Early Pregnancy In Mice Research Articles

Functional Differentiation of Uterine Stromal Cells Involves Cross-Regulation between Bone Morphogenetic Protein 2 and Krüppel-Like Factor (KLF) Family Members KLF9 and KLF13

The inability of the uterine epithelium to enter a state of receptivity for the embryo to implant is a significant underlying cause of early pregnancy loss. We previously showed that mice null for the progesterone receptor (PGR)-interacting protein Krüppel-like factor (KLF) 9 are subfertile and exhibit reduced uterine progesterone sensitivity. KLF9 expression is high in predecidual stroma, undetectable in decidua, and enhanced in uteri of mice with conditional ablation of bone morphogenetic protein 2 (BMP2). Given the individual importance of KLF9 and BMP2 for implantation success, we hypothesized that the establishment of uterine receptivity involves KLF9 and BMP2 functional cross-regulation. To address this, we used early pregnant wild-type and Klf9 null mice and KLF9 small interfering RNA-transfected human endometrial stromal cells (HESCs) induced to differentiate under standard conditions. Loss of KLF9 in mice and HESCs enhanced BMP2 expression, whereas recombinant BMP2 treatment of HESCs attenuated KLF9 mRNA levels. IGFBP1 and KLF9-related KLF13 expression were positively associated with BMP2 and inversely associated with KLF9. Prolonged, but not short-term, knockdown of KLF9 in HESCs reduced IGFBP1 expression. Mouse uterine Igfbp1 expression was similarly reduced with Klf9 ablation. PGR-A and PGR-B expression were positively associated with KLF9 in predecidual HESCs but not decidualizing HESCs. KLF13 knockdown attenuated BMP2 and PGR-B and abrogated BMP2-mediated inhibition of KLF9 expression. Results support cross-regulation among BMP2, KLF9, and KLF13 to maintain progesterone sensitivity in stromal cells undergoing differentiation and suggest that loss of this regulatory network compromises establishment of uterine receptivity and implantation success.

The contribution of Kv7 channels to pregnant mouse and human myometrial contractility

Premature birth accounts for approximately 75% of neonatal mortality and morbidity in the developed world. Despite this, methods for identifying and treating women at risk of preterm labour are limited and many women still present in preterm labour requiring tocolytic therapy to suppress uterine contractility. The aim of this study was to assess the utility of Kv7 channel activators as potential uterine smooth muscle (myometrium) relaxants in tissues from pregnant mice and women. Myometrium was obtained from early and late pregnant mice and from lipopolysaccharide (LPS)-injected mice (day 15 of gestation; model of infection in pregnancy). Human myometrium was obtained at the time of Caesarean section from women at term (38–41 weeks). RT-PCR/qRT-PCR detected KCNQ and KCNE expression in mouse and human myometrium. In mice, there was a global suppression of all KCNQ isoforms, except KCNQ3, in early pregnancy (n= 6, P < 0.001 versus late pregnant); expression subsequently increased in late pregnancy (n= 6). KCNE isoforms were also gestationally regulated (P < 0.05). KCNQ and KCNE isoform expression was slightly down-regulated in myometrium from LPS-treated-mice versus controls (P < 0.05, n= 3–4). XE991 (10 μM, Kv7 inhibitor) significantly increased spontaneous myometrial contractions in vitro in both human and mouse myometrial tissues (P < 0.05) and retigabine/flupirtine (20 μM, Kv7 channel activators) caused profound myometrial relaxation (P < 0.05). In summary, Kv7 activators suppressed myometrial contraction and KCNQ gene expression was sustained throughout gestation, particularly at term. Consequently, activation of the encoded channels represents a novel mechanism for treatment of preterm labour.

