A Mouse Model of Post-Stroke Pneumonia Induced by Intra-Tracheal Inoculation with Streptococcus pneumoniae

Background: Stroke-induced immunodeficiency increases the risk of infectious complications, which adversely affects neurological outcome. Among those, pneumonia affects as many as one third of stroke patients and is the main contributor to mortality in the post-acute phase of stroke. Experimental findings on post-stroke susceptibility to spontaneous pneumonia in mice are contradictory. Here, we established a mouse model inducing standardized bacterial pneumonia and characterized the impaired pulmonary cellular and humoral immune responses after experimental stroke. Methods: Bacterial pneumonia was induced by intra-tracheal inoculation with Streptococcus pneumoniae at different time points after transient middle cerebral artery occlusion (MCAO). Bacterial counts in lungs and blood, histological changes, and cytokine production in the lungs were assessed. Furthermore, we investigated the effect of pneumonia on stroke outcome. Results: Intra-tracheal inoculation resulted in reproducible pneumonia and bacteraemia, and demonstrated post-stroke susceptibility to streptococcal pneumonia developing with a delay of at least 24 h after MCAO. Higher bacterial counts in mice infected 3 days after stroke induction correlated with reduced neutrophil and macrophage infiltration in the lungs and lower levels of pro-inflammatory cytokines in the broncho-alveolar lavage compared to sham-operated animals. Pneumonia increased mortality without affecting brain-infiltrating leukocytes. Conclusions: In this standardized mouse model of post-stroke pneumonia, we describe attenuated leukocyte infiltration and cytokine production in response to bacterial infection in the lungs that has a profound effect on outcome.

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Abstract P746: Sex Differences in Cognitive and Psychological Outcomes of Stroke: Impact of Diabetes

Stroke ◽

10.1161/str.52.suppl_1.p746 ◽

2021 ◽

Vol 52 (Suppl_1) ◽

Author(s):

Victoria L Wolf ◽

Aunay Miller ◽

Raghavendar Chandran ◽

Weiguo Li ◽

Adviye Ergul

Keyword(s):

Artery Occlusion ◽

Psychological Outcomes ◽

Experimental Stroke ◽

Dark Light ◽

Stroke Outcomes ◽

Post Stroke ◽

Stroke Research ◽

Y Maze ◽

Male Animal ◽

The Impact

Diabetes increases risk and severity of post-stroke cognitive impairment (PSCI), a major cause of disability worldwide. While it is known that females suffer more from PSCI, psychological outcomes and underlying reasons are poorly understood. From a preclinical perspective, potential explanations include 1) use of otherwise healthy animals in experimental stroke research without integration of common comorbid diseases like diabetes into the study design, and 2) optimization of most behavioral tests for sensorimotor and cognitive functions using only male animal models. Our hypothesis is that post-stroke outcomes are sex and comorbid disease-dependent. To test this, we validated the Novel Object Recognition (NOR), Y-maze, and Passive Avoidance (PAT) behavioral paradigms in Ctrl and Diabetic (DM) male (M) and female (F) rats pre- and post-stroke (S) via 60 min. middle cerebral artery occlusion (MCAO). We tested the PAT paradigm with a multi-trial method where the animals were habituated to the dark/light chambers without foot shock and then trained in 3 trials where they received foot shock upon entering the dark. We then tested retention following MCAO for their memory of foot shock 2 weeks prior. Multitrial results suggested that there was no difference between groups in learning to associate the dark chamber with the shock, so we revised the multitrial method into a single-trial method for ongoing retention tests to compare the impact of stroke on shock memory recall. PAT revealed (Table 1) disease- and sex-dependent responses to aversive stimulus. NOR revealed that M-DM-S and F-DM-S rats have decreased exploration time, suggesting that they are unmotivated or depressed. Y-maze indicated that males displayed spatial memory recovery, while females remained impaired. In summary, we have observed numerous sex- and disease-dependent post-stroke outcomes with standard behavioral paradigms, causing us to carefully consider how we evaluate preclinical outcomes.

