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a Pacific Biomedical Research Center and
b Department of Obstetrics and Gynecology, University of Hawaii, Honolulu, Hawaii 96822
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ABSTRACT |
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 TOP ABSTRACT INTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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Quantitative Northern analysis and in situ hybridization have shown that the decidua expresses mRNA for tPA. However, the immunolocalized tPA protein was most strongly associated with the amnion and chorion, as was its receptor annexin II, suggesting that the amnion and chorion are the targets for decidual tPA.
At term, decidual tPA expression was unaffected by labor, and the tPA receptor was elevated both before and after labor. At preterm, the converse was found: decidual tPA expression was significantly (p < 0.05) up-regulated by labor, but the tPA receptor was not. The results suggest that the generation of plasmin at term would be controlled by an increased concentration of the tPA receptor in the amnion and chorion, whereas at preterm a pathological increase in plasmin would be generated by an overexpression of tPA, initiated by labor.
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INTRODUCTION |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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The plasminogen activator system is important in activation of the MMPs and degradation of the ECM. There are two plasminogen activators: tissue-type plasminogen activator (tPA) and urokinase-type plasminogen activator (uPA); both are involved in extracellular tissue remodeling [4], while tPA is additionally important in the fibrinolytic system [5]. These complex proteins are derived from distinct genes and show tissue-specific expression and biological activities. Plasminogen, a precursor or zymogen, is bound to its own cell surface receptor and is brought into close spatial proximity to tPA and uPA, which are also bound on the cell surface to their respective receptors [5]. The interaction between tPA or uPA and plasminogen at the cell surface results in the generation of plasmin, an active serine protease, from the plasminogen, which is able to degrade most components of the ECM either directly or indirectly by the activation of latent MMPs [3, 6]. This entire process is dependent upon each component's being located at the cell surface on its specific "receptor" [7], providing the spatial conformations necessary for interaction to take place.
The receptors for tPA and uPA are more precisely termed cell surface-binding proteins and have recently been identified and characterized; that for tPA has been shown to correspond to annexin II [8]. Annexin II has been previously studied by immunolocalization and shown to be located in the amniotic epithelial cells, the cells of the mesenchymal layer of the amnion and chorion, and the chorionic cytotrophoblast. No apparent change in this distribution with labor at term was found [9]. The consequence of tPA binding to annexin II in the presence of plasminogen is a focally directed proteolysis within the extracellular environment, as opposed to a classical ligand-receptor intracellular signaling system [7]. The plasmin generated by this interaction of tPA, annexin II, and plasminogen is enzymatically more active than soluble plasmin and can, in addition, activate receptor-bound urokinase (uPA), thereby amplifying plasmin generation. The plasmin is then available for the activation of several matrix-degrading MMPs that are themselves produced as zymogens, requiring activation [3]. Therefore, the presence of plasminogen, annexin II, and tPA on the cell surface is essential for the generation of a fully functional proteolytic cascade.
In the human fetal membranes, plasminogen has been immunolocalized predominantly to the amniotic epithelium and cytotrophoblast of the chorionic membrane [10]. In a previous study, we immunolocalized tPA primarily to the decidual cells and showed an increase in immunostaining intensity after spontaneous labor and delivery at term, in comparison to that in tissues obtained from cesarean section deliveries at term before labor [11]. This was subsequently confirmed by quantitative Northern analysis [12].
The aim of the present study was to advance understanding of the role of tPA in preterm birth. We therefore localized and quantitated the expression of the tPA gene in the amnion, chorion, and decidua of patients as a function of labor and delivery, both at preterm and at term. In addition, we immunolocalized the tPA protein and its receptor, annexin II, in the membranes from the same patients to identify the target tissue(s) for the secreted tPA.
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MATERIALS AND METHODS |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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Human fetal membranes and decidua were collected as soon as possible after expulsion at Kapiolani Medical Center for Women and Children (Honolulu, HI) with informed consent and approval from the University Committee on Human Experimentation and the Hospital Institutional Review Board. Small pieces (1 x 1 cm) were rolled from the membrane, frozen in liquid nitrogen for in situ hybridization, and then mounted on a chuck in OCT compound (Baxter Diagnostics, Inc., McGraw Park, IL) for sectioning or placed in Bouin's fixative for immunocytochemistry. For Northern analysis, the entire membrane was rapidly frozen in liquid nitrogen and then kept at -80°C until use.