Spatial and temporal alterations of phospholipids determined by mass spectrometry during mouse embryo implantation

Molecular events involved in successful embryo implantation are not well understood. In this study, we used MALDI imaging mass spectrometry (IMS) technologies to characterize the spatial and temporal distribution of phospholipid species associated with mouse embryo implantation. Molecular images showing phospholipid distribution within implantation sites changed markedly between distinct cellular areas during days 4-8 of pregnancy. For example, by day 8, linoleate- and docosahexaenoate-containing phospholipids localized to regions destined to undergo cell death, whereas oleate-containing phospholipids localized to angiogenic regions. Arachidonate-containing phospholipids showed different segregation patterns depending on the lipid class, revealing a strong correlation of phosphatidylethanolamines and phosphatidylinositols with cytosolic phospholipase A(2alpha) and cyclooxygenase-2 during embryo implantation. LC-ESI-MS/MS was used to validate MALDI IMS phospholipid distribution patterns. Overall, molecular images revealed the dynamic complexity of lipid distributions in early pregnancy, signifying the importance of complex interplay of lipid molecules in uterine biology and implantation.

Adam12 plays a role during uterine decidualization in mice

In mouse, decidualization is characterized by the proliferation of stromal cells and their differentiation into specialized type of cells (decidual cells) with polyploidy, surrounding the implanting blastocyst. However, the mechanisms involved in these processes remain poorly understood. Using multiple approaches, we have examined the role of Adam12 in decidualization during early pregnancy in mice. Adam12 is spatiotemporally expressed in decidualizing stromal cells in intact pregnant females and in pseudopregnant mice undergoing artificially induced decidualization. In the ovariectomized mouse uterus, the expression of Adam12 is upregulated after progesterone treatment, which is primarily mediated by nuclear progesterone receptor. In a stromal cell culture model, the expression of Adam12 gradually rises with the progression of stromal decidualization, whereas the attenuated expression of Adam12 after siRNA knockdown significantly blocks the progression of decidualization. Our study suggests that Adam12 is involved in promoting uterine decidualization during pregnancy.

CXCL14 inhibits trophoblast outgrowth via a paracrine/autocrine manner during early pregnancy in mice

CXCL14, a member of chemokine family, was previously known to participate in many pathophysiological events, such as leukocytes recruitment and tumor suppression. However, it remained largely unknown whether CXCL14 is a physiological player during early pregnancy. In this regard, our recent global gene microarray analysis has observed an implantation-specific expression profile of CXCL14 mRNA during early pregnancy in mice, showing its higher levels at implantation sites compared to inter-implantation sites, implicating a potential role of CXCL14 in the periimplantation events. In the present investigation, using Northern blot, in situ hybridization and immunostaining, we further demonstrated that uterine CXCL14 expression was specifically induced at embryo implantation site and expanded with subsequent decidualization process in a spatiotemporal manner. The implanting embryo also showed a highlighted expression of CXCL14 in the blastocyst trophectoderm and its derived ectoplacental cones (EPCs) during postimplantation development. In vitro functional study revealed that CXCL14 could significantly inhibit both primary and secondary trophoblast attachment and outgrowth, correlated with a stage-dependant downregulation of MMP-2 and/or MMP-9 activity. Moreover, it was found that biotinylated CXCL14 could specifically bind to trophoblast cells in vitro and in vivo, suggesting trophoblast cell, perhaps expressing the unidentified CXCL14 receptor, is a bioactive target of CXCL14. Collectively, our findings provide evidences supporting the contention that CXCL14 is an important paracrine/autocrine modulator regulating trophoblast outgrowth at the maternal-fetal interface during the process of pregnancy establishment. This study is clinically related since CXCL14 is also highly expressed in human receptive endometrium and trophoblasts.

Is the zona pellucida an intrinsic source of signals activating maternal recognition of the developing mammalian embryo?

Mammalian mothers undergoing embryo implantation must specifically recognize the developing embryo in a species-restricted manner. We previously observed that immune cells derived from early pregnant mice could promote endometrial differentiation and embryo implantation in blastocyst-transferred pseudopregnant mice. Although the precise mechanism remains unknown, it is suggested that the maternal immune system undergoes functional changes after recognizing developing embryos from the very early stages of pregnancy. Since it is physically impossible for immune cells to directly interact with the developing embryo while it is surrounded by the zona pellucida (ZP), it is speculated that the embryo produces certain embryo- and species-specific soluble factor(s) in the oviduct before hatching. As a candidate for this factor, we have paid attention to the ZP that is normally protected from immunological attack during oogenesis in the ovarian follicle. ZP-specific glycoproteins are known to play important roles in the species- and oocyte-specific binding of sperm, and the ZP can also be considered an abundant store of oocyte- and species-specific glycoproteins. In contrast to unfertilized oocytes, developing embryos may degrade the ZP starting just after fertilization and proceeding until hatching using enzymes that are released from cortical granules or produced by the developing embryo. Accordingly, the developing embryo might provide ZP-degradation products including oligosaccharide chains to the immune system from the very early stages. Taken together, we propose here a novel hypothesis that these ZP-derivatives can act as an intrinsic signal from the developing embryo for maternal recognition by the immune system.

