3. Results

3.1. Study Population

A total of 424 canine splenectomy cases were included. All samples consisted of splenic tissue fragments submitted for histopathological evaluation between January 2019 and June 2024. The annual case distribution was 82 cases (2019), 47 (2020), 60 (2021), 86 (2022), 113 (2023), and 36 during the first half of 2024. Most submissions originated from the department of Antioquia (92%), with additional cases from two other Colombian departments (Atlántico and Norte de Santander).

The study population comprised 222 males (52.4%) and 202 females (47.6%). Age information was unavailable in six cases, and five dogs were younger than 1 year of age. The mean age (range) of dogs with non‐neoplastic lesions was 9.5 (< 1–16) years, whereas dogs with neoplastic lesions had a mean age of 10.2 years [4–17]. Dogs aged 9–12 years showed significantly increased odds of splenic neoplasia compared with other age groups (OR: 1.85; 95% CI: 1.21–2.81; p _ = _0.004).

A total of 45 breeds were represented. Crossbreed dogs were the most frequent (26.7%), followed by Schnauzer (7.3%), Pit Bull Terrier (6.6%), Beagle (6.1%), Golden Retriever (5.4%), Bulldog (5.4%), Labrador Retriever (5%), and Poodle (4%).

3.2. Clinical Presentation

Clinical information was obtained from histopathology submission forms. This section was categorized based on reported clinical signs, whereas hemoperitoneum was evaluated as a separate clinical variable.

Abdominal distension and/or abdominal pain were the most frequently reported clinical signs (41%, 174/424). Splenic lesions were detected incidentally by ultrasonography in 33.7% of cases (143/424). Dogs with incidental findings were more likely to have non‐neoplastic lesions compared with those with non‐incidental presentations (OR: 1.75; 95% CI: 1.10–2.80; p _ = _0.019).

Other nonspecific clinical signs, including lethargy, fever, weight loss, and reduced appetite, were reported in 10.6% (45/424). In 62 cases (14.6%), clinical information was not available in the submission forms, precluding further classification.

Hemoperitoneum was reported in 12.7% (54/424) of dogs and was significantly associated with splenic neoplasia (OR: 3.92; 95% CI: 2.15–7.13; p < 0.001), particularly HSA (OR: 8.83; 95% CI: 4.70–16.58; p < 0.001).

3.3. Lesion Types

Of the 424 cases, 282 (66.5%) were classified as non‐neoplastic lesions, whereas 142 (33.5%) corresponded to splenic neoplasms. Among neoplastic cases, malignant behavior predominated (130/142; 91.5%), whereas benign neoplasms accounted for 8.5% (12/142) (Figure 1).

FIGURE 1: Distribution of cases by biological behavior and sex (%). Note: Bars represent the proportion of males and females within each lesion category. Percentages were calculated relative to the total number of dogs in each diagnostic group. Non‐neoplastic lesions include inflammatory, reactive, and degenerative conditions, whereas neoplastic lesions include benign and malignant splenic tumors.

3.4. Macroscopic Findings

Mass‐forming lesions were the predominant macroscopic presentation. A single splenic mass was the most frequent pattern (256/424; 60.4%), followed by multiple masses (76/424; 17.9%), splenomegaly (42/424; 9.9%), and nodular pattern (11/424; 2.6%). In 39 dogs (9.2%), the macroscopic pattern was not specified.

No significant association was found between macroscopic pattern and final diagnosis (non‐neoplastic vs. neoplastic) (χ 2 = 7.9, p _ = _0.10). However, analysis of individual macroscopic patterns revealed several significant associations. Dogs presenting a single splenic mass were more frequently incidentally detected (OR: 1.67; 95% CI: 1.04–2.69; p _ = _0.03) and this presentation was also associated with NLH (OR: 4.04; 95% CI: 1.86–8.77; p = 0.003), and splenic hematoma (OR: 2.48; 95% CI: 1.34–4.60; p = 0.003).

Conversely, dogs with multiple splenic masses showed increased odds of malignancy (OR: 1.76; 95% CI: 1.04–2.96; p = 0.03), and HSA (OR: 2.08; 95% CI: 1.19–3.65; p = 0.01). Additional associations are summarized in Table 1.

