Environmental Toxicant Exposure Paralyzes Human Placental Macrophage Responses to Microbial Threat.

Exposure to environmental toxicants (such as dioxins) has been epidemiologically linked to adverse reproductive health outcomes, including placental inflammation and preterm birth. However, the molecular underpinnings that govern these outcomes in gravid reproductive tissues remain largely unclear. Placental macrophages (also known as Hofbauer cells) are crucial innate immune cells that defend the gravid reproductive tract and help promote maternal-fetal tolerance. We hypothesized that exposure to environmental toxicants such as 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) could alter placental macrophage responses to inflammatory insults such as infection. To test this, placental macrophages were cultured in the presence or absence of TCDD and then infected with the perinatal pathogen Group B Streptococcus (GBS). Our results indicate that TCDD is lethal to placental macrophages at and above a 5 nM concentration and that sublethal dioxin exposure inhibits phagocytosis and cytokine production. Taken together, these results indicate that TCDD paralyzes placental macrophage responses to bacterial infection.

Impact of Metabolic Stress on BeWo Syncytiotrophoblast Function.

During placental formation, cytotrophoblasts (CTBs) fuse into multinucleate, microvilli-coated syncytiotrophoblasts (STBs), which contact maternal blood, mediating nutrient, metabolite, and gas exchange between mother and fetus, and providing a barrier against fetal infection. Trophoblasts remodel the surrounding extracellular matrix through the secretion of matrix metalloproteinases (MMPs). Maternal obesity and diabetes mellitus can negatively impact fetal development and may impair trophoblast function. We sought to model the impact of metabolic stress on STB function by examining MMP and hormone secretion. The BeWo CTB cell line was syncytialized to STB-like cells with forskolin. Cell morphology was examined by electron microscopy and immunofluorescence; phenotype was further assessed by ELISA and RT-qPCR. STBs were exposed to a metabolic stress cocktail (MetaC: 30 mM glucose, 10 nM insulin, and 0.1 mM palmitic acid). BeWo syncytialization was demonstrated by increased secretion of HCGß and progesterone, elevated syncytin gene expression (ERVW-1 and ERVFRD-1), loss of tight junctions, and increased surface microvilli. MetaC strongly suppressed syncytin gene expression (ERVW-1 and ERVFRD-1), suppressed HCGß and progesterone secretion, and altered both MMP-9 and MMP-2 production. Metabolic stress modeling diabetes and obesity altered BeWo STB hormone and MMP production in vitro.

Comparison of Published Estimates of the National Prevalence of Iron, Vitamin A, and Zinc Deficiency and Sources of Inconsistencies.

Micronutrient deficiencies result in a broad range of adverse health and functional consequences, but the true prevalence of specific deficiencies remains uncertain because limited information is available from nationally representative surveys using recommended biomarkers. The present review compares various reported national deficiency prevalence estimates for nutrients and years where the estimates overlap for individual countries that conducted nationally representative surveys and explores possible reasons for any discrepancies discovered. Nationally representative micronutrient status surveys that were conducted since 2000 among preschool-aged children or women of reproductive age and included assessment of iron, vitamin A, or zinc status based on recognized biomarkers were considered eligible for inclusion, along with any modeled deficiency prevalence estimates for these same countries and years. There was considerable variation across different published prevalence estimates, with larger inconsistencies when the prevalence estimate was based on proxies, such as hemoglobin for iron deficiency and dietary zinc availability for zinc deficiency. Numerous additional methodological issues affected the prevalence estimates, such as which biomarker and what cutoff was used to define deficiency, whether the biomarker was adjusted for inflammation, and what adjustment method was used. For some country-years, the various approaches resulted in fairly consistent prevalence estimates. For other country-years, however, the results differed markedly and changed the conclusions regarding the existence and severity of the micronutrient deficiency as a public health concern. In conclusion, to determine micronutrient status, we consider the assessment of one of the recommended biomarkers in a population representative survey as the best available information. If indicated, results should be adjusted for inflammation and generally acceptable cutoffs should be applied to facilitate comparisons, although individual countries may also apply nationally defined cutoffs to determine when and where to intervene. Global consensus is needed on best practices for presenting survey results and defining the prevalence of deficiency.

