Maternal Monocytes Research Articles

Gravidity influences distinct transcriptional profiles of maternal and fetal placental macrophages at term.

Maternal intervillous monocytes (MIMs) and fetal Hofbauer cells (HBCs) are myeloid-derived immune cells at the maternal-fetal interface. Maternal reproductive history is associated with differential risk of pregnancy complications. The molecular phenotypes and roles of these distinct monocyte/macrophage populations and the influence of gravidity on these phenotypes has not been systematically investigated. Here, we used RNA sequencing to study the transcriptional profiles of MIMs and HBCs in normal term pregnancies. Our analyses revealed distinct transcriptomes of MIMs and HBCs. Genes involved in differentiation and cell organization pathways were more highly expressed in MIMs vs. HBCs. In contrast, HBCs had higher expression of genes involved in inflammatory responses and cell surface receptor signaling. Maternal gravidity influenced monocyte programming, as expression of pro-inflammatory molecules was significantly higher in MIMs from multigravidae compared to primigravidae. In HBCs, multigravidae displayed enrichment of gene pathways involved in cell-cell signaling and differentiation. Our results demonstrated that MIMs and HBCs have highly divergent transcriptional signatures, reflecting their distinct origins, locations, functions, and roles in inflammatory responses. Furthermore, maternal gravidity influences the gene signatures of MIMs and HBCs, potentially modulating the interplay between tolerance and trained immunity. The phenomenon of reproductive immune memory may play a novel role in the differential susceptibility of primigravidae to pregnancy complications.

Association of maternal leukocyte, monocyte, and neutrophil counts with hypertensive disorders of pregnancy: the Japan Environment and Children’s Study (JECS)

Hypertensive disorders of pregnancy (HDP) increase the risk of preterm births and cesarean delivery. This study aimed to investigate whether maternal blood leukocyte, monocyte, or neutrophil counts in the first trimester are related to the development of HDP. Data were collected from the Japan Environment and Children’s Study, a large birth cohort study (n = 38,194) that recruited pregnant women in 15 Regional Centers across Japan (from January 2011 to March 2014). The odds ratios (ORs) for mild/severe HDP according to the cut-off value of leukocyte/neutrophil/monocyte counts by the receiver operating characteristic curve showed high ORs. Furthermore, pregnant women with the highest quartiles of leukocyte and monocyte counts had higher adjusted ORs (aORs) for mild (leukocyte: aOR = 1.27, 95% confidence interval [CI]: 1.02–1.58; monocyte: aOR = 1.30, 95% CI 1.04–1.63) and severe HDP (leukocyte: aOR = 1.51, 95% CI 1.08–2.13; monocyte: aOR = 1.44, 95% CI 1.03–2.01) compared with those with the lowest quartiles of those counts. In addition, pregnant women with the highest neutrophil counts had higher aOR for mild HDP (aOR = 1.26, 95% CI 1.02–1.56) compared with those with the lowest count. In conclusion, high leukocyte and monocyte counts in the first trimester are associated with the development of HDP. Thus, they may be used to predict subsequent HDP.

Elevated expression of Toll-like receptor 4 and cytokines in both serum and myometrium at term may serve as promising biomarkers for uterine activation preceding labor.

Increased inflammation and cytokine levels are considered risk factors and promoters of preterm birth (PTB). However, the regulatory mechanism of pregnancy-related inflammation remains unclear. Toll-like receptor 4 (TLR4) plays a critical role in inflammatory responses in various diseases. Therefore, our study aimed to investigate whether TLR4 is involved in the inflammatory responses during uterine activation for labor, with the goal of identifying potential biomarkers for uterine activation at term. We used flow cytometry to detect TLR4 expression on CD14+ maternal blood monocytes in the first, second, and third trimesters. ELISA was employed to measure TLR4 and cytokines levels in the maternal serum of term non-labor (TNL), term labor (TL) women and LPS induced preterm labor and PBS injected controls. TLR4siRNA was transfected into the human myometrial smooth muscle cells (HMSMCs), which were subsequently treated with IL-1β. The mRNA and protein levels of TLR4, uterine contraction-related protein connexin 43 (CX43), oxytocin receptor (OTR), MAPK/NF-κB signaling pathway, and cytokines were analyzed using qRT-PCR, western blotting, and immunohistochemistry. The study revealed TLR4 expression on CD14+ maternal blood monocytes was higher in the third trimester group compared to the first and second trimester groups (p<0.001). Maternal serum concentrations of TLR4 and cytokines were significantly higher in the TL group than the TNL group (p<0.001). TLR4, OTR, CX43, activated MAPK/NF-κB expression, and cytokines levels were upregulated in TL group, and similarly significantly higher in the LPS-induced preterm group than in the control group. Using the HMSMCs we demonstrated that TLR4siRNA transfection suppressed contractility. Interfering with TLR4 expression reduced the expression of OTR, CX43, cytokines, and MAPK/NF-κB activation. There was a significant positive relationship between TLR4 expression and the inflammatory status in the myometrium. ROC analysis indicated that TLR4 and cytokines may serve as potential biomarkers for predicting uterine activation for labor. Our data suggest that TLR4 and cytokines can act as stimulators of uterine activation for labor at term. Furthermore, the MAPK/NF-κB pathway appears to be one of the potential signaling pathways mediating TLR4's regulation of parturition initiation.

