Hypothesis

Telomerase‑high hepatocytes survive in pericentral Zone 3 only when Wnt/β‑catenin signaling simultaneously activates a ferroptosis‑defective program via upregulation of GPX4 and SLC7A11, thereby coupling regenerative capacity to lipid‑peroxide resistance.

Mechanistic Rationale

• Telomerase expression preserves replicative potential but does not intrinsically protect against iron‑dependent lipid peroxidation.
• Wnt/β‑catenin signaling in Zone 3 drives glutamine synthetase and Axin2, and recent data show it also induces the antioxidant response element (ARE) transcription factor NRF2, leading to increased GPX4 and SLC7A11 transcription.
• Consequently, telomerase‑high cells that receive Wnt input acquire a dual advantage: they can proliferate while avoiding ferroptosis, a major driver of Zone 3 hepatocyte loss in NAFLD/NASH.
• In the absence of Wnt, telomerase‑high cells proliferate but accumulate lipid peroxides, triggering ferroptosis and limiting their regenerative contribution.

Testable Predictions

1. Genetic ablation of β‑catenin specifically in telomerase‑high hepatocytes (using Tert‑CreERT2; Ctnnb1^fl/fl mice) will reduce GPX4 and SLC7A11 protein levels in Zone 3 and increase ferroptosis markers (PTGS2, 4‑HNE) after a high‑fat, high‑fructose diet, despite unchanged telomerase activity.
2. Pharmacologic Wnt activation (e.g., with a PORCN agonist) in telomerase‑high cells will rescue GPX4/SLC7A11 expression and attenuate ferroptosis even when senescence is induced by low‑dose doxorubicin.
3. Combining senolytic clearance of p21^high hepatocytes with transient Wnt agonism will expand the telomerase‑high pool and lower hepatic iron‑laden lipid droplets more effectively than senolytics alone, but only if GPX4 is intact; GPX4 heterozygous mice will not show this benefit.

Experimental Approach

• Generate Tert‑CreERT2; Rosa26‑LSL‑tdTomato; Ctnnb1^fl/fl mice to trace telomerase‑high cells and conditionally delete β‑catenin after tamoxifen induction.
• Feed mice a methionine‑choline‑deficient (MCD) diet for 8 weeks to induce NASH‑like pathology.
• Assess Zone 3 ferroptosis (PTGS2 immunoreactivity, lipid‑ROS C11‑BODIPY flow), telomerase activity (TRAP assay), and proliferation (Ki67/tdTomato co‑localization).
• Parallel arm: treat wild‑type NASH mice with dasatinib+quercetin senolytics followed by a short course of the Wnt agonist LGK974 withdrawal (to activate Wnt signaling) and measure GPX4/SLC7A11, telomerase‑high expansion, and fibrosis (Sirius Red).
• Include GPX4^+/‑ mutants to test dependency.

Potential Outcomes and Falsification

• If β‑catenin loss in telomerase‑high cells does not alter GPX4/SLC7A11 levels or ferroptosis susceptibility, the hypothesis is falsified, indicating Wnt’s role is independent of antioxidant programming.
• If Wnt agonist fails to rescue ferroptosis despite increasing telomerase‑high proliferation, the coupling is not operative.
• If senolytic plus Wnt expands telomerase‑high cells but ferroptosis markers remain high and GPX4 haploinsufficiency abolishes the protective effect, the hypothesis holds; otherwise, alternative mechanisms must be sought.

Broader Implication

Linking telomerase‑driven regenerative capacity to Wnt‑mediated ferroptosis resistance reframes Zone 3 vulnerability not as a passive metabolic bottleneck but as a regulatable checkpoint where enhancing the germline‑like editing budget requires concurrent redox shielding.