No full-text available
To read the full-text of this research,
you can request a copy directly from the authors.
Reperfusion following focal stroke hastens inflammation and resolution of ischemic injury
Abstract
Previously, we described cellular changes following Permanent Middle Cerebral Artery Occlusion (PMCAO) in spontaneously hypertensive rats. Ischemic changes following PMCAO included a time-related focal pan necrosis, inflammatory cell infiltration, gliosis, and eventual loss of necrotic tissue post PMCAO. We have now characterized changes which occur after Temporary Middle Cerebral Artery Occlusion (TMCAO; 80 or 160 min) followed by reperfusion and compared these changes to those which occur following PMCAO. TMCAO with reperfusion results in cortical infarcts which vary in size in an occlusion-time-dependent manner. After 1 h of reperfusion, ischemic changes were observed histologically, including microhemorrhages and the beginning of a slight inflammatory infiltration in and around the meningeal vasculature. This infiltrate consisted primarily of neutrophils, which by 6 h of reperfusion was significant with infiltration from deep blood vessels into brain tissue, including the presence of some monocytes adhering within blood vessels. Neutrophil infiltration occurred sooner and to a greater extent in reperfused tissues than in permanently occluded tissues, where it only began at 12 h post PMCAO. As occurred following PMCAO, increased Glial Fibrillary Acidic Protein (GFAP) immunoreactivity indicating astrogliosis was first observed at 12 h postTMCAO. Over 1-3 days of reperfusion, a heavy macrophage infiltrate was observed in the reperfused tissues in addition to a continued influx of neutrophils. Following 5 days of reperfusion, the lesion was completely replaced with inflammatory cells, of which macrophages predominated. Unlike PMCAO, which resulted in focal spots of neutrophil accumulation, neutrophils were more distributed throughout the infarcted cortex following TMCAO.(ABSTRACT TRUNCATED AT 250 WORDS)
... Lymphocytes accumulate in the brain 3-6 days after cerebral ischemia, and serve as a regulatory and neuroprotective function. 8 Adaptive immunity also exerts an immunosuppressive effect that promotes intercurrent infections. 5 Platelets participate in immune-inflammatory responses by releasing mediators to boost inflammation after stroke, which results in the release of neutrophils and lymphocytes into the vessel wall. ...
... Complications occurred in 481 of all 3402 patients (14%). Patients with complications had much higher median initial NIHSS scores (16 [8][9][10][11][12][13]) than those without complications (4 [2][3][4][5][6][7][8]; P <0.001). ...
... Complications occurred in 481 of all 3402 patients (14%). Patients with complications had much higher median initial NIHSS scores (16 [8][9][10][11][12][13]) than those without complications (4 [2][3][4][5][6][7][8]; P <0.001). ...
Purpose
We investigated the differences of clinical features, four immune-inflammatory markers, namely neutrophil counts, platelet-to-lymphocyte ratio, neutrophil-to-lymphocyte ratio (NLR), and systemic immune-inflammation index (SII), as well as outcomes between patients with in-hospital ischemic stroke (IHIS) and out-of-hospital ischemic stroke (OHIS).
Patients and Methods
We retrospectively enrolled 72 patients with IHIS and 3330 patients with OHIS.
Results
IHIS accounted for 2% of all patients with ischemic stroke and occurred more often in cardiology and orthopedic surgery wards. Infection, cardiac disease, and pulmonary disorder were the most common causes of hospitalization. Compared with those with OHIS, patients with IHIS had higher levels of immune-inflammatory markers, initial National Institute of Health Stroke Scale (NIHSS) scores, longer hospital stays, higher rates of heart disease, large-artery atherosclerosis or cardioembolism, received more intravenous thrombolysis (IVT) or endovascular thrombectomies (EVTs), more complications, unfavorable outcomes, and mortality. Every immune-inflammatory marker exhibited positive correlations with initial NIHSS scores and discharge modified Rankin Scale scores among patients with OHIS. NLR and SII were higher among patients with a fatal outcome in both groups. Among patients receiving IVT, most of treatment time intervals were shorter for those with IHIS than those with OHIS. Significant factors for mortality were NLR >5.5, atrial fibrillation, and complications, with a C-statistic of 0.897 in those with IHIS; in those with OHIS, these factors were an initial NIHSS score of >10, NLR >6.0, atrial fibrillation, prior stroke, cancer history, and complications with a C-statistic of 0.902. The results were similar after replacing the NLR with SII.
Conclusion
Patients with IHIS had more complicated clinical features, higher levels of immune-inflammatory markers, and higher rates of mortality than patients with OHIS. The most significant predictor for mortality among those with OHIS was NIHSS score >10, and the predictors among patients with IHIS were NLR >5.5 and SII >2120.
... It is one of the inflammatory processes that activates the immune system in AIS. [11] In ischemic strokes, neutrophil migration occurs in the intraparenchymal perivascular area within 6 to 24 hours following the ischemia, and neutrophils damage the blood-brain barrier with the cytokines they release into this area. [11] Lymphocytes migrate to the area 3 to 6 days after the stroke, and unlike neutrophils, undertake a regulatory function rather than acting with destructive impact. ...
... [11] In ischemic strokes, neutrophil migration occurs in the intraparenchymal perivascular area within 6 to 24 hours following the ischemia, and neutrophils damage the blood-brain barrier with the cytokines they release into this area. [11] Lymphocytes migrate to the area 3 to 6 days after the stroke, and unlike neutrophils, undertake a regulatory function rather than acting with destructive impact. In this way, lymphocytes induce neuroprotection. ...
Aim: Calculated based on platelet, neutrophil and lymphocyte counts, the systemic immune-inflammation index is thought to be associated with many malignancies in the literature. Despite the existing investigations on its diagnostic value, there have been no clear results reported regarding its diagnostic value in stroke patients. The current study is therefore intended to demonstrate the diagnostic value of the systemic immune-inflammation index and its prognostic value in cases of acute ischemic stroke.
Material and Method: A total of 150 cases of acute stroke and a control group of 150 individuals were retrospectively examined. The data recorded for each case included age, gender, history, vital findings, NIHSS, SIII, and outcome.
Results: In the current study, the group of stroke patients had significantly higher SIII than the control group. According to the diagnostic examinations, in stroke, the diagnostic value of SIII was greater than that of neutrophil-to-lymphocyte ratio at a statistically significant level. The present study also found that, compared to the SIII, the (Lymphocyte x Platelet)/Neutrophil ratio (called the reverse SIII) had a higher statistical significance in diagnosing the stroke and predicting early hospital mortality.
Conclusion: The SIII can be a good marker for both diagnostic evaluation and for predicting early hospital mortality in stroke cases. Additionally, it is approved to be a useful index since it can be calculated inexpensively and easily.
... Evolution of ischemic damage after tMCAo occurs rapidly and as a function of time, involving four key stages: necrosis, inflammation, organisation, and resolution [78,79]. Each stage follows in sequence: The infiltration of inflammatory cells is triggered by the initial necrosis. ...
... The rate of progression from neuronal necrosis to cavity formation was similar to that described by others [31,37,80]. However, this study adds to our understanding of the histological damage after stroke as it encompasses a greater number and range of timepoints (together with larger animal numbers) than previous studies (which have generally used only 3-4 animals per timepoint) [30, 31, 37, [79][80][81]. The randomised and blinded induction of ischemia and assessment of damage further enhance the value of the data presented here. ...
Key disparities between the timing and methods of assessment in animal stroke studies and clinical trial may be part of the reason for the failure to translate promising findings. This study investigates the development of ischemic damage after thread occlusion MCAo in the rat, using histological and behavioural outcomes. Using the adhesive removal test we investigate the longevity of behavioural deficit after ischemic stroke in rats, and examine the practicality of using such measures as the primary outcome for future studies. Ischemic stroke was induced in 132 Spontaneously Hypertensive Rats which were assessed for behavioural and histological deficits at 1, 3, 7, 14, 21, 28 days, 12 and 24 weeks (n>11 per timepoint). The basic behavioural score confirmed induction of stroke, with deficits specific to stroke animals. Within 7 days, these deficits resolved in 50% of animals. The adhesive removal test revealed contralateral neglect for up to 6 months following stroke. Sample size calculations to facilitate the use of this test as the primary experimental outcome resulted in cohort sizes much larger than are the norm for experimental studies. Histological damage progressed from a necrotic infarct to a hypercellular area that cleared to leave a fluid filled cavity. Whilst absolute volume of damage changed over time, when corrected for changes in hemispheric volume, an equivalent area of damage was lost at all timepoints. Using behavioural measures at chronic timepoints presents significant challenges to the basic science community in terms of the large number of animals required and the practicalities associated with this. Multicentre preclinical randomised controlled trials as advocated by the MultiPART consortium may be the only practical way to deal with this issue.
... Neutrophils are the primary cells responsible for innate immunity following a brain contusion. Within 6 to 24 hours, perivascular neutrophils migrate into the parenchyma (31,32). In the course of the illness, their numbers undergo considerable fluctuations over the course of a few days, as they have a short lifespan. ...
Background and purpose
Neutrophil-to-lymphocyte ratio (NLR) and monocyte-to-lymphocyte ratio (MLR) have been identified as potential prognostic markers in various conditions, including cancer, cardiovascular disease, and stroke. This study aims to investigate the dynamic changes of NLR and MLR following cerebral contusion and their associations with six-month outcomes.
Methods
Retrospective data were collected from January 2016 to April 2020, including patients diagnosed with cerebral contusion and discharged from two teaching-oriented tertiary hospitals in Southern China. Patient demographics, clinical manifestations, laboratory test results (neutrophil, monocyte, and lymphocyte counts) obtained at admission, 24 hours, and one week after cerebral contusion, as well as outcomes, were analyzed. An unfavorable outcome was defined as a Glasgow Outcome Score (GOS) of 0-3 at six months. Logistic regression analysis was performed to identify independent predictors of prognosis, while receiver characteristic curve analysis was used to determine the optimal cutoff values for NLR and MLR.
Results
A total of 552 patients (mean age 47.40, SD 17.09) were included, with 73.19% being male. Higher NLR at one-week post-cerebral contusion (adjusted OR = 4.19, 95%CI, 1.16 - 15.16, P = 0.029) and higher MLR at admission and at 24 h (5.80, 1.40 - 24.02, P = 0.015; 9.06, 1.45 - 56.54, P = 0.018, respectively) were significantly associated with a 6-month unfavorable prognosis after adjustment for other risk factors by multiple logistic regression. The NLR at admission and 24 hours, as well as the MLR at one week, were not significant predictors for a 6-month unfavorable prognosis. Based on receiver operating characteristic curve analysis, the optimal thresholds of NLR at 1 week and MLR at admission after cerebral contusion that best discriminated a unfavorable outcome at 6-month were 6.39 (81.60% sensitivity and 70.73% specificity) and 0.76 (55.47% sensitivity and 78.26% specificity), respectively.
Conclusion
NLR measured one week after cerebral contusion and MLR measured at admission may serve as predictive markers for a 6-month unfavorable prognosis. These ratios hold potential as parameters for risk stratification in patients with cerebral contusion, complementing established biomarkers in diagnosis and treatment. However, further prospective studies with larger cohorts are needed to validate these findings.
... Astrocytes are characteristic star-shaped glial cells in the brain and spinal cord, and play various key roles in supporting, guiding, nurturing and signalling neural structures and activities [33,34]. GFAP is mainly expressed in the astrocytes of the CNS and is relatively frequently used in the study of various diseases, such as traumatic brain injury, stroke and brain tumour [20,35]. In our study, astrocytes were activated by BCAS and the degree of activation was greater after the administration of TEX ( Figure 5). ...
Treatments to restore the balance of the temporomandibular joint (TMJ) are performed in the field of complementary and alternative medicine; however, evidence supporting this approach remains weak. Therefore, this study attempted to establish such evidence. Bilateral common carotid artery stenosis (BCAS) operation, which is commonly used for the establishment of a mouse model of vascular dementia, was performed, followed by tooth cutting (TEX) for maxillary malocclusion to promote the imbalance of the TMJ. Behavioural changes, changes in nerve cells and changes in gene expression were assessed in these mice. The TEX-induced imbalance of the TMJ caused a more severe cognitive deficit in mice with BCAS, as indicated by behavioural changes in the Y-maze test and novel object recognition test. Moreover, inflammatory responses were induced via astrocyte activation in the hippocampal region of the brain, and the proteins involved in inflammatory responses were found to be involved in these changes. These results indirectly show that therapies that restore the balance of the TMJ can be effectively used for the management of cognitive-deficit-related brain diseases associated with inflammation.
... When most vertebrates suffered from ischemia, even minutes of lacking of oxygen and nutrients leads to irreversible apoptosis and necrosis of a large number of neurons, as well as paralysis and other dysfunction (Buck & Pamenter, 2018;Sun et al., 2020). Therefore, efficient prevention measures aim to prevent neuronal apoptosis and necrosis, will be a great boon for patients suffering from hypoxic stroke (Baker et al., 1998;Bano & Nicotera, 2007;Clark et al., 1994;Lipton, 1999). ...
