Platelet Endothelial Cell Adhesion Molecule (PECAM/CD31) Blockade Modulates Neutrophil Recruitment Patterns and Reduces Infarct Size in Experimental Ischemic Stroke.

The infiltration of polymorphonuclear leukocytes (PMNs) in ischemia-reperfusion injury (I/RI) has been implicated as a critical component of inflammatory damage following ischemic stroke. However, successful blockade of PMN transendothelial migration (TEM) in preclinical studies has not translated to meaningful clinical outcomes. To investigate this further, leukocyte infiltration patterns were quantified, and these patterns were modulated by blocking platelet endothelial cell adhesion molecule-1 (PECAM), a key regulator of TEM. LysM-eGFP mice and microscopy were used to visualize all myeloid leukocyte recruitment following ischemia/reperfusion. Visual examination showed heterogeneous leukocyte distribution across the infarct at both 24 and 72 hours after I/RI. A semiautomated process was designed to precisely map PMN position across brain sections. Treatment with PECAM function-blocking antibodies did not significantly affect total leukocyte recruitment but did alter their distribution, with more observed at the cortex at both early and later time points (24 hours: 89% PECAM blocked vs. 72% control; 72 hours: 69% PECAM blocked vs. 51% control). This correlated with a decrease in infarct volume. These findings suggest that TEM, in the setting of I/RI in the cerebrovasculature, occurs primarily at the cortical surface. The reduction of stroke size with PECAM blockade suggests that infiltrating PMNs may exacerbate I/RI and indicate the potential therapeutic benefit of regulating the timing and pattern of leukocyte infiltration after stroke.

Inhibition of PECAM-1 Significantly Delays Leukocyte Extravasation into the Subcortex Post-Stroke.

BACKGROUND AND PURPOSE: Current therapies for ischemic stroke focus on reperfusion but do not address the acute inflammatory response that results in significant reperfusion injury. To advance future therapies, a thorough understanding of the precise spatiotemporal underpinnings of leukocyte extravasation and infiltration is necessary. We describe the evolution of the inflammatory response in a mouse transient middle cerebral artery occlusion (tMCAO) stroke model at several time points after reperfusion and the modulation of this response with PECAM blockade. METHODS: The transient Middle Cerebral Artery Occlusion model (90 minutes of ischemia followed by reperfusion) was used to simulate large vessel occlusion stroke and recanalization. We used wide field and confocal immunofluorescence microscopy to examine the exact distribution of neutrophils with close examination of the leukocyte position with regard to the brain vasculature and the perivascular space. Flow cytometry of single cell suspensions was used to confirm cell identity at different time points post-stroke. RESULTS: Large ischemic strokes involving both the subcortex and cortex (over 20% of the ischemic hemisphere) were induced in mice. At 12 and 24 hours, leukocyte recruitment and extravasation was primarily localized to the cortical surface. This contrasts with other organs where there is considerable migration of neutrophils deep into the inflamed tissue by 24 hours. Flow cytometry showed at 24 hours a majority of leukocytes were neutrophils. Over 48 to 72 hours, leukocytes were increasingly found deeper into the subcortex. Throughout the infarct (determined with triphenyl tetrazolium chloride staining), leukocyte recruitment was not uniform but rather organized in clusters. Disrupting leukocyte diapedesis with PECAM function-blocking monoclonal antibody restricted leukocytes to within 500 microns of the surface when compared to control; and this was still evident at 72 hours (n=3 mice per group, p<0.01, Control 46% ± 4.0 %; PECAM-1 Ab 62% ± 5.0%). High-resolution wide-field microscopy confirmed inhibition of TEM by PECAM-1 blockade at 24 hours. Flow cytometry showed approximately equal numbers of monocytes and neutrophils at 72 hours. CONCLUSIONS: Our findings demonstrate that leukocyte infiltration into a stroke evolves over several days following reperfusion. The use of PECAM blockade modulates the natural progression of leukocytes into the infarcted stroke bed. A better understanding of leukocyte spatiotemporal infiltration and its regulators could help inform the next generation of therapeutic interventions.

Elevated Cerebrospinal Fluid Protein Is Associated with Unfavorable Functional Outcome in Spontaneous Subarachnoid Hemorrhage.

