title: mRNA expression pattern of selected candidate genes differs in bovine oviductal epithelial cells in vitro compared with the in vivo state and during cell culture passages.
abstract: BACKGROUND
  The mammalian oviduct provides the optimal environment for gamete maturation including sperm capacitation, fertilization, and development of the early embryo. Various cell culture models for primary bovine oviductal epithelial cells (BOEC) were established to reveal such physiological events. The aim of this study was to evaluate 17 candidate mRNA expression patterns in oviductal epithelial cells (1) in transition from in vivo cells to in vitro cells; (2) during three consecutive cell culture passages; (3) affected by the impact of LOW or HIGH glucose content media; and (4) influenced by different phases of the estrous cycle in vivo and in vitro. In addition, the release of a metabolite and proteins from BOEC at two distinct cell culture passage numbers was estimated to monitor the functionality.
  
  
  METHODS
  BOEC from 8 animals were isolated and cultured for three consecutive passages. Total RNA was extracted from in vivo and in vitro samples and subjected to reverse transcription quantitative polymerase chain reaction to reveal mRNA expression of selected candidate genes. The release of prostaglandin E2 (PGE2), oviduct-specific glycoprotein 1 (OVGP1) and interleukin 8 (IL8) by BOEC was measured by EIA or ELISA after 24 h.
  
  
  RESULTS
  Almost all candidate genes (prostaglandin synthases, enzymes of cellular metabolism and mucins) mRNA expression pattern differed compared in vivo with in vitro state. In addition, transcription of most candidate genes was influenced by the number of cell culture passages. Different glucose medium content did not affect mRNA expression of most candidate genes. The phase of the estrous cycle altered some candidate mRNA expression in BOEC in vitro at later passages. The release of PGE2 and OVGP1 between passages did not differ. However, BOEC in passage 3 released significantly higher amount of IL8 compared with cells in passage 0.
  
  
  CONCLUSIONS
  This study supports the hypothesis that candidate mRNA expression in BOEC was influenced by transition from the in vivo situation to the new in vitro environment and during consecutive passages. The consequence of cell culture passaging on BOEC ability to release bioactive compounds should be considered.
authors.affiliation: Institute of Veterinary Biochemistry, Freie Universität Berlin, Oertzenweg 19b, 14163, Berlin, Germany.
authors.forename: Sadjad
authors.initials: S
authors.lastname: Danesh Mesgaran
authors.affiliation: Institute of Veterinary Biochemistry, Freie Universität Berlin, Oertzenweg 19b, 14163, Berlin, Germany.
authors.forename: Jutta
authors.initials: J
authors.lastname: Sharbati
authors.affiliation: Institute of Veterinary Biochemistry, Freie Universität Berlin, Oertzenweg 19b, 14163, Berlin, Germany.
authors.forename: Ralf
authors.initials: R
authors.lastname: Einspanier
authors.affiliation: Institute of Veterinary Biochemistry, Freie Universität Berlin, Oertzenweg 19b, 14163, Berlin, Germany. christoph.gabler@fu-berlin.de.
authors.forename: Christoph
authors.identifier: http://orcid.org/0000-0002-2839-2857
authors.initials: C
authors.lastname: Gabler
mesh_terms: D000818:Animals; D002417:Cattle; D018929:Cell Culture Techniques; D002478:Cells, Cultured; D015232:Dinoprostone / Q000096:biosynthesis / Q000235:genetics; D004847:Epithelial Cells / Q000378:metabolism*; D005187:Fallopian Tubes / Q000166:cytology / Q000378:metabolism*; D005260:Female; D005786:Gene Expression Regulation; D056726:Genetic Association Studies / Q000379:methods*; D006023:Glycoproteins / Q000096:biosynthesis / Q000235:genetics; D012333:RNA, Messenger / Q000096:biosynthesis* / Q000235:genetics*
chemical_list: D006023:Glycoproteins; C087523:OVGP1 protein, Bos taurus; D012333:RNA, Messenger; D015232:Dinoprostone
keywords: Cell culture passage; Culture medium; Enzymes of cellular metabolism; Mucin; Oviduct; Prostaglandin synthase; mRNA expression