Naked mole‐rats (NMRs) (Heterocephalus glaber) are highly social and subterranean rodents with large communal colonies in burrows containing low oxygen levels. The inhibition of severe hypoxic conditions is of particular interest to this study. To understand the mechanisms that facilitate neuronal preservation during hypoxia, we investigated the proteins regulating hypoxia tolerance in NMR hippocampal neurons. Caveolin‐1 (Cav‐1), a transmembrane scaffolding protein, confers prosurvival signalling in the central nervous system. The present study aimed to investigate the role of Cav‐1 in hypoxia‐induced neuronal injury. Western blotting analysis and immunocytochemistry showed that Cav‐1 expression was significantly upregulated in NMR hippocampal neurons under 8% O2 conditions for 8 h. Cav‐1 alleviates apoptotic neuronal death from hypoxia. Downregulation of Cav‐1 by lentiviral vectors suggested damage to NMR hippocampal neurons under hypoxic conditions in vitro and in vivo. Overexpression of Cav‐1 by LV‐Cav‐1 enhanced hypoxic tolerance of NMR hippocampal neurons in vitro and in vivo. Mechanistically, the levels of hypoxia inducible factor‐1α (HIF‐1α) are also increased under hypoxic conditions. After inhibiting the binding of HIF‐1α to hypoxia response elements in the DNA by echinomycin, Cav‐1 levels were downregulated significantly. Furthermore, chromatin immunoprecipitation assays showed the direct role of HIF1α in regulating the expression levels of Cav‐1 in NMR hippocampal neurons under hypoxic conditions. These findings suggest that Cav‐1 plays a critical role in modulating the apoptosis of NMR hippocampal neurons and warrant further studies targeting Cav‐1 to treat hypoxia‐associated brain diseases.
... Neutrophils, which are systemic inflammation markers, infiltrate the ischemic brain within 30 minutes to a few hours [20]; it releases several cytokines and proinflammatory mediators such as inducible nitric oxide synthase and matrix metalloproteinases to contribute to inflammation in the lesion [21]. Neutrophils also participate in the early destruction of the blood-brain barrier [22]. As one hallmark of brain ischemic injury, clinical studies have shown that the infiltration of neutrophils after ischemic stroke was correlated with the severity of the injury [23,24]. ...
Purpose:
Acute ischemic stroke (AIS) is a devastating disease and remains the leading cause of death and disability. This retrospective study aims to investigate associations between systemic immune-inflammation index (SII) and all-cause mortality in patients with AIS. Patients and Methods. We used the data from Medical Information Mart for Intensive Care IV. A total of 1,181 patients with acute ischemic stroke (AIS) were included. Systemic immune-inflammation index (SII) was calculated as platelet count (/L) × neutrophil count (/L)/lymphocyte count (/L). The main outcomes were 30-day all-cause mortality. The association between SII with mortality was evaluated using the Cox proportional hazards regression model.
Results:
After adjusting for potential covariates, the highest quartiles of SII versus the lowest quartiles of SII, the HR was 2.74 (CI 1.79-4.19, P < 0.001). Log-transformed SII was significantly associated with 30-day all-cause mortality (HR 2.44; CI 1.72-3.46, P < 0.001). Furthermore, we found that there is a nearly linear relationship (P=0.265) between logarithmic transformed SII with all-cause mortality.
Conclusion:
Elevated SII of patients with acute ischemic stroke increased the risk of 30-day all-cause mortality. SII may serve as a useful marker to elucidate the role of thrombocytosis, inflammation, and immunity interaction in the development of AIS.
... Recruitment of leukocytes into the brain is a key hallmark of numerous CNS disorders. 93,94 For example, neutrophils enter the brain parenchyma early in the progression of AD. 94,95 Following stroke, neutrophils may extravasate into the parenchyma [96][97][98] or become trapped and confined within the neurovascular unit. 99 In addition, the parenchymal infiltration of highly activated T and B cells is a probable driver of early MS pathology. ...
Brain barriers are crucial sites for cerebral energy supply, waste removal, immune cell migration, and solute exchange, all of which maintain an appropriate environment for neuronal activity. At the capillary level, where the largest area of brain-vascular interface occurs, pericytes adjust cerebral blood flow (CBF) by regulating capillary diameter and maintain the blood-brain barrier (BBB) by suppressing endothelial cell (EC) transcytosis and inducing tight junction expression between ECs. Pericytes also limit the infiltration of circulating leukocytes into the brain where resident microglia confine brain injury and provide the first line of defence against invading pathogens. Brain "waste" is cleared across the BBB into the blood, phagocytosed by microglia and astrocytes, or removed by the flow of cerebrospinal fluid (CSF) through perivascular routes-a process driven by respiratory motion and the pulsation of the heart, arteriolar smooth muscle, and possibly pericytes. "Dirty" CSF exits the brain and is probably drained around olfactory nerve rootlets and via the dural meningeal lymphatic vessels and possibly the skull bone marrow. The brain is widely regarded as an immune-privileged organ because it is accessible to few antigen-primed leukocytes. Leukocytes enter the brain via the meninges, the BBB, and the blood-CSF barrier. Advances in genetic and imaging tools have revealed that neurological diseases significantly alter immune-brain barrier interactions in at least three ways: (1) the brain's immune-privileged status is compromised when pericytes are lost or lymphatic vessels are dysregulated; (2) immune cells release vasoactive molecules to regulate CBF, modulate arteriole stiffness, and can plug and eliminate capillaries which impairs CBF and possibly waste clearance; and (3) immune-vascular interactions can make the BBB leaky via multiple mechanisms, thus aggravating the influx of undesirable substances and cells. Here, we review developments in these three areas and briefly discuss potential therapeutic avenues for restoring brain barrier functions.
... Post-stroke neuronal death is a progressive process and continues over days to weeks after the initial insult [99]. Keeping in view the brain-immune interaction, there are multiple options that could be of therapeutic interest. ...
Ischemic stroke is still among the leading causes of mortality and morbidity worldwide. Despite intensive advancements in medical sciences, the clinical options to treat ischemic stroke are limited to thrombectomy and thrombolysis using tissue plasminogen activator within a narrow time window after stroke. Current state of the art knowledge reveals the critical role of local and systemic inflammation after stroke that can be triggered by interactions taking place at the brain and immune system interface. Here, we discuss different cellular and molecular mechanisms through which brain–immune interactions can take place. Moreover, we discuss the evidence how the brain influence immune system through the release of brain derived antigens, damage-associated molecular patterns (DAMPs), cytokines, chemokines, upregulated adhesion molecules, through infiltration, activation and polarization of immune cells in the CNS. Furthermore, the emerging concept of stemness-induced cellular immunity in the context of neurodevelopment and brain disease, focusing on ischemic implications, is discussed. Finally, we discuss current evidence on brain–immune system interaction through the autonomic nervous system after ischemic stroke. All of these mechanisms represent potential pharmacological targets and promising future research directions for clinically relevant discoveries.
... Acute ischemic stroke activates the immune system. Neutrophils migrate to the intraparenchymal perivascular areas within 6-24 hours after cerebral ischemia, producing various cytokines and participating in the early destruction of the bloodbrain barrier (14) . Lymphocytes accumulate in the brain 3-6 days after stroke, considerably later than neutrophils, and are considered to have a regulatory function whereby the induce neuroprotection (15,16) . ...
Objective:
Chronic inflammatory processes involving the vascular wall may induce atherosclerosis. Immune-inflammatory processes proceed throughout all stages of acute stroke. We investigated the association of three immune-inflammatory markers, namely systemic immune-inflammation index (SII), neutrophil-to-lymphocyte ratio (NLR), and neutrophil count (NC), with prehospital delay and clinical features in patients with acute ischemic stroke.
Methods:
We retrospectively enrolled 2543 inpatients admitted within 4 days of symptom onset from May 2010 to February 2020. Patients were stratified into three groups: Group A, comprising 161 patients with tissue plasminogen activator (tPA) treatment; Group B, comprising 415 patients who were eligible for tPA treatment; and Group C, comprising all 2543 patients.
Results:
The levels of all three immune-inflammatory markers had positive linear correlations with onsetto- emergency room time, initial National Institutes of Health Stroke Scale (NIHSS) scores, and discharge modified Rankin Scale scores. In Group B, levels of follow-up, but not initial, immuneinflammatory markers were higher in patients with unfavorable outcomes. Common significant predictors of in-hospital complications and unfavorable outcomes were age > 72 years, female sex, NIHSS > 4, diabetes mellitus, and all three immune-inflammatory markers. When combined with other predictors, NC > 7.2 × 103/mL achieved optimal predictive performance (0.794) for in-hospital complications, and SII > 651, NLR > 2.9, and NC > 7.2 × 103/mL had equal predictive performance up to 0.859 for unfavorable outcomes.
Conclusions:
Immune-inflammatory markers dynamically increased from symptom onset of acute ischemic stroke in patients eligible for thrombolytic therapy. Higher levels of immune-inflammatory markers suggest more in-hospital complications and unfavorable short-term outcomes.
... Ischemic stroke can be characterized by arterial embolic or thrombotic occlusion, resulting in brain in ammation and cell death. Notably, white blood cells delivered via the blood are known to in ltrate the brain to activate in ammation, and then cell death signals are activated by the permeability of the outer mitochondrial membrane [12][13][14]. ...
Background: Stroke is a condition characterized by brain tissue damage owing to a decrease in the brain's oxygen supply due to blocked blood vessels, and 80% of all strokes are classified as cerebral infarction. Notably, the incidence rate tends to increase with increasing age. In this study, we compared the efficacy of white ginseng (WG) and red ginseng (RG) extracts (WGex and RGex, respectively) in an ischemic stroke mouse model and confirmed the underlying mechanisms of action.
Methods: Mice were orally administered WGex or RGex 1 h before performing middle cerebral artery occlusion (MCAO) for 2 h; the size of the infarct area was measured 24 h after MCAO. The neurological deficit score was evaluated, the efficacy of the two drugs was compared, and the mechanism of action was confirmed using methods such as tissue staining and protein quantification.
Results: In the MCAO-induced ischemic stroke mouse model, WGex and RGex showed neuroprotective effects in the cortical region, with RGex demonstrating a generally stronger efficacy than WGex. Furthermore, it was confirmed that ginsenoside Rg1, a representative indicator substance, was not involved in mediating the effects of WGex and RGex.
Conclusion: WGex and RGex inhibited brain injury attributed to ischemia/reperfusion, with RGex revealing a more potent effect. At 1,000 mg/kg body weight, only RGex reduced cerebral infarction and edema, and both anti-inflammatory and anti-apoptotic pathways were involved in mediating these effects.
... Acute ischemic cerebral stroke is considered the leading cause of morbidity and mortality in modern society [1][2][3][4][5]. When ischemic stroke occurs, cerebral inflammation and cell death are induced in the ischemic lesions, and inflammatory signals are activated by harmful stimuli such as arterial occlusion [6][7][8][9]. ...
Background:
The root of Angelica gigas Nakai (Apiaceae) has been traditionally used as an important herbal medicine to treat blood-deficiency-related disorders in Eastern Asian countries, and recently, it has been recognized as a potential candidate for improving cardiovascular diseases.
Methods:
In this study, the neuroprotective effect of a methanol extract of A. gigas root (RAGE) was investigated in a mouse stroke model induced by a 90 min transient middle cerebral artery occlusion (tMCAO). Infarction volumes and morphological changes in brain tissues were measured using TTC, cresyl violet, and H&E staining. The neuroprotective mechanism of RAGE was elucidated through investigation of protein expression levels using western blotting, IHC, and ELISA assays. The plasma concentrations of decursin, a major compound in RAGE, were measured after oral administration of RAGE to SD rats.
Results:
The infarction volumes in brain tissues were significantly reduced and the morphological deteriorations in the brain neuron cells were improved in tMCAO mice when pre-treated with RAGE at 1000 mg/(kg bw·d) for two consecutive days. The neuroprotective mechanism of RAGE was confirmed to attenuate ERK-related MAPK signaling pathways in the ipsilateral hippocampus hemisphere in mice. The concentrations of decursin in rat plasma samples showed peak absorption and elimination in vivo after oral administration of RAGE at 100 mg/rat.
Conclusion:
Mice administered RAGE before the tMCAO operation had less neuronal cell death than those that were not administered RAGE prior to the operation, and this study provides preclinical evidence for use of A. gigas in ischemic stroke.
... Interestingly, baseline or admission NLR or LMR had no independent predictive value for outcome in our cohort presumably because the thrombectomy treatment modified the outcome. Intraparenchymal perivascular neutrophil migration occurs within 6 to 24 h [30,31], and further accumulation of neutrophils in ischaemic and reperfused areas occurs at a higher rate after endovascular recanalisation and correlates with poor neurological outcome and brain damage severity both in humans and rodents [32]. Therefore, dynamic measurement of NLR or LMR may be a stronger predictive tool for outcome compared with single measurements. ...
Background and aim:
Neutrophil-lymphocyte ratio (NLR) and lymphocyte-monocyte ratio (LMR) are associated with clinical outcomes in malignancy, cardiovascular disease and stroke. Here we investigate their association with outcome after acute ischaemic stroke treated by mechanical thrombectomy (MT).
Methods:
Patients were selected using audit data for MT for acute anterior circulation ischaemic stroke at a UK centre from May 2016-July 2017. Clinical and laboratory data including neutrophil, lymphocyte and monocyte count tested before and 24 h after MT were collected. Poor functional outcome was defined as modified Rankin Scale (mRS) of 3-6 at 3 months. Multivariable logistic regression analyses were performed to explore the relationship of NLR and LMR with functional outcome.
