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Undifferentiated High-Grade Pleomorphic Sarcoma in a California Sea Lion (Zalophus californianus)
J. Comp. Path. 2011, Vol. 144, 200e203
Available online at www.sciencedirect.com
www.elsevier.com/locate/jcpa
SHORT PAPER
Undifferentiated High-Grade Pleomorphic Sarcoma in a California Sea Lion (Zalophus californianus) B. Martı´nez-Ferna´ndez*, M. J. Garcı´a-Iglesias*, S. Borraga´n-Santos†, J. Espinosa-Alvarez* and C. Pe´rez-Martı´nez* * Pathological Anatomy Section, Animal Health Department, Veterinary School, University of Leo´n, 24007 Leo´n and † Veterinary Services, Nature Reserve of Caba´rceno, 39693 Obrego´n, Cantabria, Spain
Summary A tumour located in the pectoral region and the left front flipper was observed in a 29-year-old female California sea lion (Zalophus californianus) that died following signs of respiratory disease and inappetence. Metastases were present in the lung and adrenal gland. The histological pattern of the tumour was variable. In some areas the tumour consisted of pleomorphic fibroblast-like cells arranged in a storiform pattern, while in other areas it comprised oval or polygonal cells with round to oval nuclei and some bizarre cells arranged in an alveolar pattern. Occasionally, multinucleated giant cells were observed. Immunohistochemically, the neoplastic cells only expressed vimentin. On the basis of the microscopical and immunohistochemical features the tumour was diagnosed as an undifferentiated high-grade pleomorphic sarcoma. This type of neoplasm with disseminated involvement of other organs is rare in all species and has never been reported in California sea lions. Ó 2010 Elsevier Ltd. All rights reserved. Keywords: histopathology; immunohistochemistry; sea lion; undifferentiated sarcoma
The current World Health Organization (WHO) classification of tumours considers the existence of an undifferentiated and unclassifiable category of pleomorphic sarcomas, defined as a group of pleomorphic high-grade sarcomas that fundamentally represents a diagnosis of exclusion (Fletcher, 2006). In the past, some cases of undifferentiated high-grade pleomorphic sarcomas (UPS) were diagnosed as pleomorphic malignant fibrous histiocytomas (MFH), a term that has been kept as being synonymous with UPS in the current WHO classification (Dei Tos, 2006). The former term will probably be removed in future classifications (Dei Tos, 2006) because current evidence shows that this tumour does not have true histiocytic origin or differentiation (Fletcher, 2006). Clinically, UPS appear as deep masses with progressive and eventually
Correspondence to: C. Pe´rez-Martınez (e-mail: [email protected]). 0021-9975/$ - see front matter doi:10.1016/j.jcpa.2010.06.005
rapid growth (Edson et al., 2008). Microscopically, this group of lesions shows a heterogeneous pattern with frequent pleomorphism (Enzinger and Weiss, 1995). Immunohistochemical findings are nonspecific, but expression of vimentin marks UPS as a mesenchymal tumour (Edson et al., 2008). Other immunohistochemical markers are rarely positive, but this does not mean that the tumour is well differentiated (Dei Tos, 2006). This type of tumour is not uncommon in man (Enzinger and Weiss, 1995), but few cases have been reported in animals (Thoolen et al., 1992; Gage et al., 1995; Pe´rez-Martı´ nez et al., 2000). The aim of the present report is to describe a case of UPS with metastases in a California sea lion (Zalophus californianus). A 29-year-old female California sea lion from a nature reserve in Spain developed multiple nodules in the pectoral region and the left front flipper. The animal died following signs of respiratory disease and inappetence. Ó 2010 Elsevier Ltd. All rights reserved.