BACKGROUND/OBJECTIVE: Subarachnoid hemorrhage (SAH) is a devastating neurologic event for which markers to assess poor outcome are needed. Elevated cerebrospinal fluid (CSF) protein may result from inflammation and blood-brain barrier (BBB) disruption that occurs during SAH. We sought to determine if CSF protein level is associated with functional outcome after SAH. METHODS: We prospectively collected single-center demographic and clinical data for consecutive patients admitted with spontaneous SAH. Inclusion required an external ventricular drain and daily CSF protein and cellular counts starting within 48 hours of symptom onset and extending through 7 days after onset. Seven-day average CSF protein was determined from daily measured values after correcting for contemporaneous CSF red blood cell (RBC) count. Three-month functional outcome was assessed by telephone interview with good outcome defined as modified Rankin score 0-3. Variables univariately associated with outcome at P less than .25 and measures of hemorrhage volume were included for binary logistic regression model development. RESULTS: The study included 130 patients (88% aneurysmal SAH, 69% female, 54.8 ± 14.8 years, Glasgow Coma Scale [GCS] 14 [7-15]). Three-month outcome assessment was complete in 112 (86%) patients with good functional outcome in 74 (66%). CSF protein was lower in good outcome (35.3 [20.4-49.7] versus 80.5 [40.5-115.5] mg/dL; P < .001). CSF protein was not associated with cerebral vasospasm, but delayed radiographic infarction on 3 to 12-month neuroimaging was associated with higher CSF protein (46.3 [32.0-75.0] versus 30.2 [20.4-47.8] mg/dL; P = .023). Good 3-month outcome was independently associated with lower CSF protein (odds ratios [OR] .39 [.23-.70] for 75th versus 25th percentile of protein; P = .001) and higher admission GCS (OR 1.23 [1.10-1.37] for good outcome per GCS point increase; P < .001). Parenchymal hematoma predicted worse outcome (OR 6.31 [1.58-25.25]; P = .009). Results were similar after excluding nonaneurysmal SAH and after including CSF RBC count, CT score, and intraventricular hemorrhage volume in models. CONCLUSIONS: Elevated average CSF protein is associated with poor outcome after spontaneous SAH. Further research should investigate if elevated CSF protein identifies patients in whom mechanisms such as BBB disruption contribute to poor outcome.

Congenital malformations in infants exposed to antiepileptic medications in utero at Boston Medical Center from 2003 to 2010.

OBJECTIVE: The aim of this study was to determine the frequency of association of major congenital malformations in pregnancy in women exposed to antiepileptic drugs (AEDs) in an inner city population. BACKGROUND: Approximately 0.3-0.5% of all pregnancies involve women with epilepsy. The risk of congenital malformations associated with AED therapy has been well documented, ranging from 2 to 10% as compared to a rate of 3% in the general population. However, the risk of these occurring in a higher risk population, such as an inner city tertiary care center, with multiple comorbidities is not as well known. DESIGN/METHODS: Using the Boston Medical Center Database between the years 2003 and 2010, a list of all infants born with major congenital malformations (MCMs) to mothers on AEDs was compiled. Major congenital malformations were defined as cleft lip and/or palate, ventricular or atrial septal defect, other cardiac malformations, and urogenital defects. During pregnancy, AED exposure including serum levels, other medication exposures, breakthrough seizure frequency, positive toxicology tests, and other maternal comorbidities were also analyzed. RESULTS: Of 17,246 live births between 2003 and 2010, 330 of those births demonstrated a MCM (malformation rate of 1.91%). Of those births, 64 mothers had epilepsy and were exposed to AED therapy during pregnancy, accounting for 0.37% of all births during this time period. Overall, three pregnancies in women with epilepsy resulted in a baby with a MCM, accounting for a 4.7% malformation rate in this patient population. In mothers on AEDs for other indications, the MCM rate was slightly higher, 5.0%, and in women on benzodiazepine monotherapy during pregnancy, the rate was quite high, 10.6%.

Evolving Clinical-Translational Investigations of Cerebroprotection in Ischemic Stroke.

Ischemic stroke is a highly morbid disease, with over 50% of large vessel stroke (middle cerebral artery or internal carotid artery terminus occlusion) patients suffering disability despite maximal acute reperfusion therapy with thrombolysis and thrombectomy. The discovery of the ischemic penumbra in the 1980s laid the foundation for a salvageable territory in ischemic stroke. Since then, the concept of neuroprotection has been a focus of post-stroke care to (1) minimize the conversion from penumbra to core irreversible infarct, (2) limit secondary damage from ischemia-reperfusion injury, inflammation, and excitotoxicity and (3) to encourage tissue repair. However, despite multiple studies, the preclinical-clinical research enterprise has not yet created an agent that mitigates post-stroke outcomes beyond thrombolysis and mechanical clot retrieval. These translational gaps have not deterred the scientific community as agents are under continuous investigation. The NIH has recently promoted the concept of cerebroprotection to consider the whole brain post-stroke rather than just the neurons. This review will briefly outline the translational science of past, current, and emerging breakthroughs in cerebroprotection and use of these foundational ideas to develop a novel paradigm for optimizing stroke outcomes.

Electroconvulsive Therapy (ECT) for Refractory Psychiatric Symptoms in Huntington's Disease: A Case Series and Review of the Literature.

BACKGROUND: Huntington's disease (HD) is a progressive neurodegenerative disease characterized by involuntary movements and neuropsychiatric decline. With suicide rates five times higher in patients with HD compared to the general population, there is a need for further research into the management of affective symptoms in these patients. Electroconvulsive therapy (ECT) has been long used as a treatment for severe or medication-refractory mood disorders and catatonia. There are some case studies demonstrating ECT's positive effect on depression and agitation in HD, but the data is limited. OBJECTIVE: In this single site case series, we review ECT use for four HD patients with medication-refractory depression and/or psychosis to better assess the utility of ECT in this population. We also compile and review the existing literature on the topic. METHODS: A single-center retrospective case series was conducted reviewing the indications, outcomes, and regimen of ECT treatments. Literature review was conducted via PubMed. RESULTS: Four patients received ECT treatment during an inpatient hospitalization with three continuing maintenance therapy as an outpatient. All four had improvements in depression, agitation, and suicidal ideation leading to successful hospital discharge. One of the four patients also demonstrated subjective improvement in cognitive and motor symptoms after ECT initiation. Nineteen reported cases were identified through the literature review and are summarized. CONCLUSIONS: This case series adds to the existing literature demonstrating the successful use of ECT for psychiatric symptoms in HD. Larger scale studies are warranted to further investigate the specific role and protocol for the use of ECT in the management of refractory depression and psychosis in this population.

