The protein levels of the three enzymes

The protein levels of the three enzymes in the organs in the three earthworm species ranked body wall > intestine > seminal receptacle ≈ seminal vesicle (Fig. 3B).The presence of significant detoxification capability in the body wall would be logically expected in a soil dwelling organism, to facilitate detoxification of contaminants directly absorbed from the environment, reducing their potential harm to the earthworm. Similar findings were previously reported for the ‘sludge worm’, Tubifex (Fischer and Horváth, 1977). The moderate CYP expression in the intestine may provide protection from possible harm from soil substrates entering the intestine. Our original hypothesis was that there would be greater CYP expression in seminal receptacles vesicles to protect the delicate sperm and ova. However, the data indicate instead that CYP enzyme expression is ‘load’ related with CYP expression at locations of xenobiotic exposure in the body wall and intestine, while the reproductive organs do not directly come in contact with soil and thus have less exposure to xenobiotics have lower CYP enzyme expression. Differences in CYP expression occurring in relation to sexual maturity have been observed in zebra fish where CYP11A1 gene expression was greatest in the middle phase of ovary development and lowest at sexual maturity (Chen, 2014). Finally, levels of the three CYP isoforms studied were approximately 1A2 < 2E1 < 3A4 (Fig. 2), whereas in human liver microsome samples, Liu et al. (2014) reported a ratio for CYP 1A2:2E1:3A4 of 39.3:9.3:54.0, so in both humans and earthworms CYP3A4 is the Tozadenant isoform, but CYP1A2 is much more strongly expressed in earthworms than in humans. This difference in comparative expression between earthworm and humans can also be hypothesized to have a functional significance that would be worth exploring in future research.
Acknowledgments This work was supported by the National Natural Science Foundation of China (No.31172360, 41471203). We also thank Education New Zealand, the T.R. Ellett Agricultural Trust, Ministry of Business, Innovation and Employment (C03X0902) and the Ministry of Education of China for funding to support the participation of the New Zealand authors.
Introduction The cytochrome P450 (CYP) superfamily consists of P450 enzymes that localize in the liver and intestine, accounting for 90% of the total drug metabolism [1]. Among the various CYP450 isozymes, CYP1A2, CYP2C19, CYP2D6 and CYP3A4 are four important human CYPs and they are responsible for most CYP-mediated metabolism [2]. Inhibition of the CYP450 isoform can increase the plasma concentration of a drug, which may lead to adverse drug reactions (ADRs) [3]. Therefore, it is important to assay CYP450 isoform enzyme activity to study drug-drug interactions (DDIs). To evaluate various CYP450 isoform activities simultaneously, a cocktail method that included several CYP probe drugs was performed [4,5]. Using this method, the studies analyzed the metabolic ratio between the metabolites and the probe drugs to assess in vivo drug-metabolism activities [6,7]. These cocktail methods have also been used to assay CYP activities to investigate the phenotyping of interindividual differences in metabolic activities [8]. The selected substrates of this method, which are also called probe drugs, are specifically catalyzed by CYP450 isozymes to form metabolites [9,10]. CYP1A2 activity can be assessed by using the rate of phenacetin (PHE) metabolism to acetaminophen (ACE) [11]. Omeprazole (OME) is used as a probe for CYP2C19 and metabolized to 5′-hydroxy omeprazole (OHOME) via CYP2C19 [12]. Metoprolol (MET) is used for phenotyping CYP2D6 activity and is metabolized to α-hydroxy metoprolol (OHMET) [13]. Midazolam (MDZ) is the gold- standard probe for CYP3A4 [14], and 1′-hydroxy midazolam (OHMDZ) is the main metabolite of CYP3A4. Thus, the four probe drugs (PHE, OME, MET and MDZ) were used to investigate CYP450 activities in the present study.