Signal transducer and activator of transcription 3 (Stat3) expression and activation in rat uterus during early pregnancy

Signal transducer and activator of transcription 3 (Stat3), a member of the Stat family, is specifically activated during mouse embryo implantation. The aim of this study was to investigate the expression, activation and regulation of Stat3 in rat uterus during early pregnancy, pseudopregnancy, delayed implantation and artificial decidualization. Stat3 mRNA was highly expressed in the luminal epithelium on day 5 and in the luminal epithelium and underlying stromal cells at implantation sites on day 6 of pregnancy. There was a strong level of Stat3 protein expression and phosphorylation in the stromal cells near the lumen and in the luminal epithelium on day 5 of pregnancy, which was similar to day 5 of pseudopregnancy. In the afternoon of day 6, the strong level of Stat3 phosphorylation was detected only in the luminal epithelium. Stat3 was highly expressed and activated in the decidual cells from days 7 to 9 of pregnancy and under artificial decidualization in the present study. Our results suggest that the strong level of Stat3 activation in the luminal epithelium and underlying stromal cells during the pre-implantation period may be important for establishing uterine receptivity as in mice, and the high level of Stat3 expression and activation in decidual cells may play a role during decidualization.

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Actin crosslinking protein filamin A during early pregnancy in the rat uterus

Reproduction Fertility and Development ◽

10.1071/rd14240 ◽

2016 ◽

Vol 28 (7) ◽

pp. 960 ◽

Cited By ~ 1

Author(s):

Romanthi J. Madawala ◽

Connie E. Poon ◽

Samson N. Dowland ◽

Christopher R. Murphy

Keyword(s):

Actin Cytoskeleton ◽

Early Pregnancy ◽

Filamin A ◽

Uterine Receptivity ◽

Rat Uterus ◽

Decidual Cells ◽

Cell Layers ◽

Important Regulator ◽

Blastocyst Implantation ◽

Major Transformation

During early pregnancy the endometrium undergoes a major transformation in order for it to become receptive to blastocyst implantation. The actin cytoskeleton and plasma membrane of luminal uterine epithelial cells (UECs) and the underlying stromal cells undergo dramatic remodelling to facilitate these changes. Filamin A (FLNA), a protein that crosslinks actin filaments and also mediates the anchorage of membrane proteins to the actin cytoskeleton, was investigated in the rat uterus at fertilisation (Day 1) and implantation (Day 6) to determine the role of FLNA in actin cytoskeletal remodelling of UECs and decidua during early pregnancy. Localisation of FLNA in UECs at the time of fertilisation was cytoplasmic, whilst at implantation it was distributed apically; its localisation is under the influence of progesterone. FLNA was also concentrated to the first two to three stromal cell layers at the time of fertilisation and shifted to the primary decidualisation zone at the time of implantation. This shift in localisation was found to be dependent on the decidualisation reaction. Protein abundance of the FLNA 280-kDa monomer and calpain-cleaved fragment (240 kDa) did not change during early pregnancy in UECs. Since major actin cytoskeletal remodelling occurs during early pregnancy in UECs and in decidual cells, the changing localisation of FLNA suggests that it may be an important regulator of cytoskeletal remodelling of these cells to allow uterine receptivity and decidualisation necessary for implantation in the rat.

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Expression and regulation of stanniocalcin 1 and 2 in rat uterus during embryo implantation and decidualization

Reproduction ◽

10.1530/rep.1.01100 ◽

2006 ◽

Vol 131 (6) ◽

pp. 1137-1149 ◽

Cited By ~ 13

Author(s):

Li-Juan Xiao ◽

Jin-Xiang Yuan ◽

Xin-Xin Song ◽

Yin-Chuan Li ◽

Zhao-Yuan Hu ◽

...

