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Biomedical subjects

L C Giudice

Publications and source records attributed to L C Giudice.

At least 55 records · Page 3Linked to original sources
PubMed1998

Hypoxia stimulates insulin-like growth factor binding protein 1 (IGFBP-1) gene expression in HepG2 cells: a possible model for IGFBP-1 expression in fetal hypoxia.

IGFBP-1 is elevated in fetuses with long-term, chronic hypoxia and intrauterine growth restriction. We investigated the hypothesis that hypoxia regulates IGFBP-1 in the human fetus in vivo and IGFBP-1 gene expression and protein in vitro. Umbilical artery IGFBP-1 levels (mean +/- SEM) from term babies with respiratory acidosis (acute hypoxia), normal babies, and those with mixed respiratory/metabolic acidosis (more profound and prolonged hypoxia) were measured using an immunoradiometric assay. IGFBP-1 levels were similar in normal (n = 12) and acutely hypoxic (n = 6) babies (189.1 +/- 71.8 vs. 175.8 +/- 45.9 ng /ml, respectively, P = 0.789). However, with more profound and prolonged hypoxia (n = 19), IGFBP-1 levels were markedly elevated (470.6 +/- 80.0 ng /ml, P = 0.044). To investigate IGFBP-1 regulation by hypoxia in vitro, HepG2 cells were incubated under hypoxia (pO2 = 2%) and normoxia (pO2 = 20%). IGFBP-1 protein and mRNA increased 8- and 12-fold, respectively, under hypoxic conditions. Hypoxia did not affect protein or mRNA levels of IGFBP-2 or -4. IGFBP-5 and -6 mRNAs, undetectable in control cells, were not induced by hypoxia, whereas minimally expressed IGFBP-3 mRNA increased twofold. Investigation into IGFBP-1 gene structure revealed three potential consensus sequences for the hypoxia response element (HRE) in the first intron. To investigate functionality, a 372-bp fragment of IGFBP-1 intron 1, containing putative HREs, was placed 5' to a heterologous hsp70 promoter in a plasmid using luciferase as a reporter gene. Under hypoxia, reporter gene activity increased up to 30-fold. Mutations in the middle HRE abolished reporter activity in response to hypoxia, suggesting that this HRE is functional in the IGFBP-1 hypoxia response. Cotransfection of HRE reporter genes with a constitutively expressing hypoxia-inducible factor 1 plasmid in HepG2 cells resulted in a fourfold induction of reporter activity, suggesting a role for hypoxia-inducible factor 1 in hypoxia induction of IGFBP-1 gene expression. These data support the hypothesis that hypoxia regulation of IGFBP-1 may be a mechanism operating in the human fetus to restrict insulin-like growth factor-mediated growth in utero under conditions of chronic hypoxia and limited substrate availability.

Animals↗
PubMed1998

Growth factor regulation of insulin-like growth factor binding protein secretion by cultured human granulosa-luteal cells.

OBJECTIVE: To examine the effects of epidermal growth factor (EGF), transforming growth factor-beta (TGF-beta), and fibroblast growth factor (FGF) on insulin-like growth factor binding protein (IGFBP) secretion by cultured human granulosa-luteal cells. DESIGN: Granulosa-luteal cells obtained at the time of oocyte harvest for IVF were cultured in serum-free medium in the presence or absence of EGF, TGF-beta, or FGF. Conditioned medium then was analyzed by Western ligand blot and immunoradiometric assays. SETTING: An academic medical center. PATIENT(S): Women undergoing IVF. INTERVENTION(S): None. MAIN OUTCOME MEASURE(S): IGFBP-1 secretion. RESULT(S): By Western ligand blot analysis, IGFBP-1 levels were 1.4-fold to 7.4-fold higher in conditioned medium from cells cultured in the presence of EGF than in control medium. By immunoradiometric assay, IGFBP-1 levels increased from 1.6 to 9.9 times over control. The TGF-beta had no apparent effect, and FGF did not consistently stimulate IGFBP-1 secretion. CONCLUSION(S): The EGF may decrease intrafollicular bioavailable IGF levels by increasing inhibitory IGFBPs, thereby leading to arrest of follicular development. Interactions between the EGF and IGF systems may be involved in the processes governing human ovarian follicle maturation and atresia.

Cells, Cultured↗
PubMed1998

Paracrine actions of insulin-like growth factors and IGF binding protein-1 in non-pregnant human endometrium and at the decidual-trophoblast interface.

