Hypothesis

Age‑dependent loss of specific gut‑derived metabolites (butyrate, secondary bile acids, low‑dose hydrogen sulfide) leads to epigenetic silencing and oxidative inactivation of mismatch repair (MMR) proteins in colonocytes, creating a field defect that precedes microsatellite instability and late‑onset colorectal cancer.

Mechanistic Basis

Butyrate inhibits histone deacetylases, keeping the MLH1 promoter open; secondary bile acids activate PPARγ, which transcriptionally upregulates MSH2 and MLH1; low‑dose H2S sustains Nrf2‑mediated antioxidant defenses that prevent oxidative MSH2 sulfenylation. Aging‑associated dysbiosis reduces these metabolites while increasing pro‑inflammatory microbes that raise nitric oxide and peroxynitrite, nitrosylating MSH6 and promoting proteasomal degradation. The combined epigenetic and redox pressure drives a coordinated decline of MMR capacity across the colonic epithelium, analogous to how gut‑derived signals set neuronal homeostasis Age-related microbiota shifts precede and shape neurological aging trajectories Aging‑induced gut dysbiosis drives enteric nervous system degeneration through inflammation and barrier dysfunction.

Testable Predictions

1. Old mice with induced dysbiosis will show decreased colonic butyrate, secondary bile acids, and H2S, accompanied by increased MLH1 promoter methylation and oxidized MSH2.
2. Restoring metabolite levels (butyrate gavage, bile‑acid supplementation, or low‑dose NaHS) will reverse MMR protein loss, reduce methylation, and lower spontaneous MSI in aged colons.
3. Fecal microbiota transplantation from young donors will rescue MMR expression and delay tumor onset in APC^Min/+ mice aged to 18 months.
4. In humans, fecal metabolomic profiles of low butyrate/secondary bile acids will predict higher blood‑based methylated MLH1 circulating DNA and increased risk of MMR‑deficient CRC in longitudinal cohorts.

Experimental Design

• Mouse model: C57BL/6 aged 20 months receive antibiotics to deplete microbiota, then colonize with either aged‑donor or young‑donor feces; parallel groups receive metabolite supplements.
• Readouts: Colonocyte isolation for MLH1/MSH2 western blot, promoter bisulfite sequencing, immunofluorescence for γH2AX, MSI PCR panel, and tumor count at 6 months post‑intervention.
• Human arm: Prospective cohort of 500 adults >60 y, serial stool metabolomics (GC‑MS/LC‑MS), plasma methylated MLH1 ELISA, and colonoscopy‑guided biopsies for MMR IHC; follow‑up for CRC incidence over 3 y.

Potential Confounds

Antibiotic pretreatment may itself affect epithelial turnover; we will include vehicle‑only controls and verify metabolite changes are microbiota‑dependent. Human observational links will need to adjust for diet, NSAID use, and comorbid inflammation.