Kidney injury represents a global concern, leading to chronic kidney disease. The organophosphate insecticide malathion (MT) demonstrates environmental disturbance and impairment of different mammalian organs, including kidneys. Likewise, gamma-irradiation (IRR) provokes destructive effects in the kidneys. Rutin is a flavonoid glycoside that exhibits nephro-protective and radio-protective properties. This manuscript focused on investigating the protective response of rutin on MT- and IRR-triggered kidney injury in rats. Rats were randomly divided into eight groups of twelve: G1 (C), control; G2 (Rutin), rutin-treated rats; G3 (IRR), gamma-irradiated rats; G4 (MT), malathion-treated rats; G5 (IRR/MT), gamma-irradiated rats treated with malathion; G6 (IRR/Rutin), gamma-irradiated rats treated with rutin; G7 (MT/Rutin), rats treated with malathion and rutin; and G8 (IRR/MT/Rutin), gamma-irradiated rats treated with malathion and rutin, every day for 30 days. The results demonstrated that rutin treatment regulated the biochemical parameters, the oxidative stress, the antioxidant status, and the inflammatory responses due to the down-regulation of the renal NF-κB p65 protein expression. Moreover, it amended the activity of acetylcholinesterase (AchE), angiotensin ACE I, and ACE II-converting enzymes. Besides, it regulated the iNOS, eNOS, miR-129-3p, miR-200c, and miR-210 gene expressions and bradykinin receptor (B1R and B2R) protein expressions. Histopathological examinations of the kidney tissue confirmed these investigated results. It could be concluded that rutin demonstrated nephro/radioprotection and counteracted the toxicological effects triggered in the kidney tissues of IRR, MT, and IRR/MT intoxicated rats, via regulating miR-129-3p, miR-200c-3p, and miR-210-3p gene expressions, which consequently regulated B2R protein expressions, ACE II activity, and HIF-1α production, respectively.Supplementary InformationThe online version contains supplementary material available at 10.1007/s11356-023-27166-z.