| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
|
|
|
|||||||
|
|||||||||
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Ovary |
Division of Reproductive Sciences,4 Oregon National Primate Research Center, Oregon Health & Science University, Beaverton, Oregon 97006
Department of Physiology and Pharmacology,5 Oregon Health & Science University, Portland, Oregon 97201
 |
ABSTRACT |
|---|
 TOP ABSTRACT INTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
|---|
corpus luteum, corpus luteum function, luteinizing hormone, ovary, progesterone
|
INTRODUCTION |
|---|
|
TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
|---|
The expression and activity of matrix metalloproteinases (MMPs) and their endogenous tissue inhibitors (TIMPs) have been implicated in the remodeling of extracellular matrix that occurs in the ovary throughout the cycle [4]. Many members of the MMP family are differentially and dynamically expressed in ovarian tissue; changes in MMP mRNA, protein, and activity levels are associated with major tissue remodeling events in the ovary [5–7]. Correlative data suggest that MMP and TIMP activity and expression are involved with formation and regression of CL in humans, rats, sheep, pigs, and cows [4, 8–11]. Recent studies in our laboratory have also elucidated expression patterns for MMP-1, -2, and -9 as well as for TIMP-1 and -2 that implicate MMP involvement in both CL formation and regression in rhesus monkeys [12]. In addition, MMPs and TIMPs are dynamically expressed in specific cell types in the CL at times of major tissue transformation that appear related to luteal structure. For example, MMP-1 and -9 as well as TIMP-1 and -2 proteins are detected in granulosa-lutein cells in the primate CL, whereas MMP-2 immunodetection is highest in cells related to the CL vasculature [12]. In addition, gelatinase mRNA and activity are present in luteal cells in the developing rat CL [13], and MMP-2 and -9 proteins are predominantly localized to large luteal cells in sheep [14].
In general, however, the factors regulating MMP and TIMP expression or activity in the CL have not been examined fully, especially in primates. We hypothesized that MMPs and TIMPs in the primate CL are regulated by pituitary and/or ovarian hormones. Gonadotropins, acting either directly or via ovarian steroids, regulate MMP expression in the macaque periovulatory follicle [7]. In addition, in humans, luteal rescue via administration of hCG can inhibit the increased expression and activity of MMP-2 that are characteristic of later luteal stages [9]. In the life span of the primate CL, P, acting through its receptor, plays an essential, local regulatory role [3, 15], perhaps in part by regulating the MMP/TIMP system. Additionally, P regulates MMPs in other tissues, reducing expression of pro-MMP-9 in the rabbit cervix and downregulating membrane type 1-MMP mRNA in the human endometrium (e.g., [16, 17]). Therefore, using an in vivo hormone ablation/replacement model to distinguish between the potential regulatory roles of LH and P in the functional primate CL, luteal mRNA expression of MMP-1, -2, and -9 as well as TIMP-1 and -2 was characterized using real-time polymerase chain reaction (PCR) and MMP/TIMP protein via immunohistochemistry or, as available, quantitative immunoassays.
|
MATERIALS AND METHODS |
|---|
|
TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
|---|
The general care and housing of rhesus monkeys (Macaca mulatta) at the Oregon National Primate Research Center (ONPRC) was described previously [18]. Menstrual cycles of adult female rhesus monkeys were monitored via assessment of daily hormone concentrations from serum samples. The first day of low serum estradiol following the midcycle estradiol peak corresponds with the day after the LH surge and is therefore termed Day 1 of the luteal phase [3, 12]. All protocols were approved by the ONPRC Animal Care and Use Committee and conducted in accordance with NIH Guidelines for the Care and Use of Laboratory Animals.