Results:
One hundred twenty-one patients (mean age 66.4 ± 16.7, 52% female) were included. Higher NLR (adjusted OR 0.022, 95% CI, 0.009-0.34, p = 0.001) and lower LMR (adjusted OR - 0.093, 95% CI (- 0.175)-(- 0.012), p = 0.025) at 24-h post-MT were significantly associated with poorer functional outcome when controlling for age, baseline NIHSS score, infarct size, presence of good collateral supply, recanalisation and symptomatic intracranial haemorrhage on multivariate logistic regression. Admission NLR or LMR were not significant predictors of mRS at 3 months. The optimal cut-off values of NLR and LMR at 24-h post-MT that best discriminated poor outcome were 5.5 (80% sensitivity and 60% specificity) and 2.0 (80% sensitivity and 50% specificity), respectively on receiver operating characteristic curve analysis.
Conclusion:
NLR and LMR tested at 24 h after ictus or intervention may predict 3-month functional outcome.
... During this complex process, certain receptors will appear in immune cells and also certain mediators will be released [80]. It has been shown that after stroke, certain inflammatory processes will occur in the ischemic region [81]. Various kinds of immune cells recruited to the ischemic region including Microglial cells, neutrophils, macrophages/monocytes and Tcells [80]. ...
Despite many advances in the treatment of stroke, this disease still causes great morbidity and mortality. For this purpose, different kinds of studies have been conducted based on different mechanisms. The research findings highlight the role of remote ischemic preconditioning, microRNAs, neurogenesis, inflammation, and oxidative stress. Nearly a quarter of patients with ischemic stroke will experience a recurrent stroke. It means not just immediate intervention, but also long term intervention is necessary to alleviate stroke patients. Therefore, it is mandatory to predict unwanted events and implement a thoughtful treatment, especially targeting high-risk patients with a high rate of mortality and morbidity. In this review, new advances in animal models have been proposed and overall, it is concluded that stroke patients may greatly benefit from multidisciplinary solutions and more studies are being conducted for timely implementing the best therapy.
... As a consequence, these pathological insults also frequently lead to persistent disability in patients surviving a stroke 2 . Therefore, neuronal protection and regeneration therapies have been urgently required, however, an efficient way to protect neurons from cell death during and after stroke has not been satisfying so far [3][4][5][6] . ...
Extracellular vesicles (EVs) including exosomes can serve as mediators of cell–cell communication under physiological and pathological conditions. However, cargo molecules carried by EVs to exert their functions, as well as mechanisms for their regulated release and intake, have been poorly understood. In this study, we examined the effects of endothelial cells-derived EVs on neurons suffering from oxygen-glucose deprivation (OGD), which mimics neuronal ischemia-reperfusion injury in human diseases. In a human umbilical endothelial cell (HUVEC)–neuron coculture assay, we found that HUVECs reduced apoptosis of neurons under OGD, and this effect was compromised by GW4869, a blocker of exosome release. Purified EVs could be internalized by neurons and alleviate neuronal apoptosis under OGD. A miRNA, miR-1290, was highly enriched in HUVECs-derived EVs and was responsible for EV-mediated neuronal protection under OGD. Interestingly, we found that OGD enhanced intake of EVs by neurons cultured in vitro. We examined the expression of several potential receptors for EV intake and found that caveolin-1 (Cav-1) was upregulated in OGD-treated neurons and mice suffering from middle cerebral artery occlusion (MCAO). Knock-down of Cav-1 in neurons reduced EV intake, and canceled EV-mediated neuronal protection under OGD. HUVEC-derived EVs alleviated MCAO-induced neuronal apoptosis in vivo. These findings suggested that ischemia likely upregulates Cav-1 expression in neurons to increase EV intake, which protects neurons by attenuating apoptosis via miR-1290.
... However, the entry of neutrophils is enhanced by a local blood brain barrier breakdown induced by ischemia [10,22] . Perivascular neutrophil migration into the intraparenchymal area occurs within 6 to 24 hours [22,23] . Lymphocytes are elevated 3-6 days after stroke in the ischaemic brain [24] . ...
... Reperfusion hastens inflammation 70 . This is why we have analyzed the effects of MLC901 against inflammation in different states of brain reperfusion. ...
Inflammation is considered as a major contributor to brain injury following cerebral ischemia. The therapeutic potential of both MLC601/MLC901, which are herbal extract preparations derived from Chinese Medicine, has been reported both in advanced stroke clinical trials and also in animal and cellular models. The aim of this study was to investigate the effects of MLC901 on the different steps of post-ischemic inflammation in focal ischemia in mice. In vivo injury was induced by 60 minutes of middle cerebral artery occlusion (MCAO) followed by reperfusion. MLC901 was administered in post-treatment 90 min after the onset of ischemia and once a day during reperfusion. MLC901 treatment resulted in a reduction in infarct volume, a decrease of Blood Brain Barrier leakage and brain swelling, an improvement in neurological scores and a reduction of mortality rate at 24 hours after MCAO. These beneficial effects of MLC901 were accompanied by an inhibition of astrocytes and microglia/macrophage activation, a drastically decreased neutrophil invasion into the ischemic brain as well as by a negative regulation of pro-inflammatory mediator expression (cytokines, chemokines, matrix metalloproteinases). MLC901 significantly inhibited the expression of Prx6 as well as the transcriptional activity of NFκB and the activation of Toll-like receptor 4 (TLR4) signaling, an important pathway in the immune response in the ischemic brain. MLC901 effects on the neuroinflammation cascade induced by cerebral ischemia probably contribute, in a very significant way, in its potential therapeutic value.
... However, overproduced cytokines will sabotage the host immune responses [44]. It is reported that host cells initiate immune responses by producing various proinflammatory cytokines during the infection of various viruses, including West Nile virus [45], SARS-CoV [46][47][48], and hepatitis (A, B, C) viruses [49]. Therefore, we studied whether GLY treatment affected the levels of proinflammatory cytokine mRNAs during PEDV infection. ...
Porcine epidemic diarrhea (PED), caused by porcine epidemic diarrhea virus (PEDV) infection, leads to significant economic losses in the swine industry worldwide. In our studies, we found that glycyrrhizin, the major component of licorice root extracts, could moderately inhibit PEDV infection in Vero cells, when analyzed by western blot, qRT-PCR and a plaque formation assay. We also revealed that glycyrrhizin inhibited the entry and replication of PEDV. In addition, we demonstrated that glycyrrhizin decreased the mRNA levels of proinflammatory cytokines. Since glycyrrhizin is a competitive inhibitor of high mobility group box-1 (HMGB1), we confirmed that TLR4 and RAGE (£ associated with PEDV pathogenesis during the infection in Vero cells. In summary, our studies provide a molecular basis for developing novel therapeutic methods to control PEDV infection, based on glycyrrhizin and its derivatives.
... Accordingly, the majority of the genes acutely regulated in the blood of stroke patients are expressed in neutrophils and, to a lesser extent, in monocytes [10]. The importance of innate immunity in ischemic stroke is further highlighted by the evidence that monocytes/macrophages and neutrophils are the first cells to infiltrate the injured brain, reaching a peak within 24-72 h after the insult [95,104,105]. Moreover, higher peripheral leukocyte and neutrophil counts, but not Polarization of microglia/macrophages towards the protective M2 or the proinflammatory M1 phenotype after ischemic stroke. ...
The innate immune system plays a pivotal role in ischemic stroke pathobiology, involving soluble and cellular mediators activated locally or recruited from the periphery. Upon injury, subtle modifications of the local environment trigger a rapid activation of microglia that peaks few days after the insult and may persist for several weeks. Initially, the alternatively activated M2 phenotype predominates, whereas, upon priming by ischemic neurons, microglia shift towards the M1 phenotype characterized by reduced phagocytic capacity and release of inflammatory cytokines. Maximally activated microglia can eventually turn into a round amoeboid phenotype morphologically indistinguishable from blood-derived macrophages.
A typical hallmark of cerebral ischaemia is the increased permeability of cerebral microvessels that, together with the upregulation of adhesion molecules on post-capillary venules and the choroid plexus, facilitate brain recruitment of leukocytes. Bone marrow-derived monocytes rapidly extravasate via a chemokine receptor 2 [CCR2]-dependent pathway and, once in the injured tissue, differentiate into non-inflammatory M2 macrophages, mediating neuroprotection and repair of the neurovascular unit. M2 macrophages peak few days after the insult in the core region, whereas the pro-inflammatory M1 phenotype predominates in the peri-infarct areas to gradually increase in number in the core, outnumbering M2 cells over time. The dualistic role exerted by microglia/macrophages suggests that a mere inhibition of their activation/recruitment might not represent a promising strategy to rescue ischemic brain injury. By contrast, as thoroughly reviewed here, a rational modulation of their polarization status, aimed at adjusting the M1/M2 ratio coherently with the spatio-temporal progression of injury, has recently been validated in animal models.
... Mice lacking CyPD display a delayed opening of the PTP when Ca 2+ overload is present and develop smaller cerebral infarcts after prolonged ischemia and reperfusion [46] . Ischemic reperfusion also triggers a sterile inflammatory response [47] . There is emerging evidence that CsA could induce neuroprotective effects via mechanisms different from the inhibition of PTP opening [48] . ...
Background:
Ischemic stroke (IS) and acute myocardial infarction require emergency reperfusion tissue in order to improve functional outcome. Intra-arterial thrombectomy recently showed very encouraging improvement in IS patients' outcome. However, endovascular methods enhancing reperfusion may expose patients to increase in ischemic reperfusion injury. Experimental evidence indicates that brain ischemic reperfusion injury may be attenuated by ischemic pre- and postconditioning. The opening of mitochondrial permeability transition pore plays a critical role in the onset of reperfusion damage. This mechanism can be inhibited by immunosuppressive drugs like cyclosporine A (CsA).
Summary:
In this review, we present existing experimental and clinical data suggesting that conditioning interventions may prevent brain ischemic reperfusion injury and future challenge for neuroprotection by CsA in acute IS.
Key messages:
The concept of conditioning has been recently investigated clinically but to a lesser extent in the realm of IS. Recent experimental and phase II clinical research has suggested potential neuroprotective properties of cyclosporine; however, further larger clinical trials are needed to demonstrate that CsA improves clinical outcome in acute IS patients.
... Tissue injury, such as cerebral ischemia and/or spinal cord damage, results in inflammation, and leukocyte infiltration 27 . The injured tissue produces inflammatory mediators, such as interleukin-6 (IL-6), tumor necrosis factor alpha (TNF-α ), ciliary neurotrophic factor (CNTF), leukemia inhibitory factor (LIF), interferon gamma (IFN-γ ), and interleukin-18 (IL-18) 28,29 .Among the mediators, monocyte chemoattractant protein-1 (MCP-1, CCL2) and stromal-derived factor-1 (SDF-1,CXCL12) regulate the proliferation, migration and neural differentiation of NSCs 9,11,30 . ...
Neural stem cells (NSCs) bear characteristics for proliferation, migration and differentiation into three main neural cell type(s): neurons, astrocytes and/or oligodendrocytes. Formylpeptide receptors (Fprs), belonging to the family of G protein-coupled chemoattractant receptors, have been detected on neurons in the central nervous system (CNS). Here, we report that Fpr1 and Fpr2 are expressed on NSCs as detected with immunohistochemistry, RT-PCR and WB assays. In addition, Fpr1 and Fpr2 promoted NSC migration through F-actin polymerization and skewed NSC differentiation to neurons. Our study demonstrates a unique role of Fpr1 and Fpr2 in NSCs and opens a novel window for cell replacement therapies for brain and spinal cord injury.
... However, there was no proof of their identity. A similar study claiming monocyte infi ltration as early as 1 day used H&E staining and showed cells accumulating in the lumen of blood vessels (Clark et al. 1994 ). The timing of extravasation was not determined and the authors stated that macrophages were abundant in the infarct by 3 days. ...
Ischemic stroke is rarely confined to gray matter, and primary white matter injury occurs in a significant proportion of human strokes. In preclinical investigations searching for stroke therapies, the emphasis is shifting away from primary neurotoxicity to the secondary injury phase, which occurs in a time window that is more amenable to treatment in hospital. This phase is characterized by a prominent inflammatory response in the brain that can last for hours to days. This chapter focuses on the intersection of inflammation and white matter injury in experimental models of ischemic stroke. We first describe the main rodent models and methods used to monitor white matter damage, and discuss we prefer some approaches. Next, we describe the main immune cells involved in animal models of ischemia, how to monitor them, and present key findings. We then summarize the limited literature addressing the intersection of ischemic stroke, white matter injury, and inflammation in adult and neonatal rodent ischemia models, and finally comment on specific needs for further research.
... [115][116][117][118][119][120][121][122][123] In embolic stroke, these ischemic changes can be exacerbated if recanalization of the obstructed vessel and restoration of the blood flow are affected, which may then lead to reperfusion injury (Ginsberg, Clark et al., Aronowski et al.). [124][125][126] However, despite strong evidence confirming the above alterations of the microcirculation and CBF in focal models of ischemia, there has been little evidence ascertaining real-time observation of vasoreactive changes in stroke. Thus, in a rat model of global ischemia and reperfusion, Takahashi et al. [6] used correlative laser Doppler flowmetry and SEM to demonstrate enhanced VSMC contractility of brain parenchymal arterioles and precapillary arterioles, which temporally coincided with the phase of hypoperfusion. ...
