Amino acids protect against methotrexate-induced hepatorenal toxicity in vitro and in vivo

Hepatorenal toxicity is one of the most life-threatening adverse effects of methotrexate (MTX). The present study investigated the potential protective effects of glutamine (GLU), glycine (GLY), methionine (MET), and leucine (LEU) in MTX-induced hepatorenal toxicity in vitro and in vivo. Freshly isolated hepatocytes and renal slices of rats were used to investigate the effect of these amino acids in vitro, while male Sprague-Dawley rats were used to evaluate their effects in vivo. Rats were assigned into 6 equal groups: a negative control, MTX (20 mg/kg/day, i.p., 3 days), MTX + (GLU 25 mg/kg/day i.p., 6 days), MTX + (GLY 0.5 gm/kg/day, i.p., 3 days), MTX + (MET 1 mg/kg/day, i.p., 6 days) and MTX + LEU (50 mg/kg/day, i.p., 6 days). These amino acids showed enhanced viability of liver and kidney cells assessed by trypan blue exclusion and lactate dehydrogenase leakage tests, respectively. Besides, pre-incubation of suspended hepatocytes or renal slices with GLU (25 mM and 10 mM, respectively), GLY (25 mM and 5 mM, respectively), MET (12.5 mM and 5 mM, respectively) or LEU (25 mM and 10 mM, respectively), 30 min before MTX, significantly down-regulated caspase-3 and TNF-α levels as compared with MTX-intoxicated groups. MTX-induced hepatorenal toxicity was manifested by increasing liver and kidney enzymes, oxidative stress and severe histopathological alterations. Amino acids pre-treatment abrogated MTX-induced alteration in hepatorenal toxicity indices as evidenced by amelioration in oxidative stress, inflammatory mediators, and histopathological changes. Amino acids may serve as a promising adjuvant therapy with MTX as it may ameliorate its hepatorenal toxicity through antioxidant, anti-inflammatory and anti-apoptotic mechanisms.