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Matters Arising article, arising from H. Bové et al. Nature Communications https://doi.org/10.1038/s41467-019-11654-3 (2019).

Insulin-like growth factors (IGFs) influence placental cell (cytotrophoblast) kinetics. We recently reported that the protein tyrosine phosphatase (PTP) SHP-2 positively regulates IGF actions in the placenta. In other systems, the closely related PTP, SHP-1, functions as a negative regulator of signaling events but its role in the placenta is still unknown. We examined the hypothesis that SHP-1 negatively regulates IGF actions in the human placenta. Immunohistochemical (IHC) analysis demonstrated that SHP-1 is abundant in cytotrophoblast. SHP-1 expression was decreased in first-trimester placental explants using siRNA; knockdown did not alter IGF-induced proliferation but it significantly enhanced proliferation in serum-free conditions, revealing that placental growth is endogenously regulated. Candidate regulators were determined by using antibody arrays, Western blotting, and IHC to examine the activation status of multiple receptor tyrosine kinases (RTKs) in SHP-1-depleted explants; amongst the alterations observed was enhanced activation of EGFR, suggesting that SHP-1 may interact with EGFR to inhibit proliferation. The EGFR tyrosine kinase inhibitor PD153035 reversed the elevated proliferation seen in the absence of SHP-1. This study demonstrates a role for SHP-1 in human trophoblast turnover and establishes SHP-1 as a negative regulator of EGFR activation. Targeting placental SHP-1 expression may provide therapeutic benefits in common pregnancy conditions with abnormal trophoblast proliferation.

Infertility and recurrent pregnancy loss (RPL) are prevalent but distinct causes of reproductive failure that often remain unexplained despite extensive investigations. Analysis of midsecretory endometrial samples revealed that SGK1, a kinase involved in epithelial ion transport and cell survival, is upregulated in unexplained infertility, most prominently in the luminal epithelium, but downregulated in the endometrium of women suffering from RPL. To determine the functional importance of these observations, we first expressed a constitutively active SGK1 mutant in the luminal epithelium of the mouse uterus. This prevented expression of certain endometrial receptivity genes, perturbed uterine fluid handling and abolished embryo implantation. By contrast, implantation was unhindered in Sgk1-/- mice, but pregnancy was often complicated by bleeding at the decidual-placental interface and fetal growth retardation and subsequent demise. Compared to wild-type mice, Sgk1-/- mice had gross impairment of pregnancy-dependent induction of genes involved in oxidative stress defenses. Relative SGK1 deficiency was also a hallmark of decidualizing stromal cells from human subjects with RPL and sensitized these cells to oxidative cell death. Thus, depending on the cellular compartment, deregulated SGK1 activity in cycling endometrium interferes with embryo implantation, leading to infertility, or predisposes to pregnancy complications by rendering the feto-maternal interface vulnerable to oxidative damage.

During pregnancy, trophoblasts invade and transform the maternal spiral arteries that supply blood to the placenta. Recent work has revealed that this process occurs in several stages, and details of the molecular and cellular mechanisms are beginning to emerge, including changes that precede or accompany trophoblastic colonization of the vascular media. Disruption and eventual loss of smooth muscle cells and their associated extracellular matrix are central to physiological transformation. Advances in understanding will lead to the identification of the causative factors involved in failure of remodeling in pathological pregnancies and suggest novel diagnostic and therapeutic avenues.

We have used subunit-specific monoclonal antibodies (MAbs) and immunohistochemistry to examine the distribution of integrin alpha 6 beta 4 in normal ectocervical epithelium and various grades of cervical intraepithelial neoplasia (CIN). Antibodies were first characterised by immunoprecipitation from two surface-labelled tumour cell lines. Monoclonal antibody G71 was found to precipitate integrin beta 4 from BeWo but not T47D cells, while other anti-beta 4 antibodies precipitated beta 4 from both cell lines. Both G71 and an antiserum to the C-terminal peptide of beta 4 precipitated free beta 4 from surface-iodinated BeWo cells. Neither antibody recognised truncated beta 4 chains observed at approximately 160 kDa. These data suggest that different isoforms of beta 4 are expressed in different tumour cell lines, and that there may be a pool of beta 4 at the cell surface that is not complexed to alpha 6. In normal cervix, both the alpha 6 and beta 4 subunits occur at the basal surface of the basal cell layer. In CIN, the distribution is markedly altered, with strong expression of alpha 6 and beta 4 in the upper cell layers of the ectocervical epithelium. All 40 cases of CIN that were studied exhibited this alteration. Furthermore, the extent of extrabasal staining appeared to correspond with the grade of CIN. The form of integrin beta 4 recognised by antibody G71 also appears in the upper cell layers in CIN, but it shows a more restricted distribution than the normal isoform. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 5

Background Remodeling of the extracellular matrix is one of the most striking features observed in the uterus during the estrous cycle and after hormone replacement. Versican (VER) is a hyaluronan-binding proteoglycan that undergoes RNA alternative splicing, generating four distinct isoforms. This study analyzed the synthesis and distribution of VER in mouse uterine tissues during the estrous cycle, in ovariectomized (OVX) animals and after 17beta-estradiol (E2) and medroxyprogesterone (MPA) treatments, either alone or in combination. Methods Uteri from mice in all phases of the estrous cycle, and animals subjected to ovariectomy and hormone replacement were collected for immunoperoxidase staining for versican, as well as PCR and quantitative Real Time PCR. Results In diestrus and proestrus, VER was exclusively expressed in the endometrial stroma. In estrus and metaestrus, VER was present in both endometrial stroma and myometrium. In OVX mice, VER immunoreaction was abolished in all uterine tissues. VER expression was restored by E2, MPA and E2+MPA treatments. Real Time PCR analysis showed that VER expression increases considerably in the MPA-treated group. Analysis of mRNA identified isoforms V0, V1 and V3 in the mouse uterus. Conclusion These results show that the expression of versican in uterine tissues is modulated by ovarian steroid hormones, in a tissue-specific manner. VER is induced in the myometrium exclusively by E2, whereas MPA induces VER deposition only in the endometrial stroma.

Implantation of the embryo in the uterus is a critical and complex event and its failure is widely considered an impediment to improved success in assisted reproduction. Depending on whether placentation is invasive or superficial (epitheliochorial), the embryo may interact transiently or undergo a prolonged adhesive interaction with the uterine epithelium. Numerous candidate interactions have been identified, and there is good progress on identifying gene networks required for early placentation. However no molecular mechanisms for the epithelial phase are yet firmly established in any species. It is noteworthy that gene ablation in mice has so far failed to identify obligatory initial molecular events.

The cell surface mucin MUC1 is expressed by endometrial epithelial cells with increased abundance in the secretory phase of the menstrual cycle, when it is found both at the apical cell surface and in secretions. This suggests the presence of a maternal cell surface glycoprotein barrier to embryo implantation, arising from the anti-adhesive property of MUC1. In previous work, we demonstrated alternatively spliced MUC1 variant forms in tumour cells. The variant MUC1/SEC lacks the transmembrane and cytoplasmic sequences found in the full-length variant. We now show that MUC1/SEC mRNA is present in endometrial carcinoma cell lines, endometrial tissue and primary cultured endometrial epithelial cells. The protein can be detected using isoform-specific antibodies in uterine flushings, suggesting release from endometrium in vivo. However, on the basis of immunolocalisation studies, MUC1/SEC also remains associated with the apical epithelial surface both in tissue and in cultured cells. Transmembrane MUC1 and MUC1/SEC are both strikingly localised to the apical surface of tubal epithelium. Thus MUC1 may contribute to the anti-adhesive character of the tubal surface, inhibiting ectopic implantation. The mechanism by which this barrier is overcome in endometrium at implantation is the subject of ongoing investigation.

Changes in the organisation and composition of extracellular matrix in human endometrium during the menstrual cycle and early pregnancy have been assessed by immunofluorescence. Amongst interstitial components, type-III and type V-collagens and fibronectin are present in endometrial stroma throughout the menstrual cycle as well as in first trimester decidua. Type V-collagen epitopes are masked early in the cycle, but become accessible in first trimester decidua. Type VI-collagen is abundant in endometrium in the proliferative phase, but is progressively lost in the secretory phase and decidua, in which it is retained only in blood vessel walls. Vitronectin is present in some blood vessels in decidua. Decidualising stromal cells also produce basement membrane components (type IV-collagen, laminin, heparan sulphate proteoglycan and a glycoprotein family recognised by monoclonal antibody G71) and these become organised into a pericellular aura.