Insulin-like growth factors, IGF-I, IGF-II, and IGF binding protein (IGFBP-1) appear to play major roles in endometrial development during the menstrual cycle and in the process of implantation. The mitogenic, differentiative, and anti-apoptotic properties of these growth factors, as well as their spatial and temporal expression in cycling endometrium, suggest that they may participate in endometrial growth, differentiation, inhibition of apoptosis, and perhaps angiogenesis. IGFBP-1 is a major protein product of non-pregnant endometrium during the mid-late secretory phase and occurs in abundance in decidua. Its roles as an IGF-binding protein and as a trophoblast integrin ligand suggest that it may have multiple roles in endometrial development and in interactions between the decidua and the invading trophoblast. Precise elucidation of the mechanisms underlying IGF and IGFBP-1 action at the decidual-trophoblast interface in early pregnancy awaits further investigation. The future also awaits elucidation of the potential predictive utility of IGFBP-1 in serum and in decidua in, for example, pre-eclampsia and perhaps implantation failure.

Animals↗
PubMed1998

Insulin-like growth factor binding protein-1 binds to placental cytotrophoblast alpha5beta1 integrin and inhibits cytotrophoblast invasion into decidualized endometrial stromal cultures.

Insulin-like growth factor binding protein-1 (IGFBP-1) can bind to the alpha5beta1 integrin and stimulate cellular migration in Chinese hamster ovary (CHO) cells. IGFBP-1 is a major product of the endometrium of pregnancy (decidua), and may interact with invading cytotrophoblasts expressing alpha5beta1 integrin to modulate their invasion. The present study investigated IGFBP-1 interaction with cytotrophoblast alpha5beta1 integrin, and its effects on trophoblast attachment to fibronectin and invasion into decidualized endometrial stromal cell multilayers. IGFBP-1 incubated with cytotrophoblast extracts was co-precipitated by an antibody to the alpha5 integrin subunit. Up to 55% of radiolabeled IGFBP-1 bound to cytotrophoblasts was displaced by excess non-radioactive IGFBP-1, but not by IGFBP-3. Cytotrophoblast attachment to fibronectin was inhibited by an RGD-containing octapeptide, by antibodies to the alpha5 subunit or the alpha5beta1 heterodimer, and by IGFBP-1. Cytotrophoblasts showed limited invasion into endometrial stromal multilayers decidualized in vitro secreting abundant IGFBP-1, but invaded multilayers when IGFBP-1 production was inhibited by insulin. Invasion into insulin-treated multilayers was prevented by addition of exogenous IGFBP-1 but not by IGFBP-3. These findings suggest IGFBP-1 may modulate trophoblast invasiveness.

Animals↗
PubMed1998

Medical management of interstitial ectopic pregnancy: a case report and literature review.

Recent reports describe successful treatment of interstitial ectopic pregnancies using methotrexate. While the number of reported cases is increasing, no consensus exists regarding the management of this complication of pregnancy. We present the successful use of combined systemic and direct intrasac injection of methotrexate for an interstitial pregnancy with the highest yet reported initial beta-human chorionic gonadotrophin concentration (102,000 mIU/ml). We also describe the use of Doppler ultrasound for monitoring treatment progression. Through a review of the current literature, we propose to facilitate management decisions and increase outcome success by summarizing previously reported treatment regimens and by describing enhanced parameters for patient selection and monitoring.

Adult↗
PubMed1998

Status of current research on endometriosis.

Endometriosis, a benign gynecologic disorder, occurs in about 10% of women of reproductive age and in up to 50% of women with infertility. Endometriosis is defined as the presence of endometrial glandular and stromal cells outside their normal location in the uterus. Commonly affected areas in the abdominopelvic cavity include the ovaries, the cul-desac and other kinds of pelvic peritoneum, bowel and diaphragm. Rarely is it found in extraabdominal sites, including the pleura and pericardium. While it is not a malignant disorder, endometriosis exhibits cellular proliferation, cellular invasion and neoangiogenesis. The steroid hormone dependence of endometriosis is underscored by its appearance during the reproductive years. Furthermore, the progress of this enigmatic disease can be tempered by administration of antiestrogens, inhibitors of endogenous estradiol production, and hormonal and surgical castration. It is a disorder that markedly affects well-being and physical and emotional health in women. Research on the pathogenesis of endometriosis currently interfaces with four areas of basic research, including the fields of genetics, environmental science, cancer biology and immunology. Here we focus on current research in the latter two disciplines and their relevance to endometriosis research.

