic process could not be completely excluded at this point.

Treatment

The horse was put on a two-week course of phenylbutazone 1 g orally once a day (BUTE Granulare; ACME, Italy) as well as topical bromfenac (Yellox; Croma-Pharma GmbH, Austria) and enrofloxacin (Floxal, Bausch & Lomb GmbH, Germany) OS twice a day. Upon representation two weeks after discharge from the hospital, the horse was in an excellent state of health. The mass of the nictitating membrane had slightly decreased in size, however marked reddening and protrusion of the third eyelid were still evident. An excisional biopsy of the mass was recommended to the owner, since there had been merely slight improvement of the condition, despite treatment with systemic and topical anti-inflammatory treatment. The owner agreed to have the mass removed under general anaesthesia, due to the horse's demeanour. Routine preanaesthetic blood tests revealed normal blood values apart from mild hypofibrinogenaemia (Fibrinogen 80 mg/dl; reference range 150–200 mg/dl). The mass of the third eyelid was excised by sharp dissection at the base of the stalk under routine general anaesthesia in lateral recumbency. The excised tissue was placed into a plastic container in 10 per cent neutral buffered formalin and submitted to pathology for further histopathological examination. The recovery phase was uneventful. Postoperative treatment consisted of systemic NSAID 1.1 mg/kg intravenously once a day (Finadyne Paste; Intervet, Germany) and topical bromfenac OS twice a day. The horse was re-examined one day post surgery. At that point the eye was comfortable, there was mild serosanguinous discharge from the surgery wound, but no fluorescein uptake of the cornea OS. The horse was discharged two days after mass removal. Postoperative treatment was continued for two weeks post discharge from the hospital.

Outcome and follow-up

Histopathological examination of the mass revealed small lymphoid round cells (slightly larger than red blood cells) with indented nuclei, dark dispersed nucleus chromatin, and minimal cytoplasm intermingled with larger lymphoid cells (2–3× the diameter of red blood cells) forming dense sheets. The larger cells showed round to oval vesicular nuclei with branched chromatin and occasionally nucleoli (Fig 2a,b). There was 0–1 mitosis per single high power field (400×). Immunohistochemistry was positive for CD 3 (T lymphocytes) (Fig 3) and CD 20 (B lymphocytes) (Fig 4). Furthermore in the periphery of the stroma follicle-like arrangements of small lymphocytes and areas, rich with plasma cells, neutrophil granulocytes, and macrophages, as well as proliferating fibroblasts were identified. The histopathological diagnosis was malignant lymphoma, subclassified by morphology and immunohistochemistry as a TCRLBCL. Based on the result of histopathology further staging was advised, however the owner declined further diagnostic workup.

FIG 2:

(a) Histopathological section stained with haematoxylin and eosin (bar=150 µm): Diffuse infiltration of the stroma of the third eyelid. (b) Histopathological section stained with haematoxylin and eosin (bar=40 µm): The stroma is infiltrated by small lymphocytes and larger neoplastic lymphoid cells

FIG 3:

Histopathological section stained with the T cell marker CD3 (bar=80 µm): many cells are strongly positive for CD3

FIG 4:

Histopathological section stained with the B-cell marker CD20 (bar=80 µm): there is strong cytoplasm CD20 labelling of many cells

The horse was re-examined by the first author (JR) 18 months post mass removal. Upon examination the horse was in an excellent state of health. No swelling of the mandibular lymph nodes was evident upon palpation. Diagnostic fine needle aspiration of the lymph nodes for further staging was declined by the owner. Mild fibrosis of the bulbar aspect of the third eyelid was evident upon eversion of the third eyelid, however there were no signs of tumour regrowth (Fig 5). The remaining findings of both eyes were identical to the initial examination.

FIG 5:

Follow-up photograph upon eversion of the third eyelid 18 months post initial presentation. No regrowth of the mass is evident

Discussion

Differential diagnoses for masses of the third eyelid which were relevant for this case include neoplasia (squamous cell carcinoma, adenocarcinoma of the accessory lacrimal gland and haemangiosarcoma) (Gearhart and others 2007, Giuliano 2011, Mathes and others 2011), granulation tissue as a result of chronic inflammation due to Thelazia lacrimalis infestation of the lacrimal duct of the accessory lacrimal gland (Giangaspero and others 1999, 2000) and prolapse of retrobulbar fat tissue (Giuliano 2011). A recent publication also describes the occurrence of a dermoid on the bulbar aspect of the third eyelid (Greenberg and others 2012). The latter differential diagnosis was not considered at the time of workup, as there was no evidence of hair follicles using biomicroscopy and the mass was clearly acquired in a later stage of life, evidenced by the history from the owner. Further diagnostics, including fine needle aspiration and incisional biopsy of the mass excluded retrobulbar fat prolapse, since no evidence of fat tissue was identified, rendering chronic inflammation and neoplasia the most likely causes of the mass. The diagnosis of T-cell-rich large B-cell lymphoma (TCRLBL) only became apparent upon excisional biopsy of the entire mass. Fine needle aspiration and incisional biopsy were both suggestive of an inflammatory process and did not indicate the presence of lymphoma. This fact corroborates the results of a publication from physician oncology, comparing the use of fine needle aspiration and excisional biopsy in 470 lymphoma patients, in which the diagnosis based on fine needle aspiration was confirmed by excisional biopsy in merely 2 per cent of cases (Hehn and others 2004).