Keyword(s):

Early Pregnancy ◽

Embryo Implantation ◽

Normal Pregnancy ◽

Implantation Site ◽

Luminal Epithelium ◽

Rat Uterus ◽

Delayed Implantation ◽

Physiological Processes ◽

Stanniocalcin 2 ◽

Weak Expression

Stanniocalcin-1 (STC-1) is a recently discovered polypeptide hormone, while stanniocalcin-2 (STC-2) is a subsequently identified homologue of stanniocalcin-1. Although previous studies have shown that both STC-1 and -2 are involved in various physiological processes, such as ion transport, reproduction and development, their expression in the uterus and roles in implantation and early pregnancy are unclear. Here we have investigated the expression and regulation of both STC-1 and STC-2 in rat uterus during early pregnancy under various physiological conditions. We show that only basal levels of STC-1 and STC-2 mRNA were detected in the uterus from day one (D1) to day five (D5) of pregnancy. STC-2 immunostaining was gradually increased in the glandular epithelium from day two (D2), with a peak occurring on D5. High levels of both STC-1 and STC-2 mRNA were observed in the stoma cells at the implantation site on day six (D6) of pregnancy, whereas their immunostaining signals were also significant in the luminal epithelium. Basal levels of both STC-1 and STC-2 mRNA and STC-1 immunostaining were detected in the uterus with delayed implantation. After the delayed implantation was terminated by estrogen treatment, both STC-1 and STC-2 mRNA signals were significantly induced in the stroma underlying the luminal epithelium at the implantation site, and STC-2 immunostaining was also observed in the luminal epithelium surrounding the implanting blastocyst. Embryo transfer experiments further confirmed that STC-1 and STC-2 expression at the implantation sites was induced by the implanting blastocyst. Both STC-1 mRNA and immunostaining were seen in the decidualized cells from day seven (D7) to day nine (D9) of pregnancy. STC-2 mRNA was also found in the whole decidua from D7 to D9 of pregnancy; STC-2 protein, however, was strictly localized to the primary deciduas on D7 and D8, with a weak expression in the whole deciduas on D9. Consistent with the normal pregnancy process, strong STC-1 and STC-2 mRNA signals were detected in the decidualized cells under artificial decidualization, whereas only basal levels of STC-1 mRNA and immunostaining were observed in the control horn. These data suggest, for the first time, that STC-1 together with STC-2 may play important roles in the processes of implantation and decidualization in the rat.

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The autophagy protein, FIP200 (RB1CC1) mediates progesterone responses governing uterine receptivity and decidualization†

Biology of Reproduction ◽

10.1093/biolre/ioz234 ◽

2020 ◽

Vol 102 (4) ◽

pp. 843-851 ◽

Cited By ~ 2

Author(s):

Arin K Oestreich ◽

Sangappa B Chadchan ◽

Alexandra Medvedeva ◽

John P Lydon ◽

Emily S Jungheim ◽

...

Keyword(s):

Stromal Cells ◽

Embryo Implantation ◽

Conditional Knockout ◽

Trophoblast Invasion ◽

Uterine Receptivity ◽

Human Escs ◽

Embryo Survival ◽

Endometrial Stromal Cells ◽

Cellular Changes ◽

Decidual Cells

Abstract Successful establishment of pregnancy depends on steroid hormone-driven cellular changes in the uterus during the peri-implantation period. To become receptive to embryo implantation, uterine endometrial stromal cells (ESCs) must transdifferentiate into decidual cells that secrete factors necessary for embryo survival and trophoblast invasion. Autophagy is a key homeostatic process vital for cellular homeostasis. Although the uterus undergoes major cellular changes during early pregnancy, the precise role of autophagy in uterine function is unknown. Here, we report that conditional knockout of the autophagy protein FIP200 in the reproductive tract of female mice results in reduced fecundity due to an implantation defect. In the absence of FIP200, aberrant progesterone signaling results in sustained uterine epithelial proliferation and failure of stromal cells to decidualize. Additionally, loss of FIP200 impairs decidualization of human ESCs. We conclude that the autophagy protein FIP200 plays a crucial role in uterine receptivity, decidualization, and fertility. These data establish autophagy as a major cellular pathway required for uterine receptivity and decidualization in both mice and human ESCs.

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Intranuclear Crosstalk between Extracellular Regulated Kinase1/2 and Signal Transducer and Activator of Transcription 3 Regulates JEG-3 Choriocarcinoma Cell Invasion and Proliferation

The Scientific World JOURNAL ◽

10.1155/2013/259845 ◽

2013 ◽

Vol 2013 ◽

pp. 1-10 ◽

Cited By ~ 10

Author(s):

Diana M. Morales-Prieto ◽

Stephanie Ospina-Prieto ◽

Wittaya Chaiwangyen ◽

Maja Weber ◽

Sebastian Hölters ◽

...

