Undifferentiated carcinoma of the nasal tissues in the common marmoset

329

J. COMP. PATH. 1984..VOL. 94.

UNDIFFERENTIATED TISSUES IN

THE

CARCINOMA COMMON

OF THE MARMOSET

NASAL

BY

M. BASKERVILLE Chemical Defence Estabfishment,

Porton Down, Sal&q,

W&shire

A. BASKERVILLE Public Health Laboratov

Service, Centre

for Applied

Microbiolo,g

and Research, Porton Down,

Salisbury,

Wiltshire

and B. W. MANKTELOW Department

of Vet&nary

Pathology and Public Health,

Massey Unirwsity,

Palmerston North, New Zealand

INTRODUCTION

The common marmoset (Cullithrix jacchus) is now being used increasingly in many fields of medical research, including that of viral oncogenesis. The marmoset is more susceptible to many oncogenic viruses than other speciesand is especially suitable for research with viral isolates of human origin (Wolfe and Deinhardt, 1978). It is therefore important that full investigation of naturally occurring tumours of marmosets is carried out. Clinical observations of a colony of some 450 breeding and experimental marmosets showed one of the commonest conditions to be facial swelling, sometimes accompanied by nasal discharge. Many of these lesions proved to be abscesses,frequently originating at the root of an upper canine tooth. However, over a period of 1 year, 3 animals developed tumours in the nasal cavity which were associated with facial swelling, nasal discharge or impaired vision. The tumours extended into the orbit and into the frontal and ethmoidal sinuses. All were undifferentiated carcinomas and all metastasized. Surveys of naturally occurring tumours of non-human primates (O’Gara and Adamson, 1972; Benirschke, Garner and Jones, 1978) show that neoplasia has been rarely recorded in marmosets and that, in other primates, tumours of the nasal tissues have only been described 3 times; 2 were adenocarcinomas and one was a squamous carcinoma (Appleby, 1969; Cicmanec, Neubauer, Wallen, Darrow and Rabin, 1974; Brown, Cole, Berg, Chiang, Chang and Banknieder, 1977). This report describes the clinical and pathological features of the nasal tumours in the marmosets. cAsE HISTORIES All 3 occurred in a group of 450 common marmosets comprising both a breeding colony and those under experimental conditions. All 3 marmosets OOZl-9975/84/030329+10

$03.00/O

0 1984 Academic Press Inc. (London) Limited

330

M.

BASKERVILLE

etal.

had been used for behavioural studies but only 2 had received drugs; Case 2 had been given physostigmine and Case 3 atropine and benactyzine intramuscularly. Two cases occurred in marmosets caged singly in the same room, whilst the third affected animal had been culled 12 months earlier. All 3 affected animals were aged 4 to 3 years, weighed 300 to 320 g and although possibly of common genetic stock they were not otherwise related. Case I Female Distortion of the bridge of the nose and left eyebrow by a hard swelling developed over a period of weeks. The conjunctiva of the left eye was swollen and protruded at the medial canthus and there was a purulent discharge from the eye and left nostril. Treatment with parenteral ampicillin for 5 days and oxytetracycline orally for 5 days was ineffective. Good general health was maintained until the animal was destroyed by injection of ketamine hydrochloride (Vetalar, Parke, Davis). Immediately before death haematological values were normal. Case 2 Female A persistent mucopurulent, often blood-stained, discharge from the left nostril for 2 weeks was the only clinical sign of ill health. The animal then rapidly became weak, the total WBC count was low (2.9 x lO”/l), and there was severe dyspnoea. Removal of the left upper canine tooth and oral treatment with oxytetracycline for 5 days caused no clinical improvement and the animal was destroyed. Case 3 Male The earliest sign of ill health was again a unilateral nasal discharge. There also appeared to be complete loss of vision of one eye, although there was no detectable eye lesion or swelling of the periorbital tissues. Three weeks later a large ulcer was found in the posterior portion of the hard palate and the animal was destroyed.