Treatment
Hormone ablation/replacement treatment was administered daily in adult rhesus monkeys on Day 9 through Day 11 of the luteal phase. These days were selected because the CL begins to regress near the end of this time period; In addition, this is the window of CL rescue in the primate by chorionic gonadotropin in fertile cycles [1]. To assess the potential regulatory role of LH and P, females were assigned randomly to one of five treatment groups (n = 4/group): control (no treatment), antide (GnRH antagonist, 3 mg/kg, s.c. injection; previously demonstrated to suppress circulating bioLH levels and luteal function [19]), antide + recombinant human LH (LH; 40 IU three times a day, intramuscular injection; Serono Reproductive Biology Institute, Rockland, MA; previously shown to restore LH function [19]), antide + LH + trilostane (TRL; a 3ß-hydroxysteroid dehydrogenase inhibitor previously demonstrated to ablate luteal P production [20]; 600 mg administered in an oral dose [8 ml of sucrose and Tang vehicle]; Sanofi Res. Div., Northumberland, UK), and antide + LH + TRL + R5020 (R5020; a nonmetabolizable progestin, 2.5 mg administered as a 2.5-ml [total volume] s.c. injection in a vehicle of sesame oil; Dupont, Boston, MA; previously shown to restore progestin function [15]). Serum estrogen and P levels were analyzed daily starting on Day 6 postmenstruation of the follicular phase through the end of treatment on Day 12 of the luteal phase. On Day 12, the CL was removed from anesthetized monkeys during an aseptic ventral midline surgery [15] and weighed, and portions were divided and either immediately frozen for subsequent RNA or protein isolation or processed for histology [19].
Hormone Assays
Serum concentrations of estradiol and P were determined by specific electrochemoluminescent assay using a Roche Elecsys 2010 analyzer (Roche, Indianapolis, IN) by the Endocrine Services Laboratory of the ONPRC. The interassay variation was 6.1% for estradiol and 5.4% for P, and the limit of sensitivity was 5 pg/ml for estradiol and 0.03 ng/ml for P. Because TRL, R5020, and high levels of the immediate precursor for P (i.e., pregnenolone) can cross-react with the P antibody [21] in the electrochemoluminescent assay, serum samples from females receiving TRL and R5020 (n = 8) were subjected to column-chromatographic analysis as reported previously [22, 23]. This technique determines P concentrations without potential cross-reaction bias. Hormone concentration values were validated against previous assays in this laboratory [24, 25].
Real-Time PCR Analysis of MMP and TIMP mRNAs
RNA Isolation Reverse transcription (RT) was carried out on 1 µg of DNase (Gibco BRL)-treated RNA using Molony Murine Leukemia Virus reverse transcriptase (Gibco BRL, Carlsbad, CA) and random hexamer primers for 2 h at 37°C as described previously [7]. The PCR was performed on individual samples of luteal cDNA generated from the RT reaction using rhesus monkey-based TaqMan primers (Perkin-Elmer Applied Biosystems, Foster City, CA) previously optimized in this laboratory for other experiments [12]. Sequence analysis was performed on the resulting PCR products. This sequence was then used to design TaqMan primer and probe sets for the real-time assay. Perkin-Elmer parameters were adhered to during probe design as described previously [12].
MMP and TIMP mRNA expression Expression of MMP and TIMP mRNA was analyzed using TaqMan PCR Core Reagent Kit with the ABI PRISM 7700 Sequence Detection System (Perkin-Elmer Applied Biosystems) as described previously [12]. To control for the amount of total RNA added to each RT reaction and to normalize target signal, 18S mRNA was used as an active endogenous control in each well. Amplifications were conducted in a final volume of 10 µl containing 250 nM TaqMan MMP/TIMP probe (labeled with the 5'-reporter dye FAM), 300–900 nM MMP/TIMP forward and reverse primers, 250 nM TaqMan 18S probe (labeled with the 5'-reporter dye VIC), 80 nM forward and reverse 18S primers, 20 ng of cDNA, and 5 µl of TaqMan Universal PCR master mix containing ROX dye as a passive reference (Perkin-Elmer Applied Biosystems). The PCR reactions were conducted as previously described [12]. The number of amplification cycles needed for the fluorescence to reach a determined threshold level (CT) was recorded for every unknown and an internal standard curve. The internal standard curve, used for relative mRNA quantification, was generated from five 10-fold dilutions of pooled cDNA from early CL samples. The CT values for unknown samples were used to extrapolate the amount of RNA equivalents from the internal standard curve. The RNA equivalent values were then divided by complimentary 18S RNA equivalent values derived from the same internal standard curve.
