Atheroprone flow-induced endothelial dysfunction, characterized by enhanced inflammatory response, is an early vascular event leading to the focal distribution of atherosclerosis. Despite the causality between atheroprone flow pattern and endothelial dysfunction, key molecular events linking mechanical stimuli to the pro-inflammatory phenotype of vascular endothelial cells (ECs) and the consequent susceptibility to atherogenesis remain poorly understood. We recently demonstrated that atheroprone flow activates sterol regulatory element binding protein 2 (SREBP2) in ECs, which induces the NOD-like receptor family, pyrin domain containing 3 (NLRP3) inflammasome through transcriptional induction (i.e., Signal 1) and higher-order assembly (i.e., Signal 2). Such unresolved EC innate immune response leads to endothelial dysfunction and ensuing atherosclerosis. We have also identified that atheroprone flow induces TRAF-interacting protein with a forkhead-associated (FHA) domain (TIFA). Newly performed experiments indicate that SREBP2 transcriptionally upregulates TIFA and inflammasome components (Signal 1), and that phosphorylated TIFA Threonine-9 (TIFA pT9) oligomerizes TIFA leading to the higher-order assembly of NLRP3 inflammasome (Signal 2). These observations provide the foundation for the guiding hypothesis that atheroprone flow, through transcriptional induction and post-translational modification of TIFA, activates NLRP3 inflammasome in the endothelium, which contributes to atherosclerosis susceptibility.
Three Specific Aims are proposed to test this hypothesis.
Specific Aim 1 will elucidate the molecular mechanism by which atheroprone flow primes the induction of the NLRP3 inflammasome through Signal 1. In vitro flow channel experiments with oscillatory shear stress (OS) simulating atheroprone flow will be used to elucidate the transcriptional cascade of SREBP2-TIFA-NLRP3 inflammasome that respond to atheroprone flow.
Specific Aim 2 will delineate the molecular basis by which atheroprone flow activates the NLRP3 inflammasome via Signal 2. Specifically, mechanobiology and FRET-based bioimaging will be used to investigate the role of TIFA pT9 in the higher-order assembly of NLRP3 inflammasome in ECs in response to atheroprone flow.
Specific Aim 3 will investigate the role of atheroprone flow-activated SREBP2-TIFA-NLRP3 inflammasome in endothelial dysfunction, leading to atherosclerosis in vivo. We will examine endothelial function, vascular inflammation, and atherosclerosis in EC-SREBP2-/-, TIFA-/-, and TIFA T9A knock-in mice with or without an ApoE-/- background. The proposed experimental approaches that combine vascular, mechanobiological, biosensor, and animal experiments will systemically analyze the mechano and molecular basis of atheroprone flow-induced innate immune response in ECs and its functional relevance to atherosclerosis susceptibility.

Public Health Relevance

Atherosclerosis is a major vascular inflammatory disease that localizes in specific regions where blood flow is disturbed. We propose to investigate the molecular mechanism by which disturbed flow causes endothelial cell dysfunction, which contributes to atherosclerosis. The obtained results will provide new information on the etiology of atherosclerosis and knowledge for disease prevention, treatment, and management.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Project (R01)
Project #
1R01HL125643-01A1
Application #
9185542
Study Section
Vascular Cell and Molecular Biology Study Section (VCMB)
Program Officer
Olive, Michelle
Project Start
2016-08-05
Project End
2020-07-31
Budget Start
2016-08-05
Budget End
2017-07-31
Support Year
1
Fiscal Year
2016
Total Cost
$437,888
Indirect Cost
$150,712
Name
University of California San Diego
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
804355790
City
La Jolla
State
CA
Country
United States
Zip Code
92093
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