Control of occlusion of middle cerebral artery in perinatal and neonatal mice with magnetic force.
ABSTRACT: Ischemic perinatal stroke (IPS) is common, resulting in significant mortality and morbidity. In such cases, the incidence of unilateral arterial cerebral infarction is often occluded in the middle cerebral artery (MCA), leading to focal ischemia. In adult rodents, blockage of MCA is the most frequently used strategy for ischemic stroke study. However, modeling MCA occlusion (MCAo) in postnatal day 0-7 (P0-7) mouse pups for IPS study has not been accomplished. Here we occluded the dMCA by inducing the accumulation of magnetic particles (MPs) administered through the superficial temporal vein of mice between P0 and P7, which we called neonatal or perinatal SIMPLE (Stroke Induced with Magnetic Particles). SIMPLE produced either permanent or transient occlusion in the dMCA of perinatal and neonatal mice. Permanent MCA occlusion with SIMPLE resulted in cerebral infarction and neuronal death in the brain. SIMPLE can also be used to reliably produce focal ischemic stroke in neonatal or perinatal mouse brains. As a result, SIMPLE allows the modeling of IPS or focal ischemic stroke for further mechanistic studies in mice, with particular utility for mimicking transient focal ischemia in human pre-term babies, which for the first time here has been accomplished in mice.
Project description:BACKGROUND: Animal models are essential to study the pathophysiological changes associated with focal occlusive stroke and to investigate novel therapies. Currently used rodent models have yielded little clinical success, however large animal models may provide a more suitable alternative to improve clinical translation. We sought to develop a model of acute proximal middle cerebral artery (MCA) ischemic stroke in sheep, including both permanent occlusion and transient occlusion with reperfusion. MATERIALS AND METHODS: 18 adult male and female Merino sheep were randomly allocated to one of three groups (n?=?6/gp): 1) sham surgery; 2) permanent proximal MCA occlusion (MCAO); or 3) temporary MCAO with aneurysm clip. All animals had invasive arterial blood pressure, intracranial pressure and brain tissue oxygen monitoring. At 4 h following vessel occlusion or sham surgery animals were killed by perfusion fixation. Brains were processed for histopathological examination and infarct area determination. 6 further animals were randomized to either permanent (n?=?3) or temporary MCAO (n?=?3) and then had magnetic resonance imaging (MRI) at 4 h after MCAO. RESULTS: Evidence of ischemic injury in an MCA distribution was seen in all stroke animals. The ischemic lesion area was significantly larger after permanent (28.8%) compared with temporary MCAO (14.6%). Sham animals demonstrated no evidence of ischemic injury. There was a significant reduction in brain tissue oxygen partial pressure after permanent vessel occlusion between 30 and 210 mins after MCAO. MRI at 4 h demonstrated complete proximal MCA occlusion in the permanent MCAO animals with a diffusion deficit involving the whole right MCA territory, whereas temporary MCAO animals demonstrated MRA evidence of flow within the right MCA and smaller predominantly cortical diffusion deficits. CONCLUSIONS: Proximal MCAO can be achieved in an ovine model of stroke via a surgical approach. Permanent occlusion creates larger infarct volumes, however aneurysm clip application allows for reperfusion.
Project description:Ischemic stroke accounts for over 80% in total human stroke which mostly affect middle cerebral artery (MCA) territory. Embolic stroke models induced by injection of homologous clots into the internal carotid artery and MCA closely mimic human stroke and have been commonly used in stroke research. Studies indicate that the size and composition of clots are critical for the reproducibility of the stroke model. In the present study, we modified the homologous clots formation by addition of thrombin and fibrinogen which produced even distribution of fibrin with tight cross linkage of red blood cells. We optimized the embolic MCA occlusion model in rats using different size of the mixed clots. A precise lodgment of the clots at the MCA bifurcation and highly reproducible ischemic lesion in the MCA territory were demonstrated in the embolic MCA occlusion model induced by injection of 10 pieces of 1-mm long mixed clots made in PE-60 catheter. We further tested the effect of recombinant tissue plasminogen activator (rtPA) in this embolic MCA occlusion model. rtPA induced thrombolysis, improved neurological outcome, and significantly reduced ischemic lesion volume when administered at 1h after embolism as compared with control. In summary, we have established a reproducible embolic MCA occlusion model using clots made of homologous blood, thrombin and fibrinogen. The mixed clots enable precise lodgment at the MCA bifurcation which is responsive to thrombolytic therapy of rtPA.