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Impact of Key Nicotinic AChR Subunits on Post-Stroke Pneumococcal Pneumonia

Vaccines ◽

10.3390/vaccines8020253 ◽

2020 ◽

Vol 8 (2) ◽

pp. 253

Author(s):

Sandra Jagdmann ◽

Claudia Dames ◽

Daniel Berchtold ◽

Katarzyna Winek ◽

Luis Weitbrecht ◽

...

Keyword(s):

Artery Occlusion ◽

Experimental Stroke ◽

Medical Complication ◽

Α7 Nicotinic Acetylcholine Receptor ◽

Post Stroke ◽

Cholinergic Signaling ◽

Cerebral Artery Occlusion ◽

Pathogen Clearance ◽

The Impact ◽

Alveolar Capillary

Pneumonia is the most frequent severe medical complication after stroke. An overactivation of the cholinergic signaling after stroke contributes to immunosuppression and the development of spontaneous pneumonia caused by Gram-negative pathogens. The α7 nicotinic acetylcholine receptor (α7nAChR) has already been identified as an important mediator of the anti-inflammatory pathway after stroke. However, whether the α2, α5 and α9/10 nAChR expressed in the lung also play a role in suppression of pulmonary innate immunity after stroke is unknown. In the present study, we investigate the impact of various nAChRs on aspiration-induced pneumonia after stroke. Therefore, α2, α5, α7 and α9/10 nAChR knockout (KO) mice and wild type (WT) littermates were infected with Streptococcus pneumoniae (S. pneumoniae) three days after middle cerebral artery occlusion (MCAo). One day after infection pathogen clearance, cellularity in lung and spleen, cytokine secretion in bronchoalveolar lavage (BAL) and alveolar-capillary barrier were investigated. Here, we found that deficiency of various nAChRs does not contribute to an enhanced clearance of a Gram-positive pathogen causing post-stroke pneumonia in mice. In conclusion, these findings suggest that a single nAChR is not sufficient to mediate the impaired pulmonary defense against S. pneumoniae after experimental stroke.

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Erythropoietin Selectively Attenuates Cytokine Production and Inflammation in Cerebral Ischemia by Targeting Neuronal Apoptosis

Journal of Experimental Medicine ◽

10.1084/jem.20021067 ◽

2003 ◽

Vol 198 (6) ◽

pp. 971-975 ◽

Cited By ~ 358

Author(s):

Pia Villa ◽

Paolo Bigini ◽

Tiziana Mennini ◽

Davide Agnello ◽

Teresa Laragione ◽

...

Keyword(s):

Mononuclear Cells ◽

Cytokine Production ◽

Inflammatory Responses ◽

Human Peripheral Blood ◽

Inflammatory Cells ◽

Artery Occlusion ◽

Experimental Stroke ◽

Peripheral Blood Mononuclear ◽

Monocyte Chemoattractant Protein 1 ◽

Cerebral Artery Occlusion

Ischemic brain injury resulting from stroke arises from primary neuronal losses and by inflammatory responses. Previous studies suggest that erythropoietin (EPO) attenuates both processes. Although EPO is clearly antiapoptotic for neurons after experimental stroke, it is unknown whether EPO also directly modulates EPO receptor (EPO-R)–expressing glia, microglia, and other inflammatory cells. In these experiments, we show that recombinant human EPO (rhEPO; 5,000 U/kg body weight, i.p.) markedly reduces astrocyte activation and the recruitment of leukocytes and microglia into an infarction produced by middle cerebral artery occlusion in rats. In addition, ischemia-induced production of the proinflammatory cytokines tumor necrosis factor, interleukin 6, and monocyte chemoattractant protein 1 concentration is reduced by >50% after rhEPO administration. Similar results were also observed in mixed neuronal-glial cocultures exposed to the neuronal-selective toxin trimethyl tin. In contrast, rhEPO did not inhibit cytokine production by astrocyte cultures exposed to neuronal homogenates or modulate the response of human peripheral blood mononuclear cells, rat glial cells, or the brain to lipopolysaccharide. These findings suggest that rhEPO attenuates ischemia-induced inflammation by reducing neuronal death rather than by direct effects upon EPO-R–expressing inflammatory cells.