Tissue PA Expression at Preterm and Term by Northern Analysis
Quantitative Northern analysis was performed on preterm fetal membranes (n = 6); all patients were between 30 and 35 completed weeks of gestation and had been through labor and delivery. These tissues were not selected; they included tissues from patients with preeclampsia and intrauterine growth retardation. Term membranes were collected from repeat cesarean sections prior to labor (n = 5) and from patients after normal spontaneous labor and delivery (n = 6).
Tissue Distribution of tPA Expression by Quantitative Northern Analysis, Quantitative In Situ Hybridization, and Immunocytochemistry
Decidua was separated from the amnion and chorion of a single normal term cesarean section tissue, obtained prior to labor and delivery, by gently but firmly scraping the cells away with a glass slide. For a twin membrane, the portion consisting of diamnion-dichorion between the two fetuses was collected, and a piece of similar size was taken from the periphery of the tissue, which consisted of a single amnion, chorion, and decidua. For quantitative in situ hybridization and qualitative immunochemistry, two groups of preterm patients (29–36 wk gestation) were studied: patients having elective preterm cesarean sections for severe preeclampsia or intrauterine growth retardation (n = 4) and patients presenting with premature uterine contractions or labor (n = 4). All preterm tissues were examined by a pathologist and discarded from the study if there was any evidence of inflammatory cells. Term tissues were collected between 37–40 wk gestation and were obtained from two groups of patients: patients having elective repeat cesarean section prior to labor and delivery (n = 4) and those after normal term spontaneous vaginal delivery (n = 4).
Preparation of RNA and Northern Analysis
Frozen fetal membranes were homogenized in guanidinium thiocyanate-phenol-chloroform, and total RNA was prepared by the method of Chomczynski and Sacchi [13]. Poly(A)+ RNA was purified by affinity chromatography using oligo(dT)-cellulose [14], denatured with 1 M glyoxal in 0.01 M phosphate buffer (pH 6.5). An aliquot (20 µg) was size fractionated by gel electrophoresis (1.4% agarose gel) and transferred onto nylon membranes (Magna Graph; MSI, Westboro, MA) in 10-strength SSPE (single-strength is 0.18 M NaCl, 10 mM NaPO4, and 1 mM EDTA, pH 7.7). The membranes were rinsed in 5-strength SSPE and baked at 80°C for 1 h prior to hybridization. The blots were prehybridized in 50% formamide, 5-strength SSPE, 5-strength Denhardt's solution (single-strength is 0.02% Ficoll, 0.02% polyvinylpyrrolidone, 0.02% BSA), 0.1% SDS, 100 µg/ml salmon sperm DNA. The tPA probe was purchased from the American Type Culture Collection (Rockville, MD) as a 2.3-kilobase (kb) PstI insert in the pBR322 vector and was digested with PstI to produce several fragments (4.3, 1.1, 0.64, and 0.58 kb). The 640-base pair (bp) PstI fragment was used as a tPA cDNA probe. This tPA cDNA probe was labeled with an [
-32P]dATP (specific activity > 3000 Ci/mmol) using a random-primer labeling kit (Gibco-BRL, Gaithersburg, MD); it was then hybridized to the Northern blot at 42°C for 16–20 h at a concentration of 5 x 106 cpm/ml of hybridization buffer (5-strength SSPE, 50% formamide, 5-strength Denhardt's solution, 0.1% SDS, 100 µg/ml salmon sperm DNA, and 5% dextran sulfate). After hybridization, filters were rinsed twice in 6-strength SSPE-0.1% SDS at room temperature for 15 min each, washed in 3-strength SSPE-0.1% SDS at 65°C for 30 min, and finally washed in single-strength SSPE at room temperature for 15 min. The filters were exposed to Fuji RX film (Fuji Photo Film, Tokyo, Japan) at -20°C for 2 days. The filters were deprobed with 50% formamide and double-strength SSPE at 65°C for 30–60 min and then reprobed with a human glyceraldehyde-3 phosphate dehydrogenase (G3PDH) cDNA probe (1.1 kb; Clontech Laboratories, Palo Alto, CA), a housekeeping gene standard.
Quantitation of the Northern Analyses
The Ambis Image Acquisition and Analysis System (Ambis, Inc., San Diego, CA) was used to quantitate the hybridization signals from the Northern blots. The G3PDH value for each sample was used to standardize the variation in sample loading, and results were expressed as a ratio to G3PDH. Statistical analysis was determined by a one-way ANOVA.