MATERIALS

AND

METHODS

At necropsy the brain was removed and the remainder of the head, with the facial skin intact, was fixed in 10 per cent buffered neutral formalin. The brain and portions of other organs were similarly fixed. The heads were later decalcified in Gooding and Stewart’s fluid and coronal slices made anteroposteriorly. All tissues were then processed by standard procedures and embedded in paraffin wax. Sections cut 5 pm thick were stained with haematoxylin and eosin (HE) and selected sections were stained by Fontana and Schmorl’s methods for melanin and subjected to Mayer’s melanin bleach technique. Others were stained by Gordon and Sweet’s method for reticulin and Verhoeff-van Gieson for connective tissue fibres. Selected portions of the paraffin blocks were also processed for electron microscopy according to a schedule devised previously (Baskerville, Thomas, Wood and Harris, 1974). After embedding in.Araldite,

1 pm-thick

sections

were

cut with

glass knives

for selection

of suitable

NASAL

CARCINOMA

OF

MARMOSETS

331

areas by light microscopy. Silver sections of these were cut with a diamond knife and stained with uranyl acetate and lead citrate.

RESULTS

Necropsy Findings Case 1. There was a firm swelling on the bridge of the nose which involved the left eye. The conjunctiva protruded across the orbit, completely obscuring the eyeball. Examination of coronal slices of the face and head showed a hard brown pigmented mass 1.5 cm in its greatest dimension occupying the dorsal aspect of the nasal cavity posterior to the nares and extending into the frontal and ethmoidal sinuses and orbit. The nasal septum was for the most part obliterated by the tumour mass, but where recognizable it was displaced laterally. Turbinate bones could not be distinguished. There were no lesions in other -organs. C’ase 2. There were no external lesions, but on making coronal slices of the decalcified head an irregular greyish-white mass up to 1.7 cm was seen occupying the entire nasal cavity. It involved the palatal, lateral and dorsal surfaces and distorted the dorsal remnants of the nasal septum. Turbinate bones could not be discerned. Tumour tissue was also invading the frontal and ethmoidal sinuses. Small areas of consolidation were present in all lobes of the lungs, but other organs were unremarkable. Case 3. On sectioning the head, greyish-white tumour tissue was found to fill the nasal cavity and protrude through the posterior wall of the frontal sinus into the cranium. Similar tissue was present at an ulcerated site on the oral aspect of the posterior edge of the hard palate. Two white nodules, 8 mm and 2 mm in diameter, were present in the liver, but other organs were normal. Histopathology C,ase 1. The tumour mass consisted of large, solid groups of basophilic cells with some peripheral palisading and a high mitotic rate (Fig. 1). Between the solid cell nests there were small clumps and ribbons of similar cells invading the fibrous tissue stroma. Many cells were columnar, with a round, vesicular nucleus, but at the periphery and in some islands the cells were polygonal (Fig. 2) or spindle shaped. Cell boundaries were often indistinct. In some areas, the polygonal cells contained brown pigment in the cytoplasm and hadhyperchromatic nuclei. The brown pigment was identified as melanin by positive reactions in the Schmorl’s and Fontana methods and by Mayer’s melanin bleach method. There was no evidence of glandular acini, keratinization, intercellular bridges or prickle cell formation in any part of the tumour. Central zones of the larger nodules contained loose myxomatous tissue and areas of necrosis and haemorrhage. At a few sites, connections were demonstrable between the tumour and the respiratory pseudostratified columnar epithelium of the remnants of the nasal septum and turbinate bones. Dense masses of multiplying tumour cells formed plaques superficial to the basement

332

Fig. 1. Nasal tumour Fig. 2. Case 1. Tumour

M.

BASKERVILLE

et

showing

solid islands of neoplastic

from Case 1 marmoset nests contain

columnar

and polygonal

al.

cells with numerous

tissue. HE mitotic

x 150.

figures.

HE

me :mbrane at several points (Figs 3 and 4). The respiratory epithelium 3liferated on either side of the plaques and was multi-layered. A few n Pr’ nu cleate giant cells were present in the tumour nodules (Fig. 5). The tur wa s surrounded by a thin, incomplete fibrous capsule, but had invaded or-1Sts, where it was causing erosion of the lachrymal and ethmoid boner in\ Jading the eyeball (Fig. 6). It had also invaded almost to the oral epithe of the hard palate and was present around the tooth roots and periods

< 300.