Complement 3 deficiency impairs early pregnancy in mice

Human oviductal cells produce complement-3 (C3) and its derivative, iC3b. These molecules are important in immune responses. Our recent study suggested that iC3b also possessed embryotrophic activity and it stimulates the blastulation and hatching rates of in vitro cultured mouse embryos. The objective is to study the impact of C3 deficiency on early pregnancy in vivo using homozygous C3-deficient (C3KO) and wild-type (C3WT) mice. C3 protein was undetectable in the reproductive tissues of C3KO mice. Deficiency in C3 is associated with significantly longer estrous cycle (P = 0.037). No significant difference was found in the ovulation rate, total cell count in blastocysts and implantation rate between the wild-type and the C3KO mice, though C3KO mice tended to have lower values in the latter two parameters. On day 15 of pregnancy, C3KO mice had fewer conceptus (P < 0.001) and higher resorption rate (P < 0.001) than that of C3WT mice. The fetal and placental weights (P < 0.001) were lower in the C3KO mice. The placenta of C3KO mice had smaller spongiotrophoblast (P = 0.001) and labyrinth (P = 0.037). Deficiency in C3 is associated with mild impairment in early pregnancy including longer estrous cycle and higher resorption rates after implantation. The impairment may be related to compromised placental development leading to under-developed fetuses.

The uterine expression of SEC63gene is up-regulated at implantation sites in association with the decidualization during the early pregnancy in mice

BackgroundSec63 is a key component of the protein translocation machinery in the mammalian endoplasmic reticulum (ER), and involved in the post-translation processing of secretory proteins. The aim of this study was to determine the expression pattern of SEC63 gene in mouse uterus during the early pregnancy.MethodsReal-time quantitative PCR and Western blot analyses were used to evaluate the alteration in levels of uterine SEC63 gene expression during the peri-implantation period in mice. Further, both in situ hybridization and immunohistochemical analyses were performed to examine the spatial localization of SEC63 gene expression in mouse uterine tissues. The presence of Sec63 protein in human uterine tissue was also detected by immunohistochemical analysis. Statistical analysis was carried out using Tukey test.ResultsUterine SEC63 gene expression was up-regulated and predominantly localized in mouse decidual cells during days 5–8 of pregnancy. More interestingly, Sec63 protein was also detected in human decidua of 10-week pregnancy, whereas was not observed in human endometrial tissues both at proliferative and secretory phases of menstrual cycle.ConclusionThe pattern of SEC63 gene expression is consistent with a possible role for SEC63 in decidualization.

Expression of Transcriptional Repressor Slug Gene in Mouse Endometrium and its Effect During Embryo Implantation

Slug, a member of the Snail family of zinc-finger transcription factors, is involved in regulating embryonic development and tumorigenesis. The aim of this study was to investigate the expression of Slug in mouse endometrium during early pregnancy and its possible role during embryo implantation. Fluorescence quantitative polymerase chain reaction and immunohistochemistry were applied to detect Slug mRNA and Slug protein expression in endometrium of nonpregnant and early pregnant mice, respectively. The expressions of Slug mRNA and its protein in pregnant group were higher than that in nonpregnant group and gradually increased from pregnancy day 1, reaching its maximum level on day 4 and then declining on days 5, 6, and 7. Immunohistochemistry showed that Slug protein was mainly present in luminal epithelium from pregnancy days 2 to 5 and in glandular epithelium from days 2 to 6 and enhanced significantly in stromal cells on days 4, 5, and 6. The number of embryos implanted was greatly decreased after Slug function in mouse endometrium was blocked by the intrauterine injection with anti-Slug polyclonal antibody on day 3 of pregnancy before implantation. These results suggested that up-regulation of Slug expression may play a key role in the embryo implantation in mice.