Streptococcus agalactiae npx Is Required for Survival in Human Placental Macrophages and Full Virulence in a Model of Ascending Vaginal Infection during Pregnancy.

Streptococcus agalactiae, also known as group B Streptococcus (GBS), is a Gram-positive encapsulated bacterium that colonizes the gastrointestinal tract of 30 to 50% of humans. GBS causes invasive infection during pregnancy that can lead to chorioamnionitis, funisitis, preterm prelabor rupture of membranes (PPROM), preterm birth, neonatal sepsis, and maternal and fetal demise. Upon infecting the host, GBS encounters sentinel innate immune cells, such as macrophages, within reproductive tissues. Once phagocytosed by macrophages, GBS upregulates the expression of the gene npx, which encodes an NADH peroxidase. GBS mutants with an npx deletion (Δnpx) are exquisitely sensitive to reactive oxygen stress. Furthermore, we have shown that npx is required for GBS survival in both THP-1 and placental macrophages. In an in vivo murine model of ascending GBS vaginal infection during pregnancy, npx is required for invading reproductive tissues and is critical for inducing disease progression, including PPROM and preterm birth. Reproductive tissue cytokine production was also significantly diminished in Δnpx mutant-infected animals compared to that in animals infected with wild-type (WT) GBS. Complementation in trans reversed this phenotype, indicating that npx is critical for GBS survival and the initiation of proinflammatory signaling in the gravid host. IMPORTANCE This study sheds new light on the way that group B Streptococcus (GBS) defends itself against oxidative stress in the infected host. The enzyme encoded by the GBS gene npx is an NADH peroxidase that, our study reveals, provides defense against macrophage-derived reactive oxygen stress and facilitates infections of the uterus during pregnancy. This enzyme could represent a tractable target for future treatment strategies against invasive GBS infections.

Streptococcus agalactiae cadD alleviates metal stress and promotes intracellular survival in macrophages and ascending infection during pregnancy.

Perinatal infection with Streptococcus agalactiae, or Group B Streptococcus (GBS), is associated with preterm birth, neonatal sepsis, and stillbirth. Here, we study the interactions of GBS with macrophages, essential sentinel immune cells that defend the gravid reproductive tract. Transcriptional analyses of GBS-macrophage co-cultures reveal enhanced expression of a gene encoding a putative metal resistance determinant, cadD. Deletion of cadD reduces GBS survival in macrophages, metal efflux, and resistance to metal toxicity. In a mouse model of ascending infection during pregnancy, the ΔcadD strain displays attenuated bacterial burden, inflammation, and cytokine production in gestational tissues. Furthermore, depletion of host macrophages alters cytokine expression and decreases GBS invasion in a cadD-dependent fashion. Our results indicate that GBS cadD plays an important role in metal detoxification, which promotes immune evasion and bacterial proliferation in the pregnant host.

Critical assessment of the current indicator for antenatal iron-containing supplementation coverage: Insights from a mixed-methods study.

Daily consumption of iron-containing supplements is recommended for all pregnant women but there is no approved global standard indicator for assessing supplementation coverage. Furthermore, the validity of commonly used coverage indicators for iron-containing supplement consumption is questionable. The WHO-UNICEF Technical Expert Advisory Group on Nutrition Monitoring, and partners, have systematically worked to identify a feasible and valid indicator of iron-containing supplement coverage for reporting by countries. In 2019, we conducted key informant interviews with respondents in eight countries, fielded an online survey (in three languages using SurveyMonkey) to which 142 nutrition professionals from 52 countries responded, and used Demographic and Health Surveys (DHS) data from four countries to assess determinants of the quality of iron-containing supplement coverage data. Less than half (45%) of online survey respondents were satisfied with the current methods for collecting iron-containing supplement coverage data in their context. Recommended changes by study respondents include recall period <5 years, adding questions about counselling, including other beneficiary groups, and assessing supply chain functionality. The DHS analysis suggested an association between time since pregnancy and data quality. Data heaping on multiples of 30 was observed in 40%-75% of data. There is a clear demand for a revised indicator and measurement guidance for coverage of iron-containing supplementation during pregnancy. Future research should continue the development and validation of a global indicator, to more precisely validate the quality of recall data, including the distinction between distribution and consumption using various question formulations.

