As the second leading cause of death and disability, ischemic stroke kills and disables millions of people each year. Tissue plasminogen activator (TPA), the only approved treatment, dissolves the culprit fibrin thrombus to restore blood flow and relieve the brain from ischemia. Unfortunately, after prolonged ischemia, TPA may cause serious or fatal complications; this restricts TPA use to a minority of stroke patients. Although TPA has provided a model for therapeutic fibrinolysis, recent data suggest a new paradigm that assigns a central role to regulatory molecules such as alpha-2-antiplasmin (a2AP) in determining outcomes after ischemic stroke. Clinical observations suggest that high a2AP levels may increase the risk of ischemic stroke and of TPA failure. In experiments that challenge the current therapeutic paradigm for fibrinolytic treatment of stroke, we have shown that a2AP markedly increases brain injury, in a dose-dependent fashion. Conversely, a2AP deficiency or monoclonal antibody inactivation of a2AP, profoundly reduces brain injury, apoptosis, hemorrhage, and swelling. Even after prolonged brain ischemia, a2AP inactivation reduces microvascular thrombosis and MMP-9 expression (a marker of acute inflammation). As a result, a2AP inactivation prevents death and disability after ischemic stroke. Thus, when compared to TPA, a2AP-inactivation appears to provide a safe and effective approach for improving stroke treatment and, through a NINDS collaboration, we are pursuing the development of a2AP inactivation therapy. This proposal seeks to determine the pathophysiologic mechanisms through which a2AP enhances ischemic brain injury after thromboembolic stroke. The organizing hypothesis is that a2AP acts through plasminogen-dependent mechanisms to enhance the development of microvascular thrombosis and impair downstream, microvascular perfusion. Through these mechanisms, a2AP promotes the development of inflammatory responses such as MMP-9 expression and neutrophil recruitment, which have acute deleterious effects.
Aim 1 will test the hypothesis that a2AP's deleterious effects in ischemic stroke are due to diminished plasmin(ogen)-dependent, endogenous fibrinolysis that impairs microvascular blood flow through its effect on the culprit thrombus and the development of downstream, thrombin-dependent, microvascular thrombosis. We also propose to examine the hypothesis (Aim 2) that a2AP regulates the endogenous fibrinolytic system to affect the development of ischemic injury, hemorrhage, swelling and survival in thromboembolic stroke through inflammation-linked pathways that require MMP-9 activity and neutrophil deposition. Finally, we will use molecular complementation with specific a2AP mutants to examine the hypothesis that specific structural elements in the a2AP molecule selectively enhance adverse outcomes (such as neuronal injury, hemorrhage, etc.) in ischemic stroke.

Public Health Relevance

Stroke affects 15 million patients each year, leaving millions disabled or dead. We have discovered a novel therapeutic strategy for preventing brain damage, disability and death after experimental stroke. We seek to determine the mechanisms through which this novel therapy works in ischemic stroke, by examining its effects on inflammation, as well as on blood flow and thrombotic obstruction of small blood vessels.

National Institute of Health (NIH)
National Institute of Neurological Disorders and Stroke (NINDS)
Research Project (R01)
Project #
Application #
Study Section
Brain Injury and Neurovascular Pathologies Study Section (BINP)
Program Officer
Koenig, James I
Project Start
Project End
Budget Start
Budget End
Support Year
Fiscal Year
Total Cost
Indirect Cost
University of Tennessee Health Science Center
Internal Medicine/Medicine
Schools of Medicine
United States
Zip Code
Agarwal, Manyoo A; Garg, Lohit; Lavie, Carl J et al. (2018) Impact of family history of coronary artery disease on in-hospital clinical outcomes in ST-segment myocardial infarction. Ann Transl Med 6:3
Wang, Dong; Gladysheva, Inna P; Sullivan, Ryan D et al. (2018) Increases in plasma corin levels following experimental myocardial infarction reflect the severity of ischemic injury. PLoS One 13:e0202571
Agarwal, Manyoo A; Shah, Mahek; Patel, Brijesh et al. (2018) Association between Pulmonary Hypertension and Clinical Outcomes in Hospitalized Patients with Sickle Cell Disease. Am J Respir Crit Care Med 198:534-537
Singh, Satish; Houng, Aiilyan K; Reed, Guy L (2018) Matrix Metalloproteinase-9 Mediates the Deleterious Effects of ?2-Antiplasmin on Blood-Brain Barrier Breakdown and Ischemic Brain Injury in Experimental Stroke. Neuroscience 376:40-47
Zaidi, Syed S; Ward, Ryan D; Ramanathan, Kodangudi et al. (2018) Possible Enzymatic Downregulation of the Natriuretic Peptide System in Patients with Reduced Systolic Function and Heart Failure: A Pilot Study. Biomed Res Int 2018:7279036
Agarwal, Manyoo; Agrawal, Sahil; Garg, Lohit et al. (2018) Impact of smoking in patients undergoing transcatheter aortic valve replacement. Ann Transl Med 6:2
Agarwal, Manyoo; Agrawal, Sahil; Garg, Lohit et al. (2017) Effect of Chronic Obstructive Pulmonary Disease on In-Hospital Mortality and Clinical Outcomes After ST-Segment Elevation Myocardial Infarction. Am J Cardiol 119:1555-1559
Reed, Guy L; Houng, Aiilyan K; Singh, Satish et al. (2017) ?2-Antiplasmin: New Insights and Opportunities for Ischemic Stroke. Semin Thromb Hemost 43:191-199
Zambetti, Benjamin R; Thomas, Fridtjof; Hwang, Inyong et al. (2017) A web-based tool to predict acute kidney injury in patients with ST-elevation myocardial infarction: Development, internal validation and comparison. PLoS One 12:e0181658
Tripathi, Ranjana; Sullivan, Ryan; Fan, Tai-Hwang M et al. (2017) Enhanced heart failure, mortality and renin activation in female mice with experimental dilated cardiomyopathy. PLoS One 12:e0189315

Showing the most recent 10 out of 14 publications