The role of T-lymphoma invasion and metastasis inducing protein 1 in early pregnancy in mice

T-lymphoma invasion and metastasis inducing protein 1 (Tiam1) is involved in tumorigenesis processes, including cell migration, adhesion and invasion, proteolysis, cytoskeleton reorganization, cell morphological transformation and intracellular signaling. These processes are also critical for embryo implantation, although the role of Tiam1 during embryo implantation remains poorly understood. The aim of this study was to investigate the spatio-temporal expression of Tiam1 in endometria of mice comparing early pregnancy and non-pregnancy. Fluorescent quantitative-PCR and immunohistochemical analysis showed that the expression of Tiam1 mRNA and protein in endometria of pregnant mice was higher than that of non-pregnant mice, and gradually increased from Day 1 of pregnancy reaching a maximum level on Day 5 and then declining on Days 6 and 7. Immunohistochemistry showed that Tiam1 protein was present in luminal epithelium from Days 3-5 of pregnancy and in gland epithelium from Days 4 to 6 and enhanced significantly in stromal cells on Day 5, regarded as the 'implantation window' period. The number of implantation sites was greatly decreased by the intrauterine injection with anti-Tiam1 polyclonal antibody in the early morning of the Day 4 of pregnancy. These findings suggest that Tiam1 might play an important role in embryo implantation in mice.

Dehydroepiandrosterone and metformin modulate progesterone-induced blocking factor (PIBF), cyclooxygenase 2 (COX2) and cytokines in early pregnant mice

The present study examined the mechanism by which metformin ( N, N′-dimethylbiguanide) prevents embryonic resorption induced in mice by dehydroepiandrosterone (DHEA). Treatment with DHEA (60 mg/kg, s.c. 24 and 48 h post-implantation) induces embryo resorption of early pregnant BALB/c mice while simultaneous treatment with metformin (240 mg/kg, oral 24 and 48 h post-implantation) prevents it. During pregnancy progesterone-induced blocking factor (PIBF) modulates prostaglandins (PGs) and cytokine production. These findings prompted us to investigate the effect of DHEA and metformin on both PIBF and cyclooxygenase 2 (COX2) expressions at the implantation sites, as well as cytokine production. PIBF and COX2 expression were detected by immunohistochemistry from DHEA and DHEA+ metformin treated 8 days-pregnant mice and serum cytokine levels of these animals were determined by ELISA. DHEA treatment both abolished PIBF expression and increased COX2 expression. Embryo resorption correlates with the lack of PIBF expression, diminished IL-6 levels and increased IL-2 concentration while metformin was able to reverse the effect of DHEA on both PIBF and COX2 expression and IL-6 levels. We concluded that hyperandrogenization induces embryo resorption in early pregnancy diminishing PIBF in implantation sites, having a pro-inflammatory effect. Metformin is able to prevent such effects.

Maternal immune activation in mice delays myelination and axonal development in the hippocampus of the offspring

Epidemiological data suggest a relationship between maternal infection and a high incidence of schizophrenia in offspring. An animal model based on this hypothesis was made by injecting double-stranded RNA, polyinosinic-polycytidylic acid (poly-I:C), into early pregnant mice, and their offspring were examined for biochemical and histological abnormalities. Mouse brains were examined with special reference to oligodendrocytes, which have been implicated in several neurodevelopmental disorders. We detected a significant decrease of myelin basic protein (MBP) mRNA and protein at early postnatal periods in poly-I:C mice. MBP immunocytochemistry and electron microscopy revealed that the hippocampus of juvenile poly-I:C mice was less myelinated than in PBS mice, with no significant loss of oligodendrocytes. In addition, axonal diameters were significantly smaller in juvenile poly-I:C mice than in control mice. These abnormalities reverted to normal levels when the animals reached the adult stage. These findings suggest that retarded myelination and axonal abnormalities in early postnatal stages caused by maternal immune activation could be related to schizophrenia-related behaviors in adulthood.