Sarcoma in a California Sea Lion
Necropsy examination revealed multiple slightly nodular, grey-white, moderately firm masses, up to 5 cm in largest dimension in the pectoral region and the left front flipper. These masses invaded the subcutis and skeletal muscle, but the skin above them was of normal gross appearance. One adrenal gland and both pulmonary lobes were markedly enlarged and contained numerous grey-white, rounded masses, ranging from 0.2 to 3 cm in diameter, within the parenchyma. The remainder of the thoracic and abdominal organs, including the urogenital tract, appeared grossly normal. Tissue specimens were collected and fixed in 10% neutral buffered formalin and referred to the School of Veterinary Medicine at the University of Leo´n (Spain). These tissue specimens were processed routinely, embedded in paraffin wax, sectioned (4 mm) and stained with haematoxylin and eosin (HE). To further characterize the mass, staining was performed with Masson’s trichrome, periodic acideSchiff (PAS) and methenamine silver. Frozen sections from fixed neoplastic tissue were stained with Sudan black B for fat. Selected sections were dewaxed and immunohistochemically labelled by the avidinebiotineperoxidase complex (ABC) method. The primary reagents used were: rabbit polyclonal antibodies specific for S-100 (DAKO, Glostrup, Denmark; Catalogue number Z0311; 1 in 1,000 dilution); glial fibrillary acidic protein (GFAP; DAKO; Catalogue number Z0334; 1 in 5,000 dilution); CD3 (DAKO; Catalogue number A0452; 1 in 100 dilution); lysozyme (DAKO; Catalogue number A0099; 1 in 200 dilution) and mouse monoclonal antibodies specific for pan cytokeratin (Progen, Heidelberg, Germany; clone AE1/AE3; 1 in 100 dilution); CD31 (DAKO; clone JC70A; 1 in 200 dilution); vimentin (DAKO; clone V9; 1 in 1,000 dilution); smooth muscle actin (Sigma Aldrich, St. Louis, Missouri; clone 1A4; 1 in 5,000 dilution); CK8, 18 (Becton Dickinson,
201
Franklin Lakes, New Jersey; clone Cam 5.2; 1 in 5 dilution) and desmin (Thermo Fisher Scientific, Fremont, California; clone D33; 1 in 100 dilution). Microscopically, the nodules in the subcutis and skeletal muscle of the pectoral region and the left front flipper showed an infiltrative and heterogeneous growth pattern. In some fields the tumour consisted of pleomorphic spindle-shaped fibroblast-like cells with storiform arrangement and bizarre giant cells (Fig. 1). In other areas there were oval or polygonal cells with round to oval nuclei and some bizarre cells with a single nucleus in an alveolar pattern (Fig. 2). Occasional multinucleated giant cells and numerous abnormal mitoses were observed. Intercellular collagen production was not prominent, although there was more collagen evident in areas with a storiform pattern. Additionally, argentophilic fibres surrounded each tumour cell or small groups of cells in regions of storiform arrangement or larger groups of cells in areas of alveolar arrangement. Other findings included extensive areas of necrosis in the centre of large tumour nodules and infiltration of lymphocytes and plasma cells. In addition, large cells with PASand Sudan black-negative vacuoles and irregular and large or bizarre nuclei appeared adjacent to the areas of necrosis. The masses in the lungs and adrenal gland appeared microscopically as poorly demarcated, infiltrative, highly cellular tissue, consisting of foci and cords of pleomorphic spindle-shaped fibroblast-like cells similar to those described above. Large tumour emboli were present in the lumina of blood vessels in the nodules located in the flipper and pectoral region, as well as in the lung and adrenal gland. The microscopical appearance was not considered diagnostic and so immunohistochemistry (IHC) was performed. Although the antibodies used had not
Fig. 1. Flipper; California sea lion. Pleomorphic spindle-shaped fibroblast-like cells arranged in a storiform pattern. Inset: detail of bizarre cells and aberrant mitoses. HE. 50, 100 (inset).
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B. Martınez-Fern andez et al.
Fig. 2. Flipper; California sea lion. Oval or polygonal cells showing an alveolar pattern. HE. 50.
been validated for pinnipeds, normal cell types adjacent to the neoplasm were appropriately labelled in each section with each antibody. Immunohistochemically, the neoplastic cells had strong and diffuse cytoplasmic expression of vimentin, but were negative for the remaining antibodies. Expression of vimentin (Fig. 3) and absence of labelling for cytokeratins (Fig. 4) suggested a mesenchymal origin. The absence of lysozyme expression suggested that the tumour was not of histiocytic origin. Lack of expression of desmin and smooth muscle actin confirmed that the tumour was not derived from muscular tissue. Negative immunoreactivity for S100 and GFAP differentiated this tumour from malignant Schwannoma. Absence of S100 expression would also suggest that the tumour in the present case was not derived from lipoblasts
Fig. 3. Flipper; California sea lion. Expression of vimentin by the neoplastic cells. IHC. 10.