Cytokines↗
PubMed1998

Dysfunctional uterine bleeding.

Abnormal uterine bleeding is a common, debilitating condition. Dysfunctional uterine bleeding (DUB) is the diagnosis given to women with abnormal uterine bleeding in whom no clear etiology can be identified. DUB has been observed in both ovulatory and anovulatory cycles. Medical treatments include nonsteroidal anti-inflammatory drugs, oral contraceptive pills, progestins, danazol (a synthetic androgen), GnRH agonists, and antifibrinolytic drugs. The drawback to medical therapy, in addition to side effects, is that the benefit lasts only while the patient takes the medication. Surgical options have concentrated mainly on endometrial ablation and hysterectomy, and it is unclear whether one is superior to the other in terms of long-term outcome and patient satisfaction. Newer and less invasive ablation techniques, such as thermal balloon ablation, offer more treatment alternatives.

Adolescent↗
PubMed1997

Multifaceted roles for IGFBP-1 in human endometrium during implantation and pregnancy.

IGFBP-1 is a major protein product of nonpregnant endometrium during the mid-late secretory phase and occurs in abundance in decidua. Its roles as an IGF-binding protein and as a trophoblast integrin ligand suggest that it may have multiple roles in endometrial development and in interactions between the decidua and the invading trophoblast. IGFBP-1 in vaginal/cervical secretions has already had clinical application as a predictor of premature rupture of fetal membranes. The future awaits elucidation of the potential utility of IGFBP-1 in serum and in decidua in predicting fetal growth restriction and preeclampsia and perhaps implantation failure.

Embryo Implantation↗
PubMed1997

Insulin-like growth factor binding protein-1 at the maternal-fetal interface and insulin-like growth factor-I, insulin-like growth factor-II, and insulin-like growth factor binding protein-1 in the circulation of women with severe preeclampsia.

OBJECTIVES: Preeclampsia is characterized by maternal hypertension, proteinuria, edema, and shallow placental invasion. Insulin-like growth factor binding protein-1, abundant in maternal decidua, is believed to play a role in limiting trophoblast invasiveness. In this study we addressed the hypothesis that this binding protein is aberrantly expressed in preeclampsia. We also investigated circulating levels of insulin-like growth factor-I and insulin-like growth factor-II in subjects with severe preeclampsia compared with controls. STUDY DESIGN: Insulin-like growth factor binding protein-1 was investigated by immunohistochemistry at the maternal-fetal interface of eight pregnancies complicated by severe preeclampsia and six controls between 21 and 34 weeks of gestation. Cell types were identified with use of cell-specific markers. Circulating levels of insulin-like growth factor binding protein-1, insulin-like growth factor-I, and insulin-like growth factor-II in 16 patients with severe preeclampsia and 29 controls at the same gestational age were determined by an immunoradiometric assay and correlated with clinical parameters. Data were analyzed by t test and Pearson's method. RESULTS: Insulin-like growth factor binding protein-1 was highly expressed on syncytiotrophoblasts, cytotrophoblasts, and decidual cells but not on placental fibroblasts. Immunostaining was greater at the maternal-fetal interface in severe preeclamptic patients compared with controls. Circulating insulin-like growth factor binding protein-1 levels in subjects with severe preeclampsia were 428.3 +/- 85.9 ng/ml compared with 76.6 +/- 11.8 in controls (p = 0.0007). Circulating insulin-like growth factor-I levels were 80.9 +/- 17.2 ng/ml compared with 179.4 +/- 28.2 ng/ml in controls (p = 0.0001). In contrast, insulin-like growth factor-II levels were not significantly different in the two groups. In subjects with severe preeclampsia insulin-like growth factor binding protein-1 levels correlated with diastolic blood pressure (r = 0.498, p 0.049) and aspartate transcarbamylase (0.621, p = 0.010). CONCLUSIONS: The abundance of insulin-like growth factor binding protein-1 at the maternal-fetal interface in severely preeclamptic pregnancies suggests that the binding protein may participate in the pathogenesis of the shallow placental invasion observed in this disorder. Low circulating insulin-like growth factor-I and elevated insulin-like growth factor binding protein-1 levels may contribute to restricted placental and therefore fetal growth.

Adolescent↗
PubMed1997

Insulin-like growth factors and insulin-like growth factor binding proteins in androgen-dominant ovarian follicles from testosterone-treated female-to-male trans-sexuals.