In Situ Hybridization Histochemistry
For the in situ experiments, frozen fetal membrane rolls were sectioned (20 µm) in a cryostat at -16°C. Tissue sections, cut 100–200 µm apart, were thaw mounted onto Superfrost/Plus microscope slides (Fisher Scientific, Pittsburgh, PA) and stored at -80°C. Prior to hybridization, sections were prewarmed to room temperature for a few minutes, fixed in 4% paraformaldehyde (5 min) in single-strength PBS solution, rinsed twice in single-strength PBS, incubated in 0.25% acetic anhydride in 0.1 M triethanolamine-HCl (pH 8.0) for 10 min, dehydrated in serial ethanol solutions, defatted in absolute chloroform (5 min), rehydrated in graded ethanols, and air dried.
In situ hybridization was performed as described previously for human relaxin [15]. Briefly, the 640-bp tPA cDNA (American Type Culture Collection 67587, 561–1145 bp) was cleaned using the QIAquick Gel Extraction Kit (Qiagen, Chatsworth, CA) and cloned into the pGEM-4Z vector by means of the One Shot Kit (Invitrogen, San Diego, CA) for use as a template to prepare a cRNA probe. This vector and its cDNA tPA insert provided a template for SP6-dependent RNA polymerase for the generation of antisense probes protecting a 640-bp tPA mRNA fragment. The cloned human tPA cDNA insert was sequenced and shown to be identical to its published sequence [16]. Prior to hybridization, the 640-bp cRNA probe was hydrolyzed in 40 mM NaHCO3, 60 mM Na2CO3 at 60°C for 46 min, producing approximately 150-bp fragments as assessed by gel electrophoresis (6% polyacrylamide gel). The hybridization procedures used in this study were as previously described [15]. In brief, the tPA cRNA probe was labeled with [35S]UTP (specific activity > 1225 Ci/mmol) using the Riboprobe System Kit (Promega, Madison, WI), which resulted in a transcription yield of 1–2 x 108 cpm/ml (final volume = 100 µl), with probe-specific activities greater than 1 x 108 cpm/ml. Optimal hybridization signals with relatively low background were obtained at probe concentrations of 1–2 x 108 cpm/ml hybridization solution when sections were incubated at 52°C for 16–20 h. After hybridization, the sections were rinsed in double-strength SSC (single-strength is 0.15 M NaCl and 0.015 M sodium citrate, pH 7.0)-10 mM dithiothreitol (DTT) treated with ribonuclease A (RNase A, 20 µg/ml) at 37°C for 30 min, washed sequentially in buffer (1 M NaCl, 10 mM Tris, and 1 mM EDTA) at 37°C for 30 min, and rinsed twice in double-strength SSC-10 mM DTT; they were then washed in double-strength SSC-10 mM DTT at room temperature for 30 min, 0.2-strength SSC-10 mM DTT at 50°C for 30 min, and 0.1-strength SSC-10 mM DTT at 50°C for 1 h. Control sections were treated similarly; the exception was an additional RNase A treatment prior to hybridization with the tPA cRNA probe.
For autoradiographic analysis, slides were dipped in NTB-2 liquid photographic emulsion (Eastman Kodak, Rochester, NY) and exposed at -20°C for 3 wk. After development, slides were stained with toluidine blue.
Quantitation of In Situ Hybridization
The quantitation method used in this study has been previously described and validated for human relaxin [15]. Digital images and quantitation of the autoradiographic signals were acquired on a Zeiss (Carl Zeiss, Thornwood, NY) research microscope equipped with a DAGE MTI (Michigan City, IN) CCD-72 video camera, and Macintosh 8100/AV computer with the Scion LG-3 frame grabber (Scion Corporation, Frederick, MD) using the NIH Image Program (developed at the U.S. National Institutes of Health and available from the Internet by anonymous FTB from zippy.nimh.nih.gov or on floppy disk from the National Technical Information Service, Springfield, VA; part number PB95–500195GEI). The first image taken with this equipment was under brightfield illumination, while the second image (identical field as for previous image) was taken under darkfield illumination, allowing for easier detection of the silver grains. The image obtained at this point was segmented to create a binary image, which represented the autoradiographic signal. The resulting image, called the grain area (GA), consisted almost exclusively of black grains. The region of interest (decidua) was outlined into groups (field area or FA) where grains were concentrated. This area was measured in micrometers, and the final results were presented as the fraction region (FR) or the ratio of GA/FA. The number of grains and the GA values were entered into a spreadsheet program for statistical analysis. The FR value of corresponding regions on the RNase A-treated (control) slides was subtracted from the previously obtained FR value to account for background and nonspecific binding. Fifteen different fields over decidua were measured per patient. Results from each patient were the average measurements from two sections cut 100–200 µm apart.