Fig. 4. Lung; California sea lion. The neoplastic cells exhibit no immunoreactivity for pan cytokeratin. Normal bronchial epithelium and glands show appropriate cytokeratin expression. IHC. 10.
(Gebhard et al., 2002; Takeuchi et al., 2007); however, normal lipoblasts also failed to show immunoreactivity for S100. The diagnosis of liposarcoma could be excluded because there were no cells filled with lipid droplets (negative Sudan black B staining). The tumour nodules located in the flipper were presumed to be the primary site of the neoplasm because these pleomorphic sarcomas in man often arise in the extremities (Fletcher, 2006) and the lungs are a common site for metastasis (Massi et al., 2004). The presentation of this tumour in a mature sea lion is consistent with the prevalence of these lesions in older people (Fletcher, 2006). Similarly, the storiform microscopical pattern of growth is also the most common histological pattern in the human tumours, accounting for almost two-thirds of all cases (Enzinger and Weiss, 1995). With regard to outcome, it has been suggested that such a histological background constitutes a negative prognostic factor (Enzinger and Weiss, 1995), and the California sea lion in this report died shortly after becoming ill, with pathological findings suggestive of a particularly aggressive tumour. Therefore, UPS appear to have a similar clinical presentation, pathological appearance and biological behaviour in man and the California sea lion. A variety of tumours have been reported in the California sea lion. These involve different organ systems, but most are of epithelial origin, while mesenchymal tumours are less commonly reported (Newman and Smith, 2006). One undifferentiated sarcoma of possible histiocytic origin has been reported affecting the brain of a California sea lion (Gage et al., 1995), but the present report is the first of a disseminated metastatic undifferentiated sarcoma in this species.
Sarcoma in a California Sea Lion
References Dei Tos AP (2006) Classification of pleomorphic sarcomas: where are we now? Histopathology, 48, 51e62. Edson A, Magalhaes de Pa¨dua J, Vicentini EL, Marchesan MA, Comelli RC et al. (2008) Oral undifferentiated high-grade pleomorphic sarcoma: report of a case. Oral Surgery, Oral Medicine, Oral Pathology, Oral Radiology and Endodontology, 105, 37e40. Enzinger FM, Weiss SW (1995) Malignant fibrohistiocytic tumours. In: Soft Tissue Tumours, 3rd Edit., FM Enzinger, SW Weiss, Eds., Mosby, St Louis, pp. 351e380. Fletcher CDM (2006) The evolving classification of soft tissue tumours: an update based on the new WHO classification. Histopathology, 48, 3e12. Gage LJ, Trupkiewicz J, Samansky T (1995) Hydrocephalus in an aged California sea lion (Zalophus californianus). Proceedings of the International Association for Aquatic Animal Medicine, 26, 9e10. Gebhard S, Coindre JM, Michels JJ, Terrier P, Bertrand G et al. (2002) Pleomorphic liposarcoma. Clinicopathologic and immunohistochemical study of 63 cases. A study of the French sarcoma group. American Journal of Surgical Pathology, 26, 601e616.
203
Massi D, Beltrami G, Capanna R, Franchi A (2004) Histopathological re-classification of extremity pleomorphic soft tissue sarcoma has clinical relevance. European Journal of Surgical Oncology, 30, 1131e1136. Newman SJ, Smith SA (2006) Marine mammal neoplasia: a review. Veterinary Pathology, 43, 865e880. Pe´rez-Martı´ nez C, Garcı´ a Ferna´ndez RA, Reyes Avila LE, Pe´rez-Pe´rez V, Gonza´lez N et al. (2000) Malignant fibrous histiocytoma (giant cell type) associated with a malignant tumour in the salivary gland of a dog. Veterinary Pathology, 37, 350e353. Takeuchi A, Nakano K, Nishimuta Y, Kayikawa S, Ito K et al. (2007) Spontaneously occurring subcutaneous tissue sarcoma in a Sprague-Dawley rat. Journal of Toxicologic Pathology, 20, 161e166. Thoolen RJMM, Vos JH, Van der Linde-Spiman JS, De Weger RA, Van Unnik JAM et al. (1992) Malignant fibrous histiocytoma in dogs and cats: an immunohistochemical study. Research in Veterinary Science, 53, 198e204.
