Kenneth Gaines

The Novel Methods for Analysis of Exosomes Released from Endothelial Cells and Endothelial Progenitor Cells

Exosomes (EXs) are cell-derived vesicles that mediate cell-cell communication and could serve as biomarkers. Here we described novel methods for purification and phenotyping of EXs released from endothelial cells (ECs) and endothelial progenitor cells (EPCs) by combining microbeads and fluorescence quantum dots (Q-dots®) techniques. EXs from the culture medium of ECs and EPCs were isolated and detected with cell-specific antibody conjugated microbeads and second antibody conjugated Q-dots by using nanoparticle tracking analysis (NTA) system. The sensitivities of the cell origin markers for ECs (CD105, CD144) and EPCs (CD34, KDR) were evaluated. The sensitivity and specificity were determined by using positive and negative markers for EXs (CD63), platelets (CD41), erythrocytes (CD235a), and microvesicles (Annexin V). Moreover, the methods were further validated in particle-free plasma and patient samples. Results showed that anti-CD105/anti-CD144 and anti-CD34/anti-KDR had the highest sensitivity and specificity for isolating and detecting EC-EXs and EPC-EXs, respectively. The methods had the overall recovery rate of over 70% and were able to detect the dynamical changes of circulating EC-EXs and EPC-EXs in acute ischemic stroke. In conclusion, we have developed sensitive and specific microbeads/Q-dots fluorescence NTA methods for EC-EX and EPC-EX isolation and detection, which will facilitate the functional study and biomarker discovery.

Analyses of Endothelial Cells and Endothelial Progenitor Cells Released Microvesicles by Using Microbead and Q-Dot Based Nanoparticle Tracking Analysis

Accurate analysis of specific microvesicles (MVs) from biofluids is critical and challenging. Here we described novel methods to purify and detect MVs shed from endothelial cells (ECs) and endothelial progenitor cells (EPCs) by combining microbeads with fluorescence quantum dots (Q-dots) coupled nanoparticle tracking analysis (NTA). In the in vitro screening systems, we demonstrated that 1) anti-CD105 (EC marker) and anti-CD34 (EPC marker) conjugated-microbeads had the highest sensitivity and specificity for isolating respective MVs, which were confirmed with negative controls, CD41 and CD235a; 2) anti-CD144 (EC marker) and anti-KDR (EPC marker) conjugated-Q-dots exhibited the best sensitivity and specificity for their respective MV NTA detection, which were confirmed with positive control, anti-Annexin V (MV universal marker). The methods were further validated by their ability to efficiently recover the known amount of EC-MVs and EPC-MVs from particle-depleted plasma, and to detect the dynamical changes of plasma MVs in ischemic stroke patients, as compared with traditional flow cytometry. These novel methods provide ideal approaches for functional analysis and biomarker discovery of ECs- and EPCs- derived MVs.

A stroke/vascular neurology service increases the volume of urgent carotid endarterectomies performed in a tertiary referral center.

BACKGROUND Increasing evidence supports that urgent carotid endarterectomy (CEA), defined as CEA during the index hospitalization, may be undertaken in select patients with acute carotid-related neurologic symptoms to prevent recurrent ischemic events. We aimed to determine the effect of a stroke/vascular neurology service on the volume of urgent CEAs performed and assess perioperative outcomes. METHODS A retrospective review from a single tertiary referral center between June 2005 through December 2011 revealed 393 patients who underwent CEA. We identified the number of urgent CEAs before (June 2005-August 2008) and after (September 2008-December 2011) a stroke/vascular neurology service was implemented, as well as asymptomatic CEAs and symptomatic but electively performed CEAs. Demographic data as well as 30-day adverse outcomes (transient ischemic attack [TIA], stroke, myocardial infarction, and mortality) were analyzed for each group. In patients undergoing urgent CEA, TIA and stroke severity were assessed by a stroke neurologist using the ABCD2 TIA score and National Institutes of Health Stroke Scale (NIHSS), respectively. The χ2 test was used to compare differences between the urgent CEA volume before and after a stroke/vascular neurology service. Fishers exact test was used to analyze perioperative outcomes. RESULTS Demographics and comorbidities were similar between the 2 groups. The proportion of urgent CEAs performed increased significantly after initiation of a vascular neurology service (4.1% [7 of 172] vs. 22.2% [49 of 221], P<0.0001). Per annum, urgent CEAs increased from 5.3% (4/75) in 2005 to 39.6% (25/63) in 2011. A vascular neurology service did not increase the number of nonurgent referrals. Urgent CEA indications were ocular ischemic events 4% (2/49), cerebral ischemic/infarction events 35% (17/49), crescendo TIAs 6% (3/49), acute stroke 45% (22/49), and stroke-in-evolution 10% (5/49). Mean NIHSS was 3.5 (range 0-24); mean TIA score was 5 (range 1-8). Although there were no statistical differences in 30-day outcomes, there was a trend toward a higher combined complication rate (stroke, death, myocardial infarction) in the urgent compared with the symptomatic but electively performed CEA group (7.1 % [3/49] vs. 2% [1/49]; P=.36). However, patients undergoing urgent CEA with an NIHSS<10 had no perioperative complications. CONCLUSIONS Collaboration with a vascular neurology team increased the volume of urgent CEAs over a 3-year period. In patients with mild-to-moderate strokes (NIHSS<10), urgent CEA perioperative outcomes approximate those for electively performed CEAs, suggesting improved care through a multidisciplinary approach.

