Circadian GLP-1 Rhythms from Duodenal L-Cells Pace Thymic Output and Delay Immunosenescence
We hypothesize that the circadian-driven pulsatile secretion of GLP-1 from duodenal L-cells acts as a systemic timing cue that preserves thymic epithelial function and naive T‑cell output, and that loss of this rhythm accelerates immunosenescence. Recent work shows that gut hormone secretion, including GLP-1, is tightly regulated by circadian rhythms and that microbial metabolites like SCFAs act as timing cues influencing enteroendocrine function and host clocks [Frontiers Microbiology 2025]. The gastrointestinal tract also exhibits circadian control over digestion, absorption, and expression of nutrient transporters such as PEPT1 and NHE3 [PMC6533073]. Yet no study has examined whether the circadian amplitude of the molecular clock in duodenal L-cells declines with age, or how that impacts GLP-1 pulsatility and downstream thymic homeostasis.
Mechanistically, we propose that GLP-1 binds to GLP-1R on thymic epithelial cells (TECs), raising cAMP and activating PKA‑CREB signaling, which sustains FOXN1 expression—a master regulator of TEC differentiation and thymopoiesis. Concurrently, GLP-1‑stimulated cAMP enhances NAD+ synthesis via SIRT1 activation, reducing oxidative stress in TECs and preserving the cortical‑medullary balance required for positive and negative selection. When duodenal L‑cell clock genes (Bmal1, Clock, Per2) lose amplitude, GLP-1 secretion becomes arrhythmic, depriving TECs of timed trophic signals. This leads to attenuated FOXN1, increased senescence markers (p16^INK4a^, SASP), and a precipitous drop in recent thymic emigrants, thereby accelerating immunosenescence independent of systemic glucose intolerance.
The hypothesis is testable and falsifiable through three predictive experiments in mice:
- Feeding‑time rescue – Young (3‑mo) mice subjected to time‑restricted feeding (TRF) aligned with the active phase will show heightened Bmal1 expression in duodenal L‑cells (qPCR, immunoblot), increased amplitude of plasma GLP-1 pulses (ELISA sampled every 2 h over 24 h), and consequently greater thymic cellularity, higher CD4^+CD8^+ double‑positive counts, and richer TCR‑β repertoire diversity (immunosequencing) compared with ad‑libitum fed controls.
- L‑cell‑specific clock ablation – Mice with L‑cell‑restricted Bmal1 knockout (using Villin‑CreERT2 crossed to Bmal1^fl/fl^) will exhibit blunted GLP-1 rhythms despite normal feeding, premature reduction in thymic output (lower CD4^+SP and CD8^+SP TECs, decreased sjTREC levels), and accelerated appearance of aged‑phenotype T cells (increased PD‑1^+, KLRG1^+ CD8^+ T cells) by 12 mo, even when glucose tolerance remains intact.
- Timed GLP-1 agonist challenge – Aged (18‑mo) mice receiving a long‑acting GLP-1 receptor agonist (e.g., semaglutide) administered at the endogenous GLP-1 peak (ZT6) will restore FOXN1 expression in TECs, increase naive T‑cell output, and improve vaccine‑response metrics; the same agonist given at the opposite phase (ZT18) will fail to rescue thymic function, demonstrating that timing, not mere drug exposure, is critical.
Readouts will include duodenal L‑cell clock gene profiling (RT‑qPCR, Western), plasma GLP-1 dynamics (micro‑ELISA), thymic flow cytometry (Lineage^−CD45^+EpCAM^+ subsets, intracellular FOXN1), senescence assays (SA‑β‑gal, p16), and functional T‑cell readouts (TCR diversity, sjTREC, in‑vivo proliferation after antigenic challenge). If TRF or timed GLP-1 agonist restores thymic output while L‑cell clock ablation accelerates decline despite normal metabolism, the hypothesis is supported. Conversely, if manipulating L‑cell circadian machinery fails to alter thymic phenotypes or if thymic rescue occurs independent of GLP-1 rhythm, the hypothesis is falsified. This work would position the duodenal L‑cell clock‑GLP-1 axis as a previously unappreciated pacemaker linking circadian health to immune longevity.