The cerebral microvessels are parenchymal branches of the brain′s penetrating vessels that include small diameter arterioles and capillaries and through which the cerebral microcirculation delivers vital metabolites to the brain. In contrast to conductance or meningeal vessels, vasomotor tone in cerebral microvessels is not dependent on the action of the sympathetic nervous system but rather on a combination of vasoreactive agents such as angiotensin, vasopressin, and purines from both brain endogenous and exogenous sources. The microvascular wall consists mainly of endothelial cells (ECs), smooth muscle cells, and pericytes (PCs) as well as the sieve-like basal lamina (BL), which together with perivascular astroglia (AS) interact dynamically to maintain the integrity and permeability of the blood-brain barrier (BBB). To ensure constant delivery of oxygen and glucose, the flow of blood through microcirculation is under autoregulatory control, both systemically and locally at the level of the microvascular wall. At the microvascular wall, endothelin (ET) and nitric oxide (NO) as well as circulating agents provide local vasopressor and vasodepressor effects that are crucial to maintain a normal vasomotor tone. Following trauma or stroke, three major pathologies occur: 1) alterations in structural integrity of microvessels and brain parenchymal cells, 2) acute edema formation, and 3) sustained hypoperfusion from vasospasm. Other pathologies that may contribute to a defective microcirculation include the formation of microthrombi and hemorrhaging, which can exacerbate the immune response. A defective microcirculation due to the loss of autoregulatory control of microvessels may contribute to the brain′s shift to anaerobic metabolism and to the formation of oxygen-free radicals, considered to be a major source of injury to nerve cells and the BBB. The diverse pathophysiologies ensuing from these events lead to nerve cell loss and poor neurological outcome. Because of these diverse pathophysiologies, monotherapeutic interventions to improve the microcirculation after trauma and stroke have had limited success both at the bench and the clinic. Recent polytherapeutic interventions aiming at improving the microcirculation as well as cell viability and neurological outcome after trauma and stroke are discussed.
... Cerebral ischemia-reperfusion injury often occurs following the restoration of blood flow in cerebral stroke patients, and causes neurological deficits (1,2). Despite the fact that great progress has been made over the years in studies of cerebral ischemia-reperfusion injury, numerous unsolved clinical problems remain. ...
The aim of the present study was to investigate whether erythropoietin (EPO) preconditioning affects the expression of glutamate transporter 1 (GLT-1) and glutamate aspartate transporter (GLAST) and protects against rat cerebral ischemia-reperfusion injury. A total of 140 Sprague Dawley rats were randomly assigned to one of the following four groups: Sham, EPO-sham, middle cerebral artery occlusion (MCAO) and EPO-MCAO. Neurological function scores were obtained 24, 36 and 72 h after reperfusion. Seventy-two hours after the induction of cerebral ischemia-reperfusion, the number of apoptotic neural cells and the cerebral infarct volume of each group were measured. The mRNA levels of GLT-1 and GLAST were determined using reverse transcription-quantitative polymerase chain reaction (RT-qPCR) analysis, while the GLT-1 and GLAST protein levels were assessed using western blotting. The cerebral infarct volume was significantly increased in the MCAO group compared with that in the sham group (P<0.01); however, the infarct volume of the EPO-MCAO group was significantly lower than that of the MCAO group (P<0.01). In addition, the number of apoptotic cells found in the MCAO group was higher than that in the sham group (P<0.01), but the number of apoptotic cells in the EPO-MCAO group was significantly lower than that in the MCAO group (P<0.01). The GLT-1 and GLAST mRNA and protein levels were significantly decreased 72 h after the cerebral ischemia-reperfusion (P<0.01) compared with those in the sham group, whereas the same levels were increased significantly in the EPO-MCAO group relative to those in the MCAO group (P<0.01). In conclusion, EPO preconditioning protected against cerebral ischemia-reperfusion injury and upregulated the GLT-1 and GLAST expression.
... In human stroke, microglia activation as a surrogate marker of neuroinflammation starts not earlier than 3 days after onset of the infarct, reaching its maximum within one week (108)(109)(110). In experimental ischemia involving reperfusion, both microglia activation as well as invasion of blood-borne cells is typically accelerated, starting as early as 22 hours after ischemia (100,(115)(116)(117)(118). Moreover, the up-regulation of cytokines released by glia cells occurs quite early in commonly used stroke models (119)(120)(121)(122)(123)(124). ...
The main challenge of stroke research is to translate promising experimental findings from the bench to the bedside. Many suggestions have been made how to achieve this goal, identifying the need for appropriate experimental animal models as one key issue. We here discuss the macrosphere model of focal cerebral ischemia in the rat, which closely resembles the pathophysiology of human stroke both in its acute and chronic phase. Key pathophysiological processes such as brain edema, cortical spreading depolarizations (CSD), neuroinflammation, and stem cell-mediated regeneration are observed in this stroke model, following characteristic temporo-spatial patterns. Non-invasive in vivo imaging allows studying the macrosphere model from the very onset of ischemia up to late remodeling processes in an intraindividual and longitudinal fashion. Such a design of pre-clinical stroke studies provides the basis for a successful translation into the clinic.
... Interestingly, the majority of the genes acutely regulated in the blood of stroke patients are expressed in neutrophils and, to a lesser extent, in macrophages (Tang et al., 2006). Accordingly, these are the first cells to infiltrate the ischemic brain, reaching a peak within 24-72 h after the insult (Clark et al., 1994;Garcia et al., 1994;Gelderblom et al., 2009). In patients, higher peripheral leukocyte and neutrophil counts, but not lymphocyte counts, are associated with larger infarct volumes (Buck et al., 2008), and brain accumulation of neutrophils correlates with poor neurological outcome and brain damage severity both in humans (Akopov et al., 1996) and in rodents (Matsuo et al., 1994a,b;Connolly et al., 1996;Atochin et al., 2000). ...
The innate immune system plays a dualistic role in the evolution of ischemic brain damage and has also been implicated in ischemic tolerance produced by different conditioning stimuli. Early after ischemia, perivascular astrocytes release cytokines and activate metalloproteases (MMPs) that contribute to blood–brain barrier (BBB) disruption and vasogenic oedema; whereas at later stages, they provide extracellular glutamate uptake, BBB regeneration and neurotrophic factors release. Similarly, early activation of microglia contributes to ischemic brain injury via the production of inflammatory cytokines, including tumor necrosis factor (TNF) and interleukin (IL)-1, reactive oxygen and nitrogen species and proteases. Nevertheless, microglia also contributes to the resolution of inflammation, by releasing IL-10 and tumor growth factor (TGF)-β, and to the late reparative processes by phagocytic activity and growth factors production. Indeed, after ischemia, microglia/macrophages differentiate toward several phenotypes: the M1 pro-inflammatory phenotype is classically activated via toll-like receptors or interferon-γ, whereas M2 phenotypes are alternatively activated by regulatory mediators, such as ILs 4, 10, 13, or TGF-β. Thus, immune cells exert a dualistic role on the evolution of ischemic brain damage, since the classic phenotypes promote injury, whereas alternatively activated M2 macrophages or N2 neutrophils prompt tissue remodeling and repair. Moreover, a subdued activation of the immune system has been involved in ischemic tolerance, since different preconditioning stimuli act via modulation of inflammatory mediators, including toll-like receptors and cytokine signaling pathways. This further underscores that the immuno-modulatory approach for the treatment of ischemic stroke should be aimed at blocking the detrimental effects, while promoting the beneficial responses of the immune reaction.
... 5,10 It has been suggested that reperfusion is associated with endothelial cell damage, an increase in edema, and an increased risk of ICH. 5,[11][12][13] It is likely that free radicals are mediators of a variety of injuries after an ischemic stroke. 14 -16 Much experimental stroke research has focused on developing neuroprotective agents to reduce secondary damage after the onset of ischemia. ...
It has been proposed that spin trap agents such as N:-t-butyl-phenylnitrone (PBN) may be useful as neuroprotective agents in the treatment of ischemia and stroke. However, to date, there is little information concerning the effectiveness of spin trap agents when administered in combination with the only Food and Drug Administration-approved pharmacological agent for the treatment of stroke, the thrombolytic tissue plasminogen activator (tPA). Thus, we determined the effects of PBN when administered before tPA on hemorrhage and infarct rate and volume. We also compared the effects of PBN with those of 2,2,6, 6-tetramethylpiperidine-N:-oxyl (TEMPO), another spin trap agent that has a different chemical structure and trapping profile, on the incidence of infarcts and hemorrhage.
One hundred sixty-five male New Zealand White rabbits were embolized by injecting a blood clot into the middle cerebral artery via a catheter. Five minutes after embolization, PBN or TEMPO (100 mg/kg) was infused intravenously. Control rabbits received saline, the vehicle required to solubilize the spin traps. In tPA studies, rabbits were given intravenous tPA starting 60 minutes after embolization. Postmortem analysis included assessment of hemorrhage, infarct size and location, and clot lysis.
In the control group, the hemorrhage rate after a thromboembolic stroke was 24%. The amount of hemorrhage was significantly increased to 77% if the thrombolytic tPA was administered. The rabbits treated with PBN in the absence of tPA had a 91% incidence of hemorrhage compared with 33% for the TEMPO-treated group. In the combination drug-treated groups, the PBN/tPA group had a 44% incidence of hemorrhage, and the TEMPO/tPA group had a 42% incidence of hemorrhage. tPA, PBN/tPA, and TEMPO/tPA were similarly effective at lysing clots (49%, 44%, and 33%, respectively) compared with the 5% rate of lysis in the control group. There was no significant effect of drug combinations on the rate or volume of infarcts.
This study suggests that certain spin trap agents may have deleterious effects when administered after an embolic stroke. However, spin trap agents such as PBN or TEMPO, when administered in combination with tPA, may improve the safety of tPA by reducing the incidence of tPA-induced hemorrhage. Overall, the therapeutic benefit of spin trap agents for the treatment of ischemic stroke requires additional scrutiny before they can be considered "safe" therapeutics.
... These effects protect ischaemic tissues from reperfusion damage, 1 which is originally caused by increasing accumulation of inflammatory cells, overproduction of oxygen free radicals and microvascular dysfunction. 25,26 The use of chlorpromazine and promethazine shortened the duration to reach the target hypothermia temperature (35u uC) by 10 minutes. This supports the notion that phenothiazines inhibit thermoregulatory processes and promote cooling, 19 thereby reinforcing the hypothermia treatment, which may enhance the neuroprotective effects. ...
Hypothermia is a well-recognised and effective neuroprotectant because of its depressive effect on metabolism. However, its application in focal ischaemic stroke is limited by delayed onset, prolonged duration, the need for extensive medical and nursing efforts and significant complications. This study combined mild hypothermia with phenothiazine drugs to enhance its neuroprotective effects, thus potentially avoiding side effects.
Fifty-four male Sprague-Dawley rats were induced with a 2-hour right middle cerebral artery (MCA) occlusion using an intraluminal filament. Five groups were tested: the stroke group without treatment (anal temperature 37.8-38.3°C), the mild hypothermia group (anal temperature 35.0°C), the drugs group (1 mg/kg chlorpromazine and 1 mg/kg promethazine, anal temperature at 37.8-38.3°C), combination therapy with the mild hypothermia and drugs group and the normal control group (anal temperature 37.8-38.3°C). The treatments immediately followed reperfusion. The extent of brain injury was evaluated by infarct volume and behaviour performance.
The combination treatment of mild hypothermia with phenothiazine drugs demonstrated salient and significant (P < 0.001) reductions in infarct volume (30.0 ± 15.14%) when compared to the stroke group (52.77 ± 8.99%). A better recovery of long-term motor performance was also observed for those receiving the combination therapy. However, when administered independently, neither the mild hypothermia therapy (53.8 ± 10.3%) nor phenothiazine regimen (55.7 ± 9.00%) had significant therapeutic effects on infarct volume (P = 0.85 and 0.61, respectively).
This study provides a novel and promising therapeutic strategy in the management of acute stroke.
... Moreover, the addition of follow-up visits would allow for the time-course evaluation of the KYN/TRP ratio and changes in inflammatory states of each participant. Even so, current animal models suggest that IDO is most active following cerebral ischemia 412,413 and may be necessary to induce sickness behaviours 208 , suggesting that basal cytokine levels are rarely sufficient enough to alter mood states via IDO induction. ...
... The first demonstration of early granulocyte infiltration into ischemic brain tissue occurred in studies showing accumulation of labeled granulocytes in ischemic brain tissue in the first few hours following focal cerebral ischemia [123]. Several studies have subsequently confirmed the findings of this initial study [124][125][126]. In addition, a phenomenon of microvasculature occlusion following cerebral ischemia referred to as "No-reflow", first described in a model of rabbit experimental ischemia [127], has been subsequently attributed to polymorphonuclear cell occlusion of micro vessels in a model of non-human primate experimental stroke [128]. ...
It is currently well established that the immune system is activated in response to transient or focal cerebral ischemia. This acute immune activation occurs in response to damage, and injury, to components of the neurovascular unit and is mediated by the innate and adaptive arms of the immune response. The initial immune activation is rapid, occurs via the innate immune response and leads to inflammation. The inflammatory mediators produced during the innate immune response in turn lead to recruitment of inflammatory cells and the production of more inflammatory mediators that result in activation of the adaptive immune response. Under ideal conditions, this inflammation gives way to tissue repair and attempts at regeneration. However, for reasons that are just being understood, immunosuppression occurs following acute stroke leading to post-stroke immunodepression. This review focuses on the current state of knowledge regarding innate and adaptive immune activation in response to focal cerebral ischemia as well as the immunodepression that can occur following stroke. A better understanding of the intricate and complex events that take place following immune response activation, to acute cerebral ischemia, is imperative for the development of effective novel immunomodulatory therapies for the treatment of acute stroke.
Background:
Interest is growing in the role played by intestinal flora in the pathogeneses of diseases and in the possibility of treating disease by altering intestinal flora compositions. Recent studies have focused on the relationship between the intestinal microbiome and brain function as proposed by the brain-gut axis hypothesis.