Overexpression of fucosyltransferase VII (FUT7) promotes embryo adhesion and implantation

To investigate the effect of increased fucosyltransferase VII (FUT7) on sialyl Lewis X (sLeX) synthesis and their impact on embryo implantation. Cell and animal study. Liaoning Provincial Core Lab of Glycobiology and Glycoengineering, Department of Biochemistry and Molecular Biology, Dalian Medical University. Adult female mice of Kunming species. A pIRES2-EGFP-FUT7 expression plasmid containing full-length human FUT7 cDNA was constructed and transiently transfected into RL95-2 or injected into the uteri of early pregnant mice. Control subjects received the mock pIRES2-EGFP or anti-sLeX antibody. The expression of FUT7 and sLeX was detected by reverse transcriptase polymerase chain reaction (RT-PCR) and indirect immunofluorescence. The embryo adhesion rate was assayed in vitro with the cocultured RL95-2 and JAR implantation model. In vivo embryo implantation was investigated using a mouse model. Expression of FUT7 was significantly increased after transfection of pIRES2-EGFP-FUT7 into RL95-2 cells compared with the parental control and mock vector transfectants, as demonstrated by fluorescent microscopy and RT-PCR. Expression of sLeX was also increased in FUT7 overexpressers, as detected by indirect immunofluorescence. Consistent with the elevated expression of FUT7 and sLeX, embryo adhesion rate and embryo implantation competence were significantly increased (P<.01) in both in vitro and in vivo models. Overexpression of FUT7 up-regulates sLeX synthesis and enhances sLeX-mediated embryo implantation.

211. Proteomic identification of caldesmon as one of the physiological substrates of proprotein convertase 6 during decidualisation

We have previously demonstrated that proprotein convertase 5/6 (PC6), a member of the proprotein convertase (PC) family, is a critical endometrial factor for implantation. PC6 is upregulated in the endometrium specifically at implantation in association with epithelial differentiation (in human and monkey) and stromal cell decidualisation (in the mouse, human and monkey). Knockdown of endometrial PC6 during early pregnancy in mice in vivo led to complete failure of implantation, while blocking of PC6 production in human endometrial stromal cells in vitro inhibited decidualisation. PCs convert a range of precursor proteins of important functions into their bioactive forms; they are thus regarded as critical ‘master switch’ molecules. We hypothesise that PC6 exerts its roles in the endometrium by regulating proteins of diverse functions essential for implantation. In this study, we utilised proteomic technology and aimed to identify proteins that are specifically cleaved by PC6 in human endometrial stromal cells (HESC) during decidualisation. HESC were decidualised with cyclic AMP, the cell lysates were treated with and without recombinant human PC6-A (rPC6-A), and the 2D Differential in Gel Electrophoresis (2D DiGE) protein profiles were compared between the two treatments. We identified several proteins which were differentially cleaved following the addition of rPC6-A. Mass spectrometric analysis confirmed that the most abundant of these were caldesmon, tropomyosin-2, tropomyosin-4, hypoxia Inducible factor-1 and chloride intracellular channel-1. These proteins showed spot shifts in hPC6-A treated HESC lysates consistent with hPC6-A cleavage. western blot analysis confirmed the specific cleavage of caldesmon by PC6 in HESCs, and immunohistochemical analysis showed co-localisation of caldesmon and PC6 in decidual cells in human endometrial tissue. Given that caldesmon is a structural protein previously found to be involved in actin filament reorganisation, our results strongly suggest that PC6 is a mediator of structural remodelling of stromal cells during decidualisation in the endometrium.

Progesterone, But Not Estrogen, Stimulates Vessel Maturation in the Mouse Endometrium

The human endometrium undergoes regular periods of growth and regression, including concomitant changes in the vasculature, and is one of the few adult tissues where significant angiogenesis and vascular maturation occurs on a routine, physiological basis. The aim of this study was to investigate the effects of estrogen and progesterone on endometrial vascular maturation in mice. Endometrial tissues were collected from early pregnant mice (d 1-4) and ovariectomized mice given a single 17beta-estradiol (100 ng) injection 24 h before dissection (short-term estrogen regime) or three consecutive daily injections of progesterone (1 mg) with/without estrogen priming (progesterone regime). Experiments were then repeated with the inclusion of mice treated concurrently with progesterone and either RU486 or a vascular endothelial growth factor-A antiserum. Proliferating vascular mural cells (PVMC) were observed on d 3-4 of pregnancy, corresponding with an increase in circulating progesterone. A significant increase in PVMC and alpha-smooth muscle actin (labels mural cells) coverage of vessel profiles were observed in mice treated with progesterone in comparison to controls; no significant change was noted in mice treated with estrogen or with vascular endothelial growth factor antiserum. RU486 treatment did not inhibit the progesterone-induced increases in PVMC and mural cell coverage, although progesterone-induced changes in endothelial and epithelial cell proliferation were inhibited. These results show that progesterone, but not estrogen, stimulates vessel maturation in the mouse endometrium. The work illustrates the relevancy of the mouse model for understanding endometrial vascular remodeling during the menstrual cycle and in response to the clinically important progesterone receptor antagonist RU486.

Heparanase Expression and Function During Early Pregnancy in Mice1

Embryo implantation is a complex process that involves interactions between cell-surface and extracellular components of the blastocyst and the uterus, including blastocyst adhesion to the uterine luminal epithelium, epithelial basement membrane penetration and stromal extracellular matrix remodeling, angiogenesis, and decidualization. These processes all involve interactions with heparan sulfate (HS) proteoglycans, which harbor various growth factors and cytokines and support cell adhesion. Heparanase (HPSE) is an endo-beta-glucuronidase that cleaves HS at specific sites. HPSE also can act as an adhesion molecule independent of its catalytic activity. Thus, HPSE is a multifunctional molecule contributing to and modulating HS-dependent processes. Exogenously added HPSE improves embryo implantation in mice; however, no information is available regarding the normal pattern of HPSE expression and activity during the implantation process in any system. Using several approaches, including real-time RT-PCR, in situ hybridization, and immunohistochemistry, we determined that uterine HPSE expression increases dramatically during early pregnancy in mice. Heparanase mRNA and protein were primarily expressed in decidua and were rapidly induced at the implantation site. Uterine HPSE activity was characterized and demonstrated to increase >40-fold during early pregnancy. Finally, we demonstrate that the HPSE inhibitor PI-88 severely inhibits embryo implantation in vivo. Collectively, these results indicate that HPSE plays a role in blastocyst implantation and complements previous studies showing a role for HS-dependent interactions in this process.

Lack of effects of 2,4-dichlorophenoxyacetic acid administration on markers of oxidative stress during early pregnancy in mice

Induction of oxidative stress by 2,4-dichlorophenoxyacetic acid (2,4-D) both as a pure compound and in commercial formulation was investigated during early pregnancy in mice. Pregnant animals were exposed to increasing doses of the herbicide (0.01, 0.1 and 100mg/kg/d) during gestation days 0–9, after which animals were euthanized and their blood analyzed for catalase activity, thiobarbituric acid reactive substances (TBARs) and total antioxidant capacity (TAC). Number of corpora lutea and uterine implantations and resorptions were also determined. Herbicide exposure did not cause any overt signs of maternal toxicity at any of the doses administered; neither did it cause an effect on developmental parameters. Catalase activity and TBARs were not modified by herbicide exposure although TAC was significantly decreased at 100mg/kg/d of both pure and formulated compound. Thus, 2,4-D does not seem to induce oxidative stress during early pregnancy in mice at the doses administered, indicating that this mechanism is probably not involved in mediating herbicide toxicity at these dose levels. Furthermore, since no manifestations of developmental toxicity were observed after administration of the herbicide, it is also possible that 2,4-D may not produce any early developmental toxicity at the low environmentally relevant doses tested in this animal model.

Nitric oxide mediates prostaglandins' deleterious effect on lipopolysaccharide-triggered murine fetal resorption.

