Dai, Daying

ABSTRACT Endoluminal flow diverters are increasingly used to treat cerebral aneurysms. Despite good aneurysm occlusion efficacy, delayed ischemic complications due to side branch or perforator occlusion remain a serious concern. However, the mechanisms leading to side branch occlusion are poorly understood. To investigate these mechanisms, a combination of animal and computational models was utilized. Flow diverters were deployed in four rabbit superior mesenteric artery (SMA) and studied with computational fluid dynamics (CFD) and a previously developed fibrin dynamics model. Subject‐specific vascular models were reconstructed from 3D rotational angiography images and flow conditions were derived from Doppler ultrasound velocity measurements. Flow diverting devices were virtually deployed within the vascular models under guidance from angiography images. Coupled flow and fibrin production, transport, and adhesion simulations were carried out and the effects of fibrin accumulation at the origin of jailed side branches after flow diverter implantation were analyzed. The results of the computational models suggest significant fibrin accumulation across the struts of all flow diverting devices jailing side branches. Larger accumulation was observed at the more distal branches. However, minor flow alterations were produced which were not detectable in angiography sequences. These results were in agreement with experimental observations. On the other hand, fibrin accumulation alone did not explain complete side branch occlusions, which are likely the result of the interaction between initial fibrin accumulation and subsequent cell migration from the peripheral parent artery.

Background Animal models are essential for evaluating endovascular device safety and efficacy. Rats offer advantages, such as genetic manipulability, lower cost, and shorter healing and breeding cycles, compared to large animals. These features support studying aneurysm healing mechanisms and enable high-throughput testing. We developed an optimal rat aneurysm model for device evaluation. Methods Saccular aneurysms were created at the origins of the left renal artery (LRA) with acetylsalicylic acid (ASA) (LRA/ASA group, n = 7); the right common iliac artery (RCIA) with ASA alone (RCIA/ASA group, n = 6) or ASA plus clopidogrel (CLP) (RCIA/ASA + CLP group, n = 7); and the left common iliac artery (LCIA) with ASA + CLP (LCIA/ASA + CLP group, n = 7). The origins of these arteries were surgically exposed. During temporary ligation of the vessel origin, the vessel was bisected and endoluminal elastase was incubated in the proximal stump for 10 min, followed by release of the proximal ligation and permanent ligation of the stump. Angiographical and histological analysis were performed 4 weeks post-procedure. Results Follow-up digital subtraction angiography revealed mean ± standard deviation aneurysm height/neck dimensions of 1.2 ± 0.4/1.2 ± 0.1, 2.9/3.1, 2.3 ± 0.6/1.9 ± 0.3, and 2.1 ± 0.6/2.0 ± 0.4 mm for the LRA/ASA, RCIA/ASA, RCIA/ASA + CLP, and LCIA/ASA + CLP groups, respectively. The survival rate was 29%, 17%, 71%, and 86% in the LRA/ASA, RCIA/ASA, RCIA/ASA + CLP, and LCIA/ASA + CLP groups, respectively. Histopathological analysis of these aneurysms confirmed the absence of the internal elastic lamina and revealed aneurysmal changes in the arterial wall, resembling the pathological findings observed in human aneurysm specimens. Conclusions Common iliac aneurysm models with ASA + CLP can be used to evaluate the safety and efficacy of endovascular devices.

Background Extracellular DNA traps (ETs) have important implications in both thrombosis and thrombolysis. Thus, developing benchtop thrombus analogs that recapitulate clinical ETs is potentially of great value for preclinical development and testing of thrombolytic agents and thrombectomy devices. In this study, we aimed to develop ETs-rich thrombus analogs for preclinical testing. Methods Red blood cell (RBC)-rich, fibrin-rich, and platelet-rich thrombus analogs were created using human whole blood, platelet-poor plasma, and platelet-rich plasma obtained from the blood bank following institutional approval. Peripheral blood mononuclear cells (9.9×106 cells/mL) isolated from human whole blood and lipopolysaccharide (1 µg/mL) were added to induce ETs. Histochemical, immunohistochemistry and immunofluorescence were used to identify thrombus components and ETs. Scanning electronic microscopy was used to investigate the ultrastructure of the thrombus analogs. The thrombus compositions, morphologic features of ETs and citrullinated histone H3 (H3Cit) expression were compared with those of thrombi retrieved from patients by thrombectomy. Results ETs-rich thrombus analogs were more compacted th-an the ETs-poor thrombus analogs. ETs were identified in both ETs-rich thrombus analogs and patient thrombi showing morphologic features including nuclear lobulation, nuclear swelling, diffused chromatin within cytoplasm, DNA/chromatin extending intracellularly and extracellularly, and extracellular chromatin patches and bundles. In the ETs-poor thrombus analogs, ETs were not observed and H3Cit expression was absent to minimal. The compositions and H3Cit expression in the ETs-rich thrombus analogs fell in the range of patient thrombi. Conclusions ETs-rich thrombus analogs can be consistently created in vitro and may benefit the preclinical development and testing of new thrombolytic agents and thrombectomy devices.