Keyword(s):

Dna Binding ◽

Binding Capacity ◽

Mapk Signaling ◽

Signaling Molecules ◽

Signal Transducer ◽

Stat3 Phosphorylation ◽

Functional Implications ◽

Choriocarcinoma Cells ◽

Cell Invasiveness ◽

Transcription 3

Invasiveness of trophoblast and choriocarcinoma cells is in part mediated via leukemia inhibitory factor- (LIF-) induced activation of signal transducer and activator of transcription 3 (STAT3). The regulation of STAT3 phosphorylation at its ser727 binding site, possible crosstalk with intracellular MAPK signaling, and their functional implications are the object of the present investigation. JEG-3 choriocarcinoma cells were cultured in presence/absence of LIF and the specific ERK1/2 inhibitor (U0126). Phosphorylation of signaling molecules (p-STAT3 (ser727 and tyr705) and p-ERK1/2 (thr 202/tyr 204)) was assessed per Western blot. Immunocytochemistry confirmed results, but also pinpointed the location of phosphorylated signaling molecules. STAT3 DNA-binding capacity was studied with a colorimetric ELISA-based assay. Cell viability and invasion capability were assessed by MTS and Matrigel assays. Our results demonstrate that LIF-induced phosphorylation of STAT3 (tyr705 and ser727) is significantly increased after blocking ERK1/2. STAT3 DNA-binding capacity and cell invasiveness are enhanced after LIF stimulation and ERK1/2 blockage. In contrast, proliferation is enhanced by LIF but reduced after ERK1/2 inhibition. The findings herein show that blocking ERK1/2 increases LIF-induced STAT3 phosphorylation and STAT3 DNA-binding capacity by an intranuclear crosstalk, which leads to enhanced invasiveness and reduced proliferation.

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Shp2 in uterine stromal cells critically regulates on time embryo implantation and stromal decidualization by multiple pathways during early pregnancy

PLoS Genetics ◽

10.1371/journal.pgen.1010018 ◽

2022 ◽

Vol 18 (1) ◽

pp. e1010018

Author(s):

Jianghong Cheng ◽

Jia Liang ◽

Yingzhe Li ◽

Xia Gao ◽

Mengjun Ji ◽

...

Keyword(s):

Gene Expression ◽

Signaling Pathways ◽

Stromal Cells ◽

Embryo Implantation ◽

Endometrial Stromal Cells ◽

Conditional Deletion ◽

Enhancer Binding Protein ◽

Epithelial Remodeling ◽

Multiple Signaling ◽

Transcription 3

Approximately 75% of failed pregnancies are considered to be due to embryo implantation failure or defects. Nevertheless, the explicit signaling mechanisms governing this process have not yet been elucidated. Here, we found that conditional deletion of the Shp2 gene in mouse uterine stromal cells deferred embryo implantation and inhibited the decidualization of stromal cells, which led to embryonic developmental delay and to the death of numerous embryos mid-gestation, ultimately reducing female fertility. The absence of Shp2 in stromal cells increased the proliferation of endometrial epithelial cells, thereby disturbing endometrial epithelial remodeling. However, Shp2 deletion impaired the proliferation and polyploidization of stromal cells, which are distinct characteristics of decidualization. In human endometrial stromal cells (hESCs), Shp2 expression gradually increased during the decidualization process. Knockout of Shp2 blocked the decidual differentiation of hESCs, while Shp2 overexpression had the opposite effect. Shp2 knockout inhibited the proliferation of hESCs during decidualization. Whole gene expression profiling analysis of hESCs during the decidualization process showed that Shp2 deficiency disrupted many signaling transduction pathways and gene expression. Analyses of hESCs and mouse uterine tissues confirmed that the signaling pathways extracellular regulated protein kinases (ERK), protein kinase B (AKT), signal transducer and activator of transcription 3 (STAT3) and their downstream transcription factors CCAAT/enhancer binding protein β (C/EBPβ) and Forkhead box transcription factor O1 (FOXO-1) were involved in the Shp2 regulation of decidualization. In summary, these results demonstrate that Shp2 plays a crucial role in stromal decidualization by mediating and coordinating multiple signaling pathways in uterine stromal cells. Our discovery possibly provides a novel key regulator of embryo implantation and novel therapeutic target for pregnancy failure.

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226.Regulated expression of adhesion and anti-adhesion molecules in mouse endometrium during early pregnancy

Reproduction Fertility and Development ◽

10.1071/srb04abs226 ◽

2004 ◽

Vol 16 (9) ◽

pp. 226 ◽

Cited By ~ 1

Author(s):

M. J. Jasper ◽

A. Stocker ◽

S. A. Robertson

Keyword(s):

Real Time ◽

Adhesion Molecules ◽

Early Pregnancy ◽

Embryo Implantation ◽

Early Embryo ◽

Luminal Epithelium ◽

Paracrine Factors ◽

Mean Values ◽

Actin Mrna ◽

Further Development

To implant and establish the connections that are vital for further development, the early embryo must attach to and then breech the barrier posed by the epithelium of the maternal tract. Expression of adhesion and anti-adhesion molecules in the luminal epithelium of the endometrium are thought to fluctuate in a temporal pattern to 'frame' the implantation site, with their expression regulated by endocrine and paracrine factors. Anti-adhesion molecules, such as members of the mucin family, provide a barrier to implantation in sites or at times unsuitable for embryo development. Expression of adhesion molecules, or specific integrins, are thought to aid in the adhesion of the embryo, allowing it to induce changes in the underlying tissue promoting embryo invasion and pregnancy. The aim of this study was to quantitate the expression of mRNA encoding the integrins αυ, α4 and β3 and MUC1 and MUC4 from Day 0 (oestrous) to Day 4 of pregnancy (implantation) using quantitative real time RT-PCR. Uterine tissues were collected at oestrous and at Days 1, 2, 3 and 4 of pregnancy (Day 1 corresponding to the presence of a vaginal plug), total RNA was extracted, DNAse treated, reverse transcribed into cDNA, and quantified by real-time PCR using SYBR Green chemistry. All specific primers were designed using GenBank sequences and data were normalised to β-actin mRNA expression. Expression of MUC1 and MUC4 mRNAs was dramatically reduced, with mean values 20-fold and 100-fold less than at oestrous respectively, by Day 4 of pregnancy. In contrast, expression of mRNAs encoding integrins αυ, α4 and β3 was detected throughout early pregnancy. These data demonstrate that adhesion and anti-adhesion molecules are differentially expressed in the murine uterus during early pregnancy and may be key mediators in embryo implantation, promoting attachment of the embryo to the luminal epithelium in an environment conducive to embryo growth and development. Supported by a Clive & Vera Ramaciotti Project Grant to MJ Jasper.

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Increased Hypothalamic Signal Transducer and Activator of Transcription 3 Phosphorylation after Hindbrain Leptin Injection

Endocrinology ◽

10.1210/en.2009-0854 ◽

2010 ◽

Vol 151 (4) ◽

pp. 1509-1519 ◽

Cited By ~ 20

Author(s):

Marieke Ruiter ◽

Patricia Duffy ◽

Steven Simasko ◽

Robert C. Ritter

Keyword(s):

Body Weight ◽

Food Intake ◽

Fourth Ventricle ◽

Leptin Receptor ◽

Janus Kinase ◽

Signal Transducer ◽

Solitary Tract ◽

Stat3 Signaling ◽

Stat3 Phosphorylation ◽

Transcription 3

Reduction of food intake and body weight by leptin is attributed largely to its action in the hypothalamus. However, the signaling splice variant of the leptin receptor, LRb, also is expressed in the hindbrain, and leptin injections into the fourth cerebral ventricle or dorsal vagal complex are associated with reductions of feeding and body weight comparable to those induced by forebrain leptin administration. Although these observations suggest direct hindbrain action of leptin on feeding and body weight, the possibility that hindbrain leptin administration also activates the Janus kinase/signal transducer and activator of transcription 3 (STAT3) signaling in the hypothalamus has not been investigated. Confirming earlier work, we found that leptin produced comparable reductions of feeding and body weight when injected into the lateral ventricle or the fourth ventricle. We also found that lateral and fourth ventricle leptin injections produced comparable increases of STAT3 phosphorylation in both the hindbrain and the hypothalamus. Moreover, injection of 50 ng of leptin directly into the nucleus of the solitary tract also increased STAT3 phosphorylation in the hypothalamic arcuate and ventromedial nuclei. Increased hypothalamic STAT3 phosphorylation was not due to elevation of blood leptin concentrations and the pattern of STAT3 phosphorylation did not overlap distribution of the retrograde tracer, fluorogold, injected via the same cannula. Our observations indicate that even small leptin doses administered to the hindbrain can trigger leptin-related signaling in the forebrain, and raise the possibility that STAT3 phosphorylation in the hypothalamus may contribute to behavioral and metabolic changes observed after hindbrain leptin injections.

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414. The contraceptive potential of a long-acting IL-11 inhibitor

Reproduction Fertility and Development ◽

10.1071/srb08abs414 ◽

2008 ◽

Vol 20 (9) ◽

pp. 94

Author(s):

E. Menkhorst ◽

L. Salamonsen ◽

L. Robb ◽

E. Dimitriadis

Keyword(s):

Nature Medicine ◽

Embryo Implantation ◽

Hormonal Contraceptive ◽

Cyclin D3 ◽

Human Reproduction ◽

Luminal Epithelium ◽

Endometrial Stromal Cells ◽

Decidual Cells ◽

Implantation Sites ◽

Long Acting

Interleukin 11 (IL-11) signalling is essential for the establishment of pregnancy in mice, through its action on the differentiation of uterine endometrial stromal cells (decidualisation), a critical process during embryo implantation. IL-11Rα deficient mice are infertile due to defective decidualisation1. IL-11 expression peaks between days (D) 4.5–9.5 of pregnancy (D0: day of plug) in mouse decidua. We examined the effect of administering (intraperitoneal [IP] injection or vaginal gel) a PEGylated IL-11 antagonist (PEGIL-11A) on decidualisation and pregnancy outcome in mice. The sera half-life of PEGIL-11A (IC50 2.8nM) following IP injection was 24h, compared with <1 h for the non-PEGylated antagonist (IC50 0.26nM). Following IP injection, PEGIL-11A localised to decidual cells and blocked the IL-11 decidual target protein, cyclin D3. IP injection of 600µg/application PEGIL-11A (or PEG control) at 1000 h and 1600 h on D3 and 1000 h on D4 (n = 4/group), resulted in smaller implantation sites than controls on D6 due to retarded mesometrial decidual formation. On D10, severe decidual destruction was visible: implantation sites contained regions of haemorrhage and the uterine luminal epithelium had reformed, suggesting a return to oestrous cycling. Following vaginal application in aqueous placebo gel, PEGIL-11A localised to decidual cells. Vaginal application of 200µg/application PEGIL-11A (or control) twice daily from D2 to D5 (n = 4/group), resulted in smaller implantation sites than controls on D6 due to partial inhibition of mesometrial decidual formation. This study demonstrates that PEGIL-11A blocked IL-11 action in the uterus, resulting in total pregnancy loss, equivalent to the IL-11Rα deficient mouse. In women, IL-11 and its receptor are produced by the uterine luminal and glandular epithelium during the period of uterine receptivity2, suggesting that IL-11 may act during initial blastocyst attachment to the luminal epithelium as well as stromal decidualisation. This study provides proof-of-principle for the development of a novel, non-hormonal contraceptive for women. (1) Robb L et al. Nature Medicine 1998; 4: 303–308. (2) Dimitriadis E et al. Molecular Human Reproduction 2000; 6: 907–914.

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Recruitment of circulating NK cells through decidual tissues: a possible mechanism controlling NK cell accumulation in the uterus during early pregnancy

Blood ◽

10.1182/blood-2007-08-105965 ◽

2008 ◽

Vol 111 (6) ◽

pp. 3108-3115 ◽

Cited By ~ 144

Author(s):

Claudia Carlino ◽

Helena Stabile ◽

Stefania Morrone ◽

Roberta Bulla ◽

Alessandra Soriani ◽

...

Keyword(s):

Cell Migration ◽

Nk Cells ◽

Chemokine Receptor ◽

Early Pregnancy ◽

Stromal Cells ◽

Nk Cell ◽

Distinct Pattern ◽

Cell Accumulation ◽

Decidual Cells ◽

Uterine Mucosa

Abstract During early pregnancy, uterine mucosa decidualization is accompanied by a drastic enrichment of CD56highCD16− natural killer (NK) cells. Decidual NK (dNK) cells differ from peripheral blood NK (pbNK) cells in several ways, but their origin is still unclear. Our results demonstrate that chemokines present in the uterus can support pbNK cell migration through human endothelial and stromal decidual cells. Notably, we observed that pregnant women's pbNK cells are endowed with higher migratory ability compared with nonpregnant women's or male donors' pbNK cells. Moreover, NK cell migration through decidual stromal cells was increased when progesterone-cultured stromal cells were used as substrate, and this correlated with the ability of progesterone to up-regulate stromal cell chemokine expression. Furthermore, we demonstrate that dNK cells migrate through stromal cells using a distinct pattern of chemokines. Finally, we found that pbNK cells acquire a chemokine receptor pattern similar to that of dNK cells when they contact decidual stromal cells. Collectively these results strongly suggest that pbNK cell recruitment to the uterus contributes to the accumulation of NK cells during early pregnancy; that progesterone plays a crucial role in this event; and that pbNK cells undergo reprogramming of their chemokine receptor profile once exposed to uterine microenvironment.

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The Role of Interleukin-11 in Pregnancy Involves Up-Regulation of α2-Macroglobulin Gene through Janus Kinase 2-Signal Transducer and Activator of Transcription 3 Pathway in the Decidua

Molecular Endocrinology ◽

10.1210/me.2006-0296 ◽

2006 ◽

Vol 20 (12) ◽

pp. 3240-3250 ◽

Cited By ~ 38

Author(s):

Lei Bao ◽

Sangeeta Devi ◽

Jennifer Bowen-Shauver ◽

Susan Ferguson-Gottschall ◽

Lorraine Robb ◽

...

Keyword(s):

Stromal Cells ◽

Janus Kinase ◽

Abnormal Development ◽

Trophoblast Invasion ◽

Signal Transducer ◽

Janus Kinase 2 ◽

Double Mutation ◽

Α2 Macroglobulin ◽

Decidual Cells ◽

In Pregnancy

Abstract IL-11 expressed by endometrial stromal cells is crucial for normal pregnancy. IL-11 receptor α (IL-11Rα) null mice are infertile due to abnormal development of the placenta. In these mice, the mesometrial decidual tissue, which is the site of trophoblast invasion, thins and disappears at mid-pregnancy. Degeneration of the decidua is accompanied by uncontrolled trophoblast invasion. In this report, we show, using IL-11Rα null mice, that a defect in IL-11 signaling in the decidua leads to severe down-regulation of α2-macroglobulin (α2-MG), a metalloproteinase inhibitor crucial for limiting trophoblast invasion. We also present evidence, using uterine stromal cells that decidualize in culture, that IL-11 robustly stimulates the endogenous α2-MG expression and enhances α2-MG promoter activity. Serial 5′ deletion and internal deletion of the promoter reveal two important signal transducer and activator of transcription (Stat) binding sites. Mutation of either one of these motifs decreases IL-11 stimulation, whereas double mutation prevents IL-11 action. We also found that IL-11 activates Janus kinase 2 (Jak2) and induces rapid phosphorylation, nuclear translocation, and promoter binding activity of Stat3 in decidual cells, whereas Jak1, Tyk2, and Stat5 activities are not affected. In addition, Jak2 inhibitor totally prevents α2-MG expression in decidual cells. Taken together, results of this investigation provide, at least in part, an explanation for the overinvasiveness of the trophoblast in IL-11Rα null mice and reveal, for the first time, that IL-11 signals through the Jak2/Stat3 pathway in decidual cells to stimulate the expression of α2-MG, a protease inhibitor essential for normal placentation in pregnancy.

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