OBJECTIVE: To determine insulin-like growth factor (IGF)-I and IGF-II levels, IGF binding protein (IGFBP) profile, and IGFBP-4 protease activity in androgen-dominant follicular fluid (FF) from female-to-male trans-sexuals and to compare with those in follicles from normocycling women. DESIGN: Follicular fluid samples were obtained from four female-to-male trans-sexuals and 15 women with normo-ovulatory cycles at the Dijkzigt Academic Hospital. Western ligand blot analysis and protease assays were used to determine IGFBP profile, and immunoradiometric assays were used to detect IGF levels. SETTING: The study was performed in two academic medical centers. PATIENT(S): Female-to-male trans-sexuals and women with normo-ovulatory cycles. INTERVENTIONS: None. MAIN OUTCOME MEASURE(S): Determination of IGF levels and IGFBP profile. RESULT(S): Insulin-like growth factor I levels in FF from female-to-male trans-sexuals were not significantly different compared with levels in androgen-dominant FF and estrogen-dominant FF. Significantly lower levels of IGF-II were observed in FF from female-to-male trans-sexuals than in estrogen-dominant FF, whereas IGF-II levels in FF from female-to-male trans-sexuals were not significantly different than those in androgen-dominant FF. Similar IGFBP profiles from FF from female-to-male trans-sexuals and androgen-dominant FF were noted, with markedly elevated levels of the 31- and 24-kd IGFBPs and a 28-kd IGFBP, compared with estrogen-dominant FF. An IGFBP-4-specific metalloserine protease that is active in estrogen-dominant FF likewise was undetected in FF from female-to-male trans-sexuals. CONCLUSION(S): Follicles developing under the influence of exogenous androgens in ovaries in female-to-male trans-sexuals appear to be similar to androgen-dominant follicles in normo-ovulatory women with regard to IGF-I and IGF-II levels, IGFBP profile, and the absence of IGFBP-4 protease activity.

Androgens↗
PubMed1997

Developmental regulation of telomerase activity in human fetal tissues during gestation.

Telomerase is a ribonucleoprotein that adds hexanucleotide repeats (telomeres) to the ends of linear chromosomes, compensating for the loss of telomeric DNA which occurs with DNA replication. In humans, telomerase has been previously detected in germ-line tissues, blastocysts, 16-20 week old fetal tissue, and most cancers, but not in mature sperm or ova, or in most normal somatic tissues. It has been hypothesized that telomerase is suppressed during somatic development and reactivated in malignancy. To test the hypothesis that telomerase is suppressed during somatic development, human fetal tissues of 8-21 weeks gestational age were assayed for telomerase activity. All tissues expressed telomerase at the earliest ages examined. Lung, liver, spleen, and testis maintained telomerase activity through the latest age assayed, namely 21 weeks. Brain and kidney telomerase activity was present up to the 16th week and was undetectable thereafter. Heart tissue did not display activity beyond the 12th week. Lysates of heart, brain, and kidney without telomerase activity did not inhibit the activity of known telomerase-positive cells, suggesting that suppression of telomerase activity during gestational development is due to a lack of active telomerase rather than to the presence of an inhibitor. These findings demonstrate tissue-specific and developmental regulation of telomerase in the human fetus, suggesting an important role for this ribonucleoprotein in human fetal tissue differentiation and development.

Brain↗
PubMed1997

Gestational age-dependent expression of insulin-like growth factor-binding protein-1 (IGFBP-1) phosphoisoforms in human extraembryonic cavities, maternal serum, and decidua suggests decidua as the primary source of IGFBP-1 in these fluids during early pregnancy.

