Stimulation of rat placental lactogen-II (rPL-II) secretion by cultured trophoblasts by insulin: development of a rat placental cell culture system and effects of peptide hormones on rPL-II secretion in vitro.
The purpose of this study was to develop a primary culture system using serum-free medium for rat placental trophoblast cells and to investigate the factors that control rat placental lactogen-II (rPL-II) secretion in vitro. The placentae of day 13 pregnant rats were dissociated in Medium 199 containing 0.1% collagenase and 0.002% DNAase. Dissociated cells were fractionated into five segments by centrifugation through a 40% Percoll density gradient and incubated on rat tail collagen bed in medium SFM-101 for up to 7 days. Fraction B at the Percoll gradient density of 1.05 g ml-1 was enriched with rPL-II-producing cells and the time course of rPL-II secretion was characterized by a rapid increase in the first 2 days, remaining at high values (mean: 14-16 ng micrograms-1 DNA) for the following 2-3 days and decreasing thereafter. The rPL-II-producing cells from faction B identified by immunocytochemical examination accounted for approximately 69% of total cultured cells and consisted of a few giant cells and polygonal cells. Growth factors (bovine insulin, 0.1-20 micrograms ml-1; recombinant human insulin-like growth factor (IGF)-I, IGF-II, 0.1-1.0 micrograms ml-1; murine epidermal growth factor (EGF), 0.001-10 micrograms ml-1), rat pituitary hormones (rat growth hormone, rat prolactin, 0.1-10 micrograms ml-1) and hypothalamic hormones (human growth hormone-releasing hormone (GHRH), corticotrophin-releasing hormone (CRH), LHRH, 0.1-10 micrograms ml-1) were individually added to the culture medium to investigate the putative factors that directly control rPL-II secretion by the trophoblast cells.(ABSTRACT TRUNCATED AT 250 WORDS)
Kishi
K
K
Kanzakigawa Laboratory, Shionogi and Co. Ltd, Osaka, Japan.
Itoh
M
M
Kanamori
S
S
Hirashiba
M
M
Kawai
M
M
D000818:Animals; D002478:Cells, Cultured; D003346:Corticotropin-Releasing Hormone / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004815:Epidermal Growth Factor / Q000494:pharmacology; D005260:Female; D007987:Gonadotropin-Releasing Hormone / Q000494:pharmacology; D013006:Growth Hormone / Q000494:pharmacology; D013007:Growth Hormone-Releasing Hormone / Q000494:pharmacology; D007150:Immunohistochemistry; D007328:Insulin / Q000494:pharmacology*; D007334:Insulin-Like Growth Factor I / Q000494:pharmacology; D007335:Insulin-Like Growth Factor II / Q000494:pharmacology; D010920:Placenta / Q000737:chemistry / Q000378:metabolism; D010928:Placental Lactogen / Q000032:analysis / Q000096:biosynthesis*; D011388:Prolactin / Q000494:pharmacology; D051381:Rats; D017207:Rats, Sprague-Dawley; D014327:Trophoblasts / Q000737:chemistry / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*
D007328:Insulin; C062303:placental lactogen II; D007987:Gonadotropin-Releasing Hormone; D004815:Epidermal Growth Factor; D007334:Insulin-Like Growth Factor I; D007335:Insulin-Like Growth Factor II; D011388:Prolactin; D013006:Growth Hormone; D003346:Corticotropin-Releasing Hormone; D013007:Growth Hormone-Releasing Hormone; D010928:Placental Lactogen