Introduction

Mast cell tumors (MCTs) are the most commonly reported cutaneous round cell tumor in dogs.¹ The bioactive substances histamine, heparin, proteases, chemotactic factors, and cytokines contained within MCTs, in conjunction with their variable biological behavior, account for the variable presentation and common paraneoplastic effects seen with these tumors.^1,2 They are reported in multiple locations including cutaneous, subcutaneous, conjunctival, oral mucosal, gastrointestinal, and intramuscular locations across the body.¹ Breed predilections reported in typically middle-aged dogs, such as Labrador retrievers, supports the idea of underlying genetic causes for these common dermal and/or subcutaneous tumors.³ Diagnosis of MCTs using fine needle aspiration (FNA) cytology and Wright’s stain is sufficient in 90% of cases.¹ Metastatic spread in cutaneous MCTs is most commonly seen with high-grade tumors and is reported in skin, sentinel lymph nodes, splenic tissue, liver, or other visceral organs.⁴ Clinical staging of MCTs routinely involves FNA sampling of the liver, spleen, and regional and draining lymph nodes.^3–5 Additional staging for metastatic assessment can include thoracic radiography for sternal lymph node monitoring, abdominal ultrasound (with or without FNA of the liver or spleen), sentinel lymph node mapping, and bone marrow evaluation.^3,6 Advanced imaging, such as computed tomography (CT), can also be considered as a substitute for radiography and ultrasonography during staging.⁶ Recent consensus statements concluded that these tests are not all essential for MCT staging.^3,7,8 Following surgical excision of cutaneous MCTs, tumor grading is performed by histopathological assessment of tissues using the Patnaik and Kiupel grading schemes.^9,10 There are only a small number of studies investigating the application of cutaneous MCT grading systems to subcutaneous MCTs.^6,11 Recent studies⁶ concluded that grading was not useful in terms of prognostication and subjectively varied between pathologists.¹¹ Despite this, many subcutaneous MCTs are classified as low grade (Kiupel system) or grade 2 (Patnaik system) when grading is applied.⁶

Prognostic indicators for survival time associated with subcutaneous MCTs are closely linked to mitotic index, the infiltrative growth pattern of the tumor, and the presence of multinucleate mast cells.^3,11 Negative prognostic factors are useful in predicting the biological behavior of subcutaneous MCTs, ranging from benign to extremely aggressive behavior with metastasis to skin, regional or draining lymph nodes, spleen, liver, and—less commonly—lung tissue.⁴ In cases of widespread systemic dissemination, neoplastic mast cells have been reported within the bone marrow and peripheral blood.¹² Surgical excision alone is curative for the majority of subcutaneous MCTs compared with their cutaneous counterparts, with local recurrence reported in 2% of completely excised cases.¹

There are currently no reports in the veterinary literature to the authors’ knowledge of metastatic or local invasion of synovial spaces or bone by subcutaneous MCTs. The identification of spinal MCTs causing polyostotic lesions in multiple vertebral bodies of the lumbar spine was reported in one dog.¹³ This report will review a case of a dog with a subcutaneous MCT, displaying evidence of local infiltration of the stifle joint and suspected metastatic invasion of the adjacent bone.

Informed consent for case publication was obtained from the patient’s owners, and the patient described in this article was clinically managed according to the contemporary standards as described in the JAAHA instructions for authors.

Case Report

A 4 yr old female neutered Labrador retriever was referred for investigation of a suspected MCT on the medial aspect of the left stifle. The patient initially presented 2 wk earlier to the referring veterinarian with a history of sudden onset, nontraumatic left hind-limb lameness and a soft tissue swelling over the medial aspect of the left stifle. Initial management with a nonsteroidal anti-inflammatory drug, meloxicam^a (0.1 mg/kg q 24 hr [PO]), and a nonopioid analgesic, amantadine^b (3.3 mg/kg q 24 hr PO), resulted in symptomatic resolution of the lameness and a minor reduction in soft tissue swelling. Radiographs were later obtained by the referring clinician owing to the ongoing persistence of the soft tissue mass. A left lateral stifle projection demonstrated an increase in soft tissue opacity within the joint space, effacing the patella fat pad and patella tendon. Well-defined lysis of the distal pole of the patella and a poorly marginated area of focal heterogeneous lysis at the cranial tibial plateau were identified, as outlined in Figure 1. Cytology from an FNA of the soft tissue mass performed by the referring clinician demonstrated nucleated cells composed predominantly of mast cells displaying mild to moderate anisocytosis and mild anisokaryosis, with a moderate to highly granulated appearance and free scattered mast cell granules, consistent with an MCT.

