title: Differential effects of thrombin and hypoxia on endometrial stromal and glandular epithelial cell vascular endothelial growth factor expression.
abstract: Ovarian steroids and/or premenstrual endometrial hypoxia are thought to restore the endometrial vasculature shed during menstruation by elevating endometrial vascular endothelial growth factor (VEGF) levels. During the luteal phase, VEGF levels peak, progesterone induces estradiol (E(2))-primed human endometrial stromal cells (HESCs) to decidualize and express tissue factor (TF), and endometrial vascular permeability is enhanced. The latter would present circulating clotting factors to decidual cell-expressed TF to form local thrombin. HESCs were incubated in serum-supplemented medium containing vehicle (control) or 10(-8) M E(2) or 10(-7) M medroxyprogesterone acetate (MPA) or E(2) + MPA for 7 d to induce decidualization, while monolayers of human endometrial glandular epithelial cells (HEGECs) formed during 4-d incubation of glands. The medium was exchanged for a defined medium containing corresponding vehicle or steroids +/- thrombin under normoxia or hypoxia (0-1% O(2)). Hypoxia enhanced secreted immunoreactive VEGF levels by severalfold in HESCs and HEGECs, but the steroids did not affect VEGF output in either cell type under normoxia or hypoxia. In E(2) + MPA-decidualized HESCs, VEGF levels were elevated by 0.1 U/ml of thrombin, and 0.5-2.5 U/ml of thrombin elicited maximum effects. The addition of 0.5 U/ml of thrombin evoked a time-dependent enhancement of VEGF levels and about an 8-fold increase at 48 h (P < 0.02; n = 6). Northern blotting indicated that E(2) + MPA-decidualized HESCs expressed VEGF(121), VEGF(165), and VEGF(189) mRNA, which were enhanced severalfold during 5- to 20-h incubation with thrombin. Moreover, TRAP, a synthetic peptide activator of the constitutively expressed protease activated receptor-1 thrombin receptor in decidualized HESCs, also elevated secreted VEGF levels. By contrast, HEGECs were unresponsive to thrombin added alone or with ovarian steroids. These results suggest that thrombin formed by progestin-augmented TF levels acts as an autocrine enhancer of VEGF expression in decidualized HESCs. Because angiogenesis occurs in a matrix of decidualized HESCs, these in vitro results provide a novel mechanism to account for both the peak in VEGF and angiogenesis in luteal phase human endometrium.
authors.affiliation: Department of Obstetrics and Gynecology, New York University School of Medicine, New York, New York 10016, USA.
authors.forename: Charles J
authors.initials: CJ
authors.lastname: Lockwood
authors.forename: Graciela
authors.initials: G
authors.lastname: Krikun
authors.forename: A Bon Chang
authors.initials: AB
authors.lastname: Koo
authors.forename: Susan
authors.initials: S
authors.lastname: Kadner
authors.forename: Frederick
authors.initials: F
authors.lastname: Schatz
mesh_terms: D015687:Cell Hypoxia / Q000502:physiology*; D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D004717:Endometrium / Q000166:cytology / Q000187:drug effects / Q000502:physiology*; D016228:Endothelial Growth Factors / Q000235:genetics* / Q000378:metabolism*; D004847:Epithelial Cells / Q000502:physiology*; D005260:Female; D006801:Humans; D007044:Hysterectomy; D007700:Kinetics; D008222:Lymphokines / Q000235:genetics* / Q000378:metabolism*; D017154:Stromal Cells / Q000502:physiology*; D013917:Thrombin / Q000494:pharmacology*; D014158:Transcription, Genetic / Q000187:drug effects; D042461:Vascular Endothelial Growth Factor A; D042442:Vascular Endothelial Growth Factors
chemical_list: D016228:Endothelial Growth Factors; D008222:Lymphokines; C467484:VEGFA protein, human; D042461:Vascular Endothelial Growth Factor A; D042442:Vascular Endothelial Growth Factors; D013917:Thrombin