Self-management of iron and folic acid supplementation during pre-pregnancy, pregnancy and postnatal periods: a systematic review.

INTRODUCTION: While the use of folic acid pre-pregnancy and iron and folic acid (IFA) during pregnancy and postnatal have been demonstrated to be effective and are recommended interventions by WHO, ensuring individuals adhere to the supplementation regimen can be a challenge. Self-care interventions that support an individual's ability to promote their own health with or without the support of health workers could help promote the uptake and adherence to supplementation. This systematic review assessed the evidence around self-management of IFA or folic acid supplementation accessed over-the-counter during pre-pregnancy, pregnancy and postnatal periods. METHODS: Peer-reviewed studies were included if they compared self-management of IFA or folic acid supplementation with health worker-initiated supplement use on maternal and/or fetal and newborn health outcomes, end-users' or health workers' values and preferences, or cost and/or cost-effectiveness. We searched PubMed, CINAHL, LILACS and EMBASE for articles published through November 2020, hand-searched clinical trial registries, reviewed databases and contacted experts in the field. Abstract screening and full-text review were conducted independently by two reviewers. RESULTS: Overall, 2344 results were identified, and 28 studies were identified for full-text review. All studies were excluded, as they were not primary research, lacked the outcomes of interest, lacked specificity in supplement type, and/or lacked a comparison group. CONCLUSION: No evidence was identified that distinguishes self-management of folic acid supplements pre-pregnancy and of IFA supplements during pregnancy and postnatal, highlighting a gap in our current understanding of self-care related to dietary supplementation in pregnancy. The findings of this review identify an area for further research to support the current movement towards self-care interventions as an added choice to help individuals more fully attain their reproductive health and rights. SYSTEMATIC REVIEW REGISTRATION NUMBER: PROSPERO CRD42020205548.

Cytotrophoblasts suppress macrophage-mediated inflammation through a contact-dependent mechanism.

PROBLEM: Gestational membrane (GM) infection provokes inflammation and can result in preterm prelabor rupture of membranes (PPROM). The choriodecidual layer of the GM includes decidual stromal cells (DSC), cytotrophoblasts (CTB), and macrophages (Mφ). Our laboratory has previously shown that DSCs suppress Mφ TNF-α production through secreted prostaglandin E2 . We hypothesized that CTBs would also inhibit Mφ cytokine expression through secreted mediators. METHOD OF STUDY: THP.1 Mφ-like cells with an NF-κB reporter construct or human blood monocyte-derived Mφ were co-cultured with the Jeg3 CTB cell line or primary human CTBs and challenged with group B streptococcus (GBS) or Toll-like receptor (TLR) agonists. Conditioned medium generated from CTB cultures was applied to Mφ cultures before infection or treatment. Alternatively, CTBs were co-incubated with, but physically separated from, Mφ and GBS or TLR-stimulated. NF-κB was assessed via alkaline phosphatase assay, and proinflammatory mediators were assessed by qRT-PCR and ELISA. RESULTS: CTBs suppressed GBS- or TLR-stimulated Mφ NF-κB activity, and TNF-α and MMP9 production. Direct physical contact between CTBs and Mφ was required for full immunosuppression. Immunosuppression could be overcome by increasing the ratio of Mφ to CTB. CONCLUSIONS: CTBs limit Mφ NF-κB activation and production of TNF-α and MMP9 through an as-yet unknown, cell-to-cell contact-mediated mechanism. This suppression is distinct from the PGE2 -mediated Mφ TNF-α suppression by DSC, suggesting that DSCs and CTBs regulate Mφ inflammation through distinct mechanisms. How Mφ integrates these signals in an intact GM will be paramount to determining causes and prevention of PPROM.

Fetal Membrane Organ-On-Chip: An Innovative Approach to Study Cellular Interactions.

OBJECTIVE: Fetal membranes, a vital component that helps maintain pregnancy and contribute to parturition signaling, are often studied in segments due to its structural complexity. Transwells are traditionally used to study cell interactions; however, their usefulness is limited. To overcome these difficulties, a fetal membrane-organ-on-chip (FM-OO-C) was created to study interactive properties of amnion epithelial cells (AECs) and decidual cells compared to transwell systems. METHODS: Primary AECs and decidual cells from term, nonlaboring fetal membranes were cultured in a 2-chamber (AEC/decidual cell) FM-OO-C device and sandwiched between a semipermeable membrane. Cells were treated with cigarette smoke extract (CSE) or dioxin, and membrane permeability and cellular senescence were measured after 48 hours. The same experiments were conducted in transwells for comparisons. RESULTS: Compared to transwell cultures, FM-OO-C model produced better membrane permeability readings regardless of the side of treatment or time point. Membrane permeabilization was higher in AECs directly treated with CSE (1.6 fold) compared to similar treatment on the decidual side (1.2 fold). In FM-OO-C, treatments forced changes between cellular layers. This was evident when CSE and dioxin-induced senescence on one side of the chamber produced similar changes on the opposite side. This effect was minimal in the transwell system. CONCLUSION: The controlled environment of an FM-OO-C allows for improved signal propagation between cells by minimizing noise and highlighting the small changes between treatments that cannot be seen in conventional transwell devices. Fetal membrane-organ-on-chip provides a better interaction between cell types that can be used to study fetal-maternal signaling during pregnancy in future studies.

Palmitate induces apoptotic cell death and inflammasome activation in human placental macrophages.

INTRODUCTION: There is an increasing prevalence of non-communicable diseases worldwide. Metabolic diseases such as obesity and gestational diabetes mellitus (GDM) increasingly affect women during pregnancy, which can harm pregnancy outcomes and the long-term health and wellbeing of exposed offspring. Both obesity and GDM have been associated with proinflammatory effects within the placenta, the critical organ governing fetal development. METHODS: The purpose of these studies was to model, in vitro, the effects of metabolic stress (high levels of glucose, insulin and saturated lipids) on placental macrophage biology, since these cells are the primary innate immune phagocyte within the placenta with roles in governing maternofetal immune tolerance and antimicrobial host defense. Macrophages were isolated from the villous core of term, human placentae delivered through nonlaboring, elective Cesarean sections and exposed to combinations of elevated glucose (30 mM), insulin (10 nM) and the saturated lipid palmitic acid (palmitate, 0.4 mM). RESULTS: We found that palmitate alone induced the activation of the nucleotide-binding oligomerization domain-like receptor (NLR) Family Pyrin Domain Containing 3 (NLRP3) inflammasome in placental macrophages, which was associated with increased interleukin 1 beta release and an increase in apoptotic cell death. Glucose and insulin neither provoked these effects nor augmented the impact of palmitate itself. DISCUSSION: Our findings confirm an impact of saturated fat on placental macrophage immune activation and could be relevant to the impact of metabolic stress in vivo.

Fetal Membrane Organ-On-Chip: An Innovative Approach to Study Cellular Interactions.

OBJECTIVE:: Fetal membranes, a vital component that helps maintain pregnancy and contribute to parturition signaling, are often studied in segments due to its structural complexity. Transwells are traditionally used to study cell interactions; however, their usefulness is limited. To overcome these difficulties, a fetal membrane-organ-on-chip (FM-OO-C) was created to study interactive properties of amnion epithelial cells (AECs) and decidual cells compared to transwell systems. METHODS:: Primary AECs and decidual cells from term, nonlaboring fetal membranes were cultured in a 2-chamber (AEC/decidual cell) FM-OO-C device and sandwiched between a semipermeable membrane. Cells were treated with cigarette smoke extract (CSE) or dioxin, and membrane permeability and cellular senescence were measured after 48 hours. The same experiments were conducted in transwells for comparisons. RESULTS:: Compared to transwell cultures, FM-OO-C model produced better membrane permeability readings regardless of the side of treatment or time point. Membrane permeabilization was higher in AECs directly treated with CSE (1.6 fold) compared to similar treatment on the decidual side (1.2 fold). In FM-OO-C, treatments forced changes between cellular layers. This was evident when CSE and dioxin-induced senescence on one side of the chamber produced similar changes on the opposite side. This effect was minimal in the transwell system. CONCLUSION:: The controlled environment of an FM-OO-C allows for improved signal propagation between cells by minimizing noise and highlighting the small changes between treatments that cannot be seen in conventional transwell devices. Fetal membrane-organ-on-chip provides a better interaction between cell types that can be used to study fetal-maternal signaling during pregnancy in future studies.

Protein kinase D mediates inflammatory responses of human placental macrophages to Group B Streptococcus.

PROBLEM: During pregnancy, Group B Streptococcus (GBS) can infect fetal membranes to cause chorioamnionitis, resulting in adverse pregnancy outcomes. Macrophages are the primary resident phagocyte in extraplacental membranes. Protein kinase D (PKD) was recently implicated in mediating pro-inflammatory macrophage responses to GBS outside of the reproductive system. This work aimed to characterize the human placental macrophage inflammatory response to GBS and address the extent to which PKD mediates such effects. METHOD: Primary human placental macrophages were infected with GBS in the presence or absence of a specific, small molecule PKD inhibitor, CRT 0066101. Macrophage phenotypes were characterized by evaluating gene expression, cytokine release, assembly of the NLRP3 inflammasome, and NFκB activation. RESULTS: GBS evoked a strong inflammatory phenotype characterized by the release of inflammatory cytokines (TNFα, IL-1ß, IL-6 (P ≤ 0.05), NLRP3 inflammasome assembly (P ≤ 0.0005), and NFκB activation (P ≤ 0.05). Pharmacological inhibition of PKD suppressed these responses, newly implicating a role for PKD in mediating immune responses of primary human placental macrophages to GBS. CONCLUSION: PKD plays a critical role in mediating placental macrophage inflammatory activation in response to GBS infection.

Streptococcus agalactiae Induces Placental Macrophages To Release Extracellular Traps Loaded with Tissue Remodeling Enzymes via an Oxidative Burst-Dependent Mechanism.

Streptococcus agalactiae, or group B Streptococcus (GBS), is a common perinatal pathogen. GBS colonization of the vaginal mucosa during pregnancy is a risk factor for invasive infection of the fetal membranes (chorioamnionitis) and its consequences such as membrane rupture, preterm labor, stillbirth, and neonatal sepsis. Placental macrophages, or Hofbauer cells, are fetally derived macrophages present within placental and fetal membrane tissues that perform vital functions for fetal and placental development, including supporting angiogenesis, tissue remodeling, and regulation of maternal-fetal tolerance. Although placental macrophages as tissue-resident innate phagocytes are likely to engage invasive bacteria such as GBS, there is limited information regarding how these cells respond to bacterial infection. Here, we demonstrate in vitro that placental macrophages release macrophage extracellular traps (METs) in response to bacterial infection. Placental macrophage METs contain proteins, including histones, myeloperoxidase, and neutrophil elastase similar to neutrophil extracellular traps, and are capable of killing GBS cells. MET release from these cells occurs by a process that depends on the production of reactive oxygen species. Placental macrophage METs also contain matrix metalloproteases that are released in response to GBS and could contribute to fetal membrane weakening during infection. MET structures were identified within human fetal membrane tissues infected ex vivo, suggesting that placental macrophages release METs in response to bacterial infection during chorioamnionitis.IMPORTANCEStreptococcus agalactiae, also known as group B Streptococcus (GBS), is a common pathogen during pregnancy where infection can result in chorioamnionitis, preterm premature rupture of membranes (PPROM), preterm labor, stillbirth, and neonatal sepsis. Mechanisms by which GBS infection results in adverse pregnancy outcomes are still incompletely understood. This study evaluated interactions between GBS and placental macrophages. The data demonstrate that in response to infection, placental macrophages release extracellular traps capable of killing GBS. Additionally, this work establishes that proteins associated with extracellular trap fibers include several matrix metalloproteinases that have been associated with chorioamnionitis. In the context of pregnancy, placental macrophage responses to bacterial infection might have beneficial and adverse consequences, including protective effects against bacterial invasion, but they may also release important mediators of membrane breakdown that could contribute to membrane rupture or preterm labor.

Global folate status in women of reproductive age: a systematic review with emphasis on methodological issues.

Inadequate folate status in women of reproductive age (WRA) can lead to adverse health consequences of public health significance, such as megaloblastic anemia (folate deficiency) and an increased risk of neural tube defect (NTD)-affected pregnancies (folate insufficiency). Our review aims to evaluate current data on folate status of WRA. We queried eight databases and the World Health Organization Micronutrients Database, identifying 45 relevant surveys conducted between 2000 and 2014 in 39 countries. Several types of folate assays were used in the analysis of blood folate, and many surveys used folate cutoffs not matched to the assay. To allow better comparisons across surveys, we attempted to account for these differences. The prevalence of folate deficiency was >20% in many countries with lower income economies but was typically <5% in countries with higher income economies. Only 11 surveys reported the prevalence of folate insufficiency, which was >40% in most countries. Overall, folate status data for WRA globally are limited and must be carefully interpreted due to methodological issues. Future surveys would benefit from using the microbiologic assay to assess folate status, along with assay-matched cutoffs to improve monitoring and evaluation of folic acid interventions, thus informing global efforts to prevent NTDs.

Decidual stromal cell-derived PGE2 regulates macrophage responses to microbial threat.

PROBLEM: Bacterial chorioamnionitis causes adverse pregnancy outcomes, yet host-microbial interactions are not well characterized within gestational membranes. The decidua, the outermost region of the membranes, is a potential point of entry for bacteria ascending from the vagina to cause chorioamnionitis. We sought to determine whether paracrine communication between decidual stromal cells and macrophages shaped immune responses to microbial sensing. METHOD OF STUDY: Decidual cell-macrophage interactions were modeled in vitro utilizing decidualized, telomerase-immortalized human endometrial stromal cells (dTHESCs) and phorbol ester-differentiated THP-1 macrophage-like cells. The production of inflammatory mediators in response to LPS was monitored by ELISA for both cell types, while phagocytosis of bacterial pathogens (Escherichia coli and Group B Streptococcus (GBS)) was measured in THP-1 cells or primary human placental macrophages. Diclofenac, a non-selective cyclooxygenase inhibitor, and prostaglandin E2 (PGE2 ) were utilized to interrogate prostaglandins as decidual cell-derived paracrine immunomodulators. A mouse model of ascending chorioamnionitis caused by GBS was utilized to assess the colocalization of bacteria and macrophages in vivo and assess PGE2 production. RESULTS: In response to LPS, dTHESC and THP-1 coculture demonstrated enhancement of most inflammatory mediators, but a potent suppression of macrophage TNF-α generation was observed. This appeared to reflect a paracrine-mediated effect of decidual cell-derived PGE2 . In mice with GBS chorioamnionitis, macrophages accumulated at sites of bacterial invasion with increased PGE2 in amniotic fluid, suggesting such paracrine effects might hold relevance in vivo. CONCLUSION: These data suggest key roles for decidual stromal cells in modulating tissue responses to microbial threat through release of PGE2 .

The PathLink Acquired Gestational Tissue Bank: Feasibility of Project PLACENTA.

BACKGROUND: The Vanderbilt Institute for Clinical and Translational Research piloted the development of Project PLACENTA (PathLink Acquired gEstatioNal Tissue bAnk). This project investigated the feasibility of a fresh gestational tissue biobank, which provides tissue linked to electronic medical records for investigators interested in maternal-fetal health. METHODS: We developed a pipeline for collection of placental tissue from Labor and Delivery within approximately 30 minutes of delivery. An email alert was developed, to signal delivery, with the ability to specifically flag patients with certain phenotypic traits. Once collected, 4 to 8 mm punch biopsy cores were snap frozen and subsequently used for DNA, RNA and protein extraction. Tissue was also collected for Formalin Fixed Paraffin Embedded (FFPE) histology, flow cytometry, and quality control measures. RESULTS: Of 60 deliveries using the email notification system, 25 (42%) were sent to Pathology or assigned to other research protocols and were not available for collection, 10 (16%) were discarded prior to arrival at Labor and Delivery, and 25 (42%) were available for collection. Twenty placentas were collected and averaged 38 minutes per collection. DNA extraction yielded an average of 53 µg/µl per sample and RNA extraction yielded 679 ng/µl on average per sample. Proteomic studies showed no degradation of protein, abundant and similar quantities of protein across samples and differentiation between the amnion, decidua, and villi. Histological studies showed good quality for interpretation and occasional pathology including multifocal chronic villitis, meconium laden macrophages, and Stage 2 acute chorioamnionitis. Flow cytometry demonstrated good cell viability after isolation.

Macrophage Extracellular Traps: A Scoping Review.

Tissue macrophages are derived from either circulating blood monocytes that originate in the bone marrow, or embryonic precursors that establish residence in tissues and are maintained independent of bone marrow progenitors. Macrophages perform diverse functions including tissue repair, the maintenance of homeostasis, and immune regulation. Recent studies have demonstrated that macrophages produce extracellular traps (ETs). ETs are an immune response by which a cell undergoes "ETosis" to release net-like material, with strands composed of cellular DNA that is studded with histones and cellular proteins. ETs are thought to immobilize and kill microorganisms, but also been implicated in disease pathology including aseptic inflammation and autoimmune disease. We conducted a scoping review to define what is known from the existing literature about the ETs produced by monocytes or macrophages. The results suggest that macrophage ETs (METs) are produced in response to various microorganisms and have similar features to neutrophil ETs (NETs), in that METs are produced by a unique cell death program (METosis), which results in release of fibers composed of DNA and studded with cellular proteins. METs function to immobilize and kill some microorganisms, but may also play a role in disease pathology.

Assessment of iron status in settings of inflammation: challenges and potential approaches.

The determination of iron status is challenging when concomitant infection and inflammation are present because of confounding effects of the acute-phase response on the interpretation of most iron indicators. This review summarizes the effects of inflammation on indicators of iron status and assesses the impact of a regression analysis to adjust for inflammation on estimates of iron deficiency (ID) in low- and high-infection-burden settings. We overviewed cross-sectional data from 16 surveys for preschool children (PSC) (n = 29,765) and from 10 surveys for nonpregnant women of reproductive age (WRA) (n = 25,731) from the Biomarkers Reflecting the Inflammation and Nutritional Determinants of Anemia (BRINDA) project. Effects of C-reactive protein (CRP) and α1-acid glycoprotein (AGP) concentrations on estimates of ID according to serum ferritin (SF) (used generically to include plasma ferritin), soluble transferrin receptor (sTfR), and total body iron (TBI) were summarized in relation to infection burden (in the United States compared with other countries) and population group (PSC compared with WRA). Effects of the concentrations of CRP and AGP on SF, sTfR, and TBI were generally linear, especially in PSC. Overall, regression correction changed the estimated prevalence of ID in PSC by a median of +25 percentage points (pps) when SF concentrations were used, by -15 pps when sTfR concentrations were used, and by +14 pps when TBI was used; the estimated prevalence of ID in WRA changed by a median of +8 pps when SF concentrations were used, by -10 pps when sTfR concentrations were used, and by +3 pps when TBI was used. In the United States, inflammation correction was done only for CRP concentrations because AGP concentrations were not measured; regression correction for CRP concentrations increased the estimated prevalence of ID when SF concentrations were used by 3 pps in PSC and by 7 pps in WRA. The correction of iron-status indicators for inflammation with the use of regression correction appears to substantially change estimates of ID prevalence in low- and high-infection-burden countries. More research is needed to determine the validity of inflammation-corrected estimates, their dependence on the etiology of inflammation, and their applicability to individual iron-status assessment in clinical settings.

Differing mechanisms of surviving phagosomal stress among group B Streptococcus strains of varying genotypes.

Group B Streptococcus (GBS), a leading cause of neonatal sepsis and meningitis, asymptomatically colonizes up to 30% of women and can persistently colonize even after antibiotic treatment. Previous studies have shown that GBS resides inside macrophages, but the mechanism by which it survives remains unknown. Here, we examined the ability of 4 GBS strains to survive inside macrophages and then focused on 2 strains belonging to sequence type (ST)-17 and ST-12, to examine persistence in the presence of antibiotics. A multiple stress medium was also developed using several stressors found in the phagosome to assess the ability of 30 GBS strains to withstand phagosomal stress. The ST-17 strain was more readily phagocytosed and survived intracellularly longer than the ST-12 strain, but the ST-12 strain was tolerant to ampicillin unlike the ST-17 strain. Exposure to sub-inhibitory concentrations of ampicillin and erythromycin increased the level of phagocytosis of the ST-17 strain, but had no effect on the ST-12 strain. In addition, blocking acidification of the phagosome decreased the survival of the ST-17 strain indicating a pH-dependent survival mechanism for the ST-17 strain. Congruent with the macrophage experiments, the ST-17 strain had a higher survival rate in the multiple stress medium than the ST-12 strain, and overall, serotype III isolates survived significantly better than other serotypes. These results indicate that diverse GBS strains may use differing mechanisms to persist and that serotype III strains are better able to survive specific stressors inside the phagosome relative to other serotypes.

Lipid profiling of polarized human monocyte-derived macrophages.

The highly orchestrated transcriptional and metabolic reprogramming during activation drastically transforms the main functions and physiology of human macrophages across the polarization spectrum. Lipids, for example, can modify protein function by acting remotely as signaling molecules but also locally by altering the physical properties of cellular membranes. These changes play key roles in the functions of highly plastic immune cells due to their involvement in inflammation, immune responses, phagocytosis and wound healing processes. We report an analysis of major membrane lipids of distinct phenotypes of resting (M0), classically activated (M1), alternatively activated (M2a) and deactivated (M2c) human monocyte derived macrophages from different donors. Samples were subjected to supercritical fluid chromatography-ion mobility-mass spectrometry analysis, which allowed separations based on lipid class, facilitating the profiling of their fatty acid composition. Different levels of arachidonic acid mobilization as well as other fatty acid changes were observed for different lipid classes in the distinct polarization phenotypes, suggesting the activation of highly orchestrated and specific enzymatic processes in the biosynthesis of lipid signaling molecules and cell membrane remodeling. Thromboxane A2 production appeared to be a specific marker of M1 polarization. These alterations to the global composition of lipid bi-layer membranes in the cell provide a potential methodology for the definition and determination of cellular and tissue activation states.