Implementing telestroke to reduce the burden of stroke in Louisiana

Cardiovascular diseases (heart diseases and stroke) are the leading cause of death in the United States. People living in rural areas have disproportionately high mortality rates due to stroke mainly due to lack of specialty services. Timely intervention is a critical factor in determining the prognosis for acute stroke and thousands of lives can be saved by recognizing/treating a stroke patient within an hour. This paper deals with telestroke, a unique intervention implemented in Louisiana that incorporated latest technology to integrate with advanced clinical protocols in treating acute stroke. People living in remote and rural areas of Louisiana were greatly benefited from this initiative and several lives saved as a result of timely intervention. Expanding these initiatives to larger populations and other states will greatly reduce the stroke mortality.

Stroke: the critically neglected first year post-stroke

Purpose Stroke is a leading cause of death and disability in the USA and worldwide. While stroke care has evolved dramatically, many new acute approaches to therapy focus only on the first 3-12 hours. Significant treatment opportunities beyond the first 12 hours can play a major role in improving outcomes for stroke patients. The purpose of this paper is to highlight the issues that affect stroke care delivery for patients and caregivers and describe an integrated care model that can improve care across the continuum. Design/methodology/approach This paper details evidence-based research that documents current stroke care and efforts to improve care delivery. Further, an innovative integrated care model is described, and its novel application to stroke care is highlighted. Findings Stroke patients and caregivers face fragmented and poorly coordinated care systems as they move through specific stroke nodes of care, from acute emergency and in-hospital stay through recovery post-discharge at a care facility or at home, and can be addressed by applying a comprehensive, technology-enabled Integrated Stroke Practice Unit (ISPU) Model of Care. Originality/value This paper documents specific issues that impact stroke care and the utilization of integrated care delivery models to address them. Evidence-based research results document difficulties of current care delivery methods for stroke and the impact of that care delivery on patients and caregivers across each node of care. It offers an innovative ISPU model and highlights specific tenets of that model for readers.

Description of a novel telemedicine-enabled comprehensive system of care: drip and ship plus drip and keep within a system of stroke care delivery.

United States (US) and worldwide telestroke programs frequently focus only on emergency room hyper-acute stroke management. This article describes a comprehensive, telemedicine-enabled, stroke care delivery system that combines “drip and ship” and “drip and keep” models with a comprehensive stroke center primary hub at Ochsner Medical Center in New Orleans, advanced stroke-capable regional hubs, and geographically-aligned, “stroke-ready” spokes. The primary hub provides vascular neurology expertise via telemedicine and monitors care for patients remaining at regional hubs and spokes using a multidisciplinary team approach. By 2014, primary hub telestroke consults grew to ≈1000/year with 16 min average door to consult initiation and 20 min to completion, and 29% of ischemic stroke patients received recombinant tissue-type plasminogen activator (rtPA), increasing 275%. Most patients remained in hospitals close to home, but neurointensive care and interventional procedures were common reasons for primary hub transfer. Given the time sensitivity and expert consultation needed for complex acute stroke care delivery paradigms, telestroke programs are effective for fulfilling unmet care needs. Combining drip and ship and drip and keep management allows more patients to stay “local,” limiting primary hub transfer unless more advanced services are required. Post admission telestroke management at spokes increases personnel efficiency and can positively impact stroke outcomes.