March 9th, 2010 ½ Received, Accepted, June 27th, 2010
Available online at www.sciencedirect.com
www.elsevier.com/locate/jcpa
SHORT PAPER
Undifferentiated High-Grade Pleomorphic Sarcoma in a California Sea Lion (Zalophus californianus) B. Martı´nez-Ferna´ndez*, M. J. Garcı´a-Iglesias*, S. Borraga´n-Santos†, J. Espinosa-Alvarez* and C. Pe´rez-Martı´nez* * Pathological Anatomy Section, Animal Health Department, Veterinary School, University of Leo´n, 24007 Leo´n and † Veterinary Services, Nature Reserve of Caba´rceno, 39693 Obrego´n, Cantabria, Spain
Summary A tumour located in the pectoral region and the left front flipper was observed in a 29-year-old female California sea lion (Zalophus californianus) that died following signs of respiratory disease and inappetence. Metastases were present in the lung and adrenal gland. The histological pattern of the tumour was variable. In some areas the tumour consisted of pleomorphic fibroblast-like cells arranged in a storiform pattern, while in other areas it comprised oval or polygonal cells with round to oval nuclei and some bizarre cells arranged in an alveolar pattern. Occasionally, multinucleated giant cells were observed. Immunohistochemically, the neoplastic cells only expressed vimentin. On the basis of the microscopical and immunohistochemical features the tumour was diagnosed as an undifferentiated high-grade pleomorphic sarcoma. This type of neoplasm with disseminated involvement of other organs is rare in all species and has never been reported in California sea lions. Ó 2010 Elsevier Ltd. All rights reserved. Keywords: histopathology; immunohistochemistry; sea lion; undifferentiated sarcoma
The current World Health Organization (WHO) classification of tumours considers the existence of an undifferentiated and unclassifiable category of pleomorphic sarcomas, defined as a group of pleomorphic high-grade sarcomas that fundamentally represents a diagnosis of exclusion (Fletcher, 2006). In the past, some cases of undifferentiated high-grade pleomorphic sarcomas (UPS) were diagnosed as pleomorphic malignant fibrous histiocytomas (MFH), a term that has been kept as being synonymous with UPS in the current WHO classification (Dei Tos, 2006). The former term will probably be removed in future classifications (Dei Tos, 2006) because current evidence shows that this tumour does not have true histiocytic origin or differentiation (Fletcher, 2006). Clinically, UPS appear as deep masses with progressive and eventually
Correspondence to: C. Pe´rez-Martınez (e-mail: [email protected]). 0021-9975/$ - see front matter doi:10.1016/j.jcpa.2010.06.005
rapid growth (Edson et al., 2008). Microscopically, this group of lesions shows a heterogeneous pattern with frequent pleomorphism (Enzinger and Weiss, 1995). Immunohistochemical findings are nonspecific, but expression of vimentin marks UPS as a mesenchymal tumour (Edson et al., 2008). Other immunohistochemical markers are rarely positive, but this does not mean that the tumour is well differentiated (Dei Tos, 2006). This type of tumour is not uncommon in man (Enzinger and Weiss, 1995), but few cases have been reported in animals (Thoolen et al., 1992; Gage et al., 1995; Pe´rez-Martı´ nez et al., 2000). The aim of the present report is to describe a case of UPS with metastases in a California sea lion (Zalophus californianus). A 29-year-old female California sea lion from a nature reserve in Spain developed multiple nodules in the pectoral region and the left front flipper. The animal died following signs of respiratory disease and inappetence. Ó 2010 Elsevier Ltd. All rights reserved.