Background & Purpose It has been hypothesized that circulating neutrophils have a direct correlation with the composition of emboli in acute ischemic stroke (AIS). The aim of this study is to evaluate the association between neutrophil-lymphocyte ratio (NLR) in peripheral blood and the expression of neutrophil extracellular traps (NETs) within stroke emboli. Methods Consecutive patients with acute ischemic stroke (AIS) due to large vessel occlusion (LVO) that underwent mechanical thrombectomy (MT) were included. Patients were divided into two groups based on NLR median value. Retrieved thrombi were histologically analyzed using Martius Scarlett Blue (MSB) for main thrombus components including red blood cells (RBCs), white blood cells (WBCs), fibrin and platelet. Immunohistochemistry staining for von Willebrand Factor (vWF) and anti-citrullinated H3 (H3Cit; NETs marker) was also performed. Results Samples from a total of 84 patients were included. The average percentage of RBCs, WBCs, fibrin, platelet, H3Cit, and vWF components in thrombi were 45.1%, 3.5%, 21.8%, 29.6%, 19.7% and 14.8% respectively. When stratifying by NLR group [low (≤3.94) versus high (>3.95)], high NLR group had significantly more WBCs (4.5%), fibrin (24.2%), H3Cit (22.7%) and vWF (17.1%) thrombus fractions compared to low NLR group. Additionally, RBC content (38.8%) was lower in the high NLR group. Conclusions NLR is correlated with the amounts of WBCs, fibrin, NETs and vWF within the thrombi retrieved from AIS patients due to LVO.

Background Flow diverters (FDs) are an effective treatment for intracranial aneurysms, though not free from hemorrhagic complications. A previous study demonstrated increased vascular contractility after FD-implantation as a potential mechanism of distal complications. Our study aimed to investigate whether L-arginine medication affects vascular contractility following FD deployment in a rabbit model. Methods FDs were implanted in the aorta of normal rabbits (+FD, n = 10), with sham-operated aorta as controls (n = 5). L-Arginine was given in the drinking water (2.25% L-arginine hydrochloride) of half of the +FD animals (+FD/+Arg). Force contraction vascular contractility studies were performed on the aortic rings proximal and distal to the FD using an organ bath. Total eNOS, eNOS(pS1177), eNOS(pT495), COX-2, and S100A4 were quantified by western analysis on total protein lysates from aortic segments, normalizing to GAPDH. Results Mean vascular contractility was 53% higher in distal relative to proximal aortic segments (P = 0.0038) in +FD animals, but were not significantly different in +FD/+Arg animals, or in sham-operated controls. The +FD animals expressed significantly reduced levels of eNOS(pS1177) than sham-operated controls (P = 0.0335), while both the +FD and +FD/+Arg groups had reduced levels of eNOS(pT495) relative to sham-operated controls (P = 0.0331 and P = 0.0311, respectively). Conclusion These results suggest that L-arginine medication reduces distal vascular contractility after FD treatment via nitric oxide production and thus might mitigate risk for downstream complications.

Background Several animal studies have demonstrated that mechanical thrombectomy (MT) for acute ischemic stroke (AIS) may cause vessel wall injury (VWI). However, the histological changes in human cerebral arteries following MT are difficult to determine. Objective To investigate the occurrence of VWI during MT by histological and immunohistochemical evaluation of AIS clots. Methods As part of the multicenter STRIP registry, 277 clots from 237 patients were analyzed using Martius Scarlett Blue stain and immunohistochemistry for CD34 (endothelial cells) and smooth muscle actin (smooth muscle cells). Results MT devices used were aspiration catheters (100 cases), stentriever (101 cases), and both (36 cases). VWI was found in 33/277 clots (12%). There was no significant correlation between VWI and MT device. The degree of damage varied from grade I (mild intimal damage, 24 clots), to grade II (relevant intimal and subintimal damage, 3 clots), and III (severe injury, 6 clots). VWI clots contained significantly more erythrocytes (p=0.006*) and less platelets/other (p=0.005*) than non-VWI clots suggesting soft thrombus material. Thrombolysis correlated with a lower rate of VWI (p=0.04*). VWI cases showed a significantly higher number of passes (2 [1–4] vs 1 [1–3], p=0.028*) and poorer recanalization outcome (p=0.01*) than cases without VWI. Conclusions Histological markers of VWI were present in 12% of AIS thrombi, suggesting that VWI might be related to MT. VWI was associated with soft thrombus consistency, higher number of passes and poorer revascularization outcome. There was no significant correlation between VWI and MT device.

