Factors affecting proliferation and dedifferentiation of primary bovine oviduct epithelial cells in vitro.
The oviduct is the physiological site for key events in reproduction, such as capacitation of spermatozoa, fertilization and early embryonic development. Interactions between oviduct epithelial cells and gametes or embryos cannot sufficiently be studied in vivo. Therefore, model systems are needed which mimic in vivo conditions most closely. In this study we optimised the method for isolating bovine oviduct cells and compared different cell support materials as well as two culture systems (perfusion vs static culture) for their ability to maintain characteristic morphological and functional features of oviduct cells. Out of nine different cell support materials tested, cellulose nitrate (0.45 micron pore size) was the most suitable to maintain cells in a manner similar to freshly isolated oviduct epithelial cells. Comparing static vs perfusion culture by electron microscopy, morphological differences of the cells were insignificant in the first days of culture, while they became more evident after 8 days. The cells in the static system lost typical characteristics such as columnar shape, cilia and secretory protrusions, while these features were still present in perfusion culture. In addition, intense ciliogenesis and cytoplasmic organelles for protein synthesis were found under perfusion conditions. These findings were underlined by differences in expression of the oviduct-specific oestrus-associated glycoprotein 85-97 kDa (GP 85-97) gene as revealed by semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR). The RNA levels of this specific gene were significantly higher in perfusion compared to the static culture system. Our data show clear advantages of perfusion vs static culture for primary bovine oviduct epithelial cells.
Reischl
J
J
Department of Molecular Animal Breeding and Genetics, Ludwig-Maximilian University, Munich, Germany.
Prelle
K
K
Schöl
H
H
Neumüller
C
C
Einspanier
R
R
Sinowatz
F
F
Wolf
E
E
D000199:Actins / Q000235:genetics; D000818:Animals; D002417:Cattle; D002448:Cell Adhesion; D018929:Cell Culture Techniques / Q000379:methods; D002454:Cell Differentiation; D002455:Cell Division; D002478:Cells, Cultured; D004847:Epithelial Cells / Q000166:cytology* / Q000502:physiology; D005187:Fallopian Tubes / Q000166:cytology* / Q000502:physiology; D005260:Female; D005786:Gene Expression Regulation; D006023:Glycoproteins / Q000235:genetics; D008871:Microvilli / Q000502:physiology / Q000648:ultrastructure; D012333:RNA, Messenger / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014158:Transcription, Genetic
D000199:Actins; D006023:Glycoproteins; D012333:RNA, Messenger; C104698:estrus-associated oviductal protein, Bos taurus