Histopathological composition of thrombus material in a large cohort of patients with acute ischemic stroke: a study of atypical clots

Abstract

Introduction: Clot composition appears to be associated with outcomes in stroke recanalization therapy. This study aims to analyze thrombus composition and assess the relationship between atypical clot composition and clinical parameters, laboratory markers, and recanalization strategies in a series of patients with acute ischemic stroke (AIS) undergoing endovascular treatment (EVT).

Methods: This is a prospective single-center registry conducted from December 2014 to July 2022. All retrieved clots were examined under an established protocol and classified as follows: red blood cell-rich clots (RBC), fibrin/platelet-rich clots (FPC), mixed clots (MC), septic emboli (SE), atheroma emboli (AE), fatty emboli (FE), and calcium emboli (CE). We categorized them into two groups: atypical clot composition (ACC: SE, AE, FE, and CE) and usual clot composition (UCC: RBC, FPC, and MC). A subgroup of 10 ACC (four SE, three AE, two FE, and one CE) and nine UCC (three RBC, three FPC, and three MC), matched by age and sex, was analyzed using immunohistochemistry to detect neutrophil extracellular traps (NETs).

Results: A total of 606 patients were assessed for EVT, with 448 (73.92%) meeting the inclusion criteria. The clot categorization was as follows: FPC 211 (47.1%), RBC 105 (23.4%), MC 104 (23.2%), SE 16 (3.6%), AE 5 (1.1%), CE 4 (0.9%), and FE 3 (0.7%). Consequently, we classified 420 (93.75%) patients into the UCC group and 28 (6.25%) into the ACC group. Bivariate analysis revealed that the ACC group had a significantly higher number of leukocytes (11.40 leukocytes/mm³ vs. 9.49, p = 0.005), a greater frequency of TICA occlusion (28.6% vs. 9.8%, p = 0.006), and higher mortality at three months (28.6% vs. 12.4%, p = 0.038). Multivariate analysis indicated that atypical clot composition was significantly associated with a higher prevalence of diabetes mellitus, smoking, occlusion of the terminal internal carotid artery, and an increased number of passes. Immunohistochemical studies showed the presence of neutrophil extracellular traps (NETs) in all 19 thrombi that were analyzed.

Conclusion: Diabetes and TICA occlusion were the strongest predictors of atypical clot composition. We also observed a significant association between atypical composition and an increased number of passes. Furthermore, the presence of NETs in all thrombi analyzed, regardless of their composition, indicates inflammatory mechanisms associated with clot formation and consolidation in AIS.

Keywords: atypical clot; histopathological; ischemic stroke; mechanical thrombectomy; septic emboli.

Figure 1

A flowchart with the results of included patients. MCA, middle cerebral artery; TICA, terminal internal carotid artery; BAS, basilar; PCA-P1, posterior cerebral artery P1 segment; PCA-P2, posterior cerebral artery P2 segment; ACA, anterior cerebral artery; TICI, thrombolysis in cerebral infarction; UCC, usual clot composition; ACC, atypical clot composition; RBC, red blood clot; and FPC, fibrin-predominant clot.

Figure 2

Histological and immunohistochemical study of usual thrombi. Hematoxylin–Eosin (H&E) stain (4× magnification, scale bar, 250 μm). Red blood clot (RBC): red blood cells ≥60%. Fibrin-predominant clot (FPC): fibrin/platelet ≥60%. Mixed clot: no clear predominance of components. Neutrophil extracellular trap (NET) components (indicated by arrows), myeloperoxidase (MPO), and citrullinated histone H3 (Cit-H3) were detected by immunohistochemistry at 10× (scale bar: 100 μm) and 20× (scale bar: 50 μm) magnification, respectively.

Figure 3

Histological features and NETs detection of a usual clot and atypical thrombi. H&E stain at 2× magnification (scale bar: 500 μm) and 4× magnification (scale bar: 250 μm) of a usual fibrin-rich clot, septic emboli, a fibrin-platelet-predominant clot with accumulated necrotic cells at the periphery (arrows), atheroma emboli with the blood vessel wall (arrow), and precipitated cholesterol crystals (arrow), and fatty emboli with fat cells (arrow). Gomori’s trichrome stain at 100× with immersion oil was used to analyze bacterial cocci accumulation (arrow). Representative images of Cit-H3 staining of different clots (20× magnification, scale bar, 50 μm), highlighting the accumulation of extracellular traps released by neutrophils (arrows).