Mechanistic evaluation of thymol in a rat model of methotrexate-induced intestinal injury: NF-κB, Nrf2/HO-1, PPARγ/SIRT1, and RIPK1/RIPK3/MLKL pathways.

e24172 Background: Thymol is a monoterpene phenol with numerous pharmacological properties, including antibacterial, antitumor, anti-inflammatory, antidiabetic, antioxidant, and antirheumatic activities. The study was designed to explore the potential of thymol to protect against methotrexate (MTX)-induced intestinal injury in rats and to explore the associated molecular mechanisms. Methods: Rats were randomly allocated into five groups: Control, thymol (100mg/kg, p.o, for 10 days), MTX (20 mg/kg; i.p. single dose on day 5), thymol (60mg/kg, p.o, for 10 days) + MTX and thymol (100mg/kg, p.o, for 10 days) + MTX. Results: Thymol administration attenuated MTx-induced severe histological alterations and restored the architecture of intestinal villi and crypts. In addition, thymol significantly reduced intestinal oxidative damage by reducing lipid peroxidation and elevating superoxide dismutase (SOD) activity and glutathione (GSH) levels, as well as enhancing intestinal protein expression of Nrf2/HO-1, PPAR-γ, and SIRT1. Furthermore, thymol markedly reduced MTX-induced intestinal inflammation by decreasing IL-6 and TNF-α levels, which was associated with inhibition of NF-κB p65 activation. Moreover, thymol effectively inhibited necroptotic signaling in the intestine through downregulation of RIPK1/RIPK3/MLKL signaling and counteracted apoptosis via reduced cleaved caspase-3 and caspase-8 protein expression. Conclusions: Thymol may present a promising strategy for mitigating MTX-induced intestinal toxicity by exerting potent antioxidant, anti-inflammatory, and anti-necroptotic effects potentially mediated by modulating NF-κB p65, Nrf2/HO-1, PPAR-γ/SIRT1, RIPK1/RIPK3/MLKL signaling, and apoptosis.