Longitudinal deep-brain imaging in mouse using visible-light optical coherence tomography through chronic microprism cranial window.

We longitudinally imaged both the superficial and deep cortical microvascular networks in brains of healthy mice and in a mouse model of stroke in vivo using visible-light optical coherence tomography (vis-OCT). We surgically implanted a microprism in mouse brains sealed by a chronic cranial window. The microprism enabled vis-OCT to image the entire depth of the mouse cortex. Following microprism implantation, we imaged the mice for 28 days and found that that it took around 15 days for both the superficial and deep cortical microvessels to recover from the implantation surgery. After the brains recovered, we introduced ischemic strokes by transient middle cerebral artery occlusion (tMCAO). We monitored the strokes for up to 60 days and observed different microvascular responses to tMCAO at different cortical depths in both the acute and chronic phases of the stroke. This work demonstrates that the combined microprism and cranial window is well-suited for longitudinal investigation of cortical microvascular disorders using vis-OCT.

The use of metacognitive strategies to decrease false memories in source monitoring in patients with mild cognitive impairment.

Patients with amnestic mild cognitive impairment (MCI) and Alzheimer's disease (AD) often demonstrate high rates of false memories, leading to stressful and frustrating situations for both patients and caregivers in everyday life. Sometimes these false memories are due to failures in monitoring the source of the information. In the current study, we examined interventions aimed to enhance the use of the metacognitive "recall-to-reject" memory strategy. Such interventions could improve source memory and decrease false memory in patients with MCI. Because the picture superiority effect (better memory for pictures compared to words) has been shown to be present in both patients with MCI and healthy older controls, we investigated whether pictures could help patients with MCI use a recall-to-reject strategy in a simulation of real-world source memory task. In this experiment, patients with MCI and healthy older adults were asked to simulate preparing for and then taking a trip to the market. Subjects first studied 30 pictures of items in their "cupboard," followed by a list of 30 words of items on their "shopping list." At test, participants saw 90 pictures (30 cupboard, 30 list, 30 new) organized as they would be if walking down the market aisles, and are provided with either standard or metacognitive instructions. With standard instructions, they were asked if they needed to buy the item. With the metacognitive instructions, they were asked a series of questions to help guide them through a recall-to-reject strategy to highlight the different sources of memories. Results showed that the metacognitive instructions did significantly reduce the false memory rates for patients with MCI. Further studies need to investigate how to best implement these practical strategies into the everyday lives of patients.

Gelsolin is an endogenous inhibitor of syncytiotrophoblast extracellular vesicle shedding in pregnancy.

BACKGROUND: Preeclampsia, a pregnancy-specific inflammatory disorder, is characterized by high levels of anti-angiogenic protein, soluble fms-like tyrosine kinase 1 (sFlt1), in the maternal circulation. sFlt1 producing molecular machinery is present in syncytiotrophoblast extracellular vesicles that are released by the placenta into maternal plasma during normal pregnancy, a process greatly accelerated in preeclampsia. We hypothesized that syncytiotrophoblast extracellular vesicles exposes cytoplasmic actin to plasma resulting in depletion of plasma gelsolin (pGSN), an abundant plasma protein that scavenges circulating actin and other pro-inflammatory mediators. OBJECTIVE: To test whether pGSN levels would be lower in preeclampsia and to assess whether recombinant human plasma gelsolin (rhpGSN) may promote placental health by decreasing shedding of syncytiotrophoblast extracellular vesicles. METHODS: We tested pGSN levels in third trimester plasma samples from women with preeclampsia and non-hypertensive pregnancies. We then assessed whether rhpGSN may act as a negative regulator of syncytial shedding in placental explant culture and dynamic mechanical stretch studies. RESULTS: pGSN levels fall in late pregnancy and decline further in preeclampsia patients. Recombinant human pGSN (rhpGSN) at 100µg/ml limits spontaneous syncytiotrophoblast vesicle release and sFlt1 protein dissemination by normal placental explants. Higher rhpGSN doses (500µg/ml) also limit syncytiotrophoblast vesicle and sFlt1 dissemination from preeclamptic placental explants. rhpGSN also mitigates syncytiotrophoblast vesicle during dynamic mechanical stretch. CONCLUSIONS: 1) pGSN, an anti-inflammatory factor of maternal origin is reduced in preeclampsia and may contribute to disease progression and 2) exogenous rhpGSN supplementation can limit the dissemination of toxic syncytiotrophoblast vesicle that characterizes the disease state.