Objectives:
To investigate the relation between ischemic stroke and plasma equol monosulfate levels (a soy isoflavone metabolite) in a middle cerebral artery occlusion (MCAO) mouse model.
Methods:
Mice (C57BL/6) were subjected to MCAO for various times (30 min to 24 h), and degrees of cerebral damage were assessed using total infarction volumes, brain edema severities and neurological deficit scores. Hematoxylin and eosin and cresyl violet staining were used to observe morphological changes in ischemic brains. Levels of equol monosulfate in plasma and the relationships between these and degree of brain injury were investigated.
Results:
Infarction volumes, brain edema severity and neurological deficit scores were significantly correlated with ischemic time, and morphological deteriorations of brain neuronal cells also increased with ischemic duration. Equol monosulfate contents were ischemic-time dependently lower in MCAO treated animals than in sham-operated controls.
Conclusion:
Ischemic stroke may time-dependently reduce plasma levels of equol monosulfate by lowering the metabolic rate of equol in MCAO-induced mice. This study provides indirect support of the brain-gut axis hypothesis.
Background: The root of Angelica gigas Nakai (Apiaceae) has been traditionally used as an important herbal medicine to treat blood-deficiency-related disorders in Eastern Asian countries, and recently, it has been recognized as a potential candidate for improving cardiovascular diseases.
Methods: In this study, the neuroprotective effect of a methanol extract of A. gigas root (RAGE) was investigated in a mouse stroke model induced by a 90 min transient middle cerebral artery occlusion (tMCAO). Infarction volumes and morphological changes in brain tissues were measured using TTC, cresyl violet, and H&E staining. The neuroprotective mechanism of RAGE was elucidated through investigation of protein expression levels using western blotting, IHC, and ELISA assays. The plasma concentrations of decursin, a major compound in RAGE, were measured after oral administration of RAGE to SD rats.
Results: The infarction volumes in brain tissues were significantly reduced and the morphological deteriorations in the brain neuron cells were improved in tMCAO mice when pre-treated with RAGE at 1,000 mg/(kg bw·d) for two consecutive days. The neuroprotective mechanism of RAGE was confirmed to attenuate ERK-related MAPK signaling pathways in the ipsilateral hippocampus hemisphere in mice. The concentrations of decursin in rat plasma samples showed peak absorption and elimination in vivo after oral administration of RAGE at 100 mg/rat.
Conclusion: Mice administered RAGE before the tMCAO operation had less neuronal cell death than those that were not administered RAGE prior to the operation, and this study provides preclinical evidence for use of A. gigas in ischemic stroke.
Despite considerable efforts in research and clinical studies, stroke is still one of the leading causes of death and disability worldwide. Originally, stroke was considered a vascular thrombotic disease without significant immune involvement. However, over the last few decades it has become increasingly obvious that the immune responses can significantly contribute to both tissue injury and protection following stroke. Recently, much research has been focused on the immune system's role in stroke pathology and trying to elucidate the mechanism used by immune cells in tissue injury and protection. Since the discovery of tissue plasminogen activator therapy in 1996, there have been no new treatments for stroke. For this reason, research into understanding how the immune system contributes to stroke pathology may lead to better therapies or enhance the efficacy of current treatments. Here, we discuss the contrasting roles of immune cells to stroke pathology while emphasizing myeloid cells and T cells. We propose that focusing future research on balancing the beneficial-versus-detrimental roles of immunity may lead to the discovery of better and novel stroke therapies.
The contribution of leukocytes to the pathogenesis of ischemic stroke has been extensively studied and thoroughly documented. In this chapter, different aspects of leukocyte involvement in the lesion formation caused by ischemic stroke are highlighted, including the inflammatory agents that mediate leukocyte recruitment to the site of injury, the primary leukocyte populations that contribute to tissue damage, and the adhesion receptors that control leukocyte–endothelial cell interactions in post-ischemic brain. Agents that interfere with leukocyte recruitment in the brain are also addressed as potential therapeutic interventions for ischemic stroke.
Biomarkers provide critical mechanistic insights to key biologic processes that occur during cerebral ischemia which, when carefully applied, can improve clinical decision-making in acute stroke management. The translation of a blood-based biomarker in ischemic stroke to clinical practice is challenging, in part, due to the complexity of ischemic stroke pathogenesis and the presence of a blood-brain barrier that restricts the release of brain-specific markers into the circulation. The pathologic and clinical aspects of ischemic stroke are described in this review, where a non-exhaustive list of biomarkers that interrogate different aspects of ischemic stroke such as oxidative damage, inflammation, thrombus formation, cardiac function and brain injury are described. The potential roles of these biomarkers are further examined under different clinical scenarios aimed at (1) averting the risk of hemorrhagic transformation, (2) identifying individuals at risk of early neurologic deterioration and malignant infarction, (3) aiding in the diagnosis of ischemic stroke and its differentiation from other stroke mimics, (4) guiding the search for stroke etiology, and (5) assessing stroke risk within the community. Researchers should explore the roles of stroke biomarkers to enhance clinical decision-making that is presently largely based on intuition and subjective reasoning.
Three general lines of evidence have been put forth to support the role of neutrophil recruitment in ischemic cell death. These include (1) the presence of neutrophils within ischemic tissue at the approximate time that substantial cell death occurs, (2) the reduction of ischemia-induced cell loss following neutropenia, and (3) the observation that treatments which prevent neutrophil trafficking can be neuroprotective. Despite the numerous studies that have been conducted, this hypothesis remains controversial, for an objective assessment of the literature reveals great uncertainty for a pathogenic role of neutrophils in the brain damage associated with ischemia. The present chapter briefly summarizes and critically discusses the available data addressing this hypothesis.
When blood flow is interrupted to regions of the brain, even for very limited periods of time, a number of pathological mechanisms are set in motion which elicit adverse sequelae. Neurons are among the cells most vulnerable to relatively brief periods of ischemia, due to their exquisite and constant need for oxygen and glucose. The causes of neuronal death are multiple, but regardless of cause, once a neuron dies, it is irrevocably lost from the substance of the brain. The focus of this chapter is on inflammatory reactions triggered in neurons or adjacent vascular cells, which contribute to the neuronal demise and tissue injury in stroke.
Experimental evidence continues to support the involvement of inflammatory/immunologic factors in the pathogenesis of ischemic brain injury. Ischemic-induced perforations in the blood-brain barrier (BBB) permit entry of humoral inflammatory mediators such as neutrophils and macrophages (Clark et al., 1994; Barone et al., 1992), whereas inherent CNS-resident microglial cells are activated following ischemic injury (Kim et al., 1995). These cells then become immunologically reactive, participating in a rapid but complex inflammatory cascade involving the release of neuroactive inflammatory mediators. These factors exert both autocrine (stimulation of further monocyte proliferation) and paracrine (effects on neuronal and endothelial cells) actions and the resulting signalling cascades have important ramifications for neurons subjected to cerebral ischemia. Soluble growth factors and cytokines are two examples of such pleiotropic signaling moieties released during inflammation that can have diverse effects on neurons under pathophysiological conditions. Both tumor necrosis factor α (TNFa) and transforming growth factor β(TGFβ) can protect neurons from metabolic, excitotoxic, and oxidative injury―modes of injury which are especially pertinent to the manifestations of ischemia. The neuroprotective mechanisms of these factors are beginning to be resolved, and involve distinct and sometimes overlapping cascades of second messenger systems leading to acute effects and long-term changes in gene expression. The further characterization of these pathways will highlight potential target sites for therapeutic pharmacological intervention, not only in stroke, but also in long term, cumulative neurodegenerative disorders, such as Alzheimer’s disease, Huntington’s disease, and amyotrophic lateral sclerosis (ALS).
The possibility of reopening an occluded cerebral artery by means of thrombolysis has renewed interest in a number of the several mechanisms that are active during acute cerebral ischemia. Over recent years, it has become apparent that leukocytes play a central role not only during the healing stage of brain infarction but also during the early phases of cerebral ischemia, when it is postulated that these cells produce harmful effects, particularly in the presence of reperfusion. This review is based on the critical analysis of more than 150 publications dealing with the role of leukocytes and some inflammatory mediators (cytokines, chemokines, and adhesion molecules) in cerebral ischemia. Animal studies indicate that leukocyte involvement is promoted by a variety of inflammatory molecules produced immediately after the onset of cerebral ischemia. Considerable experimental evidence suggests that these mediators play a key role in the progression from ischemia to irreversible injury (ie, cellular death and necrosis). However, the precise role of each molecule alone remains to be further elucidated as well as in relation to the complex network existing among different mediators. Progress in our understanding of the inflammatory mechanisms operating in cerebral ischemia has enabled the testing of new compounds with promising results in animals; in contrast, one recent controlled trial of an anti-leukocyte molecule in acute stroke patients showed negative results. This discrepancy may derive in part from our incomplete understanding of the complexity of the inflammatory mechanisms involved in cerebral ischemia. Our analysis suggests that until sufficient knowledge of the underlying disease mechanisms is acquired, more care should be taken when testing new and potentially efficacious drugs in stroke patients.
Numerous studies have consistently shown that agonist stimulation of adenosine A(1) receptors results in a significant reduction of morbidity and mortality associated with global and focal brain ischemia in animals. Based on these observations, several authors have suggested utilization of adenosine A(1) receptors as targets for the development of clinically viable drugs against ischemic brain disorders. Recent advent of adenosine A(1) receptor agonists characterized by lowered cardiovascular effects added additional strength to this argument. On the other hand, although cardioprotective, adenosine A(3) receptor agonists proved severely cerebrodestructive when administered prior to global ischemia in gerbils. Moreover, stimulation of adenosine A(3) receptors appears to reduce the efficacy of some of the neuroprotective actions mediated by adenosine A(3) receptors. The review discusses the possible role of adenosine receptor subtypes (A(1), A(2), and A(3)) in the context of their involvement in the pathology of cerebral ischemia, and analyzes putative strategies for the development of clinically useful strategies based on adenosine and its receptors. It also stresses the need for further experimental studies before definitive conclusions on the usefulness of the adenosine concept in the treatment of brain ischemia can be made.
Recent evidence indicates that thrombolysis may be an effective therapy for the treatment of acute ischemic stroke. However, the reperfusion of ischemic brain comes with a price. In clinical trials, patients treated with thrombolytic therapy have shown a 6% rate of intracerebral hemorrhage, which was balanced against a 30% improvement in functional outcome over controls. Destruction of the microvasculature and extension of the infarct area occur after cerebral reperfusion. We have reviewed the existing data indicating that an inflammatory response occurring after the reestablishment of circulation has a causative role in this reperfusion injury. The recruitment of neutrophils to the area of ischemia, the first step to inflammation, involves the coordinated appearance of multiple proteins. Intercellular adhesion molecule-1 and integrins are adhesion molecules that are up-regulated in endothelial cells and leukocytes. Tumor necrosis factor-alpha, interleukin-1, and platelet-activating factor also participate in leukocyte accumulation and subsequent activation. Therapies that interfere with the functions of these factors have shown promise in reducing reperfusion injury and infarct extension in the experimental setting. They may prove to be useful adjuncts to thrombolytic therapy in the treatment of acute ischemic stroke.
In a recent editorial, Dr. Schmid-Schönbein (1) succinctly summarized the case for inflammation in the pathogenesis of cerebral ischemia;
There is increasing evidence to suggest that reperfusion following an ischemic episode leads to an oxidative burst that serves to induce the expression of a number of proinflammatory genes, such as cytokines or leukocyte adhesion molecules. This in turn leads to an accumulation of leukocytes during reperfusion and the initiation of a proinflammatory event, with elevation of endothelial permeability, enhancement of oxygen free radical production, migration of leukocytes into the parenchyma, and expansion of tissue necrosis.
Old age is associated with an enhanced susceptibility to stroke and poor recovery from brain injury, but the cellular processes underlying these phenomena are not well understood. Potential mechanism underlying functional recovery after brain ischemia in aged subjects include neuroinflammation, changes in brain plasticity-promoting factors, unregulated expression of neurotoxic factors, or differences in the generation of scar tissue that impedes the formation of new axons and blood vessels in the infarcted region. Studies suggest that behaviorally, aged rats were more severely impaired by ischemia than were young rats and showed diminished functional recovery. Both in old and young rats, the early intense proliferative activity following stroke leads to a precipitous formation of growthinhibiting scar tissue, a phenomenon amplified by the persistent expression of neurotoxic factors. Recent evidence shows that the human brain can respond to stroke with increased progenitor proliferation in aged patients, opening the possibilities of utilizing this intrinsic attempt for neuroregeneration of the human brain as a potential therapy for ischemic stroke.
Neuroprotective treatments for acute ischemic stroke are targeted at the large array of cellular biochemical and metabolic disturbances that occur after focal brain ischemia to prevent the evolution of injury toward irreversibility. Enhanced comprehension about the pathophysiology of focal brain ischemia has expanded the number of neuroprotective modalities under development and identification of the most likely target for these therapies. Many of the neuroprotective interventions are targeted at reducing calcium influx into ischemic cells and the downstream consequences of excessive intracellular calcium. Other neuroprotective strategies include: free radical scavengers, hyperpolarization of resting transmembrane potentials, and inhibition of the inflammatory response and growth factors. Some interventions potentially may enhance recovery and have neuroprotective effects (i.e., basic fibroblast growth factor [bFGF] and citicoline). Despite the lack of proven clinical efficacy with any neuroprotective intervention, the future will hopefully yield convincing evidence that neuroprotection can be effective and then be ultimately combined with thrombolysis to maximize improvement after ischemic stroke.