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Lempel-Ziv complexity of the EEG predicts long-term functional recovery after stroke in rats

10.1101/248039 ◽

2018 ◽

Cited By ~ 1

Author(s):

Susan Leemburg ◽

Claudio L. Bassetti

Keyword(s):

Motor Function ◽

Functional Recovery ◽

Brain Function ◽

Complexity Analysis ◽

Recovery Period ◽

Artery Occlusion ◽

Multiscale Entropy ◽

Experimental Stroke ◽

Reaching Task ◽

Post Stroke

AbstractNon-linear complexity of the EEG signal can be used to detect abnormal brain function relating to behavioral deficits. Here, we compare the effects of experimental stroke on EEG complexity using Lempel-Ziv complexity analysis (LZC) and multiscale entropy analysis (SampEn).EEG was recorded in bilateral motor cortex at baseline and during a 30-day recovery period after distal middle cerebral artery occlusion in rats. Motor function was assessed using a single pellet reaching task. Stroke caused an acute drop in both LZC and SampEn in the ipsilesional hemisphere in wakefulness, NREM and REM sleep, as well as reduced pellet reaching success. SampEn reductions persisted for at least 10 days post-stroke, whereas LZC had returned to baseline levels by day 4. EEG complexity in the contralesional hemisphere and in sham-operated animals were unaffected.If EEG complexity reflects post-stroke brain function, post-stroke asymmetry could be used to predict behavioral recovery. In rats, acute LZC asymmetry was significantly correlated with the amount of motor function recovery by post-stroke day 31, but SampEn asymmetry was not. EEG LZC may thus be a useful tool for predicting functional recovery after stroke. MSE could be effective in identifying cortical dysfunction, but does not reflect behavioral outcomes.

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Abstract P733: Astroglial Activation Following Stroke Contributes to Impaired Synaptic Plasticity Through Increased Cd38-trpm2 Channel Signaling

Stroke ◽

10.1161/str.52.suppl_1.p733 ◽

2021 ◽

Vol 52 (Suppl_1) ◽

Author(s):

Amelia M Burch ◽

James E Orfila ◽

Robert Dietz ◽

Andra Dingman ◽

Danae Mitchell ◽

...

Keyword(s):

Synaptic Plasticity ◽

Memory Function ◽

Hippocampal Slices ◽

Brain Slices ◽

Artery Occlusion ◽

Experimental Stroke ◽

Post Stroke ◽

Cd38 Expression ◽

Trpm2 Channel ◽

Electrophysiological Recordings

Introduction: Post-stroke cognitive impairment (PSCI) is a major contributor to long-term disability following acute ischemic stroke. Learning and memory deficits are a common feature of PSCI and alterations in hippocampal function are a likely contributor. Interestingly, common experimental stroke models (middle cerebral artery occlusion; MCAO) cause hippocampal dysfunction, despite no direct ischemic insult to the hippocampus, suggesting perturbations in neural circuits. Thus, we utilize electrophysiological recordings of hippocampal plasticity in combination with neurobehavioral assessments of memory function. Hypothesis: Activated astrocytes in the hippocampus following MCAO increase expression of the surface enzyme CD38, which signals to neurons to impair plasticity. Methods: Extracellular field recordings of CA1 neurons were performed in acute hippocampal slices prepared 30 days after recovery from transient MCAO (60 min) in adult (6-8 week) mice. A behavioral fear conditioning paradigm (CFC) was used to evaluate contextual memory. Immunohistochemistry was performed to assess CD38 expression and slices were treated with CD38 inhibitors (78c) to assess plasticity. Results: Recordings obtained in brain slices 30 days after MCAO exhibited loss of hippocampal LTP; 134±6%, n=4 in sham and 107±12%, n=4 30 days after MCAO. Memory function, measured using CFC, was consistent with our LTP findings. MCAO decreased freezing behavior, indicating lack of memory (65±7% in sham mice (n=6) and 37±7% in MCAO mice, n=7). Immunohistochemical data indicates increased CD38 expression in activated astrocytes following MCAO in the hippocampus. Treatment of hippocampal slices with 78c, a potent CD38 inhibitor, after MCAO rescues LTP impairment. Finally, no additive increase in LTP when 78c is co-administered with a TRPM2 channel inhibitor was observed. Conclusion: These data indicate that MCAO is a reproducible model of post-stroke memory dysfunction (PSCI) and remote astrogliosis in the uninjured hippocampus may contribute to altered neuronal function (plasticity). Our data implicates increased levels of CD38 as an upstream activator of neuronal TRPM2 channel in the hippocampus following stroke, resulting in impaired synaptic plasticity.