Genital tract bacterial infections could induce abortion and are some of the most common complications of pregnancy; however, the mechanisms remain unclear. We investigated the role of prostaglandins (PGs) in the mechanism of bacterial lipopolysaccharide (LPS)-induced pregnancy loss in a mouse model, and we hypothesized that PGs might play a central role in this action. LPS increased PG production in the uterus and decidua from early pregnant mice and stimulated cyclooxygenase (COX)-II mRNA and protein expression in the decidua but not in the uterus. We also observed that COX inhibitors prevented embryonic resorption (ER). To study the possible interaction between nitric oxide (NO) and PGs, we administered aminoguanidine, an inducible NO synthase inhibitor. NO inhibited basal PGE and PGF(2alpha) production in the decidua but activated their uterine synthesis and COX-II mRNA expression under septic conditions. A NO donor (S-nitroso-N-acetylpenicillamine) produced 100% ER and increased PG levels in the uterus and decidua. LPS-stimulated protein nitration was higher in the uterus than in the decidua. Quercetin, a peroxynitrite scavenger, did not reverse LPS-induced ER. Our results suggest that in a model of septic abortion characterized by increased PG levels, NO might nitrate and thus inhibit COX catalytic activity. ER prevention by COX inhibitors adds a possible clinical application to early pregnancy complications due to infections.

Expression of angiopoietin-1/-2 in the process of mouse embryo implantation

This study examined the expression and distribution of angiopoietin-1/-2 (Ang-1/-2) in the endometrium of early pregnant mice. The expression of Ang-1/-2 was detected by immunohisto-chemical staining and in situ hybridization respectively. Computerized image analysis system was used to measure the average optical intensity of Ang-1/-2 in endometria at different time points after gestation. Mice were randomly divided into 5 groups: control group, D2 group (2 days after pregnancy), D4 group (4 days after pregnancy), D6 group (6 days after pregnancy) and D8 group (8 days after pregnancy), each containing 15 mice. The results showed that the expression of Ang-1 and Ang-2 was very different among 4 groups (P<0.01). Immunohistochemical staining revealed that Ang-1 was localized in the cytoplasma of stromal cells 2 days after pregnancy (day 2), and in luminal epithelial cells on day 4. The protein of Ang-2 was mainly expressed in the cytoplasma of glandular epithelia and stromal cells. With gestation time, the positive reactions of Ang-1/-2 were stronger in the endometria of the pregnant mice (P<0.01). In situ hybridization showed Ang-1 mRNA in stromal cells on day 2. Hybridization signal was localized in both stromal cells and vessel epithelial cells on day 4; Ang-2 mRNA was expressed in stromal cells and glandular epithelia on day 2; high mRNA levels appeared in stromal cells, glandular epithelia and vascular endothelia on day 4; an increasing in mRNA expression of Ang-1/-2 was observed on day 6 and day 8 (P<0.01). It is suggested that Ang-1/-2 may play an important role in the cross-talk between blastocyst and maternal endometrium during the process of embryo implantation.

Early pregnancy failure induced by dibutyltin dichloride in mice

In this study, we examined the adverse effects of dibutyltin on initiation and maintenance of pregnancy after maternal administration during early pregnancy in mice. Following successful mating, female ICR mice were given dibutyltin dichloride (DBTCl) at 0, 7.6, 15.2, or 30.4 mg/kg bw/day by gastric intubation on days 0-3 or days 4-7 of pregnancy. Female mice were sacrificed on day 18 of pregnancy, and the pregnancy outcome was determined. After administration of DBTCl on days 0-3, the rate of nonpregnant females and the incidence of preimplantation embryonic loss were significantly increased at 30.4 mg/kg bw/day. The incidences of postimplantation embryonic loss in females given DBTCl on days 0-3 at 15.2 mg/kg and higher and on days 4-7 at 7.6 mg/kg bw/day and higher were increased. No increase in the incidence of fetuses with external malformations was observed after the administration of DBTCl on days 0-3 or days 4-7. A decline in the serum progesterone levels was detected in mice given DBTCl at 30.4 mg/kg bw/day on days 0-3 or days 4-7 of pregnancy. The data show that DBTCl adversely affects the initiation and maintenance of pregnancy when administered during early pregnancy in mice and suggest that the decline in serum progesterone levels is responsible for pregnancy failure.