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On clinical examination, a firm, soft tissue swelling on the medial aspect of the left stifle at the level of the fat pad caudal to the patella tendon was identified, alongside a mild joint effusion in the left stifle. The mass was relatively fixed to the underlying structures, and the skin was freely movable over it. The base of the mass (medial aspect) was difficult to distinguish from the underlying tissues of the stifle joint. No visible lameness was identified at a walk or trot in the left hind limb or any other limb, and there was no palpable enlargement of the popliteal or inguinal lymph nodes. The remainder of the physical examination was unremarkable. The patient was sedated with IV medetomidine^c (0.01 mg/kg) and butorphanol^d (0.3 mg/kg) initially; desirable effects were not achieved, and thus, an additional dose of medetomidine (0.005 mg/kg) and ketamine^e (1 mg/kg) were administered to achieve adequate chemical restraint. A contrast CT scan^f (2 mL/kg) of the abdomen and hind limbs was performed using a 64-slice multidetector unit^g. Interpretation of CT images revealed a local, circumferential, soft tissue thickening centered on the left stifle with moderate joint effusion and capsule thickening creating a mass-like appearance and bulge over the medial aspect of the joint (Figure 2). After contrast, heterogeneous enhancement of the abnormal tissue was present. Additionally, mild enlargement of the left popliteal lymph node (7 mm) in comparison to the right (5 mm) was noted. Focal lysis of the cranial tibial plateau at the level of the cranial intercondylar eminence extending into the subchondral bone was noted, as was lysis of the distal pole of the left patella (Figure 3). In the abdomen, the liver and spleen had a normal appearance, and mild enlargement and rounding of the left medial iliac lymph node (maximal dimension: 7.73 mm) was noted in comparison to the right (5 mm). Ultrasound-guided FNAs of the soft tissue mass, liver, spleen, and left popliteal lymph node were obtained under sedation, alongside joint fluid aspirates for cytological and fluid analysis. Sampling by FNA of the left medial iliac lymph node was not attempted because of the risks associated with its location and vicinity to the major blood vessels.

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Cytological examination of the soft tissue mass revealed a moderate number of mast cells occurring singly and as small aggregates amid a prominent background of red blood cells (RBCs), occasional eosinophils and fibrous/stromal cells. Mast cells were moderately to well-granulated with moderate cellular pleomorphism. Analysis of the joint fluid showed a moderate number of RBCs and a high nucleated cell count (4.94 × 10⁹/L; reference range less than 3.0 × 10⁹/L) comprising variably vacuolated macrophages (53.5%), small lymphocytes (19.5%) eosinophils (17.5%), neutrophils (3.5%), and mast cells (6%) that were single and well granulated and contained a single round nuclei 1.5 RBCs in diameter. Aspirates of the liver, spleen, and left popliteal lymph node showed no evidence of metastatic disease or reactive changes.

A diagnosis of a subcutaneous MCT was made based on the pathological findings, and invasion into the joint was suspected based on imaging and joint fluid analysis. Treatment options included pelvic limb amputation with coxofemoral disarticulation or cytoreductive surgery, in conjunction with chemotherapy, radiotherapy, or electrochemotherapy. Nonsurgical treatment options such as radiation therapy and/or chemotherapy were discussed and declined by the owner. Left hind-limb amputation was elected by the owners and was performed by the referring veterinary surgeon. The excised tissue was submitted for histopathology.

