Aging-related alterations in mechanistic target of rapamycin signaling promote platelet hyperreactivity and thrombosis
Irina Portier 1) 2), Bhanu Kanth Manne 1), Yasuhiro Kosaka 1), Neal D. Tolley 1), Frederik Denorme 1) 2) 3), Özgün Babur 4), Ashok P. Reddy 5), Phillip A. Wilmarth 5), Joseph E. Aslan 6), Andrew S. Weyrich 7), Matthew T. Rondina 1) 8) 9) 10), Robert A. Campbell 1) 2) 8) 9)
1-University of Utah Molecular Medicine Program, Salt Lake City, Utah, USA
2-Department of Emergency Medicine Washington University School, St. Louis, Missouri, USA
3-Division of Vascular Neurology, Department of Neurology, University of Utah, Salt Lake City, Utah, USA
4-Department of Computer Science, University of Massachusetts Boston, Boston, Massachusetts, USA
5-Proteomics Shared Resource, Oregon Health & Science University, Portland, Oregon, USA
6-Knight Cardiovascular Institute, Oregon Health & Science University, Portland, Oregon, USA
7-Oklahoma Medical Research Foundation, Oklahoma City, Oklahoma, USA
8-Division of Microbiology and Immunology, Department of Pathology, University of Utah, Salt Lake City, Utah, USA
9-Division of Hematology and Hematologic Malignancies, Department of Internal Medicine, University of Utah, Salt Lake City, Utah, USA
10-George E. Wahlen VAMC Department of Internal Medicine and the Geriatric Research, Education and Clinical Center, Salt Lake City, Utah, USA
Abstract
Background
Aging is an independent risk factor for the development of cardiovascular, thrombotic, and other chronic diseases. However, mechanisms of platelet hyperactivation in aging remain poorly understood.
Objectives
Here, we examine whether and how aging alters intracellular signaling in platelets to support platelet hyperactivity and thrombosis.
Methods
Quantitative mass spectrometry with tandem mass tag labeling systematically measured protein phosphorylation in platelets from healthy aged (>65 years) and young human (<45 years) subjects. The role of platelet mechanistic target of rapamycin (mTOR) in aging-induced platelet hyperreactivity was assessed using pharmacologic mTOR inhibition and a platelet-specific mTOR-deficient mouse model (mTORplt−/−).
Results
Quantitative phosphoproteomics uncovered differential site-specific protein phosphorylation within mTOR, Rho GTPase, and MAPK pathways in platelets from aged donors. Western blot confirmed constitutive activation of the mTOR pathway in platelets from both aged humans and mice, which was associated with increased aggregation compared with that in young controls. Inhibition of mTOR with either Torin 1 in aged humans or genetic deletion in aged mice reversed platelet hyperreactivity. In a collagen–epinephrine pulmonary thrombosis model, aged wild-type (mTORplt+/+) mice succumbed significantly faster than young controls, while time to death of aged mTORplt−/− mice was similar to that of young mTORplt+/+ mice. Mechanistically, we noted increased Rac1 activation and levels of mitochondrial reactive oxygen species in resting platelets from aged mice, as well as increased p38 phosphorylation upstream of thromboxane generation following agonist stimulation.
Conclusion
Aging-related changes in mTOR phosphorylation enhance Rac1 and p38 activation to enhance thromboxane generation, platelet hyperactivity, and thrombosis.