Hypothesis

Restoring mitochondrial coenzyme Q10 (CoQ10) biosynthesis specifically in pericentral (zone 3) hepatocytes will prevent ferroptosis‑driven progression of metabolic dysfunction‑associated steatotic liver disease (MASLD) in aging, independent of systemic iron overload.

Rationale

• Aging hepatocytes show a preferential loss of zonation markers (Ctnnb1, Foxo1, Tcf7l2) and an expansion of aberrant bi‑zonal cells that co‑express ASS1, CYP2E1 and GS [1].
• Zone 3 cells in aged MASLD livers accumulate lipids, undergo lipid peroxidation and display a ferroptotic‑senescent hybrid state termed ferrosenescence [2].
• Recent snRNA‑seq data reveal that the Aging Hepatocyte Gene Signature (AHGS) captures this ferrosenescent population and predicts advanced fibrosis with AUC > 0.80 [2].
• Mitochondrial dysfunction is an upstream driver of ferroptosis; reduced CoQ10 compromises electron transport, elevates superoxide, and fuels phospholipid peroxidation [3].
• In young livers, Wnt/β‑catenin signaling sustains COQ2 expression in zone 3; aging‑associated β‑catenin decline therefore predicts a zone‑specific CoQ10 deficit.

Novel Mechanistic Insight

We propose that the age‑dependent drop in Wnt/β‑catenin activity directly suppresses COQ2 transcription in pericentral hepatocytes, creating a localized bottleneck in ubiquinone synthesis. This mitochondrial CoQ10 shortage forces electrons to leak onto oxygen, generating ROS that peroxidize polyunsaturated fatty acids stored in lipid droplets. The resulting lipid hydroperoxides trigger ferroptosis, which concurrently activates senescence pathways (p16^INK4a^, SASP) and drives the AHGS phenotype. Restoring COQ2 only in zone 3 should re‑establish mitochondrial antioxidant capacity, quench lipid peroxidation, and break the feed‑forward loop between ferroptosis and senescence without altering hepatic iron pools.

Testable Predictions

1. Genetic rescue – Adeno‑associated virus (AAV8) carrying a hepatocyte‑specific, zone‑3‑targeted COQ2 construct (driven by a CYP2E1 promoter) administered to 18‑month‑old mice fed a methionine‑choline‑deficient (MCD) diet will:

   • Decrease hepatic MDA and 4‑HNE levels by ≥ 30 % compared with AAV‑control.
   • Lower AHGS‑positive hepatocyte proportion from ~15 % to ≤ 5 % (flow‑sorted snRNA‑seq).
   • Reduce fibrosis (Sirius Red area) by ≥ 40 % without changing non‑heme iron concentration (Perl’s stain).

2. Pharmacological corroboration – Treatment with exogenous ubiquinol‑10 (10 mg/kg/day, i.p.) in the same aged MCD model will phenocopy the genetic rescue, confirming that the effect is due to CoQ10 replenishment rather than off‑target AAV effects.

3. Falsification – If COQ2 overexpression fails to diminish MDA/4‑HNE, AHGS, or fibrosis despite confirmed transgene expression (qPCR, immunostaining), the hypothesis is refuted. Likewise, if systemic iron chelation (deferoxamine) adds no further benefit beyond COQ2 rescue, it supports the iron‑independent nature of the mechanism.

Experimental Outline (brief)

• Animals: Wild‑type C57BL/6J mice, 18 months old, n = 10/group.
• Interventions: AAV8‑CYP2E1‑COQ2 vs. AAV8‑CYP2E1‑GFP (control); oral MCD diet 8 weeks.
• Readouts: Liver non‑heme iron, lipid peroxidation (MDA, 4‑HNE immunoblot), ferroptosis markers (GPX4, ACSL4), senescence (p16, SASP cytokines), AHGS via snRNA‑seq, histology (steatosis, fibrosis).
• Statistical plan: Two‑tailed t‑test or ANOVA with post‑hoc Tukey; power = 0.8 to detect 25 % change in AHGS.

Implications

Confirming that a zone‑specific mitochondrial antioxidant deficit drives ferrosenescence would shift therapeutic focus from broad iron chelation to pericentral metabolic reprogramming. It would also provide a mechanistic bridge between Wnt/β‑catenin zonation loss and the ferroptosis‑senescence axis, offering a precision‑medicine avenue: patients with high AHGS and low hepatic CoQ10 (measurable via mass‑spec) could be enrolled in trials of liver‑targeted CoQ10 gene therapy or mitochondrially targeted ubiquinone analogs.