Oxidative stress induced by ochratoxin A in LLC-PK1 cell line and the chemoprotective effects of carnosic acid

Mycotoxins are secondary metabolites of various moulds that contaminate several alimentary substrates. One of the most dangerous of these is ochratoxin A (OTA). Reactive oxygen species (ROS) play an important role in the toxicity mechanism of OTA, so the use of natural or synthetic free radical scavengers could be a potential chemopreventive strategy. Carnosic acid (CA) is the major polyphenolic compound present in rosemary plants. This work aimed to determine whether CA could counteract OTA-induced cell damage. Free radical scavenging properties of CA were chemically determined at pH 7.4. Cytotoxicity of CA and OTA on LLC-PK1 cells, and the protective effects of CA, were assessed using the Alamar Blue test. The effects in vitro of CA pre-treatment on the production of ROS, the DNA oxidation and the induction of apoptosis induced by OTA were studied. It was found that CA has free radical scavenging properties at both the considered pH values. Moreover, a pre-treatment of 24 h with 10, 20 and 30 µM CA is able to reduce OTA-induced cytotoxicity; a pre-treatment of 24 h with 20 and 30 µM CA achieved ROS reduction and with 30 µM CA decreased the OTA-induced increase of 8-OH-2'-deoxyguanosine and of DNA fragmentation in LLC-PK1. These findings suggest a starting point to develop alimentary strategies against OTAinduced cell damage. Moreover, our results provide further evidence that oxidative stress plays an important role in the OTA cytotoxicity mechanism.