Project description:By means of introgressing a loss-of-function mutation in the p22phox gene from the Matsumoto Eosinophilia Shinshu (MES) rat to stroke-prone spontaneously hypertensive rats (SHRSP), we constructed the SHRSP-based congenic strain lacking the P22PHOX expression (i.e., lacking NADPH oxidases [NOX] activities) (SHRSP.MES-Cyba(mes)/Izm; hereafter referred to as SP.MES). To examine the effects of Nox activities on the focal ischemic injury or stroke, we performed middle cerebral artery (MCA) occlusion in this new congenic strain; the distal MCA was occluded by 561-nm laser-driven photothrombosis. Resting mean arterial blood pressure was significantly lower in SP.MES when compared with the control PM0/SHRSP (150±11 mmHg vs. 166±11 mmHg). Cerebral blood flow decreased to 37±13% in SP.MES and 35±17% in PM0/SHRSP at 10 min after MCA occlusion (not significant). Infarct volume determined at 24 h after MCA occlusion in SP.MES was 89±39 mm3, which was not significantly different from 83±35 mm3 in PM0/SHRSP. The distal MCA pattern was more complex in SP.MES (median 3, IQR 3-5) than PM0/SHRSP (median 2, IQR 1-3) (p = 0.001). Because more complex distal MCA is known to produce larger infarction after distal MCA occlusion in SHR, we adjusted for the branching pattern in an ANCOVA. The adjusted mean of infarct volume was significantly smaller in SP.MES compared with that in PM0/SHRSP (67 [95% CI 46 to 87] mm3 vs. 100 [95% CI 82 to 118] mm3, p = 0.032). Elimination of the P22PHOX expression induced complex distal MCA, which would suggest the presence of 'loss of complexity' induced by enhanced oxidative stress in SHRSP; infarct size in SP.MES--when adjusted for distal MCA complexity--was significantly attenuated compared with that in PM0/SHRSP. Therefore, the present results suggest that Nox is harmful for ischemic brain tissue.
Project description:SMTP-7 (Stachybotrys microspora triprenyl phenol-7), a small molecule that promotes plasminogen activation through the modulation of plasminogen conformation, has excellent therapeutic activity against cerebral infarction in several rodent models. Detailed evaluations of SMTP-7 in a primate stroke model are needed for effective, safe drug development. Here we evaluated SMTP-7 in a monkey photochemical-induced thrombotic middle cerebral artery (MCA) occlusion model (n=6), in which MCA occlusion was followed by recanalization/reocclusion. SMTP-7 (10?mg/kg, intravenous infusion) significantly increased the postinfusion MCA recanalization rate (32.5-fold, P=0.043) and ameliorated the post-24-h neurologic deficit (by 29%, P=0.02), cerebral infarct (by 46%, P=0.033), and cerebral hemorrhage (by 51%, P=0.013) compared with the vehicle control animals. In normal monkeys, SMTP-7 did not affect general physiologic or hemostatic variables, including coagulation and platelet parameters. Investigations in rodent models of transient and permanent focal cerebral ischemia, as well as arterial thrombosis and bleeding tests, suggest a role for SMTP-7's regulated profibrinolytic action and neuroprotective properties in the monkey MCA occlusion model. In conclusion, SMTP-7 is effective in treating thrombotic stroke in monkeys. SMTP-7 is thus a promising candidate for the development of alternative therapy for ischemic stroke.
