title: Association between the expression of LHR, FSHR and CYP19 genes, cellular distribution of encoded proteins and proliferation of porcine granulosa cells in real-time.
abstract: The process of granulosa cell luteinization is part of the main process determining growth, differentiation and proliferation of these cells. Although the mechanisms underlying the regulation of luteinizing hormone receptor (LHR), follicle stimulating hormone receptor (FSHR) and cytochrome P450 aromatase expression in mammalian granulosa cells is well understood, still little is known about the expression of mRNA and encoded proteins in relation to cell proliferation and luteinization in vitro. Porcine granulosa cells were observed in vitro at a168-h period while undergoing real-time proliferation using an RTCA system. Furthermore, LHR, FSHR and CYP19 mRNA expression were detected using RQ-PCR after 168 h of in vitro culture (IVC) at 24-h intervals, and LHR, FSHR and P450arom were examined by confocal microscopic observation at 0 h, 24 h, 48 h, 96 h, and 168 h of IVC. We found increased expression of LHR and CYP19 mRNA at 24 h and 48 h of IVC compared to the other stages (P less than 0.01, P less than 0.001), whereas FSHR mRNA was higher only at 0 h (P less than 0.001). In contrast, LHR, FSHR and P450arom protein expression was significantly higher at the end of the 168-h IVC period compared to 0 h, 24 h, 48 h and 96 h (P less than 0.001). LHR, FSHR and P450arom were distributed in the cytoplasm of porcine GCs at each time point of IVC. When analyzing cell proliferation, differences in cell index were observed (at least P less than 0.05) between the first (0-24 h) and the last period (144-168 h) of IVC; however, soon after 24 h of IVC a logarithmic increase in proliferation was also seen. We assume that the expression of LHR, FSHR and CYP19 mRNAs depends on the period of in vitro cultivation and may be linked with the luteinization process of porcine GCs. Furthermore, the patterns of mRNA and protein expression suggest a post-transcriptional regulation of LHR, FSHR and P450arom. In summary, it can be presumed that mRNA and protein expression and in vitro luteinization and proliferation of porcine GCs are regulated by different mechanisms, because not all of these processes are correlated.
authors.affiliation: Department of Histology and Embryology, Poznań University of Medical Sciences, Poznań , Poland.
authors.forename: B
authors.initials: B
authors.lastname: Kempisty
authors.affiliation: Department of Histology and Embryology, Poznań University of Medical Sciences, Poznań , Poland.
authors.forename: A
authors.initials: A
authors.lastname: Ziółkowska
authors.affiliation: Department of Histology and Embryology, Poznań University of Medical Sciences, Poznań, Poland.
authors.forename: S
authors.initials: S
authors.lastname: Ciesiółka
authors.affiliation: Department of Toxicology, Poznań University of Medical Sciences, Poznań, Poland.
authors.forename: H
authors.initials: H
authors.lastname: Piotrowska
authors.affiliation: Department of Veterinary, PPoznań University of Life Sciences, Poznań, Poland.
authors.forename: P
authors.initials: P
authors.lastname: Antosik
authors.affiliation: Department of Veterinary, Poznań University of Life Sciences, Poznań, Poland.
authors.forename: D
authors.initials: D
authors.lastname: Bukowska
authors.affiliation: Department of Histology and Embryology, Poznań University of Medical Sciences, Poznań, Poland.
authors.forename: M
authors.initials: M
authors.lastname: Nowicki
authors.affiliation: Institute of Reproductive Biology, Leibniz Institute for Farm Animal Biology, Dummerstorf, Germany.
authors.forename: K P
authors.initials: KP
authors.lastname: Brüssow
authors.affiliation: Department of Histology and Embryology, Wroclaw Medical University, Poland.
authors.forename: M
authors.initials: M
authors.lastname: Zabel
mesh_terms: D000818:Animals; D001141:Aromatase / Q000096:biosynthesis*; D049109:Cell Proliferation; D005260:Female; D005786:Gene Expression Regulation / Q000502:physiology*; D006107:Granulosa Cells / Q000166:cytology / Q000378:metabolism*; D012333:RNA, Messenger / Q000096:biosynthesis; D011962:Receptors, FSH / Q000096:biosynthesis*; D011974:Receptors, LH / Q000096:biosynthesis*; D013552:Swine
chemical_list: D012333:RNA, Messenger; D011962:Receptors, FSH; D011974:Receptors, LH; D001141:Aromatase