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Short-duration hypothermia completed prior to reperfusion prevents intracranial pressure elevation following ischaemic stroke in rats

Scientific Reports ◽

10.1038/s41598-021-01838-7 ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Daniel Omileke ◽

Sara Azarpeykan ◽

Steven W. Bothwell ◽

Debbie Pepperall ◽

Daniel J. Beard ◽

...

Keyword(s):

Intracranial Pressure ◽

Ischaemic Stroke ◽

Short Duration ◽

Infarct Volume ◽

Arterial Occlusion ◽

Reperfusion Therapy ◽

Artery Occlusion ◽

Experimental Stroke ◽

Stroke Patients ◽

Post Stroke

AbstractReperfusion therapies re-establish blood flow after arterial occlusion and improve outcome for ischaemic stroke patients. Intracranial pressure (ICP) elevation occurs 18–24 h after experimental stroke. This elevation is prevented by short-duration hypothermia spanning the time of reperfusion. We aimed to determine whether hypothermia-rewarming completed prior to reperfusion, also prevents ICP elevation 24 h post-stroke. Transient middle cerebral artery occlusion was performed on male outbred Wistar rats. Sixty-minute hypothermia to 33 °C, followed by rewarming was induced prior to reperfusion in one group, and after reperfusion in another group. Normothermia controls received identical anaesthesia protocols. ΔICP from pre-stroke to 24 h post-stroke was measured, and infarct volumes were calculated. Rewarming pre-reperfusion prevented ICP elevation (ΔICP = 0.3 ± 3.9 mmHg vs. normothermia ΔICP = 5.2 ± 2.1 mmHg, p = 0.02) and reduced infarct volume (pre-reperfusion = 78.6 ± 23.7 mm3 vs. normothermia = 125.1 ± 44.3 mm3, p = 0.04) 24 h post-stroke. There were no significant differences in ΔICP or infarct volumes between hypothermia groups rewarmed pre- or post-reperfusion. Hypothermia during reperfusion is not necessary for prevention of ICP rise or infarct volume reduction. Short-duration hypothermia may be an applicable early treatment strategy for stroke patients prior to- during-, and after reperfusion therapy.

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Hypothermia and rewarming prior to reperfusion, prevents intracranial pressure elevation after ischaemic stroke in rats: an investigation to define the importance of cooling during reperfusion

10.21203/rs.3.rs-846595/v1 ◽

2021 ◽

Author(s):

Daniel Omileke ◽

Sara Azarpeykan ◽

Steven W Bothwell ◽

Debbie Pepperall ◽

Daniel J Beard ◽

...

Keyword(s):

