iNOS Mediates High-Fat Diet-Associated Aggravation of Complete Freund's Adjuvant-Induced Inflammatory Pain

Chronic inflammatory pain (IP) remains a therapeutic challenge under the worldwide prevalence of the high-fat dietary lifestyle. This study aimed at identifying mediators of the IP augmented by short-term high-fat diet (HFD). IP was induced on C57BL/6J mice by unilateral, intra-plantar, injection of Complete Freund’s Adjuvant (CFA). Von Frey test for mechanical hyperalgesia and Hargreaves’ test for thermal hyperalgesia were performed at pre-injection baseline and post-injection 6th h. and days 1/3/5/7/10/14. Ad libitum HFD feeding started 2 weeks pre-injection in assigned groups. Body weight and random blood glucose levels were measured. RT-qPCR and ELISA helped quantify expression levels of the selected candidate genes at manipulated hind-paws. After CFA injection, at 1400 W, a highly selective inducible nitric oxide synthase (iNOS) inhibitor was administered regularly to elicit differences in CFA-induced pain behaviors and gene expression in HFD-fed mice. Results showed that HFD-fed mice were heavier (p < 0.001) and relatively hyperglycemic (p = 0.013) at baseline. HFD aggravated CFA-induced mechanical and thermal pain (mechanical: p = 0.0004, thermal: p = 0.003), showing prolonged hyperalgesic durations and reduced pain thresholds at multiple timepoints. HFD-influenced paws showed accentuated overexpression of pro-inflammatory cytokines and iNOS (RT-qPCR for IL-1β: p = 0.015, IL-6: p = 0.019, TNF: p = 0.04; ELISA for iNOS: p = 0.011). At 1400 W, exertion of analgesic effects (mechanical: p < 0.0001, thermal: p < 0.0001) but pro-inflammatory (RT-qPCR for IL-1β: p = 0.004, IL-6: p = 0.03, TNF: p = 0.04) were exerted on the inflamed paw on day 5 post-injection. In conclusion, short-term HFD aggravated CFA-induced inflammatory pain. Pharmacological inhibition of iNOS attenuated the CFA-induced pain in HFD-fed mice. Future research might uncover signaling pathways mediating such effects, potentially benefiting obese patients with chronic IP.

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