Histopathological examination of soft tissues obtained from the mass at the level of the medial left stifle confirmed a subcutaneous MCT. The subcutaneous MCT was identified to have low mitotic count (0 in 10 high-power fields; ×400) and the infiltrative population of pleomorphic round cells with increased nucleus-to-cytoplasmic ratios, moderate anisocytosis, and anisokaryosis.¹⁴ Eight sections of joint capsule were examined and showed neoplastic cells infiltrating the adjacent subcutis and multiple sites within the joint capsule, occasionally extending into the synovial membrane (Figure 4). Giemsa staining confirmed the presence of metachromatic granules within the neoplastic round cells of the soft tissue mass (Figure 5). Assessment of the small, circular, focal tibial lesion with localization guidance using the CT report identified a focal area of bone trabeculae, lined by plump, reactive osteoblasts and increased numbers of round to polygonal cells, as seen in Figure 6A. Giemsa staining confirmed the presence of clusters and aggregates of mast cells within this distinct solitary lesion (Figure 6B), together with osteoblasts, fibroblasts, macrophages, and other round cells. The clear distinction between both the focal tibial osteolytic lesion and the Giemsa-stained mast cells in adjacent soft tissues can be seen in Figure 4. Despite their close approximation, no evidence of infiltration by neoplastic mast cells between these sites could be identified by the pathologist, either by gross examination or microscopically.

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Adjunctive chemotherapy was discussed owing to bone lysis, infiltration into the joint capsule, potential for undiagnosed metastatic disease, and limited documentation of previously encountered cases of synovial or bony invasion; however, the owner declined this treatment plan. The patient was then lost to further follow-up with the referral team. A 12 mo metastasis check performed by the referring veterinarians showed an unremarkable physical examination and no further evidence of regional iliac lymph node enlargement on ultrasonographic examination. Lymph node size was not documented by the regular clinician at this visit. Aspiration of the iliac lymph node, liver, and spleen were not repeated at this time as per owner request.

Discussion

There is limited veterinary knowledge surrounding invasive and metastatic subcutaneous MCTs in dogs. Recent studies have classified subcutaneous MCT separately from its cutaneous counterpart, with the lack of histological grading schemes for subcutaneous MCTs making them difficult to prognosticate.^11,15 Despite surgical excision being the main therapeutic option for subcutaneous MCTs, local excision was not deemed possible in this case because of the extensive involvement of the stifle joint, thus requiring pelvic limb amputation with coxofemoral disarticulation. With local tumor recurrence uncommon,¹¹ subcutaneous MCTs have been reported to metastasize to regional lymph nodes, skin, spleen, liver, and—less commonly—lungs.⁴ In this case, it would have been sensible to consider sentinel lymph node mapping as the specific draining lymph node was not clear. Risks associated with aspiration of the medial iliac lymph node prevented sampling pre- and postoperatively, limiting our ability to comment on the status of the lymph node with regard to potential metastasis.

Mast cells originate from pluripotent progenitor cells within the bone marrow. They mature within the mucosal and epithelial tissues in which they reside and do not commonly circulate within the bloodstream.¹⁵ Low numbers of individual, uniform mast cells are normally seen in tissues like bone marrow and the synovial lining of joints including loose connective tissue and adipose fat.^16,17

Differentiation between inflammatory processes and metastatic MCT relies largely on the histopathological changes identified in tissues. In inflammatory conditions like rheumatoid arthritis, the presence of other types of inflammatory cells (eosinophils, neutrophils, macrophages, and monocytes) alongside an overall increase in the number of individual mast cells is often reported, in particular around blood vessels and nerves in the synovial sublining.¹⁸ With metastatic MCTs, any atypia present within the mast cell population and the presence of dense aggregates or sheets of mast cells allows for differentiation between metastatic tumor deposits and the increased number of mast cells normally seen with reactive hyperplasia of lymph nodes.¹⁸ The limited information available pertaining to the ability to definitively distinguish between the behavior of inflammatory versus metastatic MCT enables the authors to extrapolate that the histopathological changes reported in the synovium, joint fluid, and bone are similar to those seen in lymph nodes with metastatic spread of cutaneous MCTs.

Interpretation of the histopathological changes reported in the tissues of the synovium, tibia, and patella for this case were based on the distinguishable variation in biological behavior for MCTs associated with inflammatory processes and the behavior of metastatic MCTs in lymph tissue. In this case, the presence of small clusters and aggregates of mast cells within the synovium and bone trabeculae of the tibia and patella is abnormal.¹⁹ The inability to definitively differentiate between individual reactive and neoplastic mast cells microscopically prevents a clear interpretation of the results, hence a reliance on the overall lesion appearance and mast cell behavior such as the formation of clusters and aggregates within the bone. If we interpret the above histopathological findings for this case using the World Health Organization criteria applied to canine lymph node tissue, which classifies metastasis based on the number of, distribution of, and architectural disruption by nodal mast cells, with the presence of three or more mast cells within one aggregate suggestive of metastatic spread,¹⁸ one could extrapolate that these changes could be indicative of in-transit metastasis of the MCT to the tibia and patella through exposure to diseased joint fluid.