Variable | Outcome | OR | 95% CI | p value
Demographic variables
Sex (male vs. female) | Neoplasia | 1.07 | 0.72–1.61 | 0.73
HSA | 1.05 | 0.66–1.68 | 0.93
Splenitis | 0.74 | 0.49–1.11 | 0.15
Age (9–12 years vs. other age groups) | Neoplasia | 1.85 | 1.21–2.81 | 0.004
HSA | 1.99 | 1.21–3.26 | 0.009
Splenitis | 0.83 | 0.56–1.24 | 0.36
Crossbreed vs. purebred | HSA | 1.61 | 0.97–2.65 | 0.083

Clinical presentation
Incidental detection | Non‐neoplasm | 1.75 | 1.10–2.80 | 0.019
Single mass | 1.67 | 1.04–2.69 | 0.033
Multiple masses | 1.33 | 0.78–2.26 | 0.296
Hemoperitoneum (present vs. absent) | Neoplasia | 3.92 | 2.15–7.13 | < 0.001
HSA | 8.83 | 4.70–16.58 | < 0.001
Lymphoma | 0.31 | 0.04–2.30 | 0.24
Splenitis | 0.41 | 0.21–0.80 | 0.009
NLH | 0.19 | 0.04–0.82 | 0.027

Non‐hemoperitoneum HSA
Incidental detection | HSA | 0.81 | 0.38–1.75 | 0.590
Abdominal distension and/or pain | HSA | 1.17 | 0.53–2.58 | 0.690

Macroscopic findings
Single mass (present vs. absent) | Malignant | 0.79 | 0.50–1.24 | 0.296
Splenitis | 0.77 | 0.50–1.18 | 0.23
Splenic hematoma | 2.48 | 1.34–4.60 | 0.003
NLH | 4.04 | 1.86–8.77 | < 0.001
Multiple masses (present vs. absent) | Malignant | 1.76 | 1.04–2.96 | 0.032
Splenitis | 0.77 | 0.46–1.29 | 0.32
HSA | 2.08 | 1.19–3.65 | 0.010

Histopathological findings
Splenic hematoma∗ | HSA | 1.45 | 0.90–2.33 | 0.13

3.5. Non‐Neoplastic Splenic Lesions

Non‐neoplastic lesions represented 66.5% of cases (282/424). Because multiple morphological processes could occur within the same spleen, entities were not mutually exclusive; therefore, up to three diagnoses were recorded per case, yielding a cumulative total of 451 non‐neoplastic diagnoses.

Inflammatory and reactive conditions predominated. Splenitis was the most frequent diagnosis (54.6%, 154/282 dogs) (Figure 2A), followed by splenic hematoma (33.7%, 95/282) (Figure 2B,C) and NLH (22.3%, 63/282) (Figure 2D,E). The main non‐neoplastic diagnoses and their clinicopathological characteristics are summarized in Table 2. Detailed demographic features, concurrent diagnoses, and associated histopathological findings for each morphological diagnosis are presented in Supporting Table S1.

FIGURE 2: Histopathological non‐neoplastic splenic features using hematoxylin and eosin stain. (A) Chronic hemorrhagic splenitis (4×): hemorrhage accompanied by neutrophilic infiltration, hemosiderin deposits, and areas of fibrosis. (B) Non-organized splenic hematoma (10×): presence of abundant erythrocytes and minimal cellular debris, supported by fibrin. (C) Organized splenic hematoma (4×/10×): hemorrhagic lesion characterized by fibrin deposition supporting erythrocytes, cellular debris, hemosiderin accumulation, and collagen deposition. (D) Nodular lymphoid hyperplasia (4×): circumscribed proliferation of lymphocytes organized into lymphoid follicles, occasionally with coalescence. (E) Hypertrophy of the central arteriole (10×/40×): in a dog with lymphoid nodular hyperplasia. (F–G) Severe extramedullary hematopoiesis (10×/40×): abundant erythroid islands and megakaryocytes were found. (H) Subcapsular hemorrhage and siderotic nodules (10×): extensive subcapsular hemorrhage and siderosclerosis, accompanied by mild fibroplasia and EMH.

