iturelix and trilostane

This study was designed to provide a genome-wide analysis of the effects of luteinizing hormone (LH) versus steroid ablation/replacement on gene expression in the developed corpus luteum (CL) in primates during the menstrual cycle. On Days 9-11 of the luteal phase, female rhesus monkeys were left untreated (control) or received a GnRH antagonist Antide (A), A + LH, A + LH + the 3beta-hydroxysteroid dehydrogenase inhibitor Trilostane (TRL) or A + LH + TRL + a progestin R5020. On Day 12 of the luteal phase, CL were removed and samples of RNA from individual CL were hybridized to Affymetrix rhesus macaque total genome microarrays. The greatest number of altered transcripts was associated with the ablation/replacement of LH, while steroid ablation/progestin replacement affected fewer transcripts. Replacement of LH during Antide treatment restored the expression of most transcripts to control levels. Validation of a subset of transcripts revealed that the expression patterns were similar between microarray and real-time PCR. Analyses of protein levels were subsequently determined for two transcripts. This is the first genome-wide analysis of LH and steroid regulation of gene transcription in the developed primate CL. Further analysis of novel transcripts identified in this data set can clarify the relative role for LH and steroids in CL maintenance and luteolysis.

Topics: 3-Hydroxysteroid Dehydrogenases; Animals; Blotting, Western; Cluster Analysis; Corpus Luteum; Dihydrotestosterone; Female; Gene Expression Regulation; Hormone Antagonists; Luteinizing Hormone; Macaca mulatta; Oligonucleotide Array Sequence Analysis; Oligopeptides; Polymerase Chain Reaction; Steroids

The factors regulating the dynamic expression of matrix metalloproteinases (MMPs) and their tissue inhibitors (TIMPs) in the primate corpus luteum (CL) during the menstrual cycle are unknown. We hypothesized that LH or progesterone (P) regulate interstitial-collagenase (MMP-1), the gelatinases (MMP-2 and -9), TIMP-1, and TIMP-2 in the CL. Hormone ablation/replacement was performed in rhesus monkeys on Days 9-11 of the luteal phase in five treatment groups (n = 4/group): control (no treatment), antide (GnRH antagonist), antide + LH; antide + LH + trilostane (TRL; 3beta-hydroxysteroid dehydrogenase inhibitor), and antide + LH + TRL + R5020 (nonmetabolizable progestin). On Day 12, the CL was removed and the RNA and protein isolated for real-time polymerase chain reaction and immunoassays, respectively. The MMP-1 mRNA increased 20-fold with antide, whereas LH replacement maintained MMP-1 mRNA at control levels. Likewise, TRL increased MMP-1 mRNA 54-fold, and R5020 prevented this effect. Immunodetectable MMP-1 protein also increased with antide or TRL; these increases were abated with LH or R5020. Gelatinase mRNA and/or protein levels increased with antide (e.g., 3-fold, MMP-2 mRNA), and LH replacement reduced protein levels (e.g., 11-fold, MMP-2). The TRL increased MMP-9, but not MMP-2, expression; however, R5020 replacement had no effect on mRNA or protein levels. The LH treatment increased TIMP-1 and -2 mRNA and TIMP-1 protein expression compared to controls and antide groups, whereas R5020 enhanced only immunodetectable TIMP-1. These data strongly suggest that LH suppresses MMP-1 in the primate CL via P and that it also suppresses gelatinases, either at the mRNA (MMP-2) or protein (MMP-2 and -9) levels, perhaps in part via steroids, including P. In contrast, LH promotes TIMP expression, perhaps via steroids, including P.

Topics: Animals; Corpus Luteum; Dihydrotestosterone; Enzyme Inhibitors; Female; Gene Expression; Hormone Antagonists; Immunohistochemistry; Luteal Phase; Luteinizing Hormone; Macaca mulatta; Matrix Metalloproteinase 1; Matrix Metalloproteinase 2; Matrix Metalloproteinase 9; Oligopeptides; Polymerase Chain Reaction; Progesterone; Promegestone; RNA, Messenger; Tissue Inhibitor of Metalloproteinase-1; Tissue Inhibitor of Metalloproteinase-2

Studies were designed to determine if ADAMTS-1 (a disintegrin and metalloproteinase with thrombospondin repeats-1) is expressed in the rhesus monkey corpus luteum (CL), is regulated by endocrine (LH) or local (progesterone) factors, and is correlated with tissue inhibitor of matrix metalloproteinase-3 (TIMP-3), an inhibitor of ADAMTS-1. PCR analyses indicated that ADAMTS-1 mRNA is expressed in luteinized granulosa cells during controlled ovarian stimulation cycles, and peaks in CL during the early luteal phase of the menstrual cycle, before decreasing (P<0.05) by the mid-late stage. Immunostaining for ADAMTS-1 was detected in luteal cells, peaking in early CL. LH and/or steroid depletion at mid-late luteal stage decreased (P<0.05) ADAMTS-1 mRNA levels compared to controls; LH but not progestin (R5020) replacement prevented this decrease. In contrast, LH and/or steroid ablation-replacement in the early CL did not affect ADAMTS-1 levels. TIMP-3 mRNA levels were lowest during the early CL and rose progressively (P<0.05), peaking in late CL. The divergent expression patterns during the CL lifespan suggest that an imbalance between ADAMTS-1 and TIMP-3 is important during luteal formation (ADAMTS-1 predominates) and regression (TIMP-3 predominates). Also, LH, perhaps via steroids other than progesterone, promotes ADAMTS-1 expression as a function of the stage of the CL.

Topics: ADAM Proteins; ADAMTS1 Protein; Animals; Corpus Luteum; Dihydrotestosterone; Disintegrins; Estradiol; Female; Hormone Antagonists; Luteinizing Hormone; Macaca mulatta; Menstrual Cycle; Metalloendopeptidases; Oligopeptides; Progesterone; RNA, Messenger; Tissue Inhibitor of Metalloproteinase-3