A Practical Approach, Second Edition=Ronald D. Ho.pdf

544 DEVELOPMENTAL REPRODUCTIVE TOXICOLOGY: A PRACTICAL APPROACH, SECOND EDITION27 active compounds in the presence of rat liver S 9 fraction, suggesting that rat embryonic cellspossess enzymes capable of xenobiotic activation. Several attempts by other investigators to definitivelyidentify P450s in rodent, human, and chick embryos and extraembryonic tissues have metwith limited success. Recent reviews 2,5 of the published data (most of which are indirect, conflicting,and/or negative) on this subject have finally concluded that constitutive functional P450 enzymescapable of xenobiotic oxidation have not been definitely identified in rodent or human embryosduring organogenesis. It appears that, with a possible exception of P4503A7, the constitutive formsof P4501A1, P4501A2, and probably P4502E1 and P4502B are either absent or their signal is soweak that it can not be detected by ultrasensitive probes such as reverse transcriptase–polymerasechain reaction (RT-PCR). Thus, the marginally positive results claiming P450 involvement reportedby some investigators appear to be of questionable significance and relevance to the bioactivationof teratogens during human organogenesis.b. InductionAlthough a few studies 5 have reported the inductive effects of 3-MC, β-naphthoflavone, and PCNon rodent embryonic xenobiotic oxidation, published data are too scant for other inducers. P450sin rodent embryos are refractive to induction following exposure to phenobarbital. 70,71 However,those from MC-pretreated dams were capable of measurable oxidative biotransformation of AAF,suggesting P450 induction. 70,71 In preimplantation stage mice, activation of BP, possibly byCYP1A1, can be detected on GD 4. 110 Mouse embryos preexposed to 3-MC exhibit a higherincidence of BP-induced fetal abnormality, suggesting possible induction of CYP1A1. 111 Interestingly,the incidence of fetal anomalies was much lower in noninducible littermates. The signals forP4501A1 mRNA were detectable in conceptal tissues from 3-MC–treated rats only on GD 12. 112However, the intensity of the signal was lowest in tissues of the embryo proper. This casts doubtsregarding the presumed activation of teratogenic chemicals by embryonic P450s. In light of otherknown pathways, Wells and Winn 3 have recently questioned whether the elevated levels of embryonicP450 are sufficient to mediate teratologically relevant bioactivation of xenobiotics. Treatmentof eggs with 3,3′,4,4′-tetrachloro-biphenyl has been shown to result in a 2- and 14-fold increase inECOD and AHH activities, respectively, in livers from 5-day-old chick embryos. 1132. Prostaglandin SynthasePGS activity occurs in mouse 114 and rat 115 blastocysts before implantation. The liberation ofprostaglandin 6-keto-PGF 1 , PGE, and PGF from endogenous arachidonate was noted in GD 11 ratembryo homogenates. 116 As expected, nonsteroidal antiinflammatory drugs, such as aspirin,indomethacin, salicylate, meclofenamic acid, ibuprofen, 5,8,11-eicosatriynoic acid (ETI), and5,8,11,14-eicosatetraynoic acid (ETYA), caused dose-dependent inhibition of prostaglandin synthesis.Nevertheless, the authors opined that the inhibition of PGS is not likely to be the cause ofaspirin-induced limb defects in rats. Recent reports 117,118 have described the occurrence of PGSactivity in mouse embryos. Even though, the presence of PGS in human embryos is expected, itappears that relevant published data on this subject are not currently available.A series of investigations reported by Wells and coworkers provide compelling evidence implicating,in part, embryonic PGS as one of the major enzymes responsible for activation of phenytoinand other teratogenic compounds. The results supporting the hypothesis that phenytoin teratogenicityinvolves a free radical mechanism include oxidation of DNA and covalent binding toprotein, 118,119 oxidation of GSH to GSSG, 118,120 and partial protection from phenytoin-induced cleftpalate formation in mice when dams were pretreated with either EYTA, 120 caffeic acid (a dualinhibitor of PGS and LO), and aspirin (a cyclooxygenase inhibitor) or with α-phenyl-N-t-butylnitrone (PBN, a spin trap agent for free radicals). 121 Purified PGS in the presence of arachidonategenerates reactive metabolites from phenytoin that bind covalently to proteins and can be detected© 2006 by Taylor & Francis Group, LLC