Immunohistochemistry (tPA and Annexin II)
The same tissues used for in situ hybridization were also used for immunolocalization of tPA and annexin II. Pieces of fixed tissues were embedded in Paraplast (Fisher Scientific) using conventional methods, and were cut (7 µm) and mounted on Vectabond-treated slides (Vector Laboratories, Burlingame, CA). Tissue sections were deparaffinized and hydrated in a graded series of xylenes and ethanols, rinsed in deionized distilled water (ddH2O), and treated with 0.3% hydrogen peroxide in methanol for 30 min to block endogenous peroxidase activity; only those sections used for tPA immunolocalization were incubated in 1% Triton X-100 in Tris-buffered saline (TBS is 0.05 M Tris-Cl, 0.15 M NaCl, pH 7.4) solution for 20 min to permeabilize the membrane, then rinsed in TBS for 20 min [11]. Sections used for annexin II immunolocalization were immediately washed in a PBS solution for 20 min after treatment with 0.3% hydrogen peroxide. Sections were treated with normal goat (tPA) or horse (annexin II) serum (1.5%) for 20 min to block all nonspecific binding sites; they were then incubated in either a polyclonal rabbit anti-tPA antibody (kindly provided by Dr. Stewart Cederholm-Williams of Oxford University) at a concentration of 1:7000 in 0.5% normal goat serum-TBS at 4°C for 18 h, or with a monoclonal mouse anti-annexin II antibody (Zymed Laboratories, Inc., So. San Francisco, CA) at concentrations ranging between 1:50 and 1:400 in 0.5% normal horse serum-PBS at room temperature for 45 min. Both antibodies were titrated for their optimal concentrations, which were 1:7000 for tPA and 1:50 or 1:400 for annexin II, depending upon the gestational age of the tissue. Negative controls used adjacent sections with either normal rabbit serum (1:7000) for tPA and mouse IgG (Chemicon International, Inc., Temecula, CA) at 1:50 or 1:400 for annexin II. The sections were then rinsed (three times, 3 min each) in respective buffers, incubated with a biotinylated secondary antibody for 30 min, again rinsed (three times, 3 min each), and then treated with the ABC reagent (Vector Laboratories) for 45 min and diaminobenzidine (0.5 mg/ml) at room temperature for 5 min. The sections were washed in ddH2O for 5 min, counterstained with hematoxylin (Richard-Allan Scientific, Kalamazoo, MI) for 1–2 min, dehydrated through xylene and ethanol, mounted in Pro-Texx (Baxter Scientific, Honolulu, HI), and viewed under brightfield microscopy.
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RESULTS |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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The human tPA gene transcript was quantitated as a ratio of G3PDH gene expression in poly(A)+ RNA isolated from amnion, chorion, and decidua. In this first study we simply compared the expression of tPA in preterm tissues after labor and delivery (n = 6) and tissues from cesarean section prior to labor (n = 5) with those from normal term spontaneous labor and delivery (n = 6). Figure 1 shows the means and SD of tPA expression as a ratio of G3PDH in each sample. The expression of the tPA gene transcript was significantly greater, in spite of the large variability, in these unselected preterm tissues after labor and delivery than in tissues obtained after either term cesarean section (p < 0.05) or term labor and delivery (p < 0.05). There was marginally increased expression of tPA in the tissues obtained after term labor and delivery compared to the term tissues obtained prior to labor.
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Tissue Distribution of tPA Expression: Quantitative Northern Analysis
In order to determine the relative contributions of the amnion, chorion, and decidua to tPA production, a fetal membrane from a singleton gestation was collected and decidual cells were scraped from the chorion. The decidual cells and amnion-chorion were used separately for mRNA extraction and quantitative Northern analysis. Since it is not possible using this method to obtain amnion-chorion completely free of decidua, we also collected the diamnion-dichorion separating twin fetuses, together with a similar-sized portion of full-thickness membrane from the same gestational sac. Quantitation of these Northern analyses is shown in Figure 2. There was a 9-fold greater expression of tPA in the decidua compared to the amnion-chorion of the singleton tissue, and 7-fold more tPA was expressed in the full-thickness membrane with the decidua than in the amnion and chorion of the same twin membrane (Fig. 2). This result suggests that the decidual cells are indeed the major site of tPA production.
