Aging-related alterations in mechanistic target of rapamycin signaling promote platelet hyperreactivity and thrombosis

J Thromb Haemost. 2024 Sep;22(9):2576-2588. doi: 10.1016/j.jtha.2024.05.025. Epub 2024 Jun 6.

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.

Keywords: aging; phosphoproteome; platelets; tandem mass tag; thrombosis.

MeSH terms

  • Adult
  • Age Factors
  • Aged
  • Aging*
  • Animals
  • Blood Platelets* / drug effects
  • Blood Platelets* / metabolism
  • Disease Models, Animal
  • Female
  • Humans
  • MTOR Inhibitors / pharmacology
  • Male
  • Mice
  • Mice, Inbred C57BL
  • Mice, Knockout
  • Middle Aged
  • Neuropeptides
  • Phosphorylation
  • Platelet Activation* / drug effects
  • Platelet Aggregation / drug effects
  • Proteomics / methods
  • Reactive Oxygen Species / metabolism
  • Signal Transduction*
  • TOR Serine-Threonine Kinases* / metabolism
  • Thrombosis* / blood
  • Thrombosis* / metabolism
  • rac1 GTP-Binding Protein* / metabolism

Substances

  • TOR Serine-Threonine Kinases
  • rac1 GTP-Binding Protein
  • MTOR protein, human
  • mTOR protein, mouse
  • Rac1 protein, mouse
  • RAC1 protein, human
  • Reactive Oxygen Species
  • MTOR Inhibitors
  • Neuropeptides