had ultiiour 10th and ium tntal

NASAL

CARCINOMA

OF

MARMOSETS

Fig. 3. Case 1. Area ofdeveloping carcinoma in respiratory epithelium ofinferior turbinate. Proliferation of cells has produced a nodule between the basement membrane and the ciliated epithelium. HE x 300. Fig. 4. Case 1. Early neoplastic change in respiratory tissue by cell groups. HE x 325.

epithelium

of turbinate,

with invasion

of underlying

membranes and was infiltrating the nasal and frontal bones, with destruction of osseous tissue. Tumour cell nests were also present at the base of one eyelid. Three small metastases (1 mm) were found in the lungs. These had the same histological structure as the main tumour and in each there was a number of mitotic figures. A turnout- embolus was found in the lumen of a branch of the pulmonary artery. Other organs were histologically normal.

334

Fig. 5 Multinucleate Fig. 6 Case 1, showing HE x45.

M.

giant cells in tumour a margin

BASKERVILLE

mass of Case 1. HE

of th? turnour

et al.

x 100

mass; with incomplc~

capsule,

invading

the orbital

t

appearance was similar to that described al3ove c’,ase 2. The histological and the pigmentation was much less exce :pt that there was no haemorrhage nodules were of the :solid and confined to certain cell islands. The tumour varil ety, with many mitoses and clumps and ribbons of columnar and spi ndle giant cells were present. There was cells streaming into the stroma. Occasional exte nsive erosion of bone of the turbinate, frontal, palatine, lachrymal and

NASAL

CARCINOMA

OF

335

MARMOSETS

ethmoid bones and destruction of much of the cartilage of the nasal septum. The periodontal tissues of several incisor, premolar and molar teeth were invaded by tumour. As in Case 1, connections could be seen at several sites between neoplastic tissue and carcinoma in the respiratory epithelium of the nasal septum and turbinate bones. At these and neighbouring sites the epithelium was hyperplastic and multilayered. The tumour was partially- surrounded by an incomplete fibrous capsule. which was thick in places hut absent in others. In the lungs a single almost spherical 1 mm diameter metastasis was present peripherally. The tumour nuclei contained a prominent nucleolus and there were a number of mitotic figures. There were also a few areas of hronchopneumonia with polymorphonuclear leucocytes in groups of alveoli. Lesions were not present in other tissues. Case 3. The tumour had a similar histological structure to that seen in the other 2 animals, with solid masses of poorly differentiated polyhedral cells with pale, vesicular nuclei and numerous mitoses. Tumour had invaded the nasal. frontal, lachrymal and ethmoid bones and destroyed much of the nasal septum and turbinates. It was particularly extensive in the hard palate, where it had eroded completely through the bone to the ulcerated oral surface. The appearance ofthe 2 metastases in the liver was also ofan undifferentiated carcinoma (Fig. 7). There was some palisading of tumour cells, many mitotic figures, central necrosis and compression of the surrounding hepatic tissue. A number of phagocytic histiocytes with translucent cytoplasm containing cell debris were scattered in the larger tumour nodule.

Fig.

7. Case 3; tumour metastasis tissue. HE x 500.

in liver,

composed

of solid

islands

of’rolumnar

cells

compressing

hcpatic

336

M.

BASKERVILLE

et

d.

Electron Microsco$y Tumour cells from all 3 cases had large nuclei with few indentations and the chromatin was dispersed evenly in small clumps throughout the nuclei. Cell borders were regular and, despite only moderate preservation of cellular detail, desmosomes were visible at several points along the plasma membrane of most contiguous cells. Bundles of intracytoplasmic tonofibrils were present in a high proportion of the cells of all 3 primary tumours and also in the liver metastases. DISCUSSION