The insulin-like growth factors (IGFs) and their binding proteins (IGFBPs) are important regulators of fetal and maternal tissue development during pregnancy. Posttranslational modification of IGFBP-1 yields up to six IGFBP-1 phosphovariants and a nonphosphorylated form, which in vitro, have some different properties. Nonphospho IGFBP-1 has less affinity for IGFs than the phospho isoforms and also may have IGF-independent actions. Herein, we have investigated the complement of IGFBP-1 phosphoisoforms present in extraembryonic coelomic (EEC) fluid, amniotic fluid (AF), and maternal serum (MS) throughout human gestation. Also, to determine potential tissue source(s) of IGFBP-1 in these fluids, we have quantified IGFBP-1 and examined IGFBP-1 phosphoisoforms in conditioned media (CM) from maternal decidua, fetal liver, and fetal kidney explants throughout gestation. Western immunodetection revealed that IGFBP-1, present in EEC and AF in early pregnancy and in CM from early pregnancy decidua, is primarily in the nonphosphorylated form. MS in this period contains primarily the nonphospho form and, as in nonpregnant adults, the highly phosphorylated form of IGFBP-1. The phosphorylation profile of IGFBP-1 in AF, MS, and decidua CM changes as pregnancy progresses. All the IGFBP-1 phosphoisoforms ultimately are produced by decidua and are present in midgestation MS, and all but the most highly phosphorylated form are present in AF. In late gestation, MS contains primarily the highly phosphorylated form. In contrast, profiles in CM from explants of fetal liver and kidney at different gestational ages remain unchanged. Nonphosphorylated IGFBP-1 is the primary form in fetal kidney CM, whereas fetal liver CM contains all IGFBP-1 phosphoisoforms. Concentrations of IGFBP-1 in fetal liver and kidney CM are significantly lower (482 +/- 146 and 120 +/- 32 ng/mL x 100 mg wet wt tissue, respectively) than in decidua CM (11,417 +/- 2,358 ng/mL x 100 mg wet wt tissue). The data cumulatively suggest that maternal decidua is the primary source of IGFBP-1 in EEC, AF, and MS in early pregnancy and that fetal liver and kidney are not likely significant contributors. The presence of nonphospho IGFBP-1 in AF, EEC, and MS suggests an important role for this isoform during early gestation.

Body Fluids↗
PubMed1997

Programmed cell death in human ovary is a function of follicle and corpus luteum status.

Although extensive investigation on follicular apoptosis (programmed cell death) has been conducted in the infraprimate ovary, there is little information regarding apoptosis and its relationship to follicular status in the human. In this study, apoptosis was investigated in 116 human ovarian follicles (primordial to dominant) and 5 corpora lutea from a total of 27 premenopausal women. Follicles and corpora lutea were evaluated for the presence of DNA fragmentation, characteristic of apoptosis, by two methods: in situ hybridization using 3' end-labeling of DNA with digoxigenin-labeled nucleotides and subsequent digoxigenin antibody and peroxidase staining, and/or biochemical analysis of low molecular weight DNA laddering. Follicle functional status was evaluated by determining follicle sizes and follicular fluid androgen/estrogen (A/E) ratios. No apoptosis was observed in 67 primordial, primary, or secondary follicles. Positive staining for DNA fragmentation was found in a few granulosa cells in 0.1- to 2-mm follicles, whereas abundant staining in granulosa was detected in 2.1- to 9.9-mm follicles. In contrast, no DNA fragmentation was detected in dominant follicles (10-16 mm). The frequency of apoptosis in follicles was calculated to be 37% in 0.1- to 2-mm follicles, 50% in 2.1- to 5-mm follicles, and 27% in 5.1- to 9.9-mm follicles. Abundant low molecular weight DNA laddering was only found in androgen-dominant follicles and not in estrogen-dominant follicles. Positive staining for DNA fragmentation and low molecular weight DNA laddering were observed in degenerating but not healthy-appearing corpora lutea. In the former, DNA fragmentation was found primarily in large luteal cells. These data suggest that follicular atresia in human ovary results from normal programmed cell death and primarily occurs in the granulosa cell layers of the early antral and < 10-mm antral follicles primarily. Furthermore, because apoptosis occurs as early as the 200-mm stage, follicle selection may begin as early as the initial formation of the antrum. The results also suggest that degeneration of the corpus luteum occurs by apoptotic mechanisms.

Adult↗
PubMed1997

Insulin-like growth factor binding protein-1: recent findings and new directions.

