It has been recognized that blood brain barrier (BBB) dysfunction exists in most neurological diseases, including stroke, multiple sclerosis, Alzheimer's and Parkinson's diseases, brain infections and epilepsy. Inflammation plays a significant role in BBB injury, secondary to pro-inflammatory factors produced in the brain or blood and leukocyte engagement of brain endothelium. Brain microvascular endothelial cells (BMVEC) are active participants and regulators of inflammatory processes at a site of inflammation. Inflammatory responses in brain endothelium involve hundreds of genes whose expression requires fine-tuned regulation. microRNAs (miRNAs) recently emerged as major regulators of gene expression. Very limited information exists about their participation in inflammatory responses caused by ischemia/reperfusion (I/R) events in brain endothelial cells. We propose to investigate the role of miRNAs in brain endothelium using well-established in vitro systems for functional studies of the BBB and imaging of brain microvasculature in an in vivo model of neuroinflammation. Based on our studies of BBB dysfunction during neuroinflammation, we propose that barrier protection is best achieved when the intervening agents possess anti-inflammatory properties and can stabilize tight junctions. Recently we have identified highly modified miRNAs, belonging to the let-7 miRNA family, which are important for endothelial maintenance. let-7 and miR-98 were predicted to target the inflammatory molecules, CCL2, CCL5, IL8 and IP10/CXCL10. Overexpression of let-7 and miR-98 in vitro and in vivo resulted in reduced leukocyte adhesion to and migration across brain endothelium and diminished expression of pro-inflammatory cytokines. In oxygen glucose deprivation (OGD) followed by reperfusion (OGD/R), an in vitro I/R model, overexpression of these miRNAs led to increased BBB tightness, thereby attenuating barrier `leakiness'. Overexpression of these miRNAs resulted in decreased infarct volume and neutrophil infiltration in the brain in tMCAO, an in vivo I/R stroke animal model. In our proposed study, we will test the overexpression or inhibition of selected miRNAs on BBB tightness and leukocyte-endothelial cell engagement (adhesion/migration). Using bioinformatics, we will identify other targets for miRNAs. Next, we will perform miRNA transfection in vivo and monitor how the presence of miRNAs will change leukocyte adhesion/migration in an animal model of stroke. Proposed experiments will provide identification and functional assessment of miRNAs in brain endothelium, and lead to future therapeutic developments for prevention of deleterious effects of ischemia/reperfusion on the brain.

Public Health Relevance

Endothelial dysfunction is perhaps the earliest event in the initiation of vascular disease caused by inflammation. In this work, we propose to use microRNA machinery as a therapeutic tool to prevent deleterious effects of brain ischemia/reperfusion stroke injury on endothelial dysfunction and neuroinflammation. The ability to prevent leukocyte adhesion and trafficking to the central nervous system (CNS) and to protect the blood brain barrier from increased permeability would prevent vascular malfunction, a common feature of several CNS disorders, such as stroke, Alzheimer's and Parkinson's diseases, multiple sclerosis, atherosclerosis and traumatic brain injury.

National Institute of Health (NIH)
National Institute of Neurological Disorders and Stroke (NINDS)
Research Project (R01)
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Brain Injury and Neurovascular Pathologies Study Section (BINP)
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Bosetti, Francesca
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Temple University
Schools of Medicine
United States
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Rom, Slava; Zuluaga-Ramirez, Viviana; Gajghate, Sachin et al. (2018) Hyperglycemia-Driven Neuroinflammation Compromises BBB Leading to Memory Loss in Both Diabetes Mellitus (DM) Type 1 and Type 2 Mouse Models. Mol Neurobiol :
Rom, Slava; Zuluaga-Ramirez, Viviana; Reichenbach, Nancy L et al. (2018) Secoisolariciresinol diglucoside is a blood-brain barrier protective and anti-inflammatory agent: implications for neuroinflammation. J Neuroinflammation 15:25