Thrombolysis with alteplase (recombinant tissue plasminogen activator; rtPA) has proven to be beneficial for acute stroke management, despite the narrow window of opportunity for treatment and the increased risk of haemorrhage. Because of the latter, recent studies have attempted to identify compounds that may be given concomitantly with alteplase to reduce the haemorrhage rate Matrix metalloproteinase (MMP) inhibitors have been proposed as potential combination therapy candidates because they prevent MMP-induced production of the cytokine tumour necrosis factor-α (TNFα), as well as membrane and vessel remodelling following ischaemia. Spin trap agents also have been put forward due to their free radical scavenging capabilities. In the rabbit large clot embolism model, alteplase effectively lysed blood clots, whether or not other drugs were used in combination. However, haemorrhage rate also was increased compared with that in control animals. The alteplase-induced haemorrhage rate was reduced significantly by administration of the MMP inhibitor batimastat (BB-94) or the spin trap agent α-phenyl-N-t-butylnitrone (PBN). Other rodent studies have also demonstrated that PBN is effective in decreasing the haemorrhage rate following alteplase administration. Overall, preclinical studies indicate that MMP inhibition or free radical scavenging in combination with alteplase may circumvent the high risk of haemorrhaging with alteplase.
Background Ischemic stroke is the most frequent cause of persistent neurological disability in Western societies. New treatment strategies are required and effective in vivo models are crucial to their development.
The current study establishes a novel in vivo rat model of focal striatal ischemia using the vasoconstrictive agent N5-(1-iminoethyl)-L-ornithine (L-NIO). Adult male Sprague Dawley rats received a unilateral intrastriatal infusion of L-NIO in combination with jugular vein occlusion.
L-NIO infusion was associated with zero mortality, low surgical complexity and a reproducible infarct, providing advantages over established models of focal ischemia. The mean infarct volume of 8.5±5.3% of the volume of the contralateral striatum resulted in blood-brain barrier dysfunction, neuronal hypoxia and ongoing neurodegeneration. Further characteristics of ischemic stroke were exhibited, including robust microglia/macrophage and astroglial responses lasting at least 35 days post-ischemia, in addition to chronic motor function impairment.
When compared to other models such as the MCAo models, the consistency in regions affected, high success rate, zero mortality, reduced surgical complexity and minimal welfare requirements of the L-NIO model make it ideal for initial high-throughput investigations into preclinical efficacy and proof of principle studies of acute ischemic stroke interventions.
We propose that the L-NIO rat model of focal striatal ischemia does not replace the use of other ischemic stroke models. Rather it provides a new, complementary tool for initial preclinical investigations into the treatment of ischemic stroke.
Copyright © 2015. Published by Elsevier B.V.
The cytokine tumor necrosis factor (TNF-alpha) is a pleotrophic polypeptide that plays a significant role in brain immune and inflammatory activities. TNF-alpha is produced in the brain in response to various pathological processes such as infectious agents [e.g., human immunodeficiency virus (HIV) and malaria], ischemia, and trauma. TNF-alpha mRNA is rapidly produced in response to brain ischemia within 1 h, reaches a peak at 6-12 h post ischemia, and subsides 1-2 days later. TNF-alpha mRNA expression corresponds in a temporal fashion to other cytokines such as interleukin (IL)-6, cytokine-induced neutrophil chemoattractant (KC), and IL-1 and precedes the infiltration of inflammatory cells into the injured zone. TNF-alpha is present early in neuronal cells in and around the ischemic tissue (penumbra), yet at later time points, the peptide is found in macrophages in the infarcted tissue. TNF-alpha has been demonstrated to cause expression of proadhesive molecules on the endothelium, which results in leukocyte accumulation, adherence, and migration from capillaries into the brain. Furthermore, TNF-alpha activates glial cells, thereby regulating tissue remodeling, gliosis, and scar formation. Thus, evidence is emerging in support of a role for TNF-alpha in injury induced by infectious, immune, toxic, traumatic, and ischemic stimuli. TNF-alpha promotes inflammation by stimulation of capillary endothelial cell proinflammatory responses and thereby provides leukocyte adhesion and infiltration into the ischemic brain. The evidence generated so far suggests that agents that suppress TNF-alpha's production or actions will reduce leukocyte infiltration into ischemic brain regions and thereby diminish the extent of tissue loss.
Reperfusion of ischemic tissues is associated with an enhanced production of inflammatory mediators, increased rolling, adherence, and emigration of leukocytes in postcapillary venules, and vascular protein leakage. There is a growing body of evidence that the leukocyte--endothelial cell interactions are largely responsible for the microvascular dysfunction induced by ischemia--reperfusion. Oxygen radicals, produced by xanthine oxidase and other enzymes, appear to play an important role in initiating and amplifying the inflammatory response elicited by ischemia--reperfusion, while neutrophilic proteases contribute significantly to the injury response. The magnitude of the inflammatory responses observed during ischemia and reperfusion is also influenced by adhesion forces generated by specific glycoproteins expressed on the surface of granulocytes and microvascular endothelium, as well as shear forces that are generated by the movement of blood within the microcirculation. Manipulation of free-radical production, leukocyte--endothelial cell adhesion, and (or) intravascular shear forces provides an effective means for attenuating the deleterious influences of ischemia--reperfusion on the microvasculature.
Forty-seven patients presenting with symptoms of large middle cerebral artery territory infarcts were studied prospectively using serial CT brain scanning to assess infarct extent and swelling, and transcranial Doppler ultrasound to assess middle cerebral artery patency. The single most important determinant of cerebral infarct swelling was the extent of the infarct: the larger the infarct, the greater the amount of swelling (Spearman Rank correlation coefficient 0.74, p < 0.0001). In addition, if the blood velocity in the symptomatic middle cerebral artery did not increase in the first 5 days after symptom onset (implying no early reperfusion) the odds of severe infarct swelling were increased 7.6-fold (95% confidence interval 1.2- to 46.4-fold, 2p = 0.03), and the odds of a poor clinical outcome (dead or dependent in activities of daily living at 3 months) were increased 10-fold (95% confidence interval 2.7- to 41.6-fold, 2p = 0.0007). The only patients who recovered sufficiently to be able to return home by 3 months after stroke were amongst those whose symptomatic middle cerebral artery blood velocity increased (either spontaneously or associated with fibrinolytic or antithrombotic therapy) in the first 5 days after stroke. We conclude that early reperfusion is not associated with a worsening of acute cerebral infarct swelling, and may lead to a better clinical outcome.Copyright © 1993 S. Karger AG, Basel
We investigated the contribution of neutrophils to brain edema formation using a transient focal ischemia model in rats. Rats were given antineutrophil monoclonal antibody (RP3) intraperitoneally to deplete circulating neutrophils. In RP3-treated rats, ischemic brain edema formation 1 day after reperfusion was significantly decreased compared to that of saline-treated control rats. We speculate that chemical mediators released by infiltrating neutrophils alter vascular permeability and play an important role in post-ischemic brain edema formation.
A study of the supply to the feline brain by the carotid-middle cerebral arteries was conducted using in vivo transcardiac injection with a mixture of micropaque and carbon black. Modifications in the filling pattern of the arterial vessels were visualized following short-term occlusion of a middle cerebral artery. A modified surgical method to induce occlusion of the M-l segment of the middle cerebral artery is also described.
Neutrophils are critically involved with ischemia and reperfusion injury in many tissues but have not been studied under conditions of reperfusion after focal cerebral ischemia. The present studies were conducted to confirm our previous observations quantifying neutrophils in rat permanent focal stroke using a myeloperoxidase activity assay and to extend them to transient ischemia with reperfusion. In addition, leukotriene B4 receptor binding in ischemic tissue was evaluated as a potential marker for inflammatory cell infiltration.
Histological, enzymatic, and receptor binding techniques were used to evaluate neutrophil infiltration and receptor binding in infarcted cortical tissue 24 hours after permanent middle cerebral artery occlusion (n = 25) or temporary occlusion for 80 (n = 12) or 160 (n = 22) minutes followed by reperfusion for 24 hours in spontaneously hypertensive rats.
Sham surgery (n = 26) produced no changes in any parameter measured. After permanent middle cerebral artery occlusion, neutrophil accumulation was observed histologically, but the infiltration was moderate and typically within and adjacent to blood vessels bordering the infarcted cortex. After temporary middle cerebral artery occlusion with reperfusion, marked neutrophil infiltration was observed throughout the infarcted cortex. Myeloperoxidase activity was increased (p less than 0.05) after permanent occlusion and to a greater extent after temporary occlusion with reperfusion. Myeloperoxidase activity (units per gram wet weight) in ischemic cortex was increased over that in nonischemic (control) cortex 32.2-fold, 54.6-fold, and 92.1-fold for permanent occlusion and 80 and 160 minutes of temporary occlusion with reperfusion, respectively (p less than 0.05). Sham surgery produced no changes in myeloperoxidase activity. Leukotriene B4 receptor binding also was increased (p less than 0.05) after focal ischemia and paralleled the increases in myeloperoxidase activity. Ischemic cortex-specific receptor binding (femtomoles per milligram protein) was 3.87 +/- 0.63 in sham-operated rats and 4.57 +/- 0.98, 8.98 +/- 1.11, and 11.12 +/- 1.63 for rats subjected to permanent occlusion and 80 and 160 minutes of temporary occlusion with reperfusion, respectively (all p less than 0.05 different from sham-operated). Cortical myeloperoxidase activity was significantly correlated with the degree of cortical leukotriene B4 receptor binding (r = 0.66 and r = 0.79 in two different studies, p less than 0.01).
These data indicate that neutrophils are involved in focal ischemia and that there is a dramatic accumulation of neutrophils in infarcted tissue during reperfusion that can be quantified using the myeloperoxidase activity assay. Leukotriene B4 receptor binding increases in infarcted tissue in a parallel manner, which suggests that the increased leukotriene B4 binding is to receptors located on the accumulating neutrophils.
To determine the effect of intravenous recombinant tissue plasminogen activator (rt-PA) on vascular and neurologic outcomes, we enrolled 31 patients with acute carotid artery-territory ischemic stroke within 6 hours from symptom onset in a randomized, double-blind, placebo-controlled study. We gave either rt-PA (duteplase at the dose of 20 or 30 mega-international units [MIU]) or placebo intravenously for 60 minutes in patients randomly assigned to the three groups. A comparison between the baseline and postinfusion angiograms showed that complete or partial reperfusion occurred in 50% (5/10) of patients treated with 30 MIU rt-PA, 44% (4/9) of those treated with 20 MIU rt-PA, and 17% (2/12) in the control group. In patients with middle cerebral artery occlusions, reperfusion occurred in 71% (5/7) of the 30-MIU group, in 67% (4/6) of the 20-MIU group, and in 13% (1/8) of the control group. Patients treated with 30 MIU rt-PA showed a significantly early and better clinical improvement, as measured by the neurologic scale, than did those treated with placebo. Parenchymal hemorrhage occurred in one patient in each group, and frequency of clinically insignificant hemorrhagic infarction was comparable among the treatment groups. No major systemic complications occurred in any group. These results support the efficacy of intravenous infusion of rt-PA soon after the onset of stroke in producing rapid thrombolysis and neurologic recovery; it may be of particular value in patients with thromboembolic occlusion in the middle cerebral artery.
While polymorphonuclear leukocytes may contribute to the "no-reflow" phenomenon after focal cardiac and skeletal muscle ischemia/reperfusion, their contribution to acute focal cerebral ischemia is unresolved. We have examined the role of polymorphonuclear leukocytes in microvascular perfusion defects after focal cerebral ischemia/reperfusion in a baboon model of reversible middle cerebral artery occlusion with the anti-CD18 monoclonal antibody IB4, which inhibits neutrophil adherence to endothelium.
Microvascular patency in the basal ganglia after 3-hour middle cerebral artery occlusion and 1-hour reperfusion (by india ink tracer perfusion) was quantified by computerized video imaging. Animals were randomized to receive intravenous IB4 infusion 15 minutes before reperfusion (n = 7) or to receive no treatment (n = 6). Binding of IB4 to baboon leukocytes was maximal within 5 minutes of infusion.
In the untreated group, a significant reduction in patency was observed in microvessels less than 30 microns diameter: mean percent reflow was 51% in the capillary diameter class (4.0-7.5 microns) and 39% in the precapillary arteriole and postcapillary venule diameter class (7.5-30 microns). Infusion of IB4 before middle cerebral artery reperfusion increased reflow in microvessels of all size classes, most significantly in those 7.5-30 microns (p = 0.049) and 30-50 microns (p = 0.034) in diameter.
These results suggest that CD18-mediated polymorphonuclear leukocyte-endothelium adherence contributes to no-reflow predominantly in noncapillary microvessels and at least partially to that in capillaries.
