Hypothesis

Age‑related NAD+ depletion triggers a metabolic downgrade in hepatocytes that elevates synthesis and secretion of Factor VIII (FVIII) and von Willebrand Factor (VWF) as a compensatory hemostatic set‑point shift, preparing the organism for increased bleeding risk when repair capacity is limited.

Mechanistic Rationale

NAD+ is a obligate cofactor for sirtuin‑1 (SIRT1) and SIRT6, which deacetylate transcription factors controlling hepatic acute‑phase gene expression. When NAD+ falls, SIRT activity wanes, leading to hyperacetylation of NF‑κB and STAT3, thereby increasing transcriptional drive for FVIII and VWF genes. Concurrently, low NAD+ reduces mitochondrial oxidative phosphorylation, raising the cytosolic NADH/NAD+ ratio and activating hypoxia‑inducible factor‑1α (HIF‑1α), which further stimulates VWF transcription via hypoxia‑response elements. The combined effect is a coordinated increase in procoagulant factor output that does not require inflammatory cytokines but reflects a cell‑autonomous response to diminished energetic budget.

Testable Predictions

1. Hepatic NAD+ levels will inversely correlate with plasma FVIII and VWF concentrations in cross‑sectional human cohorts aged 40‑80 y, after adjusting for CRP and fibrinogen.
2. Pharmacological restoration of NAD+ (e.g., with nicotinamide riboside) in aged mice will reduce hepatic SIRT1/6 deacetylation activity, lower FVIII/VWF mRNA, and decrease circulating factor levels without altering inflammatory markers.
3. Liver‑specific SIRT1 or SIRT6 knockout in young mice will recapitulate the aged procoagulant phenotype (elevated FVIII/VWF, increased thrombin generation) despite normal NAD+ pools.
4. Chromatin immunoprecipitation will show increased NF‑κB and STAT3 occupancy at the F8 and VWF promoters in NAD‑deficient hepatocytes, reversible by NAD+ repletion.

Experimental Approach

• Human cohort: Measure plasma NAD+ (via LC‑MS/MS), FVIII, VWF, high‑sensitivity CRP, and fibrinogen in 200 volunteers stratified by decade. Use multivariable regression to test the NAD+‑FVIII/VWF association.
• Mouse intervention: Treat 20‑month‑old C57BL/6 mice with nicotinamide riboside (400 mg/kg/day) or vehicle for 8 weeks. Collect liver tissue for NAD+ quantification, SIRT activity assays, qPCR of F8 and VWF, and plasma thrombin‑generation assays (calibrated automated thrombography).
• Genetic model: Generate Alb‑Cre;Sirt1^fl/fl and Alb‑Cre;Sirt6^fl/fl mice. At 3 months, assess the same hemostatic endpoints to determine if sirtuin loss mimics aging.
• Chromatin assays: Perform ChIP‑qPCR for acetylated‑H3K27, NF‑κB p65, and STAT3 on isolated hepatocytes from each group.

If NAD+ restoration normalizes FVIII/VWF levels independent of inflammation, it would support the hypothesis that the age‑related procoagulant shift is a metabolically programmed adaptation rather than indiscriminate damage accumulation. Conversely, a lack of effect would falsify the proposed mechanistic link, directing focus toward alternative drivers of coagulation remodeling.