TCRLBCL is the most common type of lymphoma in horses with prevalence as high as 33 per cent (Kelley and Mahaffey 1998). A more recent retrospective analysis with a large study population has revealed a prevalence of 43 per cent for TCRLBCL, followed by 22 per cent peripheral T cell lymphomas and 12.5 per cent diffuse large B-cell lymphomas. The remainder is distributed among diffuse large cell lymphomas, enteric associated T cell lymphoma, cutaneous T cell lymphoma, T-zone lymphoma, chronic lymphocytic leukaemia, anaplastic large cell lymphoma, acute lymphocytic leukaemia, T cell lymphoblastic lymphoma, follicular lymphoma, hepatosplenic lymphoma, and lymphoplasmacytic lymphoma (Durham and others 2012). TCRLBCL is also common in the cat. Prevalence of TCRLBCL in other species including cows, dogs and pigs are well below 5 per cent (Valli and others 2002). TCRLBCL account for 34 per cent of multicentric lymphomas, 71 per cent cutaneous lymphomas, and 30 per cent gastrointestinal lymphomas in horses of all lymphoma subtypes in each category (Durham and others 2012). To the authors’ knowledge, there is no information regarding the incidence of TCRLBCL of the ocular adnexa in horses.

TCRLBCL has a slow clinical course in the dog and the cat (Durham and others 2012). It seems to be the same in the horse according to the literature (Valli 2007) and the observations with no recurrence after 18 months. But the good outcome may also be due to the location of the lymphoma. For instance TCRLBCL located in the dorsolateral periorbital dermis of horses tends to recur after dissection (Valli 2007). A recent study compared the prognosis with the signalments and lesion locations of horses with extraocular lymphoma. Therein the histopathological classification of the type of lymphoma was not considered. However based on the results of that study, 54 per cent of 26 horses had lymphoma associated with the third eyelid, six of which were solely localised in the third eyelid, the remainder also affected other extraocular tissues. Horses with involvement of the eyelids or skin were 13 times more likely to have a poor outcome than horses affected merely by third eyelid, conjunctival and /or corneoscleral lymphoma (Schnoke and others 2013). This prognostic factor however was based on the anatomical location of the tumour, rather than the pathohistological classification. In physician medicine, the overall prognosis of ocular adnexal lymphomas is excellent (Jakobiec and Knowles 1989). However more recent studies have revealed an association of conjunctival lymphoid tumours with systemic lymphoma in 31 per cent of cases (Shields and others 2001). To the authors’ knowledge no study is available to compare species differences of ocular adnexal lymphomas with regards to prognosis.

There is no evidence of risk factors associated with the presence of TCRLBCL, however a recent case of Borrelia associated cutaneous pseudolymphoma in a horse, with histopathological features suggestive of TCRLBCL or cutaneous lymphoid hyperplasia was reported (Sears and others 2012). The involvement of infectious agents in the development of extraocular lymphoma has also been reported in physician medicine (Ferreri and others 2004). The excised mass of the horse in this report was therefore submitted for PCR and immunohistochemical analysis to detect Borrelia species, and Chlamydia species, the results were negative. Although an involvement of infectious agents in the development of TCRLBCL cannot be excluded, there is no reason to believe that the mass was caused by a specific pathogen in this case.

Treatment of neoplasia of the third eyelid is usually limited to surgical resection of the complete third eyelid. However tumour regression using systemic or topical steroids has been reported (Giuliano 2011). In a recent retrospective study 68 per cent of horses with surgical treatment of extraocular lymphoma had a good outcome (Schnoke and others 2013). However large representative studies, comparing surgical treatment with or without adjunctive therapy are still lacking. In this case surgical removal of the mass at the base of the stalk on the third eyelid resulted in a favourable outcome 18 months after surgery. However had there been evidence of the presence of lymphoma at the time of surgical intervention, the entire third eyelid would have been removed, to reduce the risk of tumour relapse. Upon receipt of the histopathological report, the owner decided to monitor the horse for evidence of tumour recrudescence, with no immediate removal of the remaining third eyelid or further staging. In the retrospective study by Rebhun and Del Piero, lymphosarcoma involving the eyes was diagnosed in 21 horses, 20 of which died or were euthanased within a period of six months after establishment of the diagnosis, due to systemic spread of the tumour. The remaining horse, which was free from lymphosarcoma in all other organs was treated with removal of both third eyelids and a conjunctival mass. The horse was alive upon follow-up six years after surgery (Rebhun and Del Piero 1998). However, alas no further immunophenotyping of the tumour was performed in that study.

A limitation of this report, is the lack of tumour staging, however no enlargement of the respective mandibular lymph node or presence of masses in other locations were evident at the time of hospitalisation or within the 18 month follow-up period. However the rapid onset of the mass prior to surgical removal and the long postoperative follow-up suggest a minimal risk of tumour relapse or metastasis at this stage.

Acknowledgments

The authors thank Dr Claudia Siedler for providing the photograph of the initial examination and her contribution to the case workup.

  • Received March 24, 2014.
  • Revision received April 22, 2014.
  • Accepted April 24, 2014.

References

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Footnotes

  • Contributors JR was involved in the case workup and surgery, and is the primary author of the manuscript. AF-B performed gross pathology and histopathology as well as immunohistochemistry, provided important input to the manuscript and proof-read the manuscript. BN supervised the case work-up, surgery, provided important input to the manuscript and proof-read the manuscript.

  • Competing interests None.

  • Provenance and peer review Not commissioned; externally peer reviewed.

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