Sarcoma in a California Sea Lion
Necropsy examination revealed multiple slightly nodular, grey-white, moderately firm masses, up to 5 cm in largest dimension in the pectoral region and the left front flipper. These masses invaded the subcutis and skeletal muscle, but the skin above them was of normal gross appearance. One adrenal gland and both pulmonary lobes were markedly enlarged and contained numerous grey-white, rounded masses, ranging from 0.2 to 3 cm in diameter, within the parenchyma. The remainder of the thoracic and abdominal organs, including the urogenital tract, appeared grossly normal. Tissue specimens were collected and fixed in 10% neutral buffered formalin and referred to the School of Veterinary Medicine at the University of Leo´n (Spain). These tissue specimens were processed routinely, embedded in paraffin wax, sectioned (4 mm) and stained with haematoxylin and eosin (HE). To further characterize the mass, staining was performed with Masson’s trichrome, periodic acideSchiff (PAS) and methenamine silver. Frozen sections from fixed neoplastic tissue were stained with Sudan black B for fat. Selected sections were dewaxed and immunohistochemically labelled by the avidinebiotineperoxidase complex (ABC) method. The primary reagents used were: rabbit polyclonal antibodies specific for S-100 (DAKO, Glostrup, Denmark; Catalogue number Z0311; 1 in 1,000 dilution); glial fibrillary acidic protein (GFAP; DAKO; Catalogue number Z0334; 1 in 5,000 dilution); CD3 (DAKO; Catalogue number A0452; 1 in 100 dilution); lysozyme (DAKO; Catalogue number A0099; 1 in 200 dilution) and mouse monoclonal antibodies specific for pan cytokeratin (Progen, Heidelberg, Germany; clone AE1/AE3; 1 in 100 dilution); CD31 (DAKO; clone JC70A; 1 in 200 dilution); vimentin (DAKO; clone V9; 1 in 1,000 dilution); smooth muscle actin (Sigma Aldrich, St. Louis, Missouri; clone 1A4; 1 in 5,000 dilution); CK8, 18 (Becton Dickinson,
201
Franklin Lakes, New Jersey; clone Cam 5.2; 1 in 5 dilution) and desmin (Thermo Fisher Scientific, Fremont, California; clone D33; 1 in 100 dilution). Microscopically, the nodules in the subcutis and skeletal muscle of the pectoral region and the left front flipper showed an infiltrative and heterogeneous growth pattern. In some fields the tumour consisted of pleomorphic spindle-shaped fibroblast-like cells with storiform arrangement and bizarre giant cells (Fig. 1). In other areas there were oval or polygonal cells with round to oval nuclei and some bizarre cells with a single nucleus in an alveolar pattern (Fig. 2). Occasional multinucleated giant cells and numerous abnormal mitoses were observed. Intercellular collagen production was not prominent, although there was more collagen evident in areas with a storiform pattern. Additionally, argentophilic fibres surrounded each tumour cell or small groups of cells in regions of storiform arrangement or larger groups of cells in areas of alveolar arrangement. Other findings included extensive areas of necrosis in the centre of large tumour nodules and infiltration of lymphocytes and plasma cells. In addition, large cells with PASand Sudan black-negative vacuoles and irregular and large or bizarre nuclei appeared adjacent to the areas of necrosis. The masses in the lungs and adrenal gland appeared microscopically as poorly demarcated, infiltrative, highly cellular tissue, consisting of foci and cords of pleomorphic spindle-shaped fibroblast-like cells similar to those described above. Large tumour emboli were present in the lumina of blood vessels in the nodules located in the flipper and pectoral region, as well as in the lung and adrenal gland. The microscopical appearance was not considered diagnostic and so immunohistochemistry (IHC) was performed. Although the antibodies used had not
Fig. 1. Flipper; California sea lion. Pleomorphic spindle-shaped fibroblast-like cells arranged in a storiform pattern. Inset: detail of bizarre cells and aberrant mitoses. HE. 50, 100 (inset).
202
B. Martınez-Fern andez et al.
Fig. 2. Flipper; California sea lion. Oval or polygonal cells showing an alveolar pattern. HE. 50.
been validated for pinnipeds, normal cell types adjacent to the neoplasm were appropriately labelled in each section with each antibody. Immunohistochemically, the neoplastic cells had strong and diffuse cytoplasmic expression of vimentin, but were negative for the remaining antibodies. Expression of vimentin (Fig. 3) and absence of labelling for cytokeratins (Fig. 4) suggested a mesenchymal origin. The absence of lysozyme expression suggested that the tumour was not of histiocytic origin. Lack of expression of desmin and smooth muscle actin confirmed that the tumour was not derived from muscular tissue. Negative immunoreactivity for S100 and GFAP differentiated this tumour from malignant Schwannoma. Absence of S100 expression would also suggest that the tumour in the present case was not derived from lipoblasts
Fig. 3. Flipper; California sea lion. Expression of vimentin by the neoplastic cells. IHC. 10.
