Hypothesis

Aging increases glucocorticoid receptor (GR) signaling in the ileum, which upregulates SGK1 and consequently ASBT more strongly in females than males. This sex‑biased hyperabsorption of conjugated primary bile acids raises luminal ammonia, compromises intestinal barrier integrity, and allows systemic ammonia to reach the brain, where it triggers microglial activation and synaptic loss. Simultaneous inhibition of ileal SGK1 and hepatic FGF19 signaling will restore enterohepatic bile acid homeostasis, reduce ammonia translocation, and improve cognitive function more effectively than either intervention alone.

Mechanistic Basis

1. GR‑SGK1 Axis: In aged ileum, elevated corticosterone binds GR, inducing Sgk1 transcription. SGK1 phosphorylates and stabilizes NHE3 and ASBT transporters, enhancing bile acid reuptake. Estrogen receptor‑α (ERα) can heterodimerize with GR, increasing its transcriptional activity in females, explaining the observed sex difference.
2. Bile Acid‑Ammonia Loop: Increased ASBT activity depletes luminal bile acids, shifting the equilibrium NH4⁺ ⇌ NH₃ + H⁺ toward ammonia. Higher luminal ammonia diffuses across the epithelium, especially when tight‑junction proteins (occludin, claudin‑5) are downregulated by SGK1‑mediated oxidative stress.
3. Brain Effects: Circulating ammonia crosses the blood‑brain barrier, astrocytes convert it to glutamine via glutamine synthetase, causing osmotic stress and excitotoxicity. Microglial NLRP3 inflammasome activation follows, leading to IL‑1β release and synaptic pruning in the hippocampus.
4. Hepatic Counterpart: Aging suppresses Cyp7a1 via increased FGF19 signaling from the ileum (via FGFR4), reducing bile acid synthesis and perpetuating the futile cycle.

Testable Predictions

• Female aged mice will show higher ileal SGK1 phosphorylation, ASBT expression, luminal ammonia, and serum conjugated primary bile acids than age‑matched males.
• Pharmacologic SGK1 inhibition or ileal‑specific Sgk1 knockout will normalize ASBT levels, reduce ammonia translocation, tighten the barrier (↑ ZO‑1, occludin), and lower hippocampal microglial activation in both sexes, with a greater magnitude of effect in females.
• Combining an SGK1 inhibitor with an FGF19 antagonist (to raise hepatic Cyp7a1 activity) will synergistically restore the bile acid pool, decrease serum ammonia, and improve performance in the Morris water maze beyond monotherapy.
• Administering an ERα antagonist (fulvestrant) to aged female mice will attenuate the glucocorticoid‑SGK1‑ASBT axis, reducing sex‑biased differences in bile acid absorption and cognitive decline.

Experimental Approach

1. Animal Models: Use 24‑month‑old C57BL/6J mice; generate ileal‑specific Sgk1 floxed mice crossed with Villin‑CreERT2 for inducible knockout. Include male and female cohorts, plus ERα antagonist treatment groups.
2. Interventions: (a) SGK1 inhibitor (GSK650394) via oral gavage; (b) FGF19 neutralizing antibody; (c) monotherapies and combined treatment.
3. Readouts:
   • qPCR/Western blot for Sgk1, phosphorylated SGK1, Asbt, Cyp7a1, Fgf15 (mouse FGF19 homolog).
   • Bile acid profiling (LC‑MS) of serum, feces, and ileal content.
   • Luminal ammonia assay (enzymatic) and serum ammonia.
   • Intestinal permeability (FITC‑dextran) and tight‑junction immunofluorescence.
   • Hippocampal cytokine levels (ELISA) and microglial morphology (Iba1 staining).
   • Cognitive testing (Morris water maze, novel object recognition).
4. Statistical Analysis: Two‑way ANOVA (sex × treatment) with post‑hoc Tukey; significance set at p<0.05.

If the data confirm that ileal SGK1 drives sex‑specific ASBT upregulation and that dual targeting of intestinal transport and hepatic synthesis rescues cognitive decline, the hypothesis will be supported. Conversely, if SGK1 manipulation fails to affect ammonia levels, barrier integrity, or cognition, or if combining therapies shows no added benefit, the hypothesis will be falsified.