Non‐neoplastic diagnoses | Cases n (%) | Median age years (range) | Sex (F/M) | Most affected breeds | Common macroscopic pattern | Common concurrent morphological diagnosis | Common concurrent histopathological findings
Splenitis | 154 (54.6) | 9.5 (< 1–16)a | 81/73 | Crossbreed, Beagle, Pit Bull Terrier | Single mass | Splenic hematoma, nodular lymphoid hyperplasia (NLH) | Extramedullary hematopoiesis (EMH), hemorrhage
Splenic hematoma | 95 (33.7) | 10.1 (4–16)b | 37/41 | Crossbreed, Pit Bull Terrier | Single mass | Splenitis | Hemorrhage, EMH
Nodular lymphoid hyperplasia (NLH) | 63 (22.3) | 8.9 (< 1–15)c | 28/35 | Crossbreed, Poodle, Shih Tzu | Single mass | Splenitis, splenic hematoma | Hemorrhage, EMH, inflammation
Extramedullary hematopoiesis (EMH) | 30 (10.6) | 9.6 (< 1–15)d | 17/13 | Crossbreed, Poodle, Pit Bull Terrier, Bulldog | Single mass | Splenitis, splenic hematoma, NLH | Lymphoid tissue changes∗, hemorrhage, inflammation
Siderotic nodules | 27 (9.6) | 8.4 (1–14) | 12/15 | Crossbreed, Bulldog | Single mass | Splenitis, splenic hematoma, EMH, NLH, splenic hemorrhage | EMH, hemorrhage, inflammation, lymphoid tissue changes

Among inflammatory patterns, hemorrhagic splenitis was the most frequent subtype (46.8%, 72/154), followed by congestive (22.7%, 35/154) and chronic/active splenitis (18.2%, 28/154). The remaining patterns were less common, each representing less than 5% of cases (Table 3). Dogs presenting hemoperitoneum had significantly lower odds of splenitis (OR: 0.41; 95% CI: 0.21–0.81; p = 0.009).

Inflammatory pattern | Cases n | Splenitis %
Hemorrhagic | 72 | 46.8
Congestive | 35 | 22.7
Chronic/active | 28 | 18.2
Fibrosing | 6 | 3.9
Suppurative | 5 | 3.2
Mixed cases∗ | 4 | 2.6
Granulomatous/pyogranulomatous | 2 | 1.3
Necrotic | 2 | 1.3
Total | 154 | 100.0

NLH most commonly presented as a solitary mass, consistent with the significant association between this diagnosis and single‐mass presentation described above. However, NLH was not significantly associated with hemoperitoneum (OR: 0.19; 95% CI: 0.04–0.82; p = 0.027).

Co‐occurrence among the most common diagnoses was frequent. Splenitis overlapped with splenic hematoma in 28.6% of affected dogs (44/154), whereas concurrent NLH was identified in 10.4% (16/154).

Consistent secondary histopathological lesions were observed across the major diagnoses. Hemorrhage and extramedullary hematopoiesis were the most frequent accompanying findings, particularly in cases of splenitis (79.9% and 83.1%, respectively), splenic hematoma (84.2% and 68.4%), and NLH (65.0% and 54.0%). Detailed secondary histopathological findings are provided in Supporting Table S2.

Other primary diagnoses collectively represented a minor proportion of cases (Supporting Table S1).

3.6. Neoplastic Splenic Lesions

Splenic neoplasms accounted for 33.5% of cases (142/424 dogs), of which 91.5% (130/142) exhibited malignant behavior (Table 4). Tumor grading was not systematically applied due to the limited availability and inconsistent validation of grading systems for canine splenic neoplasms [30]. Detailed demographic characteristics, macroscopic presentation, concurrent diagnoses, and associated histopathological findings for each neoplasm are presented in Supporting Table S3.

Neoplastic diagnoses | Cases n (%) | Median age years (range) | Sex (F/M) | Most affected breeds | Common macroscopic pattern | Common concurrent histopathological findings
Hemangiosarcoma (HSA)a | 90 (63.4)b | 10.6 (5–17) | 42/48 | Crossbreed, Schnauzer | Single mass | Hemorrhage, necrosis, splenic hematoma, inflammation
Lymphomac | 20 (14.1) | 9.3 (4–14) | 11/9 | Crossbreed, Bulldog, Rottweiler | Splenomegaly | Lymphoid tissue changes∗, hemorrhage, necrosis
Undifferentiated sarcomad | 8 (5.6) | 10.0 (7–12) | 3/5 | Pit Bull | Single mass | Hemorrhage, inflammation, necrosis
Myelolipoma | 8 (5.6) | 10.0 (8–13) | 2/6 | Labrador Retriever, Crossbreed | Single mass | Inflammation, extramedullary hematopoiesis (EMH), hemorrhage, lymphoid tissue changes

Concurrent diagnoses were less frequent in neoplastic cases and primarily corresponded to reactive or degenerative non‐neoplastic changes. In contrast to non‐neoplastic lesions, neoplastic diagnoses were mutually exclusive, and no dog presented more than one primary splenic neoplasm.