Fig. 4. Lung; California sea lion. The neoplastic cells exhibit no immunoreactivity for pan cytokeratin. Normal bronchial epithelium and glands show appropriate cytokeratin expression. IHC. 10.
(Gebhard et al., 2002; Takeuchi et al., 2007); however, normal lipoblasts also failed to show immunoreactivity for S100. The diagnosis of liposarcoma could be excluded because there were no cells filled with lipid droplets (negative Sudan black B staining). The tumour nodules located in the flipper were presumed to be the primary site of the neoplasm because these pleomorphic sarcomas in man often arise in the extremities (Fletcher, 2006) and the lungs are a common site for metastasis (Massi et al., 2004). The presentation of this tumour in a mature sea lion is consistent with the prevalence of these lesions in older people (Fletcher, 2006). Similarly, the storiform microscopical pattern of growth is also the most common histological pattern in the human tumours, accounting for almost two-thirds of all cases (Enzinger and Weiss, 1995). With regard to outcome, it has been suggested that such a histological background constitutes a negative prognostic factor (Enzinger and Weiss, 1995), and the California sea lion in this report died shortly after becoming ill, with pathological findings suggestive of a particularly aggressive tumour. Therefore, UPS appear to have a similar clinical presentation, pathological appearance and biological behaviour in man and the California sea lion. A variety of tumours have been reported in the California sea lion. These involve different organ systems, but most are of epithelial origin, while mesenchymal tumours are less commonly reported (Newman and Smith, 2006). One undifferentiated sarcoma of possible histiocytic origin has been reported affecting the brain of a California sea lion (Gage et al., 1995), but the present report is the first of a disseminated metastatic undifferentiated sarcoma in this species.
Sarcoma in a California Sea Lion
References Dei Tos AP (2006) Classification of pleomorphic sarcomas: where are we now? Histopathology, 48, 51e62. Edson A, Magalhaes de Pa¨dua J, Vicentini EL, Marchesan MA, Comelli RC et al. (2008) Oral undifferentiated high-grade pleomorphic sarcoma: report of a case. Oral Surgery, Oral Medicine, Oral Pathology, Oral Radiology and Endodontology, 105, 37e40. Enzinger FM, Weiss SW (1995) Malignant fibrohistiocytic tumours. In: Soft Tissue Tumours, 3rd Edit., FM Enzinger, SW Weiss, Eds., Mosby, St Louis, pp. 351e380. Fletcher CDM (2006) The evolving classification of soft tissue tumours: an update based on the new WHO classification. Histopathology, 48, 3e12. Gage LJ, Trupkiewicz J, Samansky T (1995) Hydrocephalus in an aged California sea lion (Zalophus californianus). Proceedings of the International Association for Aquatic Animal Medicine, 26, 9e10. Gebhard S, Coindre JM, Michels JJ, Terrier P, Bertrand G et al. (2002) Pleomorphic liposarcoma. Clinicopathologic and immunohistochemical study of 63 cases. A study of the French sarcoma group. American Journal of Surgical Pathology, 26, 601e616.
203
Massi D, Beltrami G, Capanna R, Franchi A (2004) Histopathological re-classification of extremity pleomorphic soft tissue sarcoma has clinical relevance. European Journal of Surgical Oncology, 30, 1131e1136. Newman SJ, Smith SA (2006) Marine mammal neoplasia: a review. Veterinary Pathology, 43, 865e880. Pe´rez-Martı´ nez C, Garcı´ a Ferna´ndez RA, Reyes Avila LE, Pe´rez-Pe´rez V, Gonza´lez N et al. (2000) Malignant fibrous histiocytoma (giant cell type) associated with a malignant tumour in the salivary gland of a dog. Veterinary Pathology, 37, 350e353. Takeuchi A, Nakano K, Nishimuta Y, Kayikawa S, Ito K et al. (2007) Spontaneously occurring subcutaneous tissue sarcoma in a Sprague-Dawley rat. Journal of Toxicologic Pathology, 20, 161e166. Thoolen RJMM, Vos JH, Van der Linde-Spiman JS, De Weger RA, Van Unnik JAM et al. (1992) Malignant fibrous histiocytoma in dogs and cats: an immunohistochemical study. Research in Veterinary Science, 53, 198e204.
March 9th, 2010 ½ Received, Accepted, June 27th, 2010