A review of the sensitivity of genetically hypertensive rats to cerebral ischemia was presented together with original data describing the systematic comparison of the effects of focal ischemia (permanent and temporary with reperfusion) performed in hypertensive and normotensive rats (i.e., blood pressures verified in conscious instrumented rats). Microsurgical techniques were used to isolate and occlude the middle cerebral artery (MCAO) of spontaneously hypertensive (SHR), Sprague-Dawley (SD) and Wistar Kyoto (WKY) rats at the level of the inferior cerebral vein. Following permanent (24 h) MCAO, persistent and similar decreases in local microvascular perfusion (i.e., to 15.6 +/- 1.7% of pre-MCAO levels) were verified in the primary ischemic zone of the cortex for all strains using Laser-Doppler flowmetry. A contralateral hemiplegia that occurred following MCAO, evidenced by forelimb flexion and muscle weakness, was greater in SHR (neurological grade = 2.0 +/- 0.1) than SD (1.0 +/- 0.4) or WKY (0.7 +/- 0.4) rats (N = 7-9, p less than 0.05). SHR also exhibited sensory motor deficits following MCAO compared to sham-operation, with decreased normal placement response of the hindlimb (% normal = 20 vs. 83, N = 23-30, p decreased rota-rod (41 +/- 7 vs. 126 +/- 19 on rod, N = 10-15, p less than 0.05) and balance beam (25 +/- 5 vs. 116 +/- 29 s on beam, N = 5-7, p less than 0.05) performance. However, an index of general motor activity was not affected by permanent MCAO. Triphenyltetrazolium-stained forebrain tissue analyzed by planimetry revealed a significantly larger and more consistent cortical infarction in SHR (hemispheric infarction = 27.9 +/- 1.5%) compared to SD (15.4 +/- 4.1%) and WKY (4.0 +/- 2.4%) rats (N = 7-9, p less than 0.05), occupying predominantly the frontal and parietal areas. Also, a significant degree of ipsilateral hemispheric swelling (4.6 +/- 0.9%, N = 7-9, p less than 0.05) and increased brain water content (78.4 +/- 0.3% to 80.4 +/- 0.2%, N = 8-9, p less than 0.05) was identified in SHR that was not observed in SD or WKY rats. A novel model of temporary MCAO also was evaluated in the hypertensive and normotensive rat strains. Initially, the effect of increasing MCAO-time followed by 24 h reperfusion in SHR was studied. During temporary MCAO (20 to 300 min), persistent and stable decreases in local microvascular perfusion (i.e., to 15-20% of pre-MCAO levels) were verified in the primary ischemic zones of the cortex.(ABSTRACT TRUNCATED AT 400 WORDS)
Two different techniques were utilized to identify the infiltration of polymorphonuclear leukocytes (PMN) into cerebral tissue following focal ischemia: histologic analysis and a modified myeloperoxidase (MPO) activity assay. Twenty-four hours after producing permanent cortical ischemia by occluding and severing the middle cerebral artery of male spontaneously hypertensive rats, contralateral hemiparalysis and sensory-motor deficits were observed due to cerebral infarction of the frontal and parietal cortex. In hematoxylin-and-eosin-stained histologic sections, PMN, predominantly neutrophils, were identified at various stages of diapedesis from deep cerebral and meningeal vessels at the periphery of the infarct, into brain parenchyma. When MPO activity in normal brain tissue was studied initially, it could not be demonstrated in normal tissues extracted from non-washed homogenates. However, if tissue was homogenized in phosphate buffer (i.e., washed), MPO activity was expressed upon extraction. Utilizing this modified assay, MPO activity was significantly increased only in the infarcted cortex compared to other normal areas of the brain. This was observed in non-perfused animals and after perfusion with isotonic saline to remove blood constituents from the vasculature prior to brain removal. The increased PMN infiltration and MPO activity were not observed in forebrain tissue of sham-operated control rats. Also, MPO activity was not increased in the ischemic cortex of MCAO rats perfused immediately after middle cerebral artery occlusion, indicating that blood was not trapped in the ischemic area. By using a leukocyte histochemical staining assay, activity of peroxidases was identified within vascular-adhering/infiltrating PMN in the infarcted cortex 24 hr after focal ischemia. An evaluation of several blood components indicated that increased MPO activity was selective for PMN. The observed increase of approximately 0.3 U MPO/g wet weight ischemic tissue vs. nonischemic cerebral tissues probably reflects the increased vascular adherance/infiltration of approximately 600,000 PMN/g wet weight infarcted cortex 24 hr after focal ischemia. This combined biochemical and histological study strongly suggests that PMN adhere within blood vessels and infiltrate into brain tissue injured by focal ischemia and that the associated inflammatory response might contribute to delayed progressive tissue damage in focal stroke. This modified MPO assay is a useful, quantitative index of PMN that can be utilized to elucidate the potential deleterious consequences of neutrophils infiltrating into the central nervous system after cerebral ischemia, trauma, or other pro-inflammatory stimuli.
We evaluated the influence of time of recanalization or degree of initial leptomeningeal collateral blood flow in cardioembolic or arterio-arterial middle cerebral artery (MCA) occlusion on infarct size and clinical outcome in a series of 34 consecutive acute stroke patients with main stem (N = 31) or major branch (N = 3) occlusions using CT, initial cerebral arteriography (N = 21), repetitive close-meshed transcranial Doppler ultrasonography, and a neurologic stroke scale. We treated 15 patients with tissue plasminogen activator intravenously within the first 6 hours. The type and size of infarction depended on the location of the occluding lesions within the MCA trunk. Proximal MCA occlusion always led to infarction involving the striatum and internal capsule. Sixty-five percent of patients showed recanalization of the occluded MCA within 1 week. Following MCA recanalization, hyperperfusion was present in 38 to 44% of cases. There was a marginally significant relation between size of infarction on CT and recanalization time within the first 24 hours. The more rapidly recanalization occurred, the smaller the size of the infarct. When recanalization time was greater than 8 hours, the lesions always extended to the cortex. An additional good leptomeningeal collateral blood flow significantly reduced the size of the infarct and improved clinical outcome after 17 days and after 10 months. Early recanalization of embolic MCA occlusions within up to 8 hours, in conjunction with good transcortical collateralization, has a favorable impact on infarct size and outcome and may constitute the therapeutic window of opportunity.
Activated leukocytes appear to be directly involved in potentiating ischemic central nervous system (CNS) injury. Adhesion of leukocytes to endothelium is essential for their migration and requires the binding of adhesion receptors of the leukocyte (CD 18) to an intercellular adhesion molecule (ICAM) on endothelium. Monoclonal antibodies to an ICAM can block leukocyte adhesion and transendothelial migration. To determine the efficacy of anti-ICAM antibody treatment in preserving neurological function after CNS ischemia, two animal models were used. A 1-mg/kg dose of anti-ICAM was given to rabbits 30 minutes before induction of ischemia either in the spinal cord using temporary aortic occlusion or in the brain using intra-arterial microspheres. In this study, treatment with anti-ICAM produced a significant reduction in neurological deficits in the reversible spinal cord ischemia model but not in the irreversible brain ischemia model. This protective effect supports the active role of leukocytes in CNS reperfusion ischemic injury and offers potential for future therapy.
Cerebral ischemia is accompanied by many of the cardinal features of acute inflammation such as neutrophil and platelet activation and accumulation. We sought to determine whether circulating neutrophils or platelets contribute to brain injury in a rabbit model of thromboembolic stroke that includes a fixed duration of superimposed systemic hypotension. We randomized 18 rabbits to receive either antineutrophil antiserum (n = 6), antiplatelet antiserum (n = 5), or nonimmune serum (n = 7). We assessed brain ischemia by measuring cerebral blood flow, intracranial pressure, and infarct size. Following the intracarotid administration of an autologous clot, cerebral blood flow in all groups fell to less than 5 ml/100 g/min during induced hypotension. After restoration of baseline blood pressure, mean cerebral blood flow in neutropenic animals recovered to 20-30 ml/100 g/min while that in control and thrombocytopenic rabbits remained at less than 10 ml/100 g/min. Intracranial pressure in control animals rose steadily to a final value of 241% of baseline, while a much smaller increase (148% of baseline) was noted in the thrombocytopenic group; no change from baseline was evident in the neutropenic group. Infarct size was significantly (p less than 0.05) reduced in the neutropenic group but not in the thrombocytopenic group. These results suggest that neutrophils may be important contributors to ischemia-induced brain injury whereas the role of platelets is more subtle.
The short-term (less than or equal to 72-hour) cerebral vascular reaction to subarachnoid injectates of various specific blood components was determined by angiography in a canine model of cerebral vasospasm. Cell-free subarachnoid clots of autologous plasma in the basal cistern were found to produce no significant reaction of the basilar artery, while whole-blood clots induced a small (15%) chronic constriction after 24 hours. Because the plasma clots were not well retained in the basal cistern, however, small beads (dextran or latex) were added to stabilize them. Injection of beads and plasma led to moderate-to-severe chronic vasoconstriction (35% to 40%) with rapid onset. Control experiments demonstrated that these foreign bodies (beads) alone induced this vascular reaction. Histological examination showed that severe inflammation followed the introduction of subarachnoid beads. The experiments demonstrate that inflammation alone, in the absence of other processes associated with subarachnoid hemorrhage, may induce persistent and severe cerebroarterial constriction and raises the possibility that inflammation in response to subarachnoid blood may play a role in clinical vasospasm.
We define the concept of reperfusion injury, and we present a background chronology of experimental work supporting and questioning this concept. We identify several new influences, such as current clinical interest in thrombolytic therapy for acute ischemia of heart and brain and the growing recognition of endothelium as a regulator of homeostasis. We propose that these influences will encourage a reexamination of reperfusion injury as a factor in the ultimate outcome of tissue exposed to reversible ischemia. We briefly discuss the major mechanisms presently implicated in reperfusion injury--loss of calcium homeostasis, free radical generation, leukocyte-mediated injury, and acute hypercholesterolemia.
Leukocytes have been postulated to contribute to cerebral ischemia and reperfusion injury. The present study implies that leukocytes have a deleterious effect in the brain following ischemia. We compared the alteration of cortical electrical activity following transient, incomplete cerebral ischemia in control and leukopenic rats by monitoring somatosensory evoked potentials and electroencephalographic activity. There was complete cessation of electroencephalographic activity, and the cortical peak of the evoked potential was abolished during ischemia in the control animals. However, when the animals were rendered leukopenic, there was maintenance of electroencephalographic activity with reduced amplitude and preservation of the cortical peak of the evoked response during the ischemic period. This indicates that when the animals are made leukopenic, even under ischemic conditions, the neurophysiologic functioning is still maintained to a certain extent.
In the present investigation, the involvement of PMNLs and oxygen free radicals was explored in rats with postischemic perfusion disturbances of the brain. Reversible forebrain ischemia was induced by bilateral clamping of both carotid arteries in combination with hemorrhagic hypotension. This procedure resulted in a reproducible DPH 1 hr after start of recirculation. Neutropenia was induced by sheep ANS. One group received ANS before and a second group immediately after termination of ischemia. Two additional groups received SOD before or immediately after ischemia. Regional postischemic CBF was determined by [14C]iodoantipyrine autoradiography. It was found that CBF significantly improved in cortical structures of animals treated with ANS before ischemia. Treatment with ANS at the end of ischemia had no effect on the postischemic CBF depression. Neither was injection of SOD effective to influence DPH, irrespective whether given before or after ischemia. It is concluded that PMNLs play a role in the development of DPH of the brain, whereas free radical mechanisms seem to be less relevant.
We subjected nine dogs with severe granulocytopenia 4 days after the administration of mechlorethamine to 1 hour of cerebral ischemia induced by the controlled, incremental injection of air into the internal carotid artery. Cortical somatosensory evoked responses and cerebral blood flow determined by [14C]iodoantipyrine autoradiography were compared with those of six control dogs that had received mechlorethamine 1 day previously and were not yet granulocytopenic. Eleven additional control dogs received no mechlorethamine but had identical ischemic insults and were followed for 4 hours after ischemia. Both control groups had identical evoked response outcomes after 1 hour of recovery from ischemia. Granulocytopenic dogs had improved evoked response recoveries compared with either control group after 1 hour of recovery. No areas of very low blood flow were observed 1 hour after ischemia in the granulocytopenic dogs, but three of five dogs in the control group receiving mechlorethamine had such areas.
Stimulation of rat astrocytes in vitro by calcium ionophore A23187 and/or lipopolysaccharide results in the generation of a cytotoxic factor that is functionally similar to the previously described macrophage-derived cytotoxic factor, tumor necrosis factor. Like the macrophage product, the astrocyte cytotoxic factor kills murine L 929 cell targets. In addition, it kills rat oligodendrocytes, the myelin-producing cells of the central nervous system. Human recombinant tumor necrosis factor also has cytotoxic activity directed against rat oligodendrocytes.
Recombinant human tumor necrosis factor (rhTNF) has been tested for its effect on myelinated cultures of mouse spinal cord tissue. As controls, recombinant human interferon gamma (rhIFN) and interleukin-2 (rhIL-2) were tested, as well as T-cell supernatants, antigalactocerebroside serum, and normal culture medium. It was found that rhTNF induced delayed-onset (18-24 hr) oligodendrocyte necrosis and a type of myelin dilatation peculiar to this system. Some nerve fibers progressed to demyelination by 72 hours. The myelin dilatation was not reversible by return to normal feeding solution for 3 days. In contrast, rhIFN, rhIL-2, T-cell supernatants, and normal medium had little or no effect on cultures. This mechanism differs from other immune-mediated mechanisms in that it appears that a physiological (not structural) demyelination occurs initially without overt destruction of the myelin sheath. These observations are relevant to the evolution of the multiple sclerosis plaque: dysfunction of ionic channels might contribute to the eventual demise of oligodendrocytes and axons in the longstanding lesion.
The circulating white blood cells of patients with brain infarction were labelled in vitro with Indium-111 tropolonate; the cells were reinjected to study the inflammatory process by gamma camera imaging. Eight patients with acute cerebral ischemic infarct were studied during the first two weeks after the onset of neurological symptoms. In seven cases a well defined area of increased radioactivity was revealed in the infarcted hemisphere indicating active migration and tracking of labelled leukocytes in cerebral infarct. This method allows monitoring of the cellular inflammatory response in human cerebral infarcts and adds another imaging technique.
Cerebrospinal fluid (CSF) samples obtained by consecutive lumbar puncture of 26 patients with presumed pale cerebral infarction, 66 with presumed hemorrhagic infarction, 16 with lobar hematoma, and 18 with cerebral infarction verified at autopsy, were examined with a cytological method permitting a total and differential cell count. A transitory outflow of polymorphonuclear neutrophilic leukocytes (PNL) was found in 70% of the patients with hemorrhagic infarction and lobar hematoma, with a peak three to four days after onset. The strongest PNL reaction was recorded in CSF from patients with lobar hematoma. In 75% of patients with pale infarction, no PNL or only a few PNL were found. In the autopsy group the PNL reaction in the brain as well as in the CSF was stronger in patients with hemorrhagic infarcts than in those with pale infarcts.
Intercellular adhesion molecule-1 (ICAM-1) is a glycoprotein expressed on endothelial cells that facilitates leukocyte adhesion. To test the hypothesis that reduction of leukocytes in an ischemic lesion reduces ischemic brain damage, we measured the effect of administration of an anti-ICAM-1 monoclonal antibody on ischemic brain damage after transient middle cerebral artery occlusion in the rat. ICAM-1 expression increased in the ischemic lesion, and the lesion volume was significantly reduced by 41% in the anti-ICAM-1 antibody group compared with the control group (p < 0.05). Numbers of polymorphonuclear leukocytes (PMNs) were significantly reduced in the cortices of the anti-ICAM-1 antibody group compared with the control animals (p < 0.05). Our data indicate that administration of anti-ICAM-1 antibody results in a significant reduction of ischemic brain damage concomitant with a reduction of PMNs in the lesion after transient focal cerebral ischemia in the rat.
Tumor necrosis factor-alpha (TNF-alpha) is a cytokine with diverse proinflammatory actions, including endothelial leukocyte adhesion molecule expression. Since leukocytes infiltrate into ischemic brain lesions, the present study was conducted to examine whether TNF-alpha messenger RNA (mRNA) and peptide are expressed in the brain after experimental focal stroke and before leukocyte accumulation.
TNF-alpha mRNA and protein expression were monitored in the ischemic and nonischemic cerebral cortex of rats after focal ischemia produced by permanent middle cerebral artery occlusion. The effect of TNF-alpha administered by microinjection into the brain cortex on leukocyte adherence to brain capillaries was also studied.
Induction of TNF-alpha mRNA, normalized to a standard reference rat macrophage TNF-alpha mRNA, was detected as early as 1 hour after middle cerebral artery occlusion. TNF-alpha mRNA was elevated by 3 hours (29 +/- 6% versus 2 +/- 1% in sham-operated rats) only in the ischemic cortex, with peak expression at 12 hours (104 +/- 8%; P < .01). Five days after middle cerebral artery occlusion, TNF-alpha mRNA levels in ischemic cortex were still significantly elevated (38 +/- 5%; P < .05). Also, TNF-alpha mRNA expression was greater in the ischemic cortex of spontaneously hypertensive rats than in normotensive rats (P < .05). Double-labeling, immunohistochemical studies revealed the presence of TNF-alpha protein localized within nerve fibers in the evolving infarct at 6 and 12 hours after ischemia and further expression in the tissues immediately adjacent to the infarct 24 hours after ischemia. After 5 days, the neuronally localized peptide had diminished greatly, but macrophages located within the infarcted tissues were immunoreactive. Cortical microinjections of TNF-alpha (10 ng in 1 microL) produced a significant neutrophil adherence/accumulation in capillaries and small blood vessels 24 hours later.
These results represent the first demonstration that focal cerebral ischemia in rats results in elevated TNF-alpha mRNA and protein in ischemic neurons. The neuronal expression of peptide appears to facilitate the infiltration of inflammatory cells that can further exacerbate tissue damage in cerebral ischemia and might contribute to increased sensitivity and risk in focal stroke.
We investigated the effect of an anti-CD11b monoclonal antibody (1B6c) on ischemic cell damage after transient middle cerebral artery occlusion. We divided animals into three groups: MAb 1 group (n = 5)--rats were subjected to 2 hours of transient occlusion and 1B6c (1 mg/kg) was administered intravenously at 0 and 22 hours of reperfusion; MAb 2 group (n = 5)--same experimental protocol as MAb 1 group, except that the initial dose of 1B6c was increased to 2 mg/kg; and control group (n = 5)--same experimental protocol as MAb 2 group, except that an isotype-matched control antibody was administered. Animals were weighed and tested for neurological function before and after occlusion of the middle cerebral artery. Forty-six hours after reperfusion, brain sections were stained with hematoxylin and eosin for histology evaluation. We observed a significant reduction of weight loss and improvement in neurological function after ischemia in the MAb 2 animals compared to MAb 1 and vehicle-treated animals (p < 0.05). The lesion volume was significantly smaller in the MAb 2 group (19.5 +/- 1.9%) compared to MAb 1 (29.9 +/- 2.6%) and vehicle-treated (34.2 +/- 5.4%) groups (p < 0.01). Tissue polymorphonuclear cell numbers were reduced in both 1B6c-administered groups. Our data demonstrate that administration of anti-CD11b antibody results in a dose-dependent, significant functional improvement and reduction of ischemic cell damage after transient focal cerebral ischemia in the rat.
Postischemic cerebral inflammation may contribute to ischemic cell damage. The CD11b/18 (Mac-1) integrin mediates stimulated neutrophil binding to endothelia. We therefore investigated the effect of administration of an anti-Mac-1 monoclonal antibody on cerebral ischemic cell damage in the rat.
Rats (n = 10) were subjected to 2 hours of middle cerebral artery occlusion; the anti-Mac-1 antibody was administered at a dose of 2 mg/kg i.v. at 1 hour of reperfusion and 1 mg/kg i.v. at 22 hours of reperfusion or an isotype-matched control antibody (n = 10) was administered using the same experimental protocol. Rats were killed at 46 hours of reperfusion, and brain sections were stained with hematoxylin and eosin for histological evaluation. In a separate population of rats given either vehicle (n = 8) or anti-Mac-1 antibodies (n = 9), intraparenchymal neutrophils were measured by means of a myeloperoxidase assay.
The lesion volume was significantly smaller (28%) in the anti-Mac-1 antibody group compared with the vehicle control group (P < .01). Numbers of intraparenchymal polymorphonuclear cells were significantly reduced (P < .05) in the cortex of the anti-Mac-1 antibody group compared with the vehicle control group.
Our data demonstrate that administration of anti-Mac-1 antibody 1 hour after onset of reperfusion results in significant reductions of ischemic cell damage and intraparenchymal neutrophils after transient (2-hour) focal cerebral ischemia in the rat.
The results of several experimental studies of focal ischemia and anecdotal observations suggest that leukocytes may contribute to the injury initiated by an arterial occlusion. The timing and the nature of leukocyte responses in evolving brain infarcts (either human or experimental) are incompletely characterized. This is a study of experimental brain lesions in 96 Wistar rats that underwent occlusion of a large intracranial artery for variable intervals ranging between 30 minutes and 7 days. The experimental model, based on the occlusion of a middle cerebral artery ostium via the insertion of a nylon monofilament through the external carotid artery, does not require opening the skull; therefore, the inflammatory response is not influenced by the effects of craniotomy and changes in intracranial pressure are only those induced by the ischemic lesion. All 96 animals having the same type of arterial occlusion developed an ischemic brain lesion (limited to the territory of the corresponding artery) that evolved into an area of extensive neuronal necrosis over a period of 6 to 12 hours followed by pan-necrosis (infarct) approximately 60 hours later. In this study, leukocytes (in particular polymorphonuclear cells) were detected in the microvessels (capillaries and venules) of the ischemic hemisphere as early as 30 minutes after the arterial occlusion. Numbers of intravascular neutrophils peaked at 12 hours, whereas intraparenchymal granulocytes were most numerous at 24 hours; a few granulocytes were visible in the brain infarct as late as day 7. Circulating monocytes were first detected within the capillaries/venules of the ischemic area after 4 to 6 hours. Platelet aggregates were more abundant in the arterial than the venous side of the circulation, and luminal obstruction of arteries by platelet aggregates became noticeable only 48 hours after the arterial occlusion. Fibrin thrombi were conspicuous for their absence. These observations provide the background for studies that will attempt to unravel the relationship between the biological responses of leukocytes and neuronal necrosis secondary to focal ischemia.
Shock increases mortality from brain injuries, but the mechanism is poorly understood. We hypothesized that brain injury followed by shock and resuscitation leads to a secondary reperfusion injury mediated in part by polymorphonuclear leukocytes (PMNs). To validate this hypothesis, we studied cerebral perfusion pressure (CPP), intracranial pressure (ICP), cerebral blood flow (CBF), cortical water content (CWC), and hemodynamic variables in a porcine model of focal cryogenic brain injury and hemorrhagic shock. Cerebral PMN accumulation (CPMN) in the injured and uninjured hemispheres was determined histologically from the total PMNs in five high-power fields (400x). Twenty-nine mature swine were randomized to four groups. Group 1, the control group, was instrumented only. Group 2 animals had a brain injury alone and were studied for 24 hours. Group 3 animals had a brain injury and hemorrhagic shock. Group 4 animals had hemorrhagic shock alone. Brain injury followed by shock caused a significantly greater ICP and a significantly lower CBF than brain injury or shock alone. There was no significant difference in CPP between groups after resuscitation. The CWC of the lesioned area was similar in both brain-injured groups but was significantly increased when compared with the controls and the shock-only group. The CWC of the nonlesioned hemisphere was higher in group 3 than in group 2. The CPMN in both hemispheres in group 3 was significantly greater than in either group 2 or group 4. There was a significant positive correlation between CPMN and both ICP and CWC, and a significant negative correlation between CPMN and CBF. These data suggest an association between CPMN accumulation and secondary brain injury.
The dynamics of leukocyte infiltration in human cerebral ischemia were studied using technetium-99m hexamethylpropyleneamine oxime (99mTc HMPAO)-labeled leukocyte brain single-photon emission computed tomography (SPECT).
Twenty-two patients diagnosed as having hemispheric ischemic stroke were examined with 99mTc HMPAO brain SPECT for cerebral blood flow study and 99mTc HMPAO-labeled leukocyte brain SPECT for the study of leukocyte infiltration. Three patients with chronic hemispheric ischemic stroke received one examination. Nineteen patients with acute hemispheric ischemic stroke received their initial examination within the first after onset. Follow-up examinations were performed at intervals of 1-3 weeks whenever possible.
In patients with chronic hemispheric ischemic stroke, leukocyte infiltration was not seen in areas of perfusion defect. In patients with acute hemispheric ischemic stroke, leukocyte infiltration was seen in areas of perfusion defect during the acute stage, which persisted for no less than 5 weeks after onset and then declined.
A new method to study and monitor the process of leukocyte infiltration in acute cerebral ischemia using 99mTc HMPAO-labeled leukocyte brain SPECT is described. This method shows that leukocyte infiltration in acute hemispheric ischemic stroke is a dynamic process that persists for no less than 5 weeks and then declines.
Polymorphonuclear leukocyte infiltration into cerebral focal ischemic tissue: Myeloperoxidase activity assay and histological verification
- R K Clark
- D E Griswold
Clark, R. K.; Griswold, D. E. Polymorphonuclear leukocyte infiltration into cerebral focal ischemic tissue: Myeloperoxidase activity assay and histological verification. J. Neurosci. Res. 29:336-345; 1991. Barone, F. C.; Hillegass, L. M.;
Leukocytes and leukotriene B4 (LTB4) receptors in stroke
- R K Clark
- D E Griswold
- H M Sarau
- W J Price
Clark, R. K.; Griswold, D. E.; Sarau, H. M. Leukocytes and leukotriene B4 (LTB4) receptors in stroke. Sot. Neurosci. Abst. 19:1057; 1993. Barone, F. C.; Price, W. J.;
Murinc models of cerebral ischemia CNS trauma: Laboratory techniques and recent advancements
- A H Nelson
Nelson, A. H.; Feuer-stein, G. Z.; Sarau. H. M.: Tzimas. M. N.: Schmidt. D. B.: Kaoadia. R. D.; Sarkar, S. K. Murinc models of cerebral ischemia. In: Ohnishi, S. T., ed. Mem~ane linked diseases, vol. 4: CNS trauma: Laboratory techniques and recent advancements. New York: CRC Press (in press).
Progression from ischemic injury to infarct following middle cerebral artery occlusion in the rat
- Garcia
Development of tissue damage, inflammation and resolution following stroke: An immunohistochemical and quantitative planimetric study
- Clark
Polymorphonuclear leukocytes occlude capillaries follwoing middle cerebral artery occlusion and reperfusion in baboons
- del Zoppo
Polymorphonuclear leukocyte infiltration into cerebral focal ischemic tissue: Myeloperoxidase activity assay and histological verification
- Barone
Leukocytes and leukotriene B4 (LTB4) receptors in stroke
- Barone
Murine models of cerebral ischemia
- F C Barone
- R N Willette
- R K Clark
- A H Nelson
- G Z Feuerstein
- H M Sarau
- M N Tzimas
- D B Schmidt
- R D Kapadia
- S K Sarkar
The role of neutrophils and platelets in a rabbit model of thromboembolic stroke
- Bednar