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Quantitative In Situ Hybridization
The localization of tPA mRNA by in situ hybridization is shown by representative brightfield micrographs and darkfield micrographs of the same sections from four tissues in Figure 3. The preterm cesarean section fetal membrane (brightfield) shows the amniotic epithelium, connective tissue, chorionic cytotrophoblast, and decidua (Fig. 3A). The darkfield micrograph of this section shows a low number of silver grains over the decidual cells (Fig. 3B). A similar section from a patient after preterm labor that started prior to membrane rupture (bright- and darkfield views, Fig. 3, C and D, respectively) shows an increased number of silver grains over the decidua. This is especially visible in the darkfield view. Fetal membrane from a term cesarean section, prior to labor or membrane rupture, shows a low signal in both brightfield and darkfield views (Fig. 3, E and F). In contrast, the tissue of a patient after normal term labor and delivery (Fig. 3, G and H) shows more silver grains of the tPA mRNA than either the preterm (Fig. 3, A and B) or term cesarean section tissue (Fig. 3, E and F), but less than the number after preterm labor and delivery (Fig. 3, C and D).
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Quantitation of Decidual Cell tPA Expression
The quantitation of tPA gene expression over the decidua was conducted as two pairs of direct comparisons because of the lengthy process involved; the results are shown as means ± SD in Figure 4. Patients with preterm labor were compared to a group of patients of the same gestational age with preterm cesarean section (Fig. 4); this showed a significantly greater amount of tPA mRNA expressed in the decidua of the patients after preterm labor (p < 0.05). A comparison of the two tissues obtained at term, both before and after labor and delivery, showed no significant difference in tPA gene expression, although the expression after labor and delivery was marginally greater than that before labor and delivery (Fig. 4); these results were similar to the results obtained by Northern analysis with a different set of tissues (Fig. 1).
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Comparison of In Situ Hybridization and Immunolocalization of tPA in the Same Tissues
All the tissues used for in situ hybridization were also used for immunolocalization of the tPA protein. Tissues from preterm delivery before labor at cesarean section (35 wk gestation) and another tissue after labor and delivery (31 wk gestation) are shown in Figure 5. Darkfield and brightfield views for in situ localization of mRNA in the fetal membrane from a patient after preterm cesarean section are shown in Figure 5, A and B, respectively. The signal was predominantly over the decidual cells (Fig. 5A). However, immunostaining of this tissue showed the protein localized predominantly in the amniotic epithelium, in the chorionic cytotrophoblast, and to a lesser extent in the decidual cells (Fig. 5C). An adjacent control section showed only very light staining (Fig. 5D). A more marked signal was obtained by in situ hybridization over the decidua from a patient after labor and delivery at preterm, shown in darkfield view (Fig. 5E) and brightfield view (Fig. 5F). The immunostaining was also stronger in the amniotic epithelium, chorionic cytotrophoblast, and decidua (Fig. 5G) compared to that in the patient prior to labor at cesarean section (Fig. 5C). An in situ control from a section adjacent to the one shown in Figure 5, E and F, after RNase treatment, shows no signal (Fig. 5H).
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Immunolocalization of Annexin II
The results suggest that the tPA protein shown associated with the amnion and chorion by immunolocalization is of decidual origin and that it targets these cells. We therefore used the same tissues to immunolocalize the tPA receptor protein, annexin II. The tPA receptor showed a distribution pattern similar to that of the tPA protein as shown in Figure 6, A–D. There was strong staining in the amniotic epithelium and chorionic cytotrophoblast, with lighter staining of the decidual cells. At the optimal antibody concentration (1:400), the staining in tissues obtained preterm, either before or after labor and delivery, was of about the same intensity; this is shown for a representative preterm cesarean section patient (Fig. 6A). The control, at the same antibody concentration, is shown in an adjacent section with absence of stain in Figure 6B. However, an example of a tissue from a term patient after labor and delivery, using the same antibody concentration (1:400), showed very dark staining of the amniotic epithelium and chorionic cytotrophoblast. The decidual cells were also darker, as was the staining in the connective tissue cells (Fig. 6C); the corresponding control is shown in Figure 6D. The staining was equally intense in all term tissues, whether before or after labor and delivery.