Project description:Occlusion of the middle cerebral artery (MCA) by an intraluminal filament is widely used to study focal brain ischemia in male Sprague-Dawley rats. However, permanent occlusion goes along with a high fatality. To overcome this drawback we designed a new filament carrying a bowling pin-shaped tip (BP-tip) and compared this with three conventionally tipped filaments. Follow-up periods were 24?h (all groups) and 72 and 120?h in BP-tip group. Ischemic damage and swelling were quantified using silver nitrate staining. Collateral flow via the posterior cerebral artery (PCA) was assessed using selective dye perfusion of the internal carotid artery. Despite a comparable decrease of brain perfusion in all groups, ischemic damage was significantly smaller in BP-tips (p?<?0.05). Moreover, BP-tip significantly reduced mortality from 60% to 12.5% and widely spared the occipital region and hypothalamus from ischemic damage. Conventional but not BP-tip filaments induced vascular distortion, measured as gross displacement of the MCA origin, which correlated with occipital infarction size. Accordingly, BP-tip occluded rats showed a significantly better collateral filling of the PCA territory. Ischemic volume significantly increased in BP-tip occlusion at 72?h follow-up. BP-tip filaments offer superior survival in permanent MCA occlusion, while mimicking the course of a malignant stroke in patients.
Project description:BACKGROUND:Previous experimental studies have shown that downstream microvascular thromboinflammation is involved in brain damage from acute ischemic stroke. Using intravital microscopy, we investigated and characterized the sequence of downstream microvascular thromboinflammation in an ischemia/reperfusion acute ischemic stroke model. METHODS AND RESULTS:Rats underwent transient monofilament middle cerebral artery (MCA) occlusion. Cerebral microcirculation in the MCA territory was exposed through a craniotomy and analyzed using real-time intravital imaging coupled with laser Doppler interferometry. Leukocytes, platelets, fibrinogen, and blood-brain barrier permeability were analyzed by intravenous injection of fluorescent antibodies and bovine serum albumin. MCA occlusion induced a sudden and profound drop in downstream microvascular blood flow associated with leukocyte margination in the venous compartment. Leukocyte margination fostered fibrinogen deposition and thrombosis in postcapillary venules. Either in venules or arterioles, blood flow was not fully restored after MCA recanalization. Furthermore, venular thrombi persisted despite MCA recanalization, and leukocyte extravasation continued to develop in venules in association with blood-brain barrier disruption. Finally, microhemorrhages were occasionally observed, colocalizing with thrombosed venules characterized by marked leukocyte margination. CONCLUSIONS:We showed that microvascular thrombosis in transient monofilament MCA occlusion and blood-brain barrier disruption are initiated immediately after occlusion and are propagated through the venous compartment in close association with marginating leukocytes. MCA occlusion-induced downstream microvascular thromboinflammation response was responsible for incomplete reperfusion after MCA recanalization and delayed microhemorrhages.
Project description:Cerebral collaterals are dynamically recruited after arterial occlusion and highly affect tissue outcome in acute ischemic stroke. We investigated the efficacy and safety of four pathophysiologically distinct strategies for acute modulation of collateral flow (collateral therapeutics) in the rat stroke model of transient middle cerebral artery (MCA) occlusion. A composed randomization design was used to assign rats (n?=?118) to receive phenylephrine (induced hypertension), polygeline (intravascular volume load), acetazolamide (cerebral arteriolar vasodilation), head down tilt (HDT) 15° (cerebral blood flow diversion), or no treatment, starting 30?min after MCA occlusion. Compared to untreated animals, treatment with collateral therapeutics was associated with lower infarct volumes (62% relative mean difference; 51.57?mm<sup>3</sup> absolute mean difference; p?<?0.001) and higher chance of good functional outcome (OR 4.58, p?<?0.001). Collateral therapeutics acutely increased cerebral perfusion in the medial (+40.8%; p?<?0.001) and lateral (+19.2%; p?=?0.016) MCA territory compared to pretreatment during MCA occlusion. Safety indicators were treatment-related mortality and cardiorespiratory effects. The highest efficacy and safety profile was observed for HDT. Our findings suggest that acute modulation of cerebral collaterals is feasible and provides a tissue-saving effect in the hyperacute phase of ischemic stroke prior to recanalization therapy.