Immunohistochemistry
Portions of the CL were fixed in 10% neutral buffered formalin (Richard-Allen Scientific, Kalamazoo, MI) for 1 wk. Tissue was then dehydrated in a series of ethanol solutions (50%, 70%, and 100%) and paraffin embedded. Subsequently, sections (thickness, 6 µm) were prepared, mounted on Superfrost Plus slides (Fisher Scientific, Pittsburgh, PA), and then deparaffinized and hydrated through xylenes and a graded series of ethanol as reported previously [26]. Sections were incubated in PBS before pressure cooker-antigen retrieval in citrate buffer (Citra; BioGenex Laboratories, Inc., San Ramon, CA). Endogenous peroxidases were quenched, and sections were placed in a blocking buffer and incubated with primary antibody in normal horse serum/PBS buffer for 1 h at room temperature and then overnight at 4°C as previously described [12]. Concentrations for anti-human MMP-1 (M35L Ab-1; Oncogene, Cambridge, MA) and TIMP-1 (MS 608; Neomarkers, Freemont, CA) antibodies were diluted to 1:200. Primary antibody was detected using a biotinylated anti-rabbit immunoglobulin G secondary antibody (1:1000; Vector Laboratories, Burlingame, CA) and the Vector ABC-Elite Kit (Vector Laboratories), visualized with Sigma Fast diaminobenzidine substrate (Sigma, St. Louis, MO), and counterstained with hematoxylin. For both MMP-1 and TIMP-1, negative controls lacking primary antibody were processed on adjacent tissue sections.
Protein Extraction and Quantitative Immunoassay
Protein was extracted from frozen portions of the CL using a tissue homogenizer with tissue protein extraction reagent (T-PER; Pierce Biotechnology, Rockford, IL) and protease inhibitors (Halt Protease Inhibitor Cocktail Kit, EDTA-free; Pierce Biotechnology) as recommended by the manufacturer. Total protein concentration was determined using the Bradford assay (Bio-Rad, Hercules, CA). Total gelatinase (MMP-2 and -9) protein content of each sample was quantified using the Quantikine human MMP-2 and -9 enzyme-linked immunosorbent assay kits (R&D Systems, Minneapolis, MN). Each sample was run in duplicate, and optical density values at 540 nm were averaged after equivalence to protein-specific standard curve.
Statistical Analysis
Statistical evaluation of mean differences among experimental groups was performed by ANOVA or ANOVA on Ranks with significance level set at 0.05 using the SigmaStat software package (SPSS, Chicago, IL). To isolate significant differences between groups, the Student-Newman-Keuls method was used for the pairwise multiple comparisons.
|
RESULTS |
|---|
|
TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
|---|
Before treatment, serum P concentrations increased in all monkeys during the early luteal phase (Days 0–6) to concentrations typical of natural menstrual cycles or those in untreated controls [12]. Treatment with antide significantly decreased systemic P levels by 24 h postadministration. Serum P concentrations were significantly less than those of controls (Fig. 1, A and B), but addition of LH maintained P concentrations at a level similar to control values. Treatment with TRL also caused a comparable (P < 0.05) reduction in serum P (Fig. 1, A and B). Serum P concentrations remained low in the R5020 group as determined by column chromatography, although assay of nonchromatographed samples confirmed the presence of high "progestin" levels (Fig. 1B). After the periovulatory estradiol surge, estradiol concentrations decreased and were low throughout the luteal phase (Table 1). Treatment with antide or TRL further reduced serum estradiol levels compared to those in animals receiving LH, (P < 0.05), but not R5020, replacement.
|
|
No differences in CL mass were observed between treatment protocols (p = 0.49) (Table 1). Control tissues displayed morphology typical of the macaque CL in the late luteal phase of the menstrual cycle [19] (Fig. 2A). However, treatment with antide (Fig. 2B) or TRL (Fig. 2D) appeared to reduce the volume of steroidogenic cells (e.g., compare the field of view in Fig. 2, C vs. D) and to increase the observance of neutrophils (Fig. 2E). The replacement of LH (Fig. 2C) or R5020 (Fig. 2F) to antide or TRL treatment groups, respectively, reversed these effects: Steroidogenic cell volume noticeably increased, and neutrophils were less apparent in these tissues. However, these observations were not quantified.
|
Real-Time PCR Analysis of MMP and TIMP mRNAs
Interstitial collagenase (MMP-1) The MMP-1 mRNA was detectable in control (Day 12) tissues, and levels increased 20-fold compared to controls with antide treatment (P < 0.05) (Fig. 3A). In vivo replacement of LH returned MMP-1 mRNA expression to control levels. The TRL treatment also increased MMP-1 mRNA expression up to 54-fold compared with controls and antide + LH tissues (P < 0.05) (Fig. 3A). Replacement of P with R5020 treatment reduced MMP-1 mRNA expression compared with antide + LH + TRL (P < 0.05) (Fig. 3A); however, MMP-1 mRNA expression remained somewhat elevated compared to controls and antide + LH-treated tissues (P < 0.05).