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In order to obtain the same intensity of immunostain for annexin II in the preterm as in the term tissues, an antibody concentration of 1:50 was needed to stain the preterm tissues, suggesting an approximately 8-fold increase of annexin II in the term compared to the preterm tissues. All sections were immunostained several times and were put through the staining procedure at the same time and always with the same reagents.
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DISCUSSION |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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Our localization of the tPA gene by in situ hybridization is in direct contrast to observations recently reported by Liu and colleagues [20]. We initially used the digoxigenin-labeled method for tPA as did they and obtained results similar to theirs. However, with larger numbers of tissues, we found the results to be highly variable and not reproducible. For this reason, we repeated the work with a radioactive probe and obtained reproducibility, increased sensitivity, and the ability to quantitate expression with this method. Since the tPA probe (346 bp) used by Liu and colleagues [20] was completely covered by our larger probe (548 bp), the differences in results are unlikely to be attributable to the use of a different probe. The differences may also be due to our different pretreatment of the tissues or different hybridization or counterstaining procedures. Using quantitative Northern analysis, we have shown that the contribution of the amnion and chorion to tPA gene expression and therefore to tPA production is minor compared with that of the maternal decidual cells.
In contrast, our results for tPA immunolocalization are in complete agreement with those of Liu et al. [20] and show the tPA protein to be most strongly localized to the amniotic epithelium and chorionic cytotrophoblast. Since the tPA protein is secreted and targets its cell surface receptor, annexin II, immunolocalization of the protein would likely detect both the cells of origin (decidua) and the target cells (amniotic epithelium, chorionic cytotrophoblast and mesenchyme of the amnion and chorion). Hence, we immunolocalized the annexin II in the same tissues used for immunolocalization and in situ hybridization of tPA. As expected from previous studies, the annexin II was found predominantly in the amniotic epithelial cells, chorionic cytotrophoblast, and mesenchyme of the amnion and chorion [9], precisely where the tPA protein was also immunolocalized by Liu et al. [20] and ourselves. The results strongly suggest, therefore, that transcription of tPA occurs predominantly in the maternal decidual cells and that its translated protein is secreted and then bound to its receptor, annexin II, at the sites of its action on the plasminogen substrate in the amnion and chorion [5]. This would provide plasmin at the site of its action on the ECM of the amnion and chorion. This suggestion is supported by the finding that amniotic fluid contains tPA and that this is produced by the fetal membranes [21].
Our results suggest that the expression of annexin II in the amnion and chorion is developmentally regulated; we found an approximate 8-fold increase in its immunostaining intensity at term compared to that in preterm tissues. We did not perform extensive studies to demonstrate the linearity of staining with antibody dilution, and this result can be taken only as an estimate. However, the result reflects similar changes reported in the uPA receptor, which increased during and after spontaneous labor and delivery at term; unfortunately, preterm tissues were not studied [22]. It is likely that these parallel systems, which both result in the generation of plasmin, are normally coordinated during development. Annexin II did not appear to be up-regulated with preterm labor and delivery, although more quantitative methods are needed to verify this. However, it was present in the amnion and chorion at preterm and may not be a limiting factor in the generation of plasmin.
These results suggest that the components of the tPA proteolytic system during normal pregnancy are extremely well regulated and controlled; results also indicate that decidual tPA expression was unaffected by term labor but that the tPA receptor was elevated both before and after labor. However, in preterm deliveries, the converse was found: decidual tPA expression was significantly up-regulated by labor, but the tPA receptor was not. Therefore, the generation of plasmin at term would be controlled by an increased concentration of the tPA receptor in the amnion and chorion, whereas at preterm a pathological increase in plasmin would be generated by an overexpression of tPA, initiated by labor.
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ACKNOWLEDGMENTS |
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FOOTNOTES |
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2 Correspondence: G.D. Bryant-Greenwood, Department of Anatomy and Reproductive Biology, University of Hawaii, 1960 East-West Road, Honolulu, HI 96822. FAX: 808 956 9481; gbg{at}pbrc.hawaii.edu
Accepted: November 24, 1998.
Received: October 8, 1998.
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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