Animal ethics
The animal was placed in shade, in standard conditions, water ad libitum, and without restriction of movement according to the guidelines of Institutional Animal Ethical Committee of the Tehran University of Animal Science, Iran. Surgery was performed under aseptic conditions and sedation by injection of Xylazine hydrochloride (0.05 mg/kg) followed by 2% Lignocaine hydrochloride.
Clinical case reports
In October 2013, a 5-year-old, 7 kg, female black terrier dog was presented to the clinic at the college of Veterinary Medicine, university of Tehran, with a growing mass around axillary and mobility problems related to the mass, and euthanasia was applied upon the owner’s request and also taking into consideration the age of the animal and the size of the mass. The mass spreading into subcutis is of proximal right front leg, was of 2.8 × 2.5 cm size and 185 g weight (Figure 1). The mass was surgically resected and an impression smear of the tissue was prepared and stained with Giemsa. The rest of the tissue was subjected to histopathologic evaluation.
One formalin-fixed, paraffin wax-embedded section of each neoplasm was stained with haematoxylin and eosin (HE). Tumour was classified independently by two pathologists in order to confirm the diagnosis according to the WHO criteria [16]. Also tumour was graded according to the criteria proposed by Patnaik et al. [14] and Kiupel et al. [17], as well, moderately or poorly differentiated (grades I, II or III, respectively) [14, 17]. These criteria included the following histomorphological features: extent of tumour (invasiveness), cellular morphology, size of cytoplasmic granules, mitotic activity and stromal reaction. Invasiveness was assessed as follows: non-invasive (tumours confined to the superficial dermis and interfollicular spaces); moderately invasive (tumours with lower dermal and limited subcutaneous tissue invasion); highly invasive (tumours with massive infiltration of subcutaneous and deep tissue). Mitotic activity was assessed on toluidine blue-stained sections, five high power fields (hpfs) being evaluated in each case with a × 40 objective. The data were expressed as mean number of mitoses per hpf (mitotic index) and tumours were classified according to the following grading system: 0 (mitotic figures absent); 1 (mitotic index ranging from 0 to 2 mitoses/hpf); 2 (mitotic index > 2/hpf). Invasiveness and mitotic activity assessment were used to define the histological grade, and also as individually evaluated parameters.
Hematological findings
A complete blood work was performed. No changes were noticed in the erythrogram, however, leukogram changes included leukocytosis (55,245 leukocytes/μL), neutrophilia (27,644 neutrophils/μL) with mild regenerative left shift (4,376 band cells/μL).
Cytological findings
Cytological examination indicated mast cells with numerous metachromatic stained granules. Nuclei were varied in size and shape with high nuclear’to’cytoplasmic ratio, prominent nucleoli, marked atypical and mitotic figures. The background was filled with granules from ruptured cells and a moderate eosinophils were also present (Figure 2A and 2B).
Histopathological findings
Histopathologically, MCT tumour cells were less well circumscribed by connective tissue and neoplastic cells often exhibited an aggressive behavior, high cellularity, cellular pleomorphism, and various morphologic patterns.
However, in some regions of the tumor tissue shows neoplastic cells extending into the subcutis. Neoplastic cells had distinct cell borders and widely variable amounts of pale to brightly eosinophilic cytoplasm. Furthermore, tumour had neoplastic cells containing numerous fine eosinophilic to basophilic cytoplasmic granules consistent with mast cell granules and also highly pleomorphic tumour cells with a high mitotic index, anisokaryosis and infiltration of the surrounding dermis and subcutis were observed (Figure 2C and 2D).
Pathology grading of cutaneous mast cell tumor
In this case is characterized by one or more of the following criteria: at least 3 mitotic figures in 5 hpf, at least 2 multinucleated (2 or more nuclei) cells in 5 hpf, at least 2 bizarre nuclei (highly atypical with marked indentations, segmentation, and irregular shape) in 5 hpf; karyomegaly (ie, nuclear diameters of at least 10% of neoplastic mast cells vary by at least 2-fold). The selected field was it that was most highly mitotically active or had the highest degree of anisokaryosis.
Canine cutaneous MCTs were graded according to the Patnaik and the Kiupel systems [14, 17]. Grade III MCT was poorly demarcated accumulations of poorly differentiated mast cells with severe nuclear pleomorphism. Invasion of the deep subcutaneous fat or cutaneous muscles were a common feature of grade III tumour. Finally, a diagnosis of grade III cutaneous mast cell tumor was made.