title: A bovine oviduct epithelial cell suspension culture system suitable for studying embryo-maternal interactions: morphological and functional characterization.
abstract: We established a short-term (24 h) culture system for bovine oviduct epithelial cells (BOECs), obtained on day 3.5 of the estrous cycle and evaluated the cells with respect to morphological criteria, marker gene expression, and hormone responsiveness. BOEC sheets were isolated mechanically from the ampulla with similar yields from oviducts ipsi- and contralateral to the ovulation site (57.9 +/- 4.6 and 56.4 +/- 8.0 x 10(6) cells). BOECs showed > 95% purity and cells cultured for 24 h maintained morphological characteristics present in vivo, as determined by light microscopy, scanning electron microscopy, and transmission electron microscopy. Both secretory cells with numerous secretory granules and ciliated cells with long, well-developed, and vigorously beating kinocilia were visible. Quantitative real-time PCR failed to detect significant differences in transcript levels between ipsi-and contralateral BOECs for the majority of marker genes (estrogen receptors alpha and beta (ESR1 and ESR2), 3-hydroxy-3-methylglutaryl-coenzyme A reductase (HMGCR), oviductal glycoprotein 1 (OVGP1), progesterone receptor (PGR), and tumor rejection antigen 1 (TRA1)) throughout the 24 h culture period. However, the combined data of all time points for glutathione peroxidase 4 (GPX4), a gene previously shown to be expressed at higher levels in the ipsilateral oviduct in vivo, also indicated significantly different mRNA levels in vitro. The expression of marker genes remained stable after 6 h cell culture, indicating only a short adaptation period. Western blot analysis confirmed ESR1 and PGR protein expression throughout the culture period. In agreement with cyclic differences in vivo, estradiol-17beta stimulation increased PGR transcript abundance in BOECs. Our novel culture system provides functional BOECs in sufficient quantities for holistic transcriptome and proteome studies, e.g. for deciphering early embryo-maternal communication.
authors.affiliation: Institute of Molecular Animal Breeding and Biotechnology, Gene Center of the Ludwig-Maximilians University Munich, Munich, Germany.
authors.forename: Regine
authors.initials: R
authors.lastname: Rottmayer
authors.forename: Susanne E
authors.initials: SE
authors.lastname: Ulbrich
authors.forename: Sabine
authors.initials: S
authors.lastname: Kölle
authors.forename: Katja
authors.initials: K
authors.lastname: Prelle
authors.forename: Christine
authors.initials: C
authors.lastname: Neumueller
authors.forename: Fred
authors.initials: F
authors.lastname: Sinowatz
authors.forename: Heinrich H D
authors.initials: HH
authors.lastname: Meyer
authors.forename: Eckhard
authors.initials: E
authors.lastname: Wolf
authors.forename: Stefan
authors.initials: S
authors.lastname: Hiendleder
mesh_terms: D000818:Animals; D015153:Blotting, Western / Q000379:methods; D002417:Cattle; D046149:Embryo Culture Techniques; D004847:Epithelial Cells / Q000378:metabolism* / Q000648:ultrastructure; D005187:Fallopian Tubes / Q000378:metabolism* / Q000648:ultrastructure; D005260:Female; D015870:Gene Expression; D006728:Hormones / Q000032:analysis / Q000235:genetics; D007150:Immunohistochemistry / Q000379:methods; D007633:Keratins / Q000032:analysis; D008855:Microscopy, Electron, Scanning; D046529:Microscopy, Electron, Transmission; D023421:Models, Animal; D011247:Pregnancy; D011270:Pregnancy, Animal / Q000502:physiology*; D012333:RNA, Messenger / Q000032:analysis; D011960:Receptors, Estrogen / Q000032:analysis / Q000235:genetics; D011980:Receptors, Progesterone / Q000032:analysis / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014746:Vimentin / Q000032:analysis
chemical_list: D006728:Hormones; D012333:RNA, Messenger; D011960:Receptors, Estrogen; D011980:Receptors, Progesterone; D014746:Vimentin; D007633:Keratins