Exosomes are from the intraluminal vesicles (ILV) of multivesicular bodies (MVBs) produced via endocytic process. Mature MVBs can fuse with lysosomes to deliver their contents for degradation and recycling or fuse with the plasma membrane where ILVs are released as exosomes. Recent studies have indicated that enhanced exosome excretion in arterial smooth muscle cells (SMCs) is an essential mechanism triggering or promoting calcifying nidus formation and extracellular matrix (ECM) mineralization in the arterial wall under different pathological conditions. However, little is known so far how MVB fate and exosome excretion are controlled by lysosomes in SMCs and whether lysosomal dysfunction increases exosome excretion from SMCs to activate or accelerate vascular calcification process. The present grant proposal will test a central hypothesis that lysosomal acid ceramidase (AC)-mediated sphingolipid metabolism plays a crucial role in the control of lysosome trafficking or fusion to MVBs and subsequent exosome excretion, maintaining the normal phenotype and function of SMCs. AC gene defect or functional deficiency may disturb lysosome degradation of MVBs, increasing exosome excretion and resulting in calcifying nidus formation and ultimate vascular calcification under pathological conditions. To test this hypothesis, three Specific Aims are proposed.
Specific Aim 1 will determine whether exosome excretion in arterial SMCs is fine controlled by lysosomal AC activity and whether the deficiency of this AC regulation causes arterial calcification in smooth muscle-specific AC gene knockout mice (Asah1fl/fl/SMcre) with analysis of exosomes, SMC phenotypes, and calcification in the arterial wall and cultured coronary arterial SMCs.
Specific Aim 2 attempts to test whether lysosomal AC- mediated sphingolipid signaling regulates lysosome trafficking to and fusion with MVBs to limit exosome excretion from SMCs with gene deletion (Asah1fl/fl/SMcre), CRISPR-Cas9 gene editing, and SM22? promoter- driven gene rescuing.
In Specific Aim 3, we will address whether the AC regulation of lysosome trafficking or fusion to MVBs is attributed to its action on lysosomal TRPML1 channel activity and associated Ca2+ release using patch clamping of isolated lysosomes and lysosome-specific Ca2+ imaging with GCaMP3-ML as an indicator. To our knowledge, these proposed studies will represent the first effort to investigate the lysosome regulation of exosome excretion from SMCs and associated pathogenic role in vascular calcification. The findings will provide new insights into the pathogenesis of arterial calcification and identify lysosomal AC as a therapeutic target for prevention or treatment of vascular calcification.

Public Health Relevance

Exosomes are microparticles released from many animal and human cells, which contain various bioactive lipids, microRNAs or proteins, which contribute to calcium deposition in the artery wall, hardening the arteries and thereby causing coronary artery disease and heart attack. This grant application seeks to examine how exosome release is controlled in these artery cells and to investigate whether dysregulation of exosome release is a cause of calcium deposition and consequent hardening of the arteries. We will demonstrate that acid ceramidase, a lipid metabolizing enzyme within cells, plays a crucial role in the control of exosome release and its defect may result in exosome accumulation and calcification in the artery wall, possibly helping develop new therapy in the future for more effective prevention or treatment of coronary arterial disease and heart attack.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Project (R01)
Project #
5R01HL057244-22
Application #
9859425
Study Section
Atherosclerosis and Inflammation of the Cardiovascular System Study Section (AICS)
Program Officer
Chen, Jue
Project Start
1997-01-01
Project End
2022-01-31
Budget Start
2020-02-01
Budget End
2021-01-31
Support Year
22
Fiscal Year
2020
Total Cost
Indirect Cost
Name
Virginia Commonwealth University
Department
Pharmacology
Type
Schools of Medicine
DUNS #
105300446
City
Richmond
State
VA
Country
United States
Zip Code
23298
Yuan, Xinxu; Bhat, Owais M; Meng, Nan et al. (2018) Protective Role of Autophagy in Nlrp3 Inflammasome Activation and Medial Thickening of Mouse Coronary Arteries. Am J Pathol 188:2948-2959
Chen, Yu; He, Xingxiang; Yuan, Xinxu et al. (2018) NLRP3 Inflammasome Formation and Activation in Nonalcoholic Steatohepatitis: Therapeutic Target for Antimetabolic Syndrome Remedy FTZ. Oxid Med Cell Longev 2018:2901871
Yuan, Xinxu; Wang, Lei; Bhat, Owais M et al. (2018) Differential effects of short chain fatty acids on endothelial Nlrp3 inflammasome activation and neointima formation: Antioxidant action of butyrate. Redox Biol 16:21-31
Bhat, Owais M; Yuan, Xinxu; Li, Guangbi et al. (2018) Sphingolipids and Redox Signaling in Renal Regulation and Chronic Kidney Diseases. Antioxid Redox Signal :
Li, Pin-Lan; Gulbins, Erich (2018) Bioactive Lipids and Redox Signaling: Molecular Mechanism and Disease Pathogenesis. Antioxid Redox Signal :
Bao, Jun-Xiang; Zhang, Qin-Fang; Wang, Mi et al. (2017) Implication of CD38 gene in autophagic degradation of collagen I in mouse coronary arterial myocytes. Front Biosci (Landmark Ed) 22:558-569
Li, Guangbi; Chen, Zhida; Bhat, Owais M et al. (2017) NLRP3 inflammasome as a novel target for docosahexaenoic acid metabolites to abrogate glomerular injury. J Lipid Res 58:1080-1090
Conley, Sabena M; Abais-Battad, Justine M; Yuan, Xinxu et al. (2017) Contribution of guanine nucleotide exchange factor Vav2 to NLRP3 inflammasome activation in mouse podocytes during hyperhomocysteinemia. Free Radic Biol Med 106:236-244
Koka, Saisudha; Xia, Min; Chen, Yang et al. (2017) Endothelial NLRP3 inflammasome activation and arterial neointima formation associated with acid sphingomyelinase during hypercholesterolemia. Redox Biol 13:336-344
Xu, Xiaoyang; Zhang, Aolin; Halquist, Matthew S et al. (2017) Simvastatin promotes NPC1-mediated free cholesterol efflux from lysosomes through CYP7A1/LXR? signalling pathway in oxLDL-loaded macrophages. J Cell Mol Med 21:364-374

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