Expression of cytochrome P4501b1 (Cyp1b1) during earlymurine development

[摘要] Purpose: To examine the embryonic expression of cytochromeP4501b1 (Cyp1b1) gene by whole mount in situ hybridization.Methods: FVB/NcrlBR mouse embryos staged at 9.5, 10.5, and 11.5 dpcwere obtained by timed breeding experiments. Antisense and sense RNAprobes labeled with digoxigenin UTP were generated by in vitrotranscription of an 848 bp Cyp1b1 cDNA fragment that was subclonedinto transcription vector pCR®II-TOPO®. The digoxigenin labeledRNA was localized using an alkaline phosphatase conjugatedanti-digoxigenin Fab fragment. Colorimetric detection of the digoxigeninlabeled probe was performed with substrate solution containing4-nitro-blue tetrazolium chloride (NBT) and 5-bromo-4-chloro-3-indolylphosphate (BCIP).Results: During early stages of murine development Cyp1b1 mRNAwas detected in the developing eye, hindbrain, branchial arches,forelimb bud, ligaments supporting the liver primordium and developingkidney. In the eye and forelimb bud Cyp1b1 displayed restrictedexpression along the axes of development. In the developing eyeCyp1b1 exhibited dorsal expression with respect to thedorso-distal/proximo-ventral axis and anterior expression with respectto the anterior-nasal/posterior-temporal axis. In the forelimb budCyp1b1 expression was localized posteriorly. The polarity ofCyp1b1 expression was lost at 11.5 dpc, at which time expression wasadditionally seen in ventral (eye) and anterior (forelimb bud) areas.Conclusions: The spatio-temporal expression patterns observed inthis study suggest that during early stages of murine development,Cyp1b1 participates in establishment and/or maintenance of polarityalong the axes of embryonic development. Expression of Cyp1b1 in thedorso-distal end of the optic cup, from which the ciliary body and irisare derived, correlates with the expression patterns seen in adulttissues and the abnormal development of these structures as part of theglaucoma phenotypes resulting from Cyp1b1 mutations.

[学科分类] 生物化学/生物物理