Role of Endometrial Extracellular Vesicles in Mediating Cell-to-Cell Communication in the Uterus: A Review.

There are several critical events that occur in the uterus during early pregnancy which are necessary for the establishment and maintenance of pregnancy. These events include blastocyst implantation, uterine decidualization, uterine neoangiogenesis, differentiation of trophoblast stem cells into different trophoblast cell lineages, and formation of a placenta. These processes involve several different cell types within the pregnant uterus. Communication between these cell types must be intricately coordinated for successful embryo implantation and the formation of a functional maternal-fetal interface in the placenta. Understanding how this intricate coordination transpires has been a focus of researchers in the field for many years. It has long been understood that maternal endometrial tissue plays a key role in intercellular signaling during early pregnancy, sending signals to nearby tissues in a paracrine manner. Recently, insights have been obtained into the mechanisms by which these signaling events occur. Notably, the endometrium has been shown to secrete extracellular vesicles (EVs) that contain crucial cargo (proteins, lipids, RNA, miRNA) that are taken up by recipient cells to initiate a response leading to the occurrence of critical events during implantation and placentation. In this review, we aim to summarize the role that endometrium-derived EVs play in mediating cell-to-cell communications within the pregnant uterus to orchestrate the events that must occur to establish and maintain pregnancy. We will also discuss how aberrant endometrial EV signaling may lead to pathophysiological conditions, such as endometriosis and infertility.

Runx1 regulates critical factors that control uterine angiogenesis and trophoblast differentiation during placental development.

During early pregnancy in humans and rodents, uterine stromal cells undergo a remarkable differentiation to form the decidua, a transient maternal tissue that supports the growing fetus. It is important to understand the key decidual pathways that orchestrate the proper development of the placenta, a key structure at the maternal-fetal interface. We discovered that ablation of expression of the transcription factor Runx1 in decidual stromal cells in a conditional Runx1-null mouse model (Runx1d/d) causes fetal lethality during placentation. Further phenotypic analysis revealed that uteri of pregnant Runx1d/d mice exhibited severely compromised decidual angiogenesis and a lack of trophoblast differentiation and migration, resulting in impaired spiral artery remodeling. Gene expression profiling using uteri from Runx1d/d and control mice revealed that Runx1 directly controls the decidual expression of the gap junction protein connexin 43 (also known as GJA1), which was previously shown to be essential for decidual angiogenesis. Our study also revealed that Runx1 controls the expression of insulin-like growth factor (IGF) 2 and IGF-binding protein 4 (IGFBP4) during early pregnancy. While Runx1 deficiency drastically reduced the production of IGF2 by the decidual cells, we observed concurrent elevated expression of the IGFBP4, which regulates the bioavailability of IGFs, thereby controlling trophoblast differentiation. We posit that dysregulated expression of GJA1, IGF2, and IGFBP4 in Runx1d/d decidua contributes to the observed defects in uterine angiogenesis, trophoblast differentiation, and vascular remodeling. This study therefore provides unique insights into key maternal pathways that control the early phases of maternal-fetal interactions within a critical window during placental development.

Runx1 regulates critical factors that control uterine angiogenesis and trophoblast differentiation during placental development.

During early pregnancy in humans and rodents, uterine stromal cells undergo a remarkable differentiation to form the decidua, a transient maternal tissue that supports the growing fetus. It is important to understand the key decidual pathways that orchestrate the proper development of the placenta, a key structure at the maternal-fetal interface. We discovered that ablation of expression of the transcription factor Runx1 in decidual stromal cells in a conditional Runx1 -null mouse model ( Runx1 d/d ) causes fetal lethality during placentation. Further phenotypic analysis revealed that uteri of pregnant Runx1 d/d mice exhibited severely compromised decidual angiogenesis, and a lack of trophoblast differentiation and migration, resulting in impaired spiral artery remodeling. Gene expression profiling using uteri from Runx1 d/d and control mice revealed that Runx1 directly controls the decidual expression of the gap junction protein connexin 43 (also known as GJA1), which was previously shown to be essential for decidual angiogenesis. Our study also revealed a critical role of Runx1 in controlling insulin-like growth factor (IGF) signaling at the maternal-fetal interface. While Runx1-deficiency drastically reduced the production of IGF2 by the decidual cells, we observed concurrent elevated expression of the IGF-binding protein 4 (IGFBP4), which regulates the bioavailability of IGFs thereby controlling trophoblast differentiation. We posit that dysregulated expression of GJA1, IGF2, and IGFBP4 in Runx1 d/d decidua contributes to the observed defects in uterine angiogenesis, trophoblast differentiation, and vascular remodeling. This study therefore provides unique insights into key maternal pathways that control the early phases of maternal-fetal interactions within a critical window during placental development. Significance: A clear understanding of the maternal pathways that ensure coordination of uterine differentiation and angiogenesis with embryonic growth during the critical early stages of placenta formation still eludes us. The present study reveals that the transcription factor Runx1 controls a set of molecular, cellular, and integrative mechanisms that mediate maternal adaptive responses controlling uterine angiogenesis, trophoblast differentiation, and resultant uterine vascular remodeling, which are essential steps during placenta development.

Extracellular Vesicles Secreted by Mouse Decidual Cells Carry Critical Information for the Establishment of Pregnancy.

The mouse decidua secretes many factors that act in a paracrine/autocrine manner to critically control uterine decidualization, neovascularization, and tissue remodeling that ensure proper establishment of pregnancy. The precise mechanisms that dictate intercellular communications among the uterine cells during early pregnancy remain unknown. We recently reported that conditional deletion of the gene encoding the hypoxia-inducible transcription factor 2 alpha (Hif2α) in mouse uterus led to infertility. Here, we report that HIF2α in mouse endometrial stromal cells (MESCs) acts via the cellular trafficking regulator RAB27b to control the secretion of extracellular vesicles (EVs) during decidualization. We also found that Hif2α-regulated pathways influence the biogenesis of EVs. Proteomic analysis of EVs secreted by decidualizing MESCs revealed that they harbor a wide variety of protein cargoes whose composition changed as the decidualization process progressed. The EVs enhanced the differentiation capacity of MESCs and the production of angiogenic factors by these cells. We also established that matrix metalloproteinase-2, a prominent EV cargo protein, modulates uterine remodeling during decidualization. Collectively, our results support the concept that EVs are central to the mechanisms by which the decidual cells communicate with each other and other cell types within the uterus to facilitate successful establishment of pregnancy.

Extracellular vesicles secreted by human uterine stromal cells regulate decidualization, angiogenesis, and trophoblast differentiation.

In humans, the uterus undergoes a dramatic transformation to form an endometrial stroma-derived secretory tissue, termed decidua, during early pregnancy. The decidua secretes various factors that act in an autocrine/paracrine manner to promote stromal differentiation, facilitate maternal angiogenesis, and influence trophoblast differentiation and development, which are critical for the formation of a functional placenta. Here, we investigated the mechanisms by which decidual cells communicate with each other and with other cell types within the uterine milieu. We discovered that primary human endometrial stromal cells (HESCs) secrete extracellular vesicles (EVs) during decidualization and that this process is controlled by a conserved HIF2α-RAB27B pathway. Mass spectrometry revealed that the decidual EVs harbor a variety of protein cargo, including cell signaling molecules, growth modulators, metabolic regulators, and factors controlling endothelial cell expansion and remodeling. We tested the hypothesis that EVs secreted by the decidual cells mediate functional communications between various cell types within the uterus. We demonstrated that the internalization of EVs, specifically those carrying the glucose transporter 1 (GLUT1), promotes glucose uptake in recipient HESCs, supporting and advancing the decidualization program. Additionally, delivery of HESC-derived EVs into human endothelial cells stimulated their proliferation and led to enhanced vascular network formation. Strikingly, stromal EVs also promoted the differentiation of trophoblast stem cells into the extravillous trophoblast lineage. Collectively, these findings provide a deeper understanding of the pleiotropic roles played by EVs secreted by the decidual cells to ensure coordination of endometrial differentiation and angiogenesis with trophoblast function during the progressive phases of decidualization and placentation.

Sequence signatures of two public antibody clonotypes that bind SARS-CoV-2 receptor binding domain.

Since the COVID-19 pandemic onset, the antibody response to SARS-CoV-2 has been extensively characterized. Antibodies to the receptor binding domain (RBD) on the spike protein are frequently encoded by IGHV3-53/3-66 with a short complementarity-determining region (CDR) H3. Germline-encoded sequence motifs in heavy chain CDRs H1 and H2 have a major function, but whether any common motifs are present in CDR H3, which is often critical for binding specificity, is not clear. Here, we identify two public clonotypes of IGHV3-53/3-66 RBD antibodies with a 9-residue CDR H3 that pair with different light chains. Distinct sequence motifs on CDR H3 are present in the two public clonotypes that seem to be related to differential light chain pairing. Additionally, we show that Y58F is a common somatic hypermutation that results in increased binding affinity of IGHV3-53/3-66 RBD antibodies with a short CDR H3. These results advance understanding of the antibody response to SARS-CoV-2.

Sequence signatures of two IGHV3-53/3-66 public clonotypes to SARS-CoV-2 receptor binding domain.

Since the COVID-19 pandemic onset, the antibody response to SARS-CoV-2 has been extensively characterized. Antibodies to the receptor binding domain (RBD) on the spike protein are frequently encoded by IGHV3-53/3-66 with a short CDR H3. Germline-encoded sequence motifs in CDRs H1 and H2 play a major role, but whether any common motifs are present in CDR H3, which is often critical for binding specificity, have not been elucidated. Here, we identify two public clonotypes of IGHV3-53/3-66 RBD antibodies with a 9-residue CDR H3 that pair with different light chains. Distinct sequence motifs on CDR H3 are present in the two public clonotypes that appear to be related to differential light chain pairing. Additionally, we show that Y58F is a common somatic hypermutation that results in increased binding affinity of IGHV3-53/3-66 RBD antibodies with a short CDR H3. Overall, our results advance fundamental understanding of the antibody response to SARS-CoV-2.