In 1988, insulin-like growth factor-binding protein-1 (IGFBP-1) became the first characterized member of a group of structurally related soluble proteins which specifically bind and modulate the actions of the IGFs. Since then, a wealth of information has accumulated regarding the physiology of this dynamic serum protein. In this review, we update our 1993 summary (Lee PDK et al. Proc Soc Exp Biol Med 204:4-29) of the status of IGFBP-1 research. The IGFBP-1 protein sequence contains 12 N-terminal and 6 C-terminal cysteine residues which are conserved in other mammalian IGFBP-1 sequences and amongst other IGFBPs; both of the cysteine-rich regions are required for optimal IGF binding. The nonconserved IGFBP-1 midregion may act as both a hinge which defines ligand binding characteristics and as a specific target for protease activity. Integrin-binding and phosphorylation sites within the IGFBP-1 sequence have functional significance in vitro, but their physiologic relevance in vivo have not been defined. The human IGFBP-1 and IGFBP-3 genes are contiguous and located in close proximity to the homeobox A (HOXA) gene cluster on chromosome 7. The other IGFBP genes, located on chromosomes 2, 12, and 17, are also associated with HOX clusters, suggesting evolutionary linkage of the IGFBP and HOX gene families. Similarities between the hIGFBP-1 and phosphoenolpyruvate kinase (PEPCK) promoters, including regions conferring insulin, glucocorticoid, and cyclic adenosine-monophosphate responses, are consistent with our previous hypothesis that IGFBP-1 is involved in regulation of glucose metabolism. The tissue-specific patterns of IGFBP-1 gene expression in liver, kidney, decidua, and ovary may be due to stimulation of IGFBP-1 transcription by hepatic nuclear factor 1 (HNF1) proteins. Clinical and basic studies of IGFBP-1 physiology have been aided by several recently developed assay methods. Numerous investigations have confirmed that insulin, via inhibition of IGFBP-1 transcription, is the primary determinant of IGFBP-1 expression both in vitro and in vivo. IGF-I and IGF-II also have specific inhibitory effects on IGFBP-1 expression. Glucocorticoids and cAMP stimulate IGFBP-1 transcription, but these effects are observed only in conditions of low or absent insulin effect. Other stimulants of IGFBP-1 expression include thyroid hormones and epidermal growth factor. Phorbol ester stimulation of IGFBP-1 expression can supersede the effects of insulin in vitro;however, the mechanism and in vivo correlates of this effect have not been determined. Cytokines and, perhaps, growth hormones may affect IGFBP-1 expression, perhaps by altering the regulatory actions of insulin; this effect may have important clinical relevance. IGFBP-1 expression is upregulated in liver and (nonhuman) kidney during postinjury regeneration. The IGF-inhibitory actions of IGFBP-1 has been confirmed by numerous in vitro studies and several in vivo animal investigations, including administration of recombinant IGFBP-1 and IGFBP-1 transgenic models. IGFBP-1 has been shown to inhibit somatic linear growth, weight gain, tissue growth, and glucose metabolism. Moreover, IGFBP-1 appears to be a primary determinant of free IGF-I levels in serum. Excess levels of IGFBP-1 may contribute to growth failure in intrauterine growth restriction and in pediatric chronic renal failure, while low IGFBP-1 levels are associated with obesity and with cardiovascular risk factors in insulin resistance syndromes. Serum IGFBP-1 measurements may be useful biochemical marker in these pathologic conditions. IGFBP-1 is expressed in decidualized stromal cells of the uterine endometrium and in ovarian granulosa cells. IGFBP-1, together with IGFs, insulin, ovarian steroids, cytokines, and other factors, is involved in a complex system which regulates menstrual cycles, ovulation, decidualization, blastocyst implantation, and fetal growth. (ABSTRACT TRUNCATED)

Amino Acid Sequence↗
PubMed1996

Human endometrial matrix metalloproteinase-2, a putative menstrual proteinase. Hormonal regulation in cultured stromal cells and messenger RNA expression during the menstrual cycle.

Proteinases are likely effectors of endometrial menstrual breakdown. We have investigated proteinase production by human endometrial stromal cells subjected in vitro to progesterone (P) withdrawal, the physiologic stimulus for menstruation. Culture media of cells exposed to estradiol, P, or estradiol plus P had low levels of proteolytic activity similar to cultures maintained in the absence of steroids. P withdrawal, or addition of RU486 to P-treated cultures, stimulated proteinase secretion. The stromal cell proteinase was characterized by gelatin zymography, inhibitor profile, and organomercurial activation, as a metalloproteinase present mostly as a 66-kD proenzyme with lower levels of a 62-kD active form. The P withdrawal-induced metalloproteinase was identified as matrix metalloproteinase-2 (MMP-2) by Western blotting. The increase of MMP-2 induced by P withdrawal was associated with the metalloproteinase-dependent breakdown of stromal cultures, involving dissolution of extracellular matrix and dissociation of stromal cells. Northern analysis showed the differential expression of MMP-2 mRNA in late secretory phase endometrium. These findings are consistent with the involvement of stromal cell-derived MMP-2 in the proteolysis of extracellular matrix promoting cyclic endometrial breakdown and the onset of menstrual bleeding.

Blotting, Western↗