Intracranial Pressure ◽

Ischaemic Stroke ◽

Short Duration ◽

Infarct Volume ◽

Arterial Occlusion ◽

Reperfusion Therapy ◽

Artery Occlusion ◽

Experimental Stroke ◽

Stroke Patients ◽

Post Stroke

Abstract Reperfusion therapies re-establish blood flow after arterial occlusion and improve outcome for ischaemic stroke patients. Intracranial pressure (ICP) elevation occurs 18–24 h after experimental stroke. This elevation is prevented by short-duration hypothermia spanning the time of reperfusion. We aimed to determine whether hypothermia-rewarming completed prior to reperfusion, also prevents ICP elevation 24 h post-stroke. Transient middle cerebral artery occlusion was performed on male outbred Wistar rats. Sixty-minute hypothermia to 33℃, followed by rewarming was induced prior to reperfusion in one group, and after reperfusion in another group. Normothermia controls received identical anaesthesia protocols. ΔICP from pre-stroke to 24 h post-stroke was measured, and infarct volumes were calculated. Rewarming pre-reperfusion prevented ICP elevation (ΔICP = 0.3 ± 3.9 mmHg vs. normothermia ΔICP = 5.2 ± 2.1 mmHg, p = 0.02) and reduced infarct volume (pre-reperfusion = 78.6 ± 23.7 mm3 vs. normothermia = 125.1 ± 44.3 mm3, p = 0.04) 24 h post-stroke. There were no significant differences in ΔICP or infarct volumes between hypothermia groups rewarmed pre-or post-reperfusion. Hypothermia during reperfusion is not necessary for prevention of ICP rise or infarct volume reduction. Short-duration hypothermia is a broadly applicable potential early treatment strategy for stroke patients prior to- during-, and after reperfusion therapy.

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Inadequate food and water intake determine mortality following stroke in mice

Journal of Cerebral Blood Flow & Metabolism ◽

10.1177/0271678x16660986 ◽

2016 ◽

Vol 37 (6) ◽

pp. 2084-2097 ◽

Cited By ~ 7

Author(s):

Athanasios Lourbopoulos ◽

Uta Mamrak ◽

Stefan Roth ◽

Matilde Balbi ◽

Joshua Shrouder ◽

...

Keyword(s):

Middle Cerebral Artery ◽

Middle Cerebral Artery Occlusion ◽

Water Intake ◽

Cerebral Artery ◽

Stroke Care ◽

Artery Occlusion ◽

Experimental Stroke ◽

Post Stroke ◽

Food And Water Intake ◽

Cerebral Artery Occlusion

Experimental stroke models producing clinically relevant functional deficits are often associated with high mortality. Because the mechanisms that underlie post-stroke mortality are largely unknown, results obtained using these models are often difficult to interpret, thereby limiting their translational potential. Given that specific forms of post-stroke care reduce mortality in patients, we hypothesized that inadequate food and water intake may underlie mortality following experimental stroke. C57BL/6 mice were subjected to 1 h of intraluminal filament middle cerebral artery occlusion. Nutritional support beginning on the second day after filament middle cerebral artery occlusion reduced the 14-day mortality rate from 59% to 15%. The surviving mice in the post-stroke support group had the same infarct size as non-surviving control mice, suggesting that post-stroke care was not neuroprotective and that inadequate food and/or water intake are the main reasons for filament middle cerebral artery occlusion–induced mortality. This notion was supported by the presence of significant hypoglycemia, ketonemia, and dehydration in control mice. Taken together, these data suggest that post–filament middle cerebral artery occlusion mortality in mice is not primarily caused by ischemic brain damage, but secondarily by inadequate food and/or water intake. Thus, providing nutritional support following filament middle cerebral artery occlusion greatly minimizes mortality bias and allows the study of long-term morphological and functional sequelae of stroke in mice.

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Abstract WP151: Rapamycin Improves Post-Recanalization Blood Flow After Acute Experimental Stroke in Rats

Stroke ◽

10.1161/str.51.suppl_1.wp151 ◽

2020 ◽

Vol 51 (Suppl_1) ◽

Author(s):

Anna Maria Schneider ◽

Daniel Beard ◽

Alastair Buchan

Keyword(s):