Impact of Maternal Monocyte to High-density Lipoprotein Cholesterol Ratio on the Incidence of Large-for-gestational-age Newborns: An Observational Cohort Study

Monocyte to high-density lipoprotein cholesterol ratio (MHR) has recently been identified as a new marker of inflammation and oxidative stress. However, it is unknown whether maternal MHR is associated with fetal weight at birth. Therefore, our objective was to analyze the association between maternal MHR and the frequency of small/large for gestational age (SGA/LGA) newborns in this retrospective cohort study. We retrospectively analyzed hospitalization records and laboratory data and obtained results from consecutive pregnant women in whom the blood lipid level had been investigated along with the blood cell count. Linear regression and logistic regression analyses were performed to estimate the associations of maternal MHR with birth weight and SGA/LGA. Monocyte counts and MHR were positively associated with birth weight/LGA risk (monocyte [1-109/L increase] for birth weight: β: 170.24, 95% confidence interval [CI]: 41.72-298.76, LGA: odds ratio [OR]: 7.67; 95% CI: 2.56-22.98; MHR [1-109/mmol increase] for birth weight: β: 294.84, 95% CI: 170.23-419.44, LGA: OR: 7.97; 95% CI: 3.06-20.70), whereas high-density lipoprotein cholesterol (HDL-C) levels were negatively associated with birth weight/LGA risk [1 mmol/L increase for birth weight (β: -99.83, 95% CI: -130.47 to -69.19), for LGA: (OR: 0.57, 95% CI: 0.45-0.73). Obese pregnant women (body mass index [BMI] ≥30 kg/m2) with higher MHR (tertile 3: >0.33 109/mmol) significantly increased LGA risk by 6.39 fold (95% CI: 4.81, 8.49) compared to those with low MHR (tertile 1-2: ≤0.33 109/mmol) and normal weight (BMI <25 kg/m2). Maternal MHR is associated with LGA risk, and this association might be further modified by BMI.

Correction: Maternal Monocytes Respond to Cell-Free Fetal DNA and Initiate Key Processes of Human Parturition.

Kazemi, N.Y., B. Fedyshyn, S. Sutor, Y. Fedyshyn, S. Markovic, and E.A.L. Enninga. 2022. Maternal monocytes respond to cell-free fetal DNA and initiate key processes of human parturition. J. Immunol. 207: [2433–2444][1]. In the original article, the keys for the histograms in [Fig. 3B][2] and [3E

Discrete role for maternal stress and gut microbes in shaping maternal and offspring immunity

Psychosocial stress is prevalent during pregnancy, and is associated with immune dysfunction, both for the mother and the child. The gut microbiome has been implicated as a potential mechanism by which stress during pregnancy can impact both maternal and offspring immune function; however, the complex interplay between the gut microbiome and the immune system is not well-understood. Here, we leverage a model of antimicrobial-mediated gut microbiome reduction, in combination with a well-established model of maternal restraint stress, to investigate the independent effects of and interaction between maternal stress and the gut microbiome in shaping maternal and offspring immunity. First, we confirmed that the antimicrobial treatment reduced maternal gut bacterial load and altered fecal alpha and beta diversity, with a reduction in commensal microbes and an increase in the relative abundance of rare taxa. Prenatal stress also disrupted the gut microbiome, according to measures of both alpha and beta diversity. Furthermore, prenatal stress and antimicrobials independently induced systemic and gastrointestinal immune suppression in the dam with a concomitant increase in circulating corticosterone. While stress increased neutrophils in the maternal circulation, lymphoid cells and monocytes were not impacted by either stress or antimicrobial treatment. Although the fetal immune compartment was largely spared, stress increased circulating neutrophils and CD8 T cells, and antibiotics increased neutrophils and reduced T cells in the adult offspring. Altogether, these data indicate similar, but discrete, roles for maternal stress and gut microbes in influencing maternal and offspring immune function.

The landscape of circular RNA in preterm birth

BackgroundPreterm birth (PTB) is a multifactorial syndrome that seriously threatens the health of pregnant women and babies worldwide. Recently, circular RNAs (circRNAs) have been understood as important regulators of various physiological and pathological processes. However, the expression pattern and potential roles of circRNAs in PTB are largely unclear.MethodsIn this study, we extracted and analyzed the circRNA expression profiles in maternal and fetal samples of preterm and term pregnancies, including maternal plasma, maternal monocytes, myometrium, chorion, placenta, and cord blood. We identified the circRNAs which is associated with PTB in different tissues and explored their relationships from the perspective of the overall maternal-fetal system. Furthermore, co-expression analysis of circRNAs and mRNAs, target microRNAs (miRNAs), and RNA-binding proteins (RBPs), provided new clues about possible mechanisms of circRNA function in PTB. In the end, we investigated the potential special biofunctions of circRNAs in different tissues and their common features and communication in PTB.ResultsSignificant differences in circRNA types and expression levels between preterm and term groups have been proved, as well as between tissues. Nevertheless, there were still some PTB-related differentially expressed circRNAs (DECs) shared by these tissues. The functional enrichment analysis showed that the DECs putatively have important tissue-specific biofunctions through their target miRNA and co-expressed mRNAs, which contribute to the signature pathologic changes of each tissue within the maternal-fetal system in PTB (e.g., the contraction of the myometrium). Moreover, DECs in different tissues might have some common biological activities, which are mainly the activation of immune-inflammatory processes (e.g., interleukin1/6/8/17, chemokine, TLRs, and complement).ConclusionsIn summary, our data provide a preliminary blueprint for the expression and possible roles of circRNAs in PTB, which lays the foundation for future research on the mechanisms of circRNAs in PTB.

Extracellular vesicles of first trimester trophoblast cell line overexpressing VPAC2 induce anti-inflammatory signals in HB cell and maternal monocytes

Extracellular vesