Corrigendum: Deciphering decidual leukocyte traffic with serial intravascular staining.

[This corrects the article DOI: 10.3389/fimmu.2023.1332943.].

Decidual leukocytes respond to African lineage Zika virus infection with mild anti-inflammatory changes during acute infection in rhesus macaques.

Zika virus (ZIKV) can be vertically transmitted during pregnancy resulting in a range of adverse pregnancy outcomes. The decidua is commonly found to be infected by ZIKV, yet the acute immune response to infection remains understudied in vivo. We hypothesized that in vivo African-lineage ZIKV infection induces a pro-inflammatory response in the decidua. To test this hypothesis, we evaluated the decidua in pregnant rhesus macaques within the first two weeks following infection with an African-lineage ZIKV and compared our findings to gestationally aged-matched controls. Decidual leukocytes were phenotypically evaluated using spectral flow cytometry, and cytokines and chemokines were measured in tissue homogenates from the decidua, placenta, and fetal membranes. The results of this study did not support our hypothesis. Although ZIKV RNA was detected in the decidual tissue samples from all ZIKV infected dams, phenotypic changes in decidual leukocytes and differences in cytokine profiles suggest that the decidua undergoes mild anti-inflammatory changes in response to that infection. Our findings emphasize the immunological state of the gravid uterus as a relatively immune privileged site that prioritizes tolerance of the fetus over mounting a pro-inflammatory response to clear infection.

Cotyledon-Specific Flow Evaluation of Rhesus Macaque Placental Injury Using Ferumoxytol Dynamic Contrast-Enhanced MRI.

BACKGROUND: Recently, dynamic contrast-enhanced (DCE) MRI with ferumoxytol as contrast agent has recently been introduced for the noninvasive assessment of placental structure and function throughout. However, it has not been demonstrated under pathological conditions. PURPOSE: To measure cotyledon-specific rhesus macaque maternal placental blood flow using ferumoxytol DCE MRI in a novel animal model for local placental injury. STUDY TYPE: Prospective animal model. SUBJECTS: Placental injections of Tisseel (three with 0.5 mL and two with 1.5 mL), monocyte chemoattractant protein 1 (three with 100 µg), and three with saline as controls were performed in a total of 11 rhesus macaque pregnancies at approximate gestational day (GD 101). DCE MRI scans were performed prior (GD 100) and after (GD 115 and GD 145) the injection (term = GD 165). FIELD STRENGTH/SEQUENCE: 3 T, T1-weighted spoiled gradient echo sequence (product sequence, DISCO). ASSESSMENT: Source images were inspected for motion artefacts from the mother or fetus. Placenta segmentation and DCE processing were performed for the dynamic image series to measure cotyledon specific volume, flow, and normalized flow. Overall placental histopathology was conducted for controls, Tisseel, and MCP-1 animals and regions of tissue infarctions and necrosis were documented. Visual inspections for potential necrotic tissue were conducted for the two Tisseelx3 animals. STATISTICAL TESTS: Wilcoxon rank sum test, significance level P < 0.05. RESULTS: No motion artefacts were observed. For the group treated with 1.5 mL of Tisseel, significantly lower cotyledon volume, flow, and normalized flow per cotyledon were observed for the third gestational time point of imaging (day ~145), with mean normalized flow of 0.53 minute-1 . Preliminary histopathological analysis shows areas of tissue necrosis from a selected cotyledon in one Tisseel-treated (single dose) animal and both Tisseelx3 (triple dose) animals. DATA CONCLUSION: This study demonstrates the feasibility of cotyledon-specific functional analysis at multiple gestational time points and injury detection in a placental rhesus macaque model through ferumoxytol-enhanced DCE MRI. LEVEL OF EVIDENCE: NA TECHNICAL EFFICACY: Stage 2.

Proteogenomic and V(D)J Analysis of Human Decidual T Cells Highlights Unique Transcriptional Programming and Clonal Distribution.

Immunological tolerance toward the semiallogeneic fetus is one of many maternal adaptations required for a successful pregnancy. T cells are major players of the adaptive immune system and balance tolerance and protection at the maternal-fetal interface; however, their repertoire and subset programming are still poorly understood. Using emerging single-cell RNA sequencing technologies, we simultaneously obtained transcript, limited protein, and receptor repertoire at the single-cell level, from decidual and matched maternal peripheral human T cells. The decidua maintains a tissue-specific distribution of T cell subsets compared with the periphery. We find that decidual T cells maintain a unique transcriptome programming, characterized by restraint of inflammatory pathways by overexpression of negative regulators (DUSP, TNFAIP3, ZFP36) and expression of PD-1, CTLA-4, TIGIT, and LAG3 in some CD8 clusters. Finally, analyzing TCR clonotypes demonstrated decreased diversity in specific decidual T cell populations. Overall, our data demonstrate the power of multiomics analysis in revealing regulation of fetal-maternal immune coexistence.

Immune Dysfunction in Polycystic Ovary Syndrome.

Polycystic ovary syndrome (PCOS) is the most common endocrine disorder in reproductive-aged individuals with ovaries. It is associated with anovulation and increased risk to fertility and metabolic, cardiovascular, and psychological health. The pathophysiology of PCOS is still inadequately understood, although there is evidence of persistent low-grade inflammation, which correlates with associated visceral obesity. Elevated proinflammatory cytokine markers and altered immune cells have been reported in PCOS and raise the possibility that immune factors contribute to ovulatory dysfunction. Because normal ovulation is modulated by immune cells and cytokines in the ovarian microenvironment, the endocrine and metabolic abnormalities associated with PCOS orchestrate the accompanying adverse effects on ovulation and implantation. This review evaluates the current literature on the relationship between PCOS and immune abnormalities, with a focus on emerging research in the field.

Mononuclear Cells Negatively Regulate Endothelial Ca2+ Signaling.

Endothelial Ca2+ signaling has important roles to play in maintaining pregnancy associated vasodilation in the utero-placenta. Inflammatory cytokines, often elevated in vascular complications of pregnancy, negatively regulate ATP-stimulated endothelial Ca2+ signaling and associated nitric oxide production. However, the role of direct engagement of immune cells on endothelial Ca2+ signaling and therefore endothelial function is unclear. To model immune-endothelial interactions, herein, we evaluate the effects of peripheral blood mononuclear cells (PBMCs) in short-term interaction with human umbilical vein endothelial cells (HUVECs) on agonist-stimulated Ca2+ signaling in HUVECs. We find that mononuclear cells (10:1 and 25:1 mononuclear: HUVEC) cause decreased ATP-stimulated Ca2+ signaling; worsened by activated mononuclear cells possibly due to increased cytokine secretion. Additionally, monocytes, natural killers, and T-cells cause decrease in ATP-stimulated Ca2+ signaling using THP-1 (monocyte), NKL (natural killer cells), and Jurkat (T-cell) cell lines, respectively. PBMCs with Golgi-restricted protein transport prior to interaction with endothelial cells display rescue in Ca2+ signaling, strongly suggesting that secreted proteins from PBMCs mediate changes in HUVEC Ca2+ signaling. We propose that endothelial cells from normal pregnancy interacting with PBMCs may model preeclamptic endothelial-immune interaction and resultant endothelial dysfunction.

Continuous positive airway pressure treatment of obstructive sleep apnea and hypertensive complications in high-risk pregnancy.

PURPOSE: To evaluate whether or not continuous positive airway pressure (CPAP) treatment in pregnancies complicated by obstructive sleep apnea (OSA) is associated with a decrease in hypertensive disorders of pregnancy. METHODS: This was a retrospective cohort study of perinatal outcomes in women who underwent objective OSA testing and treatment as part of routine clinical care during pregnancy. Where diagnostic criteria for OSA were reached (respiratory event index (REI) ≥ 5 events per hour), patients were offered CPAP therapy. Obstetrical outcomes were compared between the control group (no OSA), the group with untreated OSA (OSA diagnosed, not CPAP compliant), and the group with treated OSA (OSA diagnosed and CPAP compliant), with CPAP compliance defined as CPAP use ≥ 4 h, 70% of the time or greater. A composite hypertension outcome combined diagnoses of gestational hypertension (gHTN) and preeclampsia (PreE) of any severity. RESULTS: The study comprised outcomes from 177 completed pregnancies. Our cohort was characterized by obesity, with average body mass indices > 35 kg/m2, and average maternal age > 30 years old. CPAP was initiated at an average gestational age of 23 weeks (12.1-35.3 weeks), and average CPAP use was 5.9 h (4-8.5 h). The composite hypertension outcome occurred in 43% of those without OSA (N = 77), 64% of those with untreated OSA (N = 77), and 57% of those with treated OSA, compliant with CPAP (N = 23) (p = 0.034). CONCLUSION: Real-world data in this small study suggest that CPAP therapy may modulate the increased risk of hypertensive complications in pregnancies complicated by OSA.

Deciphering decidual leukocyte traffic with serial intravascular staining.

The decidual immunome is dynamic, dramatically changing its composition across gestation. Early pregnancy is dominated by decidual NK cells, with a shift towards T cells later in pregnancy. However, the degree, timing, and subset-specific nature of leukocyte traffic between the decidua and systemic circulation during gestation remains poorly understood. Herein, we employed intravascular staining in pregnant C57BL/6J mice and cynomolgus macaques (Macaca fascicularis) to examine leukocyte traffic into the decidual basalis during pregnancy. Timed-mated or virgin mice were tail-vein injected with labelled αCD45 antibodies 24 hours and 5 minutes before sacrifice. Pregnant cynomolgus macaques (GD155) were infused with labelled αCD45 at 2 hours or 5 mins before necropsy. Decidual cells were isolated and resulting suspensions analyzed by flow cytometry. We found that the proportion of intravascular (IVAs)-negative leukocytes (cells labeled by the 24h infusion of αCD45 or unlabeled) decreased across murine gestation while recent immigrants (24h label only) increased in mid- to late-gestation. In the cynomolgus model our data confirmed differential labeling of decidual leukocytes by the infused antibody, with the 5 min infused animal having a higher proportion of IVAs+ cells compared to the 2hr infused animal. Decidual tissue sections from both macaques showed the presence of intravascularly labeled cells, either in proximity to blood vessels (5min infused animal) or deeper into decidual stroma (2hr infused animal). These results demonstrate the value of serial intravascular staining as a sensitive tool for defining decidual leukocyte traffic during pregnancy.

Multiparameter Flow Cytometry for Detailed Characterization of Peritoneal Immune Cells from Patients with Ovarian Cancer.

Multiparameter flow cytometry is a convenient and efficient method for thorough phenotyping of cells, and especially immune cells from various tissues. We have successfully used multiparameter flow cytometry to characterize immune cells from patients with ovarian cancer and leveraged dimensionality reduction and machine learning for optimized visualization and analysis. Herein, we describe our optimized and established protocols for the labeling of cells with fluorophore-conjugated antibody panels, followed by details on data acquisition. Finally, we describe methods for analysis of the flow cytometry data using both FlowJo as well as R package, Cytofkit, for multidimensional data visualization.

Multiomic analysis reveals decidual-specific transcriptional programing of MAIT cells.

PROBLEM: Mucosal-Associated Invariant T (MAIT) cells have been recently identified at the maternal-fetal interface. However, transcriptional programming of decidual MAIT cells in pregnancy remains poorly understood. METHOD OF STUDY: We employed a multiomic approach to address this question. Mononuclear cells from the decidua basalis and parietalis, and control PBMCs, were analyzed via flow cytometry to investigate MAIT cells in the decidua and assess their transcription factor expression. In a separate study, both decidual and matched peripheral MAIT cells were analyzed using Cellular Indexing of Transcriptomes and Epitopes by Sequencing (CITE-seq) coupled with gene expression analysis. Lastly, decidual MAIT cells were stimulated with E.coli and expression of MR1 by antigen presenting cells was measured to evaluate decidual MAIT cell function. RESULTS: First, we identified MAIT cells in both the decidua basalis and parietalis. CITE-seq, coupled with scRNA-seq gene expression analysis, highlighted transcriptional programming differences between decidual and matched peripheral MAIT cells at a single cell resolution. Transcription factor expression analysis further highlighted transcriptional differences between decidual MAIT cells and non-matched peripheral MAIT cells. Functionally, MAIT cells are skewed towards IFNγ and TNFα production upon stimulation, with E.coli leading to IFNγ production. Lastly, we demonstrate that MR1, the antigen presenting molecule restricting MAIT cells, is expressed by decidual APCs. CONCLUSION: MAIT cells are present in the decidua basalis and obtain a unique gene expression profile. The presence of MR1 on APCs coupled with in vitro activation by E.coli suggests that MAIT cells might be involved in tissue-repair mechanisms at the maternal-fetal interface.

Identifying novel ovarian tumor biomarkers through mining of the transcriptome of circulating immune cells: A proof-of-concept study.

OBJECTIVE: Treatment of high-grade serous ovarian cancer (HGSOC) will benefit from early detection of cancer. Here, we provide proof-of-concept data supporting the hypothesis that circulating immune cells, because of their early recognition of tumors and the tumor microenvironment, can be considered for biomarker discovery. METHODS: Longitudinal blood samples from C57BL/6 mice bearing syngeneic ovarian tumors and peripheral blood mononuclear cells (PBMC) from healthy postmenopausal women and newly diagnosed for HGSOC patients were subjected to RNASeq. The results from human immune cells were validated using Affymetrix microarrays. Differentially expressed transcripts in immune cells from tumor-bearing mice and HGSOC patients were compared to matching controls. RESULTS: A total of 1282 transcripts (798 and 484, up- and downregulated, respectively) were differentially expressed in the tumor-bearing mice as compared with controls. Top 100 genes showing longitudinal changes in gene expression 2, 4, 7, and 18 days after tumor implantation were identified. Analysis of the PBMC from healthy post-menopausal women and HGSOC patients identified 4382 differentially expressed genes and 519 of these were validated through Affymetrix microarray analysis. A total of 384 genes, including IL-1R2, CH3L1, Infitm1, FP42, CXC42, Hdc, Spib, and Sema6b, were differentially expressed in the human and mouse datasets. CONCLUSION: The PBMC transcriptome shows longitudinal changes in response to the progressing tumor. Several potential biomarker transcripts were identified in HGSOC patients and mouse models. Monitoring their expression in individual PBMC subsets can serve as additional discriminator for the diagnosis of HGSOC.

The promise of placental extracellular vesicles: models and challenges for diagnosing placental dysfunction in utero†.

Monitoring the health of a pregnancy is of utmost importance to both the fetus and the mother. The diagnosis of pregnancy complications typically occurs after the manifestation of symptoms, and limited preventative measures or effective treatments are available. Traditionally, pregnancy health is evaluated by analyzing maternal serum hormone levels, genetic testing, ultrasonographic imaging, and monitoring maternal symptoms. However, researchers have reported a difference in extracellular vesicle (EV) quantity and cargo between healthy and at-risk pregnancies. Thus, placental EVs (PEVs) may help to understand normal and aberrant placental development, monitor pregnancy health in terms of developing placental pathologies, and assess the impact of environmental influences, such as infection, on pregnancy. The diagnostic potential of PEVs could allow for earlier detection of pregnancy complications via noninvasive sampling and frequent monitoring. Understanding how PEVs serve as a means of communication with maternal cells and recognizing their potential utility as a readout of placental health have sparked a growing interest in basic and translational research. However, to date, PEV research with animal models lags behind human studies. The strength of animal pregnancy models is that they can be used to assess placental pathologies in conjunction with isolation of PEVs from fluid samples at different time points throughout gestation. Assessing PEV cargo in animals within normal and complicated pregnancies will accelerate the translation of PEV analysis into the clinic for potential use in prognostics. We propose that appropriate animal models of human pregnancy complications must be established in the PEV field.

Different Human Immune Lineage Compositions Are Generated in Non-Conditioned NBSGW Mice Depending on HSPC Source.

NBSGW mice are highly immunodeficient and carry a hypomorphic mutation in the c-kit gene, providing a host environment that supports robust human hematopoietic expansion without pre-conditioning. These mice thus provide a model to investigate human hematopoietic engraftment in the absence of conditioning-associated damage. We compared transplantation of human CD34+ HSPCs purified from three different sources: umbilical cord blood, adult bone marrow, and adult G-CSF mobilized peripheral blood. HSPCs from mobilized peripheral blood were significantly more efficient (as a function of starting HSPC dose) than either cord blood or bone marrow HSPCs at generating high levels of human chimerism in the murine blood and bone marrow by 12 weeks post-transplantation. While T cells do not develop in this model due to thymic atrophy, all three HSPC sources generated a human compartment that included B lymphocytic, myeloid, and granulocytic lineages. However, the proportions of these lineages varied significantly according to HSPC source. Mobilized blood HSPCs produced a strikingly higher proportion of granulocyte lineage cells (~35% as compared to ~5%), whereas bone marrow HSPC output was dominated by B lymphocytic cells, and cord blood HSPC output was enriched for myeloid lineages. Following transplantation, all three HSPC sources showed a shift in the CD34+ subset towards CD45RA+ progenitors along with a complete loss of the CD45RA-CD49f+ long-term HSC subpopulation, suggesting this model promotes mainly short-term HSC activity. Mice transplanted with cord blood HSPCs maintained a diversified human immune compartment for at least 36 weeks after the primary transplant, although mice given adult bone marrow HSPCs had lost diversity and contained only myeloid cells by this time point. Finally, to assess the impact of non-HSPCs on transplantation outcome, we also tested mice transplanted with total or T cell-depleted adult bone marrow mononuclear cells. Total bone marrow mononuclear cell transplants produced significantly lower human chimerism compared to purified HSPCs, and T-depletion rescued B cell levels but not other lineages. Together these results reveal marked differences in engraftment efficiency and lineage commitment according to HSPC source and suggest that T cells and other non-HSPC populations affect lineage output even in the absence of conditioning-associated inflammation.

Benign ovarian thecoma with markedly elevated serum inhibin B levels mimicking adult granulosa cell tumor.

INTRODUCTION: Elevated serum inhibin B is a classic marker of adult granulosa cell tumors. Here we discuss an extremely rare and informative case of elevated inhibin B associated with an ovarian thecoma. CASE: A 57 year-old postmenopausal female presented with recurrent bleeding and was found to have an adnexal mass with an elevated serum inhibin B level of 1,915 pg/mL (normal range 10-200 pg/mL). With a preoperative diagnosis of adult granulosa cell tumor, she underwent surgical management for what was ultimately a benign ovarian thecoma. The diagnosis of thecoma was confirmed by a pericellular pattern of reticulin staining and the lack of a FOXL2 mutation by molecular testing. CONCLUSION: This case demonstrates that inhibin B lacks specificity as a tumor marker for adult granulosa cell tumor, even at very high levels. Knowledge of benign alternative explanations for this finding can facilitate improved preoperative patient counseling. Pertinent literature is reviewed, with an emphasis on proposed hypotheses for inhibin overproduction.

Identification of unique clusters of T, dendritic, and innate lymphoid cells in the peritoneal fluid of ovarian cancer patients.

PROBLEM: We hypothesize that activated peritoneal immune cells can be redirected to target ovarian tumors. Here, we obtain fundamental knowledge of the peritoneal immune environment through deep immunophenotyping of T cells, dendritic cells (DC), and innate lymphoid cells (ILC) of ovarian cancer patients. METHOD OF STUDY: T cells, DC, and ILC from ascites of ovarian cancer patients (n = 15) and peripheral blood of post-menopausal healthy donors (n = 6) were immunophenotyped on a BD Fortessa cytometer using three panels-each composed of 16 antibodies. The data were analyzed manually and by t-SNE/DensVM. CA125 levels were obtained from patient charts. RESULTS: We observed decreased CD3+ T cells and a higher proportion of activated CD4+ and effector memory CD4+ /CD8+ T cells, plasmacytoid DC, CD1c+ and CD141+ myeloid DC and CD56Hi NK cells in ascites. t-SNE/DensVM identified eight T cell, 17 DC, and 17 ILC clusters that were unique in the ascites compared to controls. Hierarchical clustering of cell frequency distinctly segregated the T-cell and ILC clusters from controls. Increased CA125 levels were associated with decreased CD8+ /CD45RA+ /CD45RO- /CCR7- T cells. CONCLUSION: The identified immune clusters serve as the basis for interrogation of the peritoneal immune environment and the development of novel immunologic modalities against ovarian cancer.

The role of RORγt at maternal-fetal interface during murine pregnancy.

PROBLEM: Innate lymphoid cells (ILCs, including NK cells) and their subsets are the most frequent lymphocytes at the maternal-fetal interface (decidua). Recent recognition of extensive ILC subset diversity at mucosal sites and the possible role they might play at different stages of pregnancy poses questions about their composition and lineage stability. Namely, RORγt-dependent ILC3s have been recognized as a key cellular mediator of tissue organization in the gut and secondary lymphoid organs, prompting examination of their distribution and role in decidua during pregnancy. METHOD OF STUDY: We employed highly polychromatic flow cytometry with conventional and machine learning-aided analysis to map ILC subsets and dissected the role of canonical transcription factor RORγt using fate-mapping animals and RORγt-/- animals. RESULTS: We demonstrate a comprehensive immunome map of ILCs/NKs, revealing a dynamic interface even in the absence of antigenic or allogeneic challenge. Strikingly, we demonstrate plasticity of RORγt expression in decidual ILCs with across gestation. However, gross reproductive efficiency is not affected in RORγt-/- animals. CONCLUSION: These results indicated that RORγt+ ILCs are highly plastic at the maternal-fetal interface, but dispensable for normal pregnancy, revealing a novel mechanism of transcriptional immunoregulation in pregnancy.

Single-cell technologies in reproductive immunology.

The maternal-fetal interface represents a unique immune privileged site that maintains the ability to defend against pathogens while orchestrating the necessary tissue remodeling required for placentation. The recent discovery of novel cellular families (innate lymphoid cells, tissue-resident NK cells) suggests that our understanding of the decidual immunome is incomplete. To understand this complex milieu, new technological developments allow reproductive immunologists to collect increasingly complex data at a cellular resolution. Polychromatic flow cytometry allows for greater resolution in the identification of novel cell types by surface and intracellular protein. Single-cell RNA-seq coupled with microfluidics allows for efficient cellular transcriptomics. The extreme dimensionality and size of data sets generated, however, requires the application of novel computational approaches for unbiased analysis. There are now multiple dimensionality reduction (tSNE, SPADE) and visualization tools (SPICE) that allow researchers to efficiently analyze flow cytometry data. Development of computational tools has also been extended to RNA-seq data (including scRNA-seq), which requires specific analytical tools. Here, we provide an overview and a brief primer for the reproductive immunology community on data acquisition and computational tools for the analysis of complex flow cytometry and RNA-seq data.

Transcriptional and Functional Programming of Decidual Innate Lymphoid Cells.

A successful pregnancy requires many physiological adaptations from the mother, including the establishment of tolerance toward the semiallogeneic fetus. Innate lymphoid cells (ILCs) have arisen as important players in immune regulation and tissue homeostasis at mucosal and barrier surfaces. Dimensionality reduction and transcriptomic analysis revealed the presence of two novel CD56Bright decidual ILCs that express low T-bet and divergent Eomes levels. Transcriptional correlation with recently identified first trimester decidual dNKs suggests that these novel decidual ILCs might be present throughout pregnancy. Functional testing with permutation analysis revealed production of multiple factors by individual cells, with a preference for IFNγ and VEGF. Overall, our data suggests continuity of a unique decidual innate lymphocytes across pregnancy with a polyfunctional functional profile conducive for pregnancy.

Decidual-Placental Immune Landscape During Syngeneic Murine Pregnancy.

Adaptive immune system, principally governed by the T cells-dendritic cells (DCs) nexus, is an essential mediator of gestational fetal tolerance and protection against infection. However, the exact composition and dynamics of DCs and T cell subsets in gestational tissues are not well understood. These are controlled in human physiology by a complex interplay of alloantigen distribution and presentation, cellular/humoral active and passive tolerance, hormones/chemokines/angiogenic factors and their gradients, systemic and local microbial communities. Reductive discrimination of these factors in physiology and pathology of model systems and humans requires simplification of the model and increased resolution of interrogative technologies. As a baseline, we have studied the gestational tissue dynamics in the syngeneic C57BL/6 mice, as the simplest immunological environment, and focused on validating the approach to increased data density and computational analysis pipeline afforded by highly polychromatic flow cytometry and machine learning interpretation. We mapped DC and T cell subsets, and comprehensively examined their maternal (decidual)-fetal (placental) interface dynamics. Both frequency and composition of decidual DCs changed across gestation, with a dramatic increase in myeloid DCs in early pregnancy, and exclusion of plasmacytoid DCs. CD4+ T cells, in contrast, were lower at all gestational ages and an unusual CD4-CD8-TCRαß+group was prominent at mid-pregnancy. Dimensionality reduction with machine learning-aided clustering revealed that CD4-CD8- T cells were phenotypically different from CD4+ and CD8+ T cells. Additionally, divergence between maternal decidual and fetal placental compartment was prominent, with absence of DCs from the placenta, but not decidua or embryo. These results provide a novel framework and a syngeneic baseline on which the specific role of alloantigen/tolerance, polymicrobial environment, and models of pregnancy pathology can be precisely modeled and analyzed.

Computational flow cytometry analysis reveals a unique immune signature of the human maternal-fetal interface.

PROBLEM: Decidual immune dysregulation is thought to underlie major pregnancy disorders; however, incomplete understanding of the decidual immune interface has hampered the mechanistic investigation. METHOD OF STUDY: Human term decidua was collected, and single-cell phenotypic information was acquired by highly polychromatic flow cytometry. Cellular identity analysis was performed with t-distributed stochastic neighbor embedding, DensVM clustering, and matched to CellOntology database. RESULTS: Traditional analytical methods validated known cellular T and dendritic cell subsets in human term decidua. Computational analysis revealed a complex and tissue-specific decidual immune signature in both the innate and adaptive immune compartments. CONCLUSION: Polychromatic flow cytometry with a streamlined computational analysis pipeline is a feasible approach to comprehensive immunome mapping of human term decidua. As an unbiased, standardized method of investigation, computational flow cytometry promises to unravel the immune pathology of pregnancy disorders.