Background and aim Mechanical thrombectomy (MT) for large vessel occlusion often requires multiple passes to retrieve the entire thrombus load. In this multi-institutional study we sought to examine the composition of thrombus fragments retrieved with each pass during MT. Methods Patients who required multiple passes during thrombectomy were included. Histopathological evaluation of thrombus fragments retrieved from each pass was performed using Martius Scarlet Blue staining and the composition of each thrombus component including RBC, fibrin and platelet was determined using image analysis software. Results 154 patients underwent MT and 868 passes was performed which resulted in 263 thrombus fragments retrieval. The analysis of thrombus components per pass showed higher RBC, lower fibrin and platelet composition in the pass 1 and 2 when compared to pass 3 and passes 4 or more combined (P values <0.05). There were no significant differences between thrombus fragments retrieved in pass 1 and pass 2 in terms of RBC, WBC, fibrin, and platelet composition (P values >0.05). Similarly, when each composition of thrombus fragments retrieved in pass 3 and passes 4 or more combined were compared with each other, no significant difference was noted (P values >0.05). Conclusion Our findings confirm that thrombus fragments retrieved with each pass differed significantly in histological content. Fragments in the first passes were associated with lower fibrin and platelet composition compared to fragments retrieved in passes three and four or higher. Also, thrombus fragments retrieved after failed pass were associated with higher fibrin and platelet components.

Background To develop a preclinical thromboembolic occlusion model for studying revascularization strategies. Methods Clot analog with barium sulfate was injected into the distal aorta in 9 New Zealand white rabbits. The situation of aorta occlusion was compared among fibrin-rich (n=4), red blood cell (RBC)-rich (n=3), and whole blood clot analogs (n=2) using digital subtraction angiography. Arterial geometries, histologic features and circumferential stretch of the distal aorta in rabbits were compared with the common carotid artery in swine and the distal internal carotid artery (ICA) in humans. Aspiration thrombectomy and mechanical thrombectomy using a stent retriever were performed in two rabbits. Results The aortic bifurcation was occluded after a single delivery of clot in 4 cases. It was occluded after the second clot injection in the 5 remaining rabbits. Fragmentation of RBC-rich clots occurred during clot injection in 2 cases. The mean diameters of the distal aorta and right common iliac artery in rabbits were 3.7±0.4 and 2.8±0.3 mm, respectively; the mean diameters of human ICA, and first and second segments of the middle cerebral artery (M1, M2) were 3.6±0.4, 3.1±0.4, and 2.4±0.4 mm, respectively. Arterial revascularization was achieved in both rabbits. Geometric, mechanical and histological factors of the distal aorta in rabbit were more close to human distal ICA than swine carotid artery. Conclusion Arterial occlusion can be achieved at the aortic bifurcation in rabbits, which is comparable to human ICA bifurcation. This thrombectomy model has the potential to be used for testing of thrombectomy devices.

Background Characterization of acute ischemic stroke (AIS) clots has typically focused on two-dimensional histological analysis of the thrombus. The three-dimensional (3D) architecture and distribution of components within emboli have not been fully investigated. The aim of this study was to examine the composition and microstructure of AIS clots using histology and serial block-face scanning electron microscopy (SBFSEM). Methods As part of the multi-institutional STRIP registry, 10 consecutive AIS emboli were collected from 10 patients treated by mechanical thrombectomy. Histological and immunohistochemical analysis was performed to determine clot composition. SBFSEM was used to assess the ultrastructural organization of the clots and specific features of individual components. Results Quantification of Martius Scarlett Blue stain identified fibrin (44.4%) and red blood cells (RBCs, 32.6%) as the main components. Immunohistochemistry showed a mean platelet and von Willebrand factor content of 23.9% and 11.8%, respectively. The 3D organization of emboli varied greatly depending on the region analyzed. RBC-rich areas were composed mainly of tightly packed RBCs deformed into polyhedrocytes with scant fibrin fibers interwoven between cells. The regions with mixed composition showed thick fibrin fibers along with platelets, white blood cells and RBC clusters. Fibrin-rich areas contained dense fibrin masses with sparse RBC. In three cases, the fibrin formed a grid-like or a sponge-like pattern, likely due to thrombolytic treatment. Segmentation showed that fibrin fibers were thinner and less densely packed in these cases. Conclusions 3D-SEM provides novel and potentially clinically relevant information on clot components and ultrastructure which may help to inform thrombolytic treatment and medical device design.