Among malignant neoplasms, HSA (Figure 3A,B) predominated (69.2%, 90/130). Dogs aged 9–12 years showed significantly increased odds of developing this neoplasm (OR: 1.99; 95% CI: 1.21–3.26; p = 0.009). Hemoperitoneum and multiple splenic masses were also significantly associated with HSA, as described above. However, among dogs with available clinical information, 57.9% (44/76) of HSA cases did not present with abdominal bleeding at diagnosis. Within this subgroup, splenic lesions were most commonly detected incidentally by ultrasonography (38.6%, 17/44), whereas others presented with abdominal distension and/or pain (34.1%, 15/44). Additional nonsignificant clinical and demographic associations are summarized in Table 1. Lymphoma ranked second (Figure 3C,D), accounting for 15.4% (20/130) of malignant tumors.

FIGURE 3: Histopathological neoplastic splenic features using hematoxylin and eosin stain. (A) Well-differentiated hemangiosarcoma (10×/40×): composed of spindle‐shaped cells arranged in vascular spaces containing erythrocytes; cells exhibit oval, hyperchromatic nuclei, moderate anisokaryosis, and fibrillar cytoplasm. (B) Poorly differentiated hemangiosarcoma (4×/40×): composed of cells with moderate pleomorphism arranged in a solid pattern or irregular channels containing erythrocytes. This neoplasm is surrounded by an extensive hematoma: spindle‐shaped and ballooniform cells with oval nuclei and cytoplasm exhibiting indistinct cell borders. (C) Small cell lymphoma (10×/40×): a neoplasm consisting of round cells, diffusely distributed and supported by scant fibrous stroma: small cells with marked atypia, spherical, euchromatic nuclei, prominent nucleoli, moderate anisokaryosis, and scant cytoplasm. (D) Follicular lymphoma (10×/40×): neoplastic lymphocytes organized into follicles: medium‐sized cells with spherical, euchromatic nuclei containing 1 to 3 prominent nucleoli, moderate anisokaryosis, and scant cytoplasm. (E) Non-angiogenic, non-lymphogenic sarcoma (10×/40×): a mesenchymal neoplasm exhibiting a solid pattern, with histological origin yet to be clarified: cells displaying marked pleomorphism and anisokaryosis, variable chromatin, prominent nucleoli, and variable eosinophilic cytoplasm. (F) Undifferentiated neoplasm (10×/40×): focus of non‐encapsulated neoplasia with a pseudoacinar arrangement, bordered by areas of hemorrhage and fibroplasia: cells with large nuclei containing a prominent central nucleolus, vacuolated to fibrillar cytoplasm with relatively well‐defined cell borders. Histological features did not allow definitive determination of histogenetic origin based on routine histology alone. (G) Fibrosarcoma (10×/40×): abundant fusiform cells organized in interwoven fascicles: spindle‐shaped cells with oval to elongated, euchromatic nuclei, moderate anisokaryosis, and fibrillar cytoplasm. (H) Histiocytic sarcoma (10×/40×): organized in a diffuse pattern of large cells with abundant cytoplasm, marked atypia, and a high degree of pleomorphism in mononuclear cells: frequent presence of multinucleated giant cells.

Consistent secondary histopathological lesions were observed across the major malignant neoplasms. Hemorrhage and necrosis were the most frequent, particularly in cases of HSA (78.9% and 56.7%, respectively) and lymphoma (65.0% and 40.0%). Histological features consistent with splenic hematoma were identified in 40.0% (36/90) of HSA cases compared with 31.4% (105/334) of non‐HSA cases; however, this difference was not statistically significant (OR: 1.45; 95% CI: 0.90–2.33; p = 0.13). Detailed secondary histopathological findings are provided in Supporting Table S4.

The remaining malignant neoplasms collectively accounted for 15.4% (20/130) and included undifferentiated sarcomas (Figure 3E–F), followed by fibrosarcoma (Figure 3G), histiocytic sarcoma (Figure 3H), liposarcoma, myxosarcoma, and a single mast cell tumor. Three cases originally described as fibrohistiocytic nodules and one case reported as malignant fibrous histiocytoma were reclassified as undifferentiated sarcomas according to contemporary veterinary tumor classification criteria [31] and the absence of immunohistochemical characterization. Additionally, one case originally diagnosed as an undifferentiated neoplasm was included within this category for descriptive purposes, although its definitive lineage could not be established. Benign neoplasms represented 8.5% of splenic tumors (12/142). Myelolipoma was the most frequent benign tumor (66.7%, 8/12), followed by plasmacytoma (25.0%, 3/12) and cavernous hemangioma (8.3%, 1/12).