|
Gelatinases (MMP-2 and -9) The MMP-2 (gelatinase A) mRNA was moderately expressed in control tissues, and levels increased 3-fold with antide treatment (P < 0.05) (Fig. 3B). However, neither LH replacement, TRL treatment, nor P replacement had further effects on MMP-2 mRNA expression. The MMP-9 (gelatinase B) mRNA levels were substantial in controls and unchanged by antide or LH replacement. Expression increased 3-fold with TRL treatment (P < 0.05) (Fig. 3C); however, replacement of P with R5020 did not reverse the increase in MMP-9 mRNA levels by TRL.
Tissue Inhibitors of MMPs (TIMP- and -2) Luteal TIMP-1 mRNA expression in antide-treated animals was similar to the substantial levels observed in controls (Fig. 4A). However, LH replacement increased TIMP-1 mRNA at least 2-fold compared with all other groups (P < 0.05) (Fig. 4A). The TRL treatment reduced the LH-induced increase; however, P replacement with R5020 did not restore high TIMP-1 mRNA expression levels (P < 0.05) (Fig. 4A). Alternatively, TIMP-2 mRNA expression increased (P < 0.05) compared to controls with both antide (2.7-fold) and LH treatment (4-fold) (Fig. 4B). The TRL treatment again reduced TIMP-2 mRNA levels compared to antide + LH, but R5020 did not restore TMP-2 expression.
|
Immunohistochemistry for MMP-1 and TIMP-1
Immunolabeling of MMP-1 was detected at low levels in granulosa-lutein cells in control tissues (Fig. 5A). Immunostaining for MMP-1 was not noted in stroma, endothelial cells, or theca-lutein cells for all groups investigated. Administration of antide increased staining intensity. Immunolabeling of MMP-1 was strongest in the perinuclear region of granulosa-lutein cells; however, it was observed throughout the cytoplasm (Fig. 5B, see insets for details). Restoration of LH prompted a decline in MMP-1 immunodetection; few strongly labeled steroidogenic cells were noted (Fig. 5C). Addition of TRL also increased MMP-1 immunodetection; granulosa-lutein cells with intense staining were observed (Fig. 5D). Staining of MMP-1 decreased with replacement of progestin in the R5020-treated tissues (Fig. 5E). No staining was noted in tissues processed without primary antibody (Fig. 5F).
|
Immunolabeling of TIMP-1 was detected in all groups. Staining appeared to concentrate in granulosa-lutein cells, whereas stroma, theca-lutein, and endothelial cells did not label (Fig. 6B). Control tissue displayed moderate to low labeling, as did antide-treated tissues (Fig. 6, A and B). The LH replacement increased TIMP-1 immunodetection among granulosa-lutein cells; labeling was diffuse throughout the cytoplasm (Fig. 6C). The TRL treatment decreased TIMP-1 immunodetection to levels similar to those of control tissues (Fig. 6D). Replacement of P with R5020 resulted in appreciable levels of TIMP-1 immunolabeling in granulosa-lutein cells (Fig. 6E). No staining was noted in tissues sections processed without primary antibody (Fig. 6F).
|
Quantitative Immunoassays for MMP-2 and -9
Total protein concentrations for MMP-2 were high in control tissues (Fig. 7A). In vivo treatment with antide did not alter MMP-2 protein levels in the CL. However, LH replacement reduced (P < 0.05) total MMP-2 protein levels in the CL by 8-fold compared to both control and antide tissues. After LH replacement, TRL without R5020 replacement failed to alter the lowered levels of MMP-2 protein (P < 0.05) (Fig. 7A); no significant differences were observed among antide + LH, antide + LH + TRL, and antide + LH + TRL + R5020 groups (Fig. 7A).
|
Total MMP-9 protein was detected in the CL of controls and increased significantly with antide treatment (Fig. 7B). The LH replacement significantly decreased total MMP-9 protein levels compared to the antide-treated group (P < 0.05) (Fig. 7B). The TRL treatment tended (P = 0.07) to increase total MMP-9 protein levels, with P replacement returning MMP-9 protein to levels comparable to those in LH-treated or control groups (Fig. 7B).
|
DISCUSSION |
|---|
|
TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
|---|
The potential regulatory roles of LH and ovarian steroid hormones on MMP/TIMP expression in the primate CL are complex. The clearest relationship appears to involve MMP-1, because the effects of antide in enhancing MMP-1 mRNA levels and immunostaining were mimicked by TRL in LH-clamped (antide + LH) animals. Moreover, the increases in MMP-1 mRNA and protein immunolabeling were reversed by replacement of either LH or R5020, respectively. Thus, the suppression of MMP-1 mRNA and protein by LH is likely indirect, via P produced from LH-stimulated steroidogenesis. The inhibition of MMP-1 by P is reminiscent of our earlier evidence that P suppresses estrogen receptor ß in the developed CL in rhesus monkeys during the menstrual cycle [15]. In addition, P negatively regulates proMMP-1 protein in stromal cells from both human and macaque endometrium [27]. However, this effect of P is opposite that observed in granulosa cells of periovulatory follicles during controlled ovarian stimulation cycles in macaques. The TRL treatment blocked MMP-1 expression following the ovulatory hCG bolus, whereas R5020 replacement mimicked hCG induction of MMP-1 mRNA [7]. Thus, P appears to promote MMP-1 expression in the ovulatory luteinizing follicle but to suppress its expression in the developed CL. We previously reported that MMP-1 mRNA and protein expression peaks in the developing CL at the early luteal phase [12] and declines markedly by the mid-luteal phase of the menstrual cycle in rhesus monkeys. Further studies are warranted to examine the dynamics of P regulation of MMP-1 expression at other stages in the life span of the primate CL, especially during luteal development postovulation and luteal rescue in early pregnancy.
In contrast to MMP-1, the effects of antide and TRL treatment, as well as of LH or P replacement, differed markedly for the gelatinases (MMP-2 and -9). Expression of MMP-2 in the developed CL appears to be negatively regulated by LH, because gonadotropin depletion by antide increased MMP-2 mRNA levels and LH replacement decreased MMP-2 protein levels. The lack of comparable effects on both mRNA and protein by antide and LH may reflect the variability between individual/samples or temporal differences in the optimal time for analyzing MMP-2 expression. Nevertheless, the remarkable suppression of MMP-2 protein levels by LH replacement, to levels well below those of control animals, is similar to the reported decrease in MMP-2 expression in the human CL during hCG-mediated luteal rescue during early simulated pregnancy [9]. Because no effects of TRL or TRL + R5020 were noted in the LH-clamped (antide + LH) model, it is unlikely that steroids, including P, play a role in regulating MMP-2 in the macaque CL. Thus, the LH/chorionic gonadotropin suppression of MMP-2 protein in primate luteal tissue is not mediated by LH-stimulated steroidogenesis. Regulation by steroids may differ among tissue types: MMP-2 appears to be positively regulated in vitro by estrogen in the retina [28] and negatively regulated by estrogen exposure in heart tissue [29]. In addition, in vitro administration of medroxyprogesterone acetate, but not estradiol, to endometrial cancer cell lines decreased the secretion of proMMP-9, proMMP-2, and MMP-2 [30]. Examination of other nonsteroidogenic factors, such as cytokines or tissue inhibitors, should also be performed to complement these studies of steroid regulation.
Analyses of MMP-9 mRNA and protein suggest that LH and steroids act at different levels to control expression of this gelatinase in the developed CL. Antide treatment and LH replacement had no effect on MMP-9 mRNA levels. However, antide markedly increased MMP-9 protein, and LH replacement reduced MMP-9 protein back to control levels. In contrast, TRL treatment in LH-clamped (antide + LH) animals substantially increased MMP-9 mRNA levels, but changes were less robust at the protein level. Notably, P replacement restored MMP-9 protein to levels observed with antide + LH treatment but was less effective at the mRNA level. These data suggest that MMP-9 is differentially regulated in the developed macaque CL. Progesterone or other ovarian steroids may suppress transcription, whereas LH may also act to suppress translation of this gelatinase. Importantly, P also suppresses MMP-9 expression and activity in endometrial explants and the rabbit cervix [16, 31]. It is possible that LH and steroids act synergistically to suppress MMP-9 expression in a number of reproductive tissues, including the primate CL. Such synergistic action is consistent with the dynamics of MMP-9 and, to a lesser extent, MMP-2 in the macaque CL, which peak at the late to very late luteal phase when sensitivity to gonadotropins and P secretion declines (reviewed in [32]) and, thereby, reduces this suppressive action on gelatinase production.