The tumours described here are unusual in several respects. Tumours of the nasal cavity are uncommon in animals (Cotchin, 1967 ; Brodey, 1970; Stunzi and Hauser, 1976; Moulton, 1978) and only 3 have been recorded in the nasal tissues in non-human primates, none in marmosets; 2 of these were adenocarcinomas and one a squamous carcinoma (Appleby, 1969 ; Cicmanec et al., 1974; Brown et al., 1977). Also, although C. jacchus has been kept in captivity for some years, there are very few reports of any naturally occurring tumours in this species (Benirschke et al., 1978). Nasal tumours in man are usually squamous carcinomas, transitional carcinomas, adenocarcinomas or undifferentiated carcinomas (Payling-Wright and Symmers, 1966; Shanmugaratnam and Muir, 1967; Osborn, 1970; Ashley, 1978) and the undifferentiated variety has formed a high proportion of some reported series (Shanmugaratnam and Muir, 1967). In our marmoset specimens the cells were predominantly undifferentiated ; there were no keratohyalin granules or keratin and no glandular acini, thereby excluding classical squamous carcinoma and adenocarcinoma as a diagnosis. The absence of areas of obvious squamous differentiation and keratin whorls also excluded basi-squamous tumour. There were, however, readily demonstrable points of continuity between carcinoma in the epithelium and underlying neoplasm. In addition, electron microscopy showed that many contiguous cells were joined by desmosomes and had bundles of cytoplasmic tonofibrils. This confirms their epithelial origin, since both squamous cells and the basal cells of the respiratory epithelium contain tonofibrils. The tumours appeared to arise from the ciliated pseudostratified epithelium covering the turbinate bones and nasal septum. As stated by Shanmugaratnam and Muir ( 1967)) undifferentiated carcinoma may arise from any of the types of epithelium normally present in the nasal cavity. Since the different types occur in close proximity in the nasal mucosa it is impossible to be certain of the exact site of origin. The presence of melanin pigment in parts of 2 of the tumours does not invalidate the diagnosis, since melanin is present normally in cells of the oronasal stratified squamous and respiratory epithelia of the marmoset. The tumour in Case 1 was so heavily pigmented that it was thought initially to be a melanoma. The behaviour of the tumours was similar to that of nasal carcinomas in man, since all metastasized, as well as being highly invasive. The relatively small size

NASAL

CARCINOMA

OF

MARMOSETS

337

of the marmoset’s nasal cavity explains the ease with which the tumours eroded the surrounding bones and invaded the orbits and sinuses. The skull of the marmoset is of a relatively primitive simian type; the orbital cavities are long and occupy the entire lateral boundaries of the very narrow nasal cavity (Beattie, 1927). The fact that 2 of these cases occurred within a short time is interesting and suggests a common cause. However, apart from sharing the same room and food the animals had nothing in common and were not related. Histological examination of the nasal tissues of 6 other marmosets of similar age which had been housed in the same rooms as Cases 1 to 3 did not show any pathological changes in the mucosa other than a localized mild rhinitis in one animal. SUMMARY

The occurrence of undifferentiated carcinoma of the nasal tissues in 3 adult marmosets is described. The tumours appeared to arise from the respiratory epithelium of the turbinate bones and nasalseptum. They were poorly differentiated, invaded the paranasal sinuses and hard palate and metastasized to the lungs or liver.

ACKNOWLEDGMENTS

We are grateful to Mr A. B. Dowsett, Mrs Iris Francis, Mrs E. EIphick, Diane Harwood for technical assistance.

and Miss

REFERENCES

Appleby, E. C. (1969). Tumours in captive wild animals: some observations and comparisons. Acta
338

M.

et al.

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Moulton, J. E. (1978). Turnours in Domestic Animals. IJniversity of California Press, Berkley. O’Gara, R. W., and Adamson, R. H. (1972). Spontaneous and induced neoplasmsin nonhuman primates. In Pathology of Simian Primates. Part I. R. N. T. W. Fiennes, Ed., Karger, Basle, pp. 190-238. Osborn, D. A. (1970). Nature and behaviour of transitional tumours in the upper respiratory tract. Cancer, 25, 50-60. Payling-Wright, G., and Symmers, W. St. C. (1966). Systemic Pathology. Vols I and II Longmans, London, pp. 303 and 1490. Shanmugaratnam, K., and Muir, C. S. (1967). Nasopharyngeal carcinoma origin and structure. In Cancer of the Nusoljharynx. C. S. Muir and K. Shanmugaratnam, Eds, Munksgaard, Copenhagen, pp. 153-162. Stunzi, H., and Hauser, B. (1976). Tumours of the nasal cavity. In WHO Bulletin. International Histological Classification of Tumows of Domestic Animals. Part 2, 53, 257-263. Wolfe, L. G., and Deinhardt, F. (1978). Overview of viral oncology studiesin Saguinus and Callithrix species.Primates in Medicine, 10, 96-I 18. [Receivedfor publication,

April

lst, 19831