Aging and chronic inflammation are independent risk factors for the development of atherothrombosis and cardiovascular disease. We hypothesized that aging-associated inflammation promotes the development of platelet hyperreactivity and increases thrombotic risk during aging. Functional platelet studies in aged-frail adults and old mice demonstrated that their platelets are hyperreactive and form larger thrombi. We identified tumor necrosis factor α (TNF-α) as the key aging-associated proinflammatory cytokine responsible for platelet hyperreactivity. We further showed that platelet hyperreactivity is neutralized by abrogating signaling through TNF-α receptors in vivo in a mouse model of aging. Analysis of the bone marrow compartments showed significant platelet-biased hematopoiesis in old mice reflected by increased megakaryocyte-committed progenitor cells, megakaryocyte ploidy status, and thrombocytosis. Single-cell RNA-sequencing analysis of native mouse megakaryocytes showed significant reprogramming of inflammatory, metabolic, and mitochondrial gene pathways in old mice that appeared to play a significant role in determining platelet hyperreactivity. Platelets from old mice (where TNF-α was endogenously increased) and from young mice exposed to exogenous TNF-α exhibited significant mitochondrial changes characterized by elevated mitochondrial mass and increased oxygen consumption during activation. These mitochondrial changes were mitigated upon TNF-α blockade. Similar increases in platelet mitochondrial mass were seen in platelets from patients with myeloproliferative neoplasms, where TNF-α levels are also increased. Furthermore, metabolomics studies of platelets from young and old mice demonstrated age-dependent metabolic profiles that may differentially poise platelets for activation. Altogether, we present previously unrecognized evidence that TNF-α critically regulates megakaryocytes resident in the bone marrow niche and aging-associated platelet hyperreactivity and thrombosis.

Classic homocystinuria (HCU) is an inherited disorder characterized by elevated homocysteine (Hcy) in plasma and tissues resulting from cystathionine β‐synthase (CBS) deficiency. There is no cure, and patients are predominantly managed by methionine‐restricted diet (MRD) to limit the production of Hcy. In this study, we used the I278T mouse model of HCU to evaluate the long‐term impact of a novel enzyme replacement therapy [truncated human CBS C15S mutant modified with linear 20‐kDa N‐hydroxysuccinimide ester polyethylene glycol (OT‐58)] on clinical end points relevant to human patients with HCU. In addition, we compared its efficacy on a background of either MRD or normal methionine intake [regular diet (REG)] to that of MRD alone. We found that, compared with untreated I278T mice, OT‐58 treatment of I278T mice fed with the REG diet resulted in a 90% decrease in plasma Hcy concentrations and correction of learning/cognition, endothelial dysfunction, hemostasis, bone mineralization, and body composition. On background of the MRD, OT‐58 performed equally well with plasma Hcy entirely normalized. The MRD alone decreased plasma Hcy by 67% and corrected the HCU phenotype in I278T mice. However, the MRD increased anxiety and reduced bone mineral content in both I278T mice and wild‐type controls. This study shows that OT‐58 is a highly efficacious novel treatment for HCU on the background of either normal or restricted methionine intake.—Majtan, T., Park, I., Cox, A., Branchford, B. R., di Paola, J., Bublil, E. M., Kraus, J. P. Behavior, body composition, and vascular phenotype of homocystinuric mice on methionine‐restricted diet or enzyme replacement therapy. FASEB J. 33, 12477–12486 (2019). http://www.fasebj.org

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Severely elevated plasma homocysteine and methionine lead to thromboembolic events and strokes in homocystinuric (HCU) patients. Mouse models of HCU failed to exhibit prothrombotic phenotype, presumably due to lack of hypermethioninemia. We evaluated the impact of hypermethioninemia together with hyperhomocysteinemia on murine HCU phenotype and compared the efficacy of the current and novel therapies for HCU. High methionine intake decreased survival of I278T mice, which died from intestinal bleeding with hepatic and pancreatic failure. I278T mice on normal or increased methionine intake developed endothelial dysfunction, but paradoxically demonstrated delayed occlusion in an induced arterial thrombosis model. RNA-seq analysis suggested that expression of coagulation factor XI (FXI) is downregulated in livers of I278T mice. Indeed, plasma concentrations of FXI were decreased in I278T mice on normal diet and further reduced by increased methionine intake. Dietary methionine restriction normalized the observed phenotype. Similarly, treatment with OT-58, a novel enzyme therapy for HCU, corrected the phenotype in I278T mice regardless of their dietary methionine intake. Hypermethioninemia does not contribute to prothrombotic phenotype in murine HCU. Downregulation of FXI may contribute to the lack of prothrombotic tendency in I278T mice. Methionine restriction or treatment with OT-58 corrects vascular disease in the I278T mouse model of HCU.

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