Role of protein synthesis, prostaglandins, and estrogen in rat ovarian metalloproteinase inhibitor production.
Matrix metalloproteinases and their inhibitors regulate follicular connective tissue remodeling associated with ovulation. We examined the control of metalloproteinase inhibitor activity and gene expression by various treatments in cultured rat granulosa cells and intact whole ovaries. Cells were isolated from preovulatory follicles of immature eCG-primed rats and cultured with various treatments for 24 h. Metalloproteinase inhibitor activity was measured in the media. The addition of LH or the phorbol ester 12-O-tetradecanoylphorbol 13-acetate (TPA) increased inhibitor activity 2.5- and 2.6-fold above that in control cultures, respectively. Cycloheximide treatment blocked basal and LH- and TPA-stimulated inhibitor activity, suggesting that the increase in granulosa cell inhibitor activity resulted from de novo protein synthesis. Indomethacin, a prostaglandin synthase inhibitor, had no effect on basal or LH-induced granulosa cell inhibitor activity. Addition of the antiestrogen tamoxifen citrate or an aromatase inhibitor, 10-propargylestr-4-ene-3,17-dione, did not affect basal or LH-stimulated inhibitor activity, implying that estrogen is not involved in the signal transduction pathway leading to increased inhibitor activity. Northern analysis demonstrated the presence of mRNA for a tissue-derived metalloproteinase inhibitor, TIMP-1, which increased with LH stimulation, in rat granulosa cells. Similarly, an hCG stimulus increased TIMP-1 mRNA in periovulatory ovaries to the highest levels prior to ovulation. Neither cycloheximide nor indomethacin altered hCG-stimulated TIMP mRNA levels in periovulatory ovaries. The present study demonstrates that the LH- and TPA-induced increase in inhibitor activity resulted from de novo protein synthesis; however, de novo protein synthesis does not appear necessary for the increase in TIMP mRNA.(ABSTRACT TRUNCATED AT 250 WORDS)
Mann
J S
JS
Department of Anatomy and Neurobiology, University of Kentucky, Lexington 40536.
Kindy
M S
MS
Hyde
J F
JF
Clark
M R
MR
Curry
T E
TE
D000818:Animals; D047072:Aromatase Inhibitors; D002478:Cells, Cultured; D006063:Chorionic Gonadotropin / Q000494:pharmacology; D003513:Cycloheximide / Q000494:pharmacology; D004965:Estrogen Antagonists / Q000494:pharmacology; D004967:Estrogens / Q000502:physiology*; D005260:Female; D015870:Gene Expression; D006023:Glycoproteins / Q000096:biosynthesis* / Q000235:genetics; D006107:Granulosa Cells / Q000187:drug effects / Q000378:metabolism; D007213:Indomethacin / Q000494:pharmacology; D008666:Metalloendopeptidases / Q000037:antagonists & inhibitors*; D010053:Ovary / Q000187:drug effects / Q000378:metabolism*; D011453:Prostaglandins / Q000502:physiology*; D014176:Protein Biosynthesis; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D013629:Tamoxifen / Q000494:pharmacology; D019714:Tissue Inhibitor of Metalloproteinases
D047072:Aromatase Inhibitors; D006063:Chorionic Gonadotropin; D004965:Estrogen Antagonists; D004967:Estrogens; D006023:Glycoproteins; D011453:Prostaglandins; D012333:RNA, Messenger; D019714:Tissue Inhibitor of Metalloproteinases; D013629:Tamoxifen; D003513:Cycloheximide; D008666:Metalloendopeptidases; D007213:Indomethacin