Fine-Needle Aspirate Cytology of Cutaneous Mast Cell Tumors

Introduction

Cutaneous mast cell tumors (MCTs) represent one of the most frequently diagnosed neoplasms in canine dermatology [1]. Their biological behavior ranges from benign, indolent lesions to aggressive, metastatic malignancies, making accurate diagnostic characterization essential for prognosis and therapeutic planning [2, 3]. Fine-needle aspirate (FNA) cytology has become a cornerstone of the initial diagnostic workup for cutaneous MCTs due to its rapidity, minimal invasiveness, and high diagnostic yield [4, 5]. This article provides an exhaustive, publication-grade review of the principles, techniques, cytomorphological features, grading systems, and ancillary diagnostic applications of FNA cytology for cutaneous MCTs in dogs.

Biological Basis and Clinical Presentation

Mast cell tumors arise from neoplastic transformation of mast cells, which are granulated immune effector cells resident in connective tissues [3]. The precise etiopathogenesis of MCTs involves dysregulation of the KIT receptor tyrosine kinase, often driven by activating mutations in the c-kit proto-oncogene [6]. These mutations lead to ligand-independent receptor autophosphorylation and uncontrolled cellular proliferation [6]. Clinically, cutaneous MCTs present as solitary or multiple dermal or subcutaneous nodules, often with a characteristic "Darier's sign" (wheal formation upon palpation) due to local histamine release [3]. Breed predispositions have been documented, with certain breeds such as Boxers, Labrador Retrievers, and Bulldogs exhibiting higher incidence rates [1]. A retrospective study analyzing cytology reports from 2019 to 2021 identified breed, age, and sex as significant risk factors for the occurrence of cutaneous neoplasms, including MCTs [1].

Fine-Needle Aspiration Technique

The FNA procedure for cutaneous MCTs is performed using a 22- to 25-gauge needle attached to a 6- to 12-mL syringe, often with the aid of a syringe holder for one-handed aspiration [4]. The technique involves inserting the needle into the mass, applying negative pressure, and redirecting the needle multiple times within the lesion to dislodge cellular material [4]. Negative pressure is released before needle withdrawal to prevent aspiration of the sample into the syringe barrel [4]. The aspirated material is then expelled onto glass slides and smeared using a squash or blood smear technique [7]. Sample preparation quality is critical; thick or bloody smears can obscure cellular detail and compromise diagnostic accuracy [8, 7]. A study comparing different sample preparation techniques for nodal mast cell quantification found that the quality of the smear directly influenced the ability to accurately enumerate mast cells [8].

Staining Protocols

Rapid Romanowsky-type stains, such as Diff-Quik, are the most commonly employed stains for FNA cytology of MCTs in clinical practice [7]. These stains allow for rapid assessment of cellular morphology and granule characteristics. The fixation time for Diff-Quik staining has been evaluated, with evidence indicating that prolonged fixation in methanol can alter staining intensity and granule visualization [7]. Specifically, a study demonstrated that fixation times exceeding the manufacturer's recommendation resulted in decreased staining quality and potential misinterpretation of granule content [7]. For optimal results, adherence to standardized fixation and staining protocols is recommended [7]. In research settings, Papanicolaou and May-Grünwald-Giemsa stains may also be used, though they require longer processing times [3].

Cytomorphological Features

The cytological diagnosis of MCTs relies on the identification of neoplastic mast cells, which are characterized by moderate to abundant amounts of basophilic cytoplasm containing numerous metachromatic cytoplasmic granules [2, 3]. These granules stain purple to magenta with Romanowsky-type stains and may vary in size, number, and distribution [3]. Neoplastic mast cells often exhibit anisocytosis and anisokaryosis, with nuclei that may be round to oval and contain coarsely clumped chromatin [2, 9]. Binucleation and multinucleation are occasionally observed [2]. The background of the smear typically contains variable numbers of eosinophils, which are attracted by chemotactic factors released from mast cell granules [3]. Fibroblasts, collagen bundles, and inflammatory cells may also be present, particularly in aspirates from lesions with associated stromal reaction or ulceration [3].

Morphometric analysis of canine cutaneous MCTs has been performed to quantify nuclear and cellular dimensions [9]. A study utilizing image analysis software demonstrated that nuclear area, nuclear perimeter, and nuclear-to-cytoplasmic ratio are significantly different between low-grade and high-grade MCTs [9]. These quantitative parameters may provide objective criteria for cytological grading, though they are not yet routinely applied in clinical practice [9].

Cytological Grading Systems

Histological grading systems, such as the Patnaik and Kiupel systems, are well-established for predicting the biological behavior of canine cutaneous MCTs [10, 5]. However, cytological grading has gained traction as a preoperative tool to guide clinical decision-making [10, 5]. A 2-tier cytological grading system, analogous to the Kiupel histological system, has been proposed and validated [10]. This system classifies MCTs as low-grade or high-grade based on the presence of cytological criteria including marked anisokaryosis, nuclear pleomorphism, prominent nucleoli, and the presence of mitotic figures [10]. A study evaluating the concordance between cytological and histological grading using this 2-tier system reported substantial agreement, supporting the utility of cytology for preoperative grading [10].

