Empagliflozin and neohesperidin mitigate methotrexate hepatotoxicity via Nrf2/PPARγ/HO-1 signalling initiation and suppression of NF-κB/Keap1/HSP70/caspase-3 axis in rats.

Abstract Hepatotoxicity is the main adverse effect of methotrexate (MTX), which limits its clinical use and effectiveness. Both empagliflozin (EMPA) and neohesperidin dihydrochalcone (NHD) have promising criteria for suppressing oxidative stress, inflammation and apoptosis. In this current study, we suggested that EMPA and NHD exhibit protective effects against MTX-triggered liver injury, considering N-acetylcysteine (NAC) as a reference standard. In order to inspect our suggestion, An experimental rat model comprising 70 male adult rats (7 groups, 10 rats in each) was implemented to investigate the effects of MTX (20 mg/kg, i.p. once), alone or with EMPA (10 and 30 mg/kg/day, p.o.), NHD (40 and 80 mg/kg/day, p.o.), and NAC (150 mg/kg/day, p.o.) compared to the normal control animals (1%CMC, p.o.). Pre-treatment with EMPA and NHD showed significant attenuation in liver function abnormalities, pathological tissue deteriorations, hepatic oxidative stress parameters, and the level of expression of pro-inflammatory cytokines TNF-α and IL-6. Also, EMPA and NHD showed significant decreases in NF-κB/Keap1/HSP70/caspase-3 and increases in Nrf2/PPARγ/HO-1 expression levels. In addition, EMPA and NHD showed a marked enhancement of the anti-tumour activity of MTX against HepG2 and lung (A549) cancer cells. This research reveals that both EMPA and NHD can inhibit oxidation, inflammatory reactions, and apoptosis in the liver tissues of MTX-treated rats, mainly through Nrf2/PPARγ/HO-1 signalling initiation and suppression of NF-κB/Keap1/HSP70/caspase-3 axis, considered a unique class of drugs that attenuates or at least delays the onset of MTX-induced toxicity and serves as an innovative therapeutic target for future clinical application in humans.