2. Materials and Methods

2.1. Study Population

A retrospective review of histopathology reports describing splenic lesions in dogs was conducted. Cases were obtained from three veterinary histopathology laboratories based in Medellín, Antioquia, Colombia, between January 2019 and June 2024. These laboratories receive samples from multiple veterinary clinics and hospitals within the region. All samples consisted of spleen fragments obtained by total splenectomy. Incisional biopsies were excluded.

2.2. Data Collection

Pathology submission forms and corresponding histopathology reports were reviewed. Extracted data included the year of the report (2019–2024), animal demographics (breed, age, and sex), clinical signs, presence of hemoabdomen, and gross lesion pattern. Macroscopic presentations were classified into four categories: single mass, multiple masses, splenomegaly, and nodular pattern. A single mass was defined as one splenic lesion > 1 cm in diameter and multiple masses as more than one lesion > 1 cm. Splenomegaly was defined as diffuse enlargement of the spleen without formation of a nodule or mass, whereas the nodular pattern referred to lesions < 1 cm in diameter.

Lesions were categorized as non‐neoplastic or neoplastic. Splenitis cases within the non‐neoplastic group were further subclassified according to established histopathological criteria for inflammatory lesions. Neoplastic cases were subsequently classified according to biological behavior as malignant or benign.

Histopathological diagnoses were recorded following standard morphological criteria. In non‐neoplastic cases, more than one diagnosis could be present within the same case; therefore, all reported histopathological diagnoses were recorded independently without hierarchical prioritization. Up to three diagnoses per case were documented when applicable. For neoplastic lesions, each case was assigned a single diagnosis based on the pathologist’s report.

Common associated histopathological findings, including necrosis, hemorrhage, extramedullary hematopoiesis, inflammation, hematoma, siderotic plaques, lymphoid tissue changes (alterations in the splenic white pulp, including lymphoid hyperplasia, atrophy, and depletion), and central arteriolar thickening, were recorded when documented in the original histopathology reports.

All histopathological evaluations were performed by board‐certified veterinary pathologists at the participating diagnostic laboratories.

2.3. Histological Processing and Staining

Splenic samples were fixed and processed following standard histopathological protocols routinely used in the participating diagnostic laboratories. Tissues were fixed in 10% neutral buffered formalin, embedded in paraffin, sectioned at 4–5 μm, and stained with hematoxylin and eosin (H&E) for microscopic evaluation. No immunohistochemical procedures were performed; only H&E‐stained sections were reviewed for this study.

2.4. Data Analysis

Data were entered into Excel (Microsoft Corp., Redmond, WA, USA) and analyzed using SAS software Version 9.4 (SAS Institute Inc., Cary, NC, USA). Descriptive statistics were used to summarize the data. Categorical variables were expressed as absolute and relative frequencies, whereas continuous variables were summarized as mean (range). Normality was assessed using the Shapiro–Wilk test. Age was analyzed both as a continuous variable and grouped into categories (< 5, 5–8, 9–12, and > 12 years) to allow comparative analysis. Dogs younger than 1 year (n = 5) were included in categorical analyses but excluded from mean age calculations because exact age in months was unavailable.

Associations between categorical variables were evaluated using chi‐square or Fisher’s exact tests, as appropriate. Odds ratios (ORs) and corresponding 95% confidence intervals (95% CI) were calculated using cases with complete data for the variable of interest. All statistical tests were two‐sided, and statistical significance was set at p < 0.05.

2.5. Ethical Considerations

This study analyzed data obtained during routine histopathological diagnosis. Prior informed consent was obtained from pet owners for academic and scientific use of data, and laboratories authorized access to records. Ethical committee approval was deemed unnecessary, as no additional interventions were performed on animals.