Immunologic identification of the aromatase enzyme system in human endometrium.
Confluent human endometrial stromal cells were cultured in medium with no hormone or supplemented with medroxyprogesterone acetate (MPA), estradiol (E2), and porcine relaxin (RLX) for 5 days. These stromal cells were then labeled with [35S]methionine for 3 h. The radioactive proteins in the particulate fraction of cell homogenate were extracted by detergent and incubated with antisera to purified placental aromatase cytochrome P-450 (P-450arom) and NADPH-cytochrome P-450 reductase to isolate the radio-labeled aromatase enzyme components. Analysis of the radio-labeled protein, isolated by antibody to the cytochrome P-450arom from different preparations (P45FBIII or R-8-2) showed a major band at molecular weight 54k on SDS polyacrylamide gel electrophoresis (SDS-PAGE). The intensity of 54k band was stronger in hormone treated stromal cells than that of control in parallel with the increase of aromatase activity. The radio-labeled protein isolated by anti-NADPH cytochrome P-450 reductase, REDFBIV, showed a major band at the molecular weight 73k on SDS-PAGE with comparable intensity in control and hormone treated samples. Thus, the apparent molecular weights of endometrial cytochrome P-450arom and cytochrome P-450 reductase were identical to placental aromatase enzyme system. When a secretory endometrium and a decidua were labeled with [35S]methionine, the cytochrome P-450arom was detected only in the decidua. NADPH cytochrome P-450 reductase was detected both in the endometrium and the decidua. These results show that antisera to placental aromatase enzyme system cross reacts with the endometrial aromatase enzyme components. The synthesis of cytochrome P-450arom was stimulated by MPA, E2 and RLX while the synthesis of the NADPH-cytochrome P-450 reductase aromatase component was not affected by the hormone.
Huang
J R
JR
Department of Obstetrics and Gynecology, School of Medicine, State University of New York, Stony Brook 11794.
Bellino
F L
FL
Osawa
Y
Y
Tseng
L
L
D001141:Aromatase / Q000276:immunology / Q000378:metabolism*; D001345:Autoradiography; D002478:Cells, Cultured; D003577:Cytochrome P-450 Enzyme System / Q000096:biosynthesis / Q000302:isolation & purification; D004591:Electrophoresis, Polyacrylamide Gel; D004717:Endometrium / Q000166:cytology / Q000187:drug effects / Q000201:enzymology*; D004790:Enzyme Induction; D004958:Estradiol / Q000494:pharmacology; D005260:Female; D006801:Humans; D008525:Medroxyprogesterone / Q000031:analogs & derivatives / Q000494:pharmacology; D017258:Medroxyprogesterone Acetate; D009251:NADPH-Ferrihemoprotein Reductase / Q000096:biosynthesis / Q000302:isolation & purification; D012065:Relaxin / Q000494:pharmacology
D004958:Estradiol; D012065:Relaxin; D003577:Cytochrome P-450 Enzyme System; D017258:Medroxyprogesterone Acetate; D001141:Aromatase; D009251:NADPH-Ferrihemoprotein Reductase; D008525:Medroxyprogesterone