In contrast to the MMPs, gonadotropin and steroids appear to promote luteal TIMP-1 and -2 expression. Antide treatment had little effect on TIMP-1 mRNA or protein levels, but LH replacement markedly enhanced both parameters. Because TRL treatment reduced both mRNA and protein expression to levels comparable to those with antide alone, one can hypothesize that ovarian steroids mediate, at least in part, LH action to promote TIMP-1 expression. Because R5020 replacement increased TIMP-1 immunostaining in TRL-treated, LH-clamped animals, P may be one such mediator. However, R5020 treatment did not increase TIMP-1 mRNA levels, which suggests that other steroids, such as estrogens or androgens, may influence TIMP-1 expression. Thus, promotion of TIMP-1 production by LH may be indirect, via P and/or other steroids acting differentially at the protein and mRNA level, respectively.
Administration of both antide and TRL decreased estrogen and, presumably, androgens compared to LH-treated tissues in the CL, and TIMP-2 as well as TIMP-1 mRNA likely are regulated by a gonadal steroid other than P. Future studies will be necessary that add back estradiol or a potent androgen (dihydrotestosterone; e.g., [22]) to identify the steroid(s) regulating TIMP-2 expression in the primate CL. Although TIMP-2 protein was not analyzed in the present study, the effects of various treatments on TIMP-2 mRNA levels were qualitatively similar to those for TIMP-1. One difference was the modest increase in TIMP-2 mRNA expression by antide alone, which was reminiscent of the effect of antide alone on MMP-2 mRNA. Because the addition of LH either further increased (TIMP-2) or had no effect (MMP-2), as opposed to reversing the antide effect, we cannot rule out that the GnRH antagonist elicits other effects independent of blocking LH secretion. Some evidence points toward GnRH receptors in the primate CL [33, 34], but the physiologic relevance of GnRH actions on luteal tissue remains controversial. Nevertheless, the data collected to date from the periovulatory follicle [7] and the developed CL (present study) suggest that LH promotes TIMP-1 and -2 expression, at least in part, via local steroid action, including that of P. These data are consistent with reports that TIMP levels decline in the regressing CL in rhesus monkeys and other species (e.g., [11, 12]).
Hormone ablation and replacement in the developed CL affected the intensity and frequency, but not the cell-specific localization, of immunostaining for both MMP-1 and TIMP-1. As reported previously [12], both MMP-1 and TIMP-1 protein were detected primarily in granulosa-lutein cells of the macaque CL during the natural menstrual cycle. The current evidence for LH and steroid regulation of luteal MMP-1 as well as TIMP-1 and -2 is consistent with reports that granulosa-lutein cells in primates contain receptors for LH, P, estrogen, and androgen, and are targets for these factors. However, it is not clear whether LH and steroid regulation of luteal gelatinases is direct, because these MMPs are detected exclusively in cells of microvasculature (MMP-2 [12]) or in lutein and microvasculature cells (MMP-9 [12]). In addition, because neutrophils can stimulate MMP-2 and -9 expression in other tissues [35] and neutrophil accumulation in macaque luteal tissue appears to be regulated by LH and P [1; present study], a possible role for immune cells in the hormonal and local control of luteal MMPs/TIMPs warrants evaluation. Further studies combining markers of LH/steroid receptor-signal transduction with MMP/TIMP expression or, importantly, enzyme activity are needed to define the cell sites and mechanisms controlling proteases in the primate CL.
In conclusion, the present study presents, to our knowledge, the first evidence in support of hormonal (LH) and local (steroid) control of the MMP/TIMP family of proteases/inhibitors in the developed CL of primates during the menstrual cycle. The action of LH to suppress MMP-1 mRNA and protein levels appears to be indirect, via LH-stimulated P production. In addition, LH suppresses gelatinase expression, perhaps in part via steroids, including P. The actions of LH and steroids at different levels (e.g., steroids suppress MMP-9 mRNA expression, whereas LH suppresses MMP-9 protein levels) may synergize to control MMP/TIMP action. In contrast, LH promotes TIMP-1 and -2 expression, at least in part via steroids, including P. The divergent effects on MMP and TIMP patterns may be important for controlling tissue remodeling in the primate CL. However, further studies of MMP/TIMP expression and, importantly, activity are needed at other stages in the luteal life span, with attention to the various cell types in the CL, to better define the regulation and role of proteases in primate luteal structure and function.
|
ACKNOWLEDGMENTS |
|---|
|
FOOTNOTES |
|---|
2 Correspondence: Richard L. Stouffer, Division of Reproductive Sciences, Oregon National Primate Research Center, Oregon Health & Science University, 505 NW 185th Ave., Beaverton, OR 97006. FAX: 503 690 5563; stouffri{at}ohsu.edu
3 Current address: Department of Biological Sciences, California State University, Long Beach, Long Beach, CA 90840
Received: 6 August 2003.
First decision: 28 August 2003.
Accepted: 11 September 2003.
|
REFERENCES |
|---|
|
TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
|---|
and ß in the rhesus monkey corpus luteum during the menstrual cycle: regulation by luteinizing hormone and progesterone. Endocrinology 2000 141:1711-1717-induced, but not spontaneous, luteal regression in rhesus monkeys. Biol Reprod 1990 43:507-516[Abstract]This article has been cited by other articles:
|
|
 |
|
  J. W. Wright, T. Pejovic, J. Fanton, and R. L. Stouffer Induction of proliferation in the primate ovarian surface epithelium in vivo Hum. Reprod., January 1, 2008; 23(1): 129 - 138. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 G. Irusta, M.J. Murphy, W.D. Perez, and J.D. Hennebold Dynamic expression of epoxyeicosatrienoic acid synthesizing and metabolizing enzymes in the primate corpus luteum Mol. Hum. Reprod., August 1, 2007; 13(8): 541 - 548. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 K. Miyakoshi, M. J. Murphy, R. R. Yeoman, S. Mitra, C. J. Dubay, and J. D. Hennebold The Identification of Novel Ovarian Proteases Through the Use of Genomic and Bioinformatic Methodologies Biol Reprod, December 1, 2006; 75(6): 823 - 835. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 J. M. Castellano, M. Gaytan, J. Roa, E. Vigo, V. M. Navarro, C. Bellido, C. Dieguez, E. Aguilar, J. E. Sanchez-Criado, A. Pellicer, et al. Expression of KiSS-1 in Rat Ovary: Putative Local Regulator of Ovulation? Endocrinology, October 1, 2006; 147(10): 4852 - 4862. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 M. C Peluffo, L. Bussmann, R. L Stouffer, and M. Tesone Expression of caspase-2, -3, -8 and -9 proteins and enzyme activity in the corpus luteum of the rat at different stages during the natural estrous cycle. Reproduction, September 1, 2006; 132(3): 465 - 475. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. Bu, C. Cao, Y. Yang, C. Miao, Z. Hu, Y. Cao, Q. A. Sang, and E. Duan Localization and temporal regulation of tissue inhibitor of metalloproteinases-4 in mouse ovary. Reproduction, June 1, 2006; 131(6): 1099 - 1107. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. Tesone, R. L. Stouffer, S. M. Borman, J. D. Hennebold, and T. A. Molskness Vascular Endothelial Growth Factor (VEGF) Production by the Monkey Corpus Luteum During the Menstrual Cycle: Isoform-Selective Messenger RNA Expression In Vivo and Hypoxia-Regulated Protein Secretion In Vitro Biol Reprod, November 1, 2005; 73(5): 927 - 934. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
A. McDonnel Smedts and T.E. Curry Jr. Expression of Basigin, an Inducer of Matrix Metalloproteinases, in the Rat Ovary Biol Reprod, July 1, 2005; 73(1): 80 - 87. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
F. Xu and R. L. Stouffer Local Delivery of Angiopoietin-2 into the Preovulatory Follicle Terminates the Menstrual Cycle in Rhesus Monkeys Biol Reprod, June 1, 2005; 72(6): 1352 - 1358. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 M. C. Peluffo, K. A. Young, and R. L. Stouffer Dynamic Expression of Caspase-2, -3, -8, and -9 Proteins and Enzyme Activity, But Not Messenger Ribonucleic Acid, in the Monkey Corpus Luteum during the Menstrual Cycle J. Clin. Endocrinol. Metab., April 1, 2005; 90(4): 2327 - 2335. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
J. Xu, R.L. Stouffer, R.P. Searles, and J.D. Hennebold Discovery of LH-regulated genes in the primate corpus luteum Mol. Hum. Reprod., March 1, 2005; 11(3): 151 - 159. [Abstract] [Full Text] [PDF]
|
|
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||