Blood Flow ◽

Infarct Volume ◽

Mammalian Target Of Rapamycin ◽

Volume Measurement ◽

Artery Occlusion ◽

Experimental Stroke ◽

Stroke Outcome ◽

Doppler Flowmetry ◽

Rapamycin Treatment ◽

Post Stroke

Background and Purpose: In the era of thrombectomy, there is evidence suggesting that successful recanalization is not always accompanied by complete reperfusion, the so called “no-reflow phenomenon”. Given the importance of reperfusion as a predictor of stroke outcome, this represents a potential target for stroke therapy. Rapamycin, a clinically approved inhibitor of Mammalian Target of Rapamycin (mTOR) has been shown to improve cerebral blood flow (CBF) in Alzheimer’s disease. However, there has been little investigation into the effect of rapamycin on post-stroke microvascular perfusion. The aim of this study was to investigate the effects of rapamycin treatment on post-recanalisation CBF and stroke outcome in an experimental animal model of stroke. Methods: Male Wistar rats (300-350g) were subjected to 90min of transient middle cerebral artery occlusion (tMCAo) followed by randomized administration of 250μg/kg intravenous rapamycin (n=8) or vehicle (n=6), 30min after the onset of MCAo. Laser Doppler flowmetry was used to continuously measure changes in MCA perfusion during MCAo and for min after recanalisation. Neurobehavioral tests were performed 24hrs after MCAo before tissue was collected for infarct volume measurement. Results: MCAo was confirmed by a 70% reduction in MCA perfusion. Rapamycin treatment significantly improved post-recanalization CBF at 55min after recanalization (p<0.01). Rapamycin significantly increased average CBF during the 60min post-recanalization period (p<0.01). Rapamycin showed a trend towards reduced final infarct volume. Rapamycin significantly improved neuroscores (p<0.05). Post-recanalization CBF was significantly inversely correlated with infarct volume (R 2 =0.4994, p<0.05). Conclusion: Rapamycin significantly improved post-recanalization CBF and behavioural outcomes after MCAo. These results suggest that rapamycin may be an effective acute intervention to improve post-recanalisation blood flow to improve stroke outcome. However, further studies are need to determine the mechanism of improved CBF and if improvements in post-stroke CBF and neurological outcome are sustained long-term post-stroke.

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Decreased Intracranial Pressure Elevation and Cerebrospinal Fluid Outflow Resistance: A Potential Mechanism of Hypothermia Cerebroprotection Following Experimental Stroke

Brain Sciences ◽

10.3390/brainsci11121589 ◽

2021 ◽

Vol 11 (12) ◽

pp. 1589

Author(s):

Daniel Omileke ◽

Steven W. Bothwell ◽

Debbie Pepperall ◽

Daniel J. Beard ◽

Kirsten Coupland ◽

...

Keyword(s):

Cerebrospinal Fluid ◽

Intracranial Pressure ◽

Ischaemic Stroke ◽

Infarct Volume ◽

Artery Occlusion ◽

Experimental Stroke ◽

Outflow Resistance ◽

Hypothermia Treatment ◽

Post Stroke ◽

Intracranial Pressure Elevation

Background: Elevated intracranial pressure (ICP) occurs 18–24 h after ischaemic stroke and is implicated as a potential cause of early neurological deterioration. Increased resistance to cerebrospinal fluid (CSF) outflow after ischaemic stroke is a proposed mechanism for ICP elevation. Ultra-short duration hypothermia prevents ICP elevation 24 h post-stroke in rats. We aimed to determine whether hypothermia would reduce CSF outflow resistance post-stroke. Methods: Transient middle cerebral artery occlusion was performed, followed by gradual cooling to 33 °C. At 18 h post-stroke, CSF outflow resistance was measured using a steady-state infusion method. Results: Hypothermia to 33 °C prevented ICP elevation 18 h post-stroke (hypothermia ∆ICP = 0.8 ± 3.6 mmHg vs. normothermia ∆ICP = 4.4 ± 2.0 mmHg, p = 0.04) and reduced infarct volume 24 h post-stroke (hypothermia = 78.6 ± 21.3 mm3 vs. normothermia = 108.1 ± 17.8 mm3; p = 0.01). Hypothermia to 33 °C did not result in a significant reduction in CSF outflow resistance compared with normothermia controls (0.32 ± 0.36 mmHg/µL/min vs. 1.07 ± 0.99 mmHg/µL/min, p = 0.06). Conclusions: Hypothermia treatment was protective in terms of ICP rise prevention, infarct volume reduction, and may be implicated in CSF outflow resistance post-stroke. Further investigations are warranted to elucidate the mechanisms of ICP elevation and hypothermia treatment.

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