Our hypothesis is that lymphocytes, PMNs and endothelial cells exhibit a """"""""proteolytic thermostat"""""""" which, in part, regulates the expression of cellular proteolytic activity via a series of diverse, complex sensing mechanisms comprised, in part, of selected MMP tethering molecules including MT1-MMP and CD44 and selected adhesion molecules including the Ig family member PECAM-1. In addition we postulate that these regulatory systems affect the functions of these cells at sites of inflammation. The goals of this project are to: 1. Characterize the signaling cascade(s) involved in the CD31 (PECAM-1)-mediated modulation of the induction of MMPs (MMP-2, MMP-14 [MT1-MMP] and MMP-9) in T lymphocytes and endothelial cells. We will determine the signaling cascade(s) initiated following PECAM-1 homophilic and heterophilic engagement. We will use WT and PECAM-1 null cells, PECAM-1 cytoplasmic domain truncation and exon deletion mutants and selected Y to F and S to C and S to D mutants known to affect PECAM-1-adaptor/ signaling molecule interactions. We will determine the levels of selected transcription factors known to be involved in modulating these MMPs. 2. Elucidate the roles of cell surface tethering molecules (MT1-MMP and CD44) as modulators of MMP-2 and MMP-9 expression. We will determine the signaling pathways following tethering of MMP-2 to the MT1- MMP complex and MMP-9 to CD44. We will determine MMP-2 and MMP-9 expression and activity levels in WT, MMP-2 null, MMP-9 null, MT1-MMP &null and CD44 null mice and endothelial cells and splenocytes isolated from these animals. Analyses will be performed on endothelial cells and lymphocytes following transfection with full-length, truncated and site-mutagenized MT1-MMP and CD 44 constructs. Signaling pathways will be elucidated using standard pharmacological and genetic approaches. We will determine the levels of selected transcription factors known to be involved in modulating these MMPs. 3. Elucidate the role(s) of MMPs as modulators of selected chemokine activities in experimental autoimmune encephalomyelitis (EAE). We will assess the abilities of MMP-2, -9 and 14 to proteolyze selected chemokines during the development of EAE in vivo and in lymphocyte and endothelial cell cultures. 4. We will test conclusions drawn from our in vitro data in our in vivo murine model of antigen-specific inflammation in, EAE. Specifically we will use WT, CD31 KO, MMP-2 KO, MMP-9 KO, MMP-14 and -/- and CD44 KO mice, and bone marrow chimeric mice generated from these mice.

National Institute of Health (NIH)
National Heart, Lung, and Blood Institute (NHLBI)
Research Project (R01)
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Study Section
Atherosclerosis and Inflammation of the Cardiovascular System Study Section (AICS)
Program Officer
Gao, Yunling
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Yale University
Schools of Medicine
New Haven
United States
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Flynn, Kelly M; Michaud, Michael; Madri, Joseph A (2013) CD44 deficiency contributes to enhanced experimental autoimmune encephalomyelitis: a role in immune cells and vascular cells of the blood-brain barrier. Am J Pathol 182:1322-36
Flynn, Kelly M; Michaud, Michael; Canosa, Sandra et al. (2013) CD44 regulates vascular endothelial barrier integrity via a PECAM-1 dependent mechanism. Angiogenesis 16:689-705
Li, Qi; Canosa, Sandra; Flynn, Kelly et al. (2013) Modeling the neurovascular niche: unbiased transcriptome analysis of the murine subventricular zone in response to hypoxic insult. PLoS One 8:e76265
Williams, Cicely; Rauch, Millicent Ford; Michaud, Michael et al. (2012) Short term interactions with long term consequences: modulation of chimeric vessels by neural progenitors. PLoS One 7:e53208
Murugesan, Nivetha; Demarest, Tyler G; Madri, Joseph A et al. (2012) Brain regional angiogenic potential at the neurovascular unit during normal aging. Neurobiol Aging 33:1004.e1-16
Frey, Merideth A; Michaud, Michael; VanHouten, Joshua N et al. (2012) Phosphorus-31 MRI of hard and soft solids using quadratic echo line-narrowing. Proc Natl Acad Sci U S A 109:5190-5
Li, Qi; Michaud, Michael; Canosa, Sandra et al. (2011) GSK-3ýý: a signaling pathway node modulating neural stem cell and endothelial cell interactions. Angiogenesis 14:173-85
Madri, J A (2009) Modeling the neurovascular niche: implications for recovery from CNS injury. J Physiol Pharmacol 60 Suppl 4:95-104
Li, Qi; Liu, Jaimei; Michaud, Michael et al. (2009) Strain differences in behavioral and cellular responses to perinatal hypoxia and relationships to neural stem cell survival and self-renewal: Modeling the neurovascular niche. Am J Pathol 175:2133-46
Long, Jennifer B; Jay, Steven M; Segal, Steven S et al. (2009) VEGF-A and Semaphorin3A: modulators of vascular sympathetic innervation. Dev Biol 334:119-32

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