Oxidative stress and enhanced paracellular permeability in the small intestine of methotrexate-treated rats.
OBJECTIVE
We previously demonstrated the increase of reactive oxygen species (ROS) production and myeloperoxidase (MPO) activity in the small intestine of methotrexate (MTX)-treated rats. In the present study, we investigated the role of ROS modulating intestinal mucosal permeability in this damage.


METHODS
MTX (20 mg/kg body weight) was administered to rats intravenously. N-Acetylcysteine (NAC; 80 mg/kg body wt), an antioxidant and a precursor of glutathione (GSH) was administered to rats intraperitoneally to investigate the contribution of ROS to the intestinal permeability enhancement. Intestinal permeability was evaluated by determining that of a poorly absorbable marker, fluorescein isothiocyanate-labeled dextran (FD-4; average molecular mass, 4.4 kDa) using the in vitro everted intestine technique. The occurrence of oxidative stress in the small intestine was assayed by measuring chemiluminescence and thiobarbituric acid reactive substances (TBARS) productions in mucosal homogenates of the small intestine.


RESULTS
The mucosal permeability of FD-4 significantly (p < 0.01) increased in MTX-treated rats compared with control rats, as demonstrated by a twofold increase of FD-4 permeation clearance. This suggests an increase in paracellular permeability. Interestingly, the ROS production was observed preceding the increase of paracellular permeability. Treatment with NAC prevented the MTX-induced ROS production and the increase of paracellular permeability.


CONCLUSIONS
NAC protected the small intestine of rats from MTX-induced change in paracellular permeability, suggesting that ROS played an important role in the enhanced paracellular permeability.
Maeda
Tomoko
T
Department of Biopharmaceutics, Graduate School of Pharmaceutical Sciences, Chiba University, 1-8-1 Inohana, Chuo-ku, Chiba, 260-8675, Japan.
Miyazono
Yuko
Y
Ito
Kousei
K
Hamada
Kazuma
K
Sekine
Shuichi
S
Horie
Toshiharu
T
D000111:Acetylcysteine / Q000008:administration & dosage / Q000494:pharmacology; D000818:Animals; D000964:Antimetabolites, Antineoplastic / Q000494:pharmacology; D001835:Body Weight / Q000187:drug effects; D003911:Dextrans / Q000493:pharmacokinetics; D016650:Fluorescein-5-isothiocyanate / Q000031:analogs & derivatives / Q000493:pharmacokinetics; D016166:Free Radical Scavengers / Q000008:administration & dosage / Q000494:pharmacology; D005978:Glutathione / Q000008:administration & dosage / Q000494:pharmacology; D007274:Injections, Intraperitoneal; D007275:Injections, Intravenous; D007408:Intestinal Absorption / Q000187:drug effects*; D007413:Intestinal Mucosa / Q000378:metabolism; D007421:Intestine, Small / Q000187:drug effects* / Q000378:metabolism; D008297:Male; D008727:Methotrexate / Q000008:administration & dosage / Q000494:pharmacology*; D018384:Oxidative Stress / Q000187:drug effects*; D010539:Permeability / Q000187:drug effects; D051381:Rats; D017208:Rats, Wistar; D017382:Reactive Oxygen Species / Q000378:metabolism; D017392:Thiobarbituric Acid Reactive Substances / Q000378:metabolism; D013997:Time Factors
D000964:Antimetabolites, Antineoplastic; D003911:Dextrans; D016166:Free Radical Scavengers; D017382:Reactive Oxygen Species; D017392:Thiobarbituric Acid Reactive Substances; C015219:fluorescein isothiocyanate dextran; D005978:Glutathione; D016650:Fluorescein-5-isothiocyanate; D000111:Acetylcysteine; D008727:Methotrexate