As shown in Figure 6, the levels of 5(S)-HETE were similar to the calcium ionophore alone, indicating

that the 5(S)-HETE was largely formed by the 5-LOX pathway. Interestingly, 5(S)-HETE concentrations were decreased approximately 25% when vitamin C was added to the media in addition to the ionophore. It is well known that vitamin C is a mediator of lipid Inhibitors,research,lifescience,medical hydroperoxide decomposition [124,125]. To further investigate the route of the 5(S)-HETE decomposition, a DNA adduct specific for lipid peroxidation was quantified in the same conditions. It was previously shown that in vitro reaction of HPETEs with 2’-deoxyguanosine (dGuo) leads to formation of DNA adducts [126,127,128] (Figure 5). Two of the DNA adducts [etheno-dGuo (εdGuo) and heptanone-etheno-dGuo (HεdGuo)] were detected in the CESS Inhibitors,research,lifescience,medical cells. Interestingly, there was a significant increase in the HεdGuo formation when the CESS cells were treated with vitamin C

and the calcium ionophore when compared with the calcium ionophore alone. The amount of the HεdGuo was dramatically decreased if the LOX pathway was inhibited by MK886. The addition of aspirin (a Inhibitors,research,lifescience,medical non-specific COX inhibitor) to the CESS cells activated with calcium ionophore had no effect on HεdGuo adduct levels. In contrast, in epithelial cells that stably express COX, the addition of aspirin reduced the HεdGuo levels to basal levels [118]. These studies provided convincing evidence that HεdGuo arose from a LOX- rather than a COX-mediated pathway. Figure 6 Amount of lipid peroxidation metabolites from CESS cells. A, 5-HETEs. B, LTB4. C, PGE2, PGD2, and PGF2α. D, 13-HODEs. NT, no treatment; CA, treated with 1.0 μm “type”:”entrez-nucleotide”,”attrs”:”text”:”A23187″,”term_id”:”833253″,”term_text”:”A23187″ Inhibitors,research,lifescience,medical … The formation Inhibitors,research,lifescience,medical of LTB4 by the CESS cells followed a similar pattern to the formation of 5(S)-HETE after calcium ionophore treatment (Figure 6). However, the addition of vitamin C did not reduce the levels of the LTB4. This selleck chemicals llc supported the hypothesis that vitamin C

was a inducing the decomposition of the lipid hydroperoxides. PGE2, PGD2, and PGF2α were the major lipid peroxidation products derived from COX-1-mediated arachidonic acid metabolism. Their levels were increased by the calcium ionophore and were not affected by vitamin C or the MK866 inhibitor (Figure 6). All three PGs Entinostat were reduced to levels lower than the NT level when aspirin was added together with calcium ionophore. Therefore, the targeted chiral lipidomics method was useful for the analysis of enantioselective pathways of cellular LOX and COX mediated arachidonic acid oxidation, being able to differentiate from the racemic mixture formed by a ROS mediated pathway. Additional data provided clear evidence that DNA damage was a result of 5-LOX-mediated arachidonic acid metabolism. 4.2.