Another cytological grading scheme, developed by Scarpa et al., incorporates additional parameters such as cytoplasmic granularity, cell density, and the presence of eosinophils [5]. This system demonstrated a significant correlation with histological grade and clinical outcome [5]. The predictive ability of FNA cytology for assessing incompletely resected MCT surgical sites has also been investigated [4]. In a study of dogs with incompletely excised MCTs, cytological evaluation of the surgical bed was found to have moderate sensitivity and high specificity for detecting residual neoplastic cells, suggesting its potential role in postoperative monitoring [4].

Ancillary Diagnostic Techniques

Immunocytochemistry for CD117

Immunocytochemical staining for CD117 (KIT receptor) can be performed on FNA cytology specimens to assess KIT expression patterns [6]. The staining pattern is classified as membranous, focal cytoplasmic, or diffuse cytoplasmic, with the latter two patterns being associated with c-kit mutations and a higher risk of aggressive biological behavior [6]. A study correlating CD117 immunocytochemistry patterns with mutational analysis of exon-11 of c-kit using PCR found that diffuse cytoplasmic staining was significantly associated with the presence of exon-11 mutations [6]. This technique can be performed on archived cytology slides, allowing for retrospective analysis and prospective risk stratification [6].

Molecular Analysis for c-kit Mutations

DNA extracted from FNA samples can be used for PCR-based detection of c-kit mutations, particularly in exons 8, 9, and 11 [6]. The presence of these mutations has prognostic significance and may predict response to tyrosine kinase inhibitor therapy [6]. The ability to obtain sufficient DNA from FNA specimens makes this approach feasible in a clinical setting, avoiding the need for more invasive biopsy procedures [6].

Sentinel Lymph Node Mapping and Aspiration

Regional lymph node assessment is a critical component of MCT staging [11]. FNA cytology of locoregional lymph nodes can detect metastatic mast cells, but the sensitivity of this approach is influenced by the sampling technique and the criteria used for diagnosis [8, 11]. Sentinel lymph node mapping and biopsy, which involves identification and selective removal of the first draining lymph node, has been shown to improve the accuracy of disease stage assignment compared to routine lymph node aspiration [11]. A study comparing these two methods found that sentinel lymph node biopsy identified metastatic disease in a higher proportion of cases, leading to upstaging in a significant number of dogs [11]. However, FNA cytology remains a valuable initial screening tool due to its simplicity and low cost [11].

Abdominal Imaging

For high-grade MCTs or those with documented lymph node metastasis, abdominal ultrasonography or computed tomography (CT) is recommended to evaluate the liver and spleen for metastatic disease [12]. However, a study evaluating abdominal CT for MCT staging found that hepatic and splenic changes were nonspecific and did not reliably differentiate metastatic disease from benign conditions [12]. Therefore, cytological or histopathological confirmation of suspected metastatic lesions is warranted [12].

Diagnostic Algorithm

The following Mermaid diagram illustrates a diagnostic workflow integrating FNA cytology for cutaneous MCTs.

flowchart TD
    A[Cutaneous nodule identified], > B[FNA cytology performed]
    B, > C{Cytological diagnosis}
    C, >|Mast cell tumor| D[Assign cytological grade]
    C, >|Other neoplasm or inflammation| E[Further diagnostics as indicated]
    D, > F{Grade}
    F, >|Low-grade| G[Local excision with histopathology]
    F, >|High-grade| H[Regional lymph node FNA]
    H, > I{Lymph node status}
    I, >|Negative| J[Wide surgical excision + histopathology]
    I, >|Positive| K[Abdominal imaging + staging]
    K, > L[Consider sentinel lymph node mapping]
    L, > M[Treatment planning: surgery, +/- tyrosine kinase inhibitor]
    G, > N[Post-excision histopathology for grade confirmation]
    N, > O[Surveillance]
    M, > O

Limitations and Considerations

While FNA cytology is a powerful diagnostic tool, it has inherent limitations. Sampling error can occur, particularly in heterogeneous lesions, leading to misclassification of tumor grade [4, 5]. The presence of marked inflammation or necrosis may obscure neoplastic mast cells and complicate interpretation [3]. Additionally, cytological grading systems, while useful, are not a substitute for histopathological grading, which remains the gold standard for definitive prognostication [10, 5]. The predictive ability of cytology for residual disease at surgical sites is moderate, and false-negative results can occur [4]. Therefore, cytological findings should always be interpreted in the context of the complete clinical picture, including signalment, physical examination findings, and results of other diagnostic tests [1, 4].

Frequently Asked Questions

What is the sensitivity and specificity of FNA cytology for diagnosing cutaneous MCTs?

FNA cytology has high sensitivity and specificity for the diagnosis of cutaneous MCTs, with reported values exceeding 90% in most studies when performed by experienced cytopathologists [4, 3]. The presence of characteristic metachromatic granules in neoplastic mast cells allows for definitive identification in the majority of cases [2, 3].

Can cytological grading replace histological grading for MCTs?

