NSAID-mediated silencing of nociceptive tone accelerates age‑associated UPR maladaptation

Chronic low‑grade pain signaling acts as a hormetic cue that keeps the PERK arm of the unfolded protein response (UPR) in a primed state, enabling selective translational attenuation and efficient chaperone induction when proteotoxic stress arises. Non‑steroidal anti‑inflammatory drugs (NSAIDs) blunt this tonic nociceptive input by inhibiting cyclooxygenase‑derived prostaglandins that normally stimulate EP2‑cAMP‑PKA signaling, a pathway known to enhance PERK autophosphorylation and eIF2α‑Ser51 phosphorylation[1]2]. In aged tissues, loss of this priming converts an adaptive UPR into an exaggerated but ineffective response: BiP induction becomes supra‑physiological (>3‑fold) while translational control shifts from selective to broad, diminishing the capacity to refold or degrade misfolded proteins.

Hypothesis – Long‑term NSAID exposure in middle‑aged mice will exacerbate the age‑related shift toward a maladaptive UPR, measured as increased BiP breadth, loss of translational selectivity, and accelerated accumulation of ubiquitinated protein aggregates, ultimately shortening healthspan.

Experimental design

• Groups (n=20 per group, male C57BL/6J, 12 months old): vehicle control, low‑dose ibuprofen (30 mg/kg/day), high‑dose ibuprofen (100 mg/kg/day), and a prostaglandin EP2 agonist (butaprost) co‑treated with high‑dose ibuprofen to rescue signaling.
• Duration – 6 months of continuous treatment.
• Readouts
  1. UPR activation – Western blot of lysates from liver and skeletal muscle for phospho‑PERK, total PERK, phospho‑eIF2α, ATF4, and BiP; quantify fold‑change vs. young (3‑month) baseline.
  2. Translational selectivity – Ribosome profiling to calculate the ratio of translationally attenuated mRNAs (UPR‑target chaperones) to globally suppressed transcripts; expect a decreased selectivity index in NSAID groups.
  3. Proteostasis capacity – Assay of chymotrypsin‑like proteasome activity and measurement of insoluble ubiquitinated proteins via filter‑trap assay.
  4. Physiological outcomes – Grip strength, treadmill endurance, frailty index, and survival curves.

Predictions

• Ibuprofen‑treated mice will show a statistically significant increase in BiP induction (>3‑fold vs. control) accompanied by a reduction in the translational selectivity index (<0.5 of control).
• Proteasome activity will decline and insoluble aggregates will rise in a dose‑dependent manner.
• Functional frailty will accelerate, and median lifespan will be reduced by ~15 % relative to vehicle.
• Co‑administration of the EP2 agonist will restore translational selectivity and attenuate BiP hyper‑induction, confirming that the effect proceeds via loss of prostaglandin‑mediated PERK priming.

Falsifiability – If NSAID treatment does not alter the BiP fold‑change, translational selectivity, or proteasome activity compared with vehicle, or if EP2 agonism fails to rescue the phenotype, the hypothesis is refuted. This design directly tests whether pharmacological silencing of nociceptive prostaglandin signaling compromises the hormetic UPR mechanism that sustains proteostasis during aging.