To the authors’ knowledge, MCT identification in the synovial fluid of dogs has not been previously documented. Research suggests that their presence in high concentrations in other types of effusions is suspicious of tumor activity, such as pleural effusions in the presence of systemic mastocytosis.²⁰ Similarly, MCTs extending across the joint capsule have not been previously reported in veterinary literature. Canine neoplastic lesions arising from joints are primarily associated with synovial structures or adjacent soft tissues, with secondary invasion of the synovial tissue and subchondral bone.²¹ As per the histological behavior reported for invasive joint neoplasia, the presence of neoplastic mast cells infiltrating the stifle synovial membrane and joint capsule across multiple sites in this case report is identical to the behavior of infiltrative synovial tumors.²¹ The mixed active eosinophilic inflammation associated with the low numbers of mast cells in the synovial fluid of the stifle could suggest neoplastic effusion (associated with synovial infiltration) given the above soft tissue findings. The presence of a neoplastic effusion in conjunction with metastatic bone lesions, or local bony invasion, would indicate aggressive behavior similar to that seen with high-grade MCTs.³

Human literature shows that the presence of increased numbers of mast cells in bone accelerates bone loss because of the production of byproducts, such as histamine and cytokines, responsible for increasing bone resorption.²² Bone disorders including rheumatoid arthritis and osteoporosis are associated with a high prevalence of mast cells within joint fluid and bone marrow in humans. Bone resorption rates in the presence of neoplastic cells are also increased in the presence of inflammatory cells such as osteoblasts, fibroblasts, and macrophages,¹⁹ as were reported in the circular lesion of the tibia in this current case_. The behavior of these cells in humans enables us to theorize that the abnormal presence of mast cells in the synovial fluid in conjunction with the clustered mast cells and the presence of osteoblasts, fibroblasts, and macrophages within the abnormal bone trabeculae of the tibia is likely due to MCT metastasis to the tibia and patella in this case, although local invasion also cannot be excluded.

Mild enlargement of the medial iliac lymph node at time of surgery was considered to be a negative prognostic factor by the authors based on the possibility this indicated distant metastasis instead of local inflammation.⁵ The adjunctive treatment advised was based on the aggressive features of the tumor described earlier in this report. There is no standardized chemotherapy protocol for MCTs with current literature based on controlled and retrospective studies.³ Chemotherapy protocols commonly used for the management of MCTs without evidence of gross disease include vinblastine and prednisolone over an eight-dose protocol. The single agent lomustine is an alternative chemotherapy agent that can be given every 2–3 wk, although it is less desirable because of its myelosuppressive and hepatotoxic side effects.³ The tyrosine kinase inhibitors masitinib and toceranib phosphate are small molecule inhibitors that are common oral therapies used against bulky MCTs.^3,4 The use of systemic therapies was considered in this case based on the features described above and the significant improvement in median survival time that is seen with the use of additional therapies in other specific scenarios relating to MCTs.³

Patient prognosis currently appears favorable given the patient’s good clinical health and the absence of gross metastatic disease on ultrasound assessment of medial iliac lymph nodes, liver, and spleen by the referring veterinarian at a 12 mo postoperative recheck.

Conclusion

To the authors’ knowledge, this is the first reported case of a subcutaneous MCT demonstrating local invasion to areas of the stifle joint and suspected metastasis to the adjacent tibial plateau and distal patella by suspected in-transit exposure to diseased joint fluid. Local infiltration to the tibia and patella cannot be completely excluded in this case based on the absence of distant site metastasis and the close proximity of the tissues involved with the primary tumor. Despite displaying hallmarks of aggressive tumor behavior, radical surgery appears to have been surgically curative in this case, owing to the absence of apparent metastasis to regional lymph nodes at annual health checks. Additional studies are required to further understand the significance of local bone or joint invasion with subcutaneous MCTs.

The authors would like to thank Dr. Phillipa Weston for taking the time to produce still images of the CT report and Dr. Tom Vicek for his initial reporting of this case.