Cytological grading provides valuable preoperative information but does not replace histological grading [10, 5]. The concordance between cytological and histological grading using a 2-tier system is substantial but not perfect, and histopathology remains the gold standard for definitive grading and prognostication [10].

How should lymph node FNA results be interpreted in MCT staging?

Lymph node FNA cytology is a useful screening tool for detecting nodal metastasis [8, 11]. The presence of mast cells in a lymph node must be interpreted cautiously, as small numbers of mast cells can be found in reactive nodes [8]. Quantitative criteria, such as the number of mast cells per high-power field, can aid in distinguishing reactive from metastatic populations [8]. Sentinel lymph node mapping and biopsy provide more accurate staging information [11].

What is the role of CD117 immunocytochemistry on FNA samples?

CD117 immunocytochemistry can be performed on FNA cytology slides to assess KIT expression patterns [6]. Diffuse cytoplasmic staining is associated with c-kit mutations and a higher risk of aggressive behavior, providing prognostic information that can guide therapeutic decisions [6].

Are there any special sample preparation requirements for molecular testing on FNA samples?

For molecular testing, such as PCR for c-kit mutations, FNA samples should be collected using sterile technique and placed in an appropriate DNA stabilization medium or directly onto slides for subsequent DNA extraction [6]. Air-dried, unstained slides can be used for DNA extraction, but the yield may be lower than from fresh samples [6].

References

[1] García-Reynoso IC, Flores-Dueñas CA, Castro-Del Campo N, et al. Risk Factors for the Occurrence of Cutaneous Neoplasms in Dogs: A Retrospective Study by Cytology Reports, 2019-2021. Animals (Basel). 2025. https://pubmed.ncbi.nlm.nih.gov/40723532/

[2] Suneja P, Nambiar K, Tanveer N, et al. Solitary Cutaneous Mastocytoma in an Adult Diagnosed on Cytology: A Rare Case Report. Diagn Cytopathol. 2025. https://pubmed.ncbi.nlm.nih.gov/39235109/

[3] Hosseini E, Pedram B, Bahrami AM, et al. Cutaneous mast cell tumor (Mastocytoma): cyto- histopathological and haematological investigations. Diagn Pathol. 2014. https://pubmed.ncbi.nlm.nih.gov/24444100/

[4] Lee CE, Lindley SS, Smith AN, et al. Predictive ability of fine-needle aspirate cytology for incompletely resected mast cell tumor surgical sites. Can Vet J. 2021. https://pubmed.ncbi.nlm.nih.gov/33542552/

[5] Scarpa F, Sabattini S, Bettini G. Cytological grading of canine cutaneous mast cell tumours. Vet Comp Oncol. 2016. https://pubmed.ncbi.nlm.nih.gov/24717019/

[6] Sailasuta A, Ketpun D, Piyaviriyakul P, et al. The Relevance of CD117-Immunocytochemistry Staining Patterns to Mutational Exon-11 in c-kit Detected by PCR from Fine-Needle Aspirated Canine Mast Cell Tumor Cells. Vet Med Int. 2014. https://pubmed.ncbi.nlm.nih.gov/24701365/

[7] Jackson DE, Selting KA, Spoor MS, et al. Evaluation of fixation time using Diff-Quik for staining of canine mast cell tumor aspirates. Vet Clin Pathol. 2013. https://pubmed.ncbi.nlm.nih.gov/23278320/

[8] Buzzi G, Gambini M, Recordati C, et al. Cytological Quantification of Nodal Mast Cells in Dogs Affected by Non-Neoplastic Condition and Mast Cell Tumor Using Different Sample Preparation Techniques: An Explorative Study. Animals (Basel). 2023. https://pubmed.ncbi.nlm.nih.gov/37627425/

[9] Strefezzi Rde F, Xavier JG, Catão-Dias JL. Morphometry of canine cutaneous mast cell tumors. Vet Pathol. 2003. https://pubmed.ncbi.nlm.nih.gov/12724567/ *** Disclaimer: This article is for educational and informational purposes only. It is not intended to substitute for professional veterinary advice, diagnosis, treatment, or regulatory guidance. Always consult a licensed veterinarian or qualified specialist regarding animal health, disease diagnosis, and therapeutic decisions.

[10] Hergt F, von Bomhard W, Kent MS, et al. Use of a 2-tier histologic grading system for canine cutaneous mast cell tumors on cytology specimens. Vet Clin Pathol. 2016. https://pubmed.ncbi.nlm.nih.gov/27483044/

[11] Lapsley J, Hayes GM, Janvier V, et al. Influence of locoregional lymph node aspiration cytology vs sentinel lymph node mapping and biopsy on disease stage assignment in dogs with integumentary mast cell tumors. Vet Surg. 2021. https://pubmed.ncbi.nlm.nih.gov/33169849/

[12] Hughes JR, Szladovits B, Drees R. Abdominal CT evaluation of the liver and spleen for staging mast cell tumors in dogs yields nonspecific results. Vet Radiol Ultrasound. 2019. https://pubmed.ncbi.nlm.nih.gov/30786323/