Supplementary Material for: Alpinia galanga essential oil mitigates gentamycin-induced renal injury: in silico and in vivo studies

Introduction: Alpinia galanga (L.) rhizome has been widely consumed as a spice and food-flavoring agent, and in traditional medicines for the treatment of various diseases such as stomach pain, vomiting, diarrhea, infections and renal disorders. The clinical use of Gentamicin (Genta), an aminoglycoside antibiotic, is constrained due to nephrotoxicity. The existing investigation intended to estimate the mechanistic antioxidant, anti-inflammatory, and antiapoptotic actions of Galangal essential oil (Gal EO) against nephrotoxicity induced by Genta. Materials and methods: Gal EO was isolated and subjected to GC-MS for analysis. The oil components were in-silico investigated against Genta-induced renal toxicity targets using network pharmacology and molecular docking approaches. In vivo studies involved the alienation of rats into four groups. Control and Genta groups rats received 0.5% CMC orally by gavage for two weeks and saline or Genta (100 mg/kg) I.P. injection on the 8th to the 14th day. Genta + Gal EO (50 mg/kg) and Genta + Gal EO (100 mg/kg) groups received Gal EO (50 or 100 mg/kg, P.O.) daily for two weeks and Genta (100 mg/kg) I.P. injection. Renal histopathological and kidney function tests, lipid peroxidation, oxidative, inflammatory mediators and apoptotic markers were assessed. Results: Network Pharmacology suggested Toll-like receptors 4 (TLR4), and Interleukin-1beta (IL-1β) as potential targets of Gal EO components in Genta induced renal toxicity. Gal EO significantly decreased Cr, uric acid, BUN, CysC, NGAL and Kim-1 and the urine albumin/creatinine ratio. Gal EO reduced MDA and NO levels with an upsurge in the GSH content, GPx, GSH-R, catalase and SOD levels. Gal EO lessened the gene expression of TLR4/ MYD88/ NF-κB/ IL-1β with subsequent reduction in ICAM-1 and MCP-1 expression and the levels of MPO, TNF-α and IL-6 while intensified IL-10. Gal EO diminished caspase 3, 9 and Bax while amplified Bcl2. Conclusion: Genta induced nephrotoxicity was mitigated by the anti-inflammatory, anti-oxidant and anti-apoptotic effect of Gal EO through decreasing TLR4/ MYD88/ NF-κB/ IL-1β signaling pathway.