Project description:Vitamin E consists of tocopherols and tocotrienols, in which ?-tocotrienol is the most potent neuroprotective form that is also effective in protecting against stroke in rodents. As neuroprotective agents alone are insufficient to protect against stroke, we sought to test the effects of tocotrienol on the cerebrovascular circulation during ischemic stroke using a preclinical model that enables fluoroscopy-guided angiography. Mongrel canines (mean weight=26.3±3.2 kg) were supplemented with tocotrienol-enriched (TE) supplement (200 mg b.i.d, n=11) or vehicle placebo (n=9) for 10 weeks before inducing transient middle cerebral artery (MCA) occlusion. Magnetic resonance imaging was performed 1 hour and 24 hours post reperfusion to assess stroke-induced lesion volume. Tocotrienol-enriched supplementation significantly attenuated ischemic stroke-induced lesion volume (P<0.005). Furthermore, TE prevented loss of white matter fiber tract connectivity after stroke as evident by probabilistic tractography. Post hoc analysis of cerebral angiograms during MCA occlusion revealed that TE-supplemented canines had improved cerebrovascular collateral circulation to the ischemic MCA territory (P<0.05). Tocotrienol-enriched supplementation induced arteriogenic tissue inhibitor of metalloprotease 1 and subsequently attenuated the activity of matrix metalloproteinase-2. Outcomes of the current preclinical trial set the stage for a clinical trial testing the effects of TE in patients who have suffered from transient ischemic attack and are therefore at a high risk for stroke.
Project description:Anterior circulation large artery occlusion (AC-LAO) related acute ischemic stroke (AIS) is particularly common in clinics in China. We retrospectively analyzed 787 consecutively hospitalized AIS patients with AC-LAO in Hebei Province, China. AC-LAO was defined as a complete occlusion of at least one intracranial internal carotid artery (ICA) or middle cerebral artery (MCA) based on computed tomography or magnetic resonance angiography. Among eight subtypes of AC-LAO, unilateral MCA occlusion is the most common one (49.8%, n?=?392), while bilateral ICA/unilateral MCA occlusion is the least (0.3%, n?=?2). Compared with unilateral MCA and unilateral ICA occlusion, patients with tandem ICA/MCA and bilateral ICA/MCA occlusion had poor outcomes after suffering AIS. Age (OR 1.022; 95%CI, 1.007 to 1.036) was an independent risk factor for single artery progressed to multiple artery occlusion, while ApoA1 (OR 0.453; 95% CI, 0.235 to 0.953) was a protective factor. Patients with unilateral MCA occlusion were prone to artery-to-artery embolism infarction subtype, unilateral ICA occlusion group were the most vulnerable to hypoperfusion/impaired emboli clearance subtype. Our results suggested various AC-LAO subtypes have different clinical characteristics and prognosis and were prone to different subtypes of infarction. Customized preventive measures based on AC-LAO subtypes may be more targeted preventions of stroke recurrences for AIS patients and could improve their prognoses.
Project description:Background and objective: The aim was to evaluate the clinical significance of prominent fluid-attenuated inversion recovery (FLAIR) vascular hyperintensity (FVH) on the prognosis of mild acute ischemic stroke with middle cerebral artery (MCA) occlusion. Methods: We recruited consecutive stroke patients with initial National Institutes of Health Stroke Scale (NIHSS) scores ≤5 and MCA occlusion on magnetic resonance angiography within 24 h of stroke onset. Prominent distal FVH was defined as an extension to more than one-third of the MCA territory. We compared clinical outcomes between prominent and non-prominent FVH groups in patients who had and had not received reperfusion therapy. Results: Of 112 participants [43 women; median age, 67 years [Interquartile range, 54-79]], prominent FVH was identified in 80 (71.4%). For 75 patients who had not received reperfusion therapy, the prominent FVH group had a more unfavorable outcome (modified Rankin Scale score >1) at 3 months than the non-prominent FVH group (44.4 vs. 15.0%, P = 0.029). In multivariate analysis, a higher NIHSS score [odd ratio [OR] = 1.67; 95% confidence interval [CI], 1.16-2.41; P = 0.006], proximal MCA occlusion [OR = 7.31; 95% CI, 1.68-31.9; P = 0.008], and prominent FVH [OR = 5.49; 95% CI, 1.29-23.4; P = 0.021], were independently associated with an unfavorable outcome. There was no association between prominent FVH and the clinical outcome in the reperfusion therapy group. Conclusions: For acute stroke patients with mild symptoms and MCA occlusion who do not receive reperfusion therapy, prominent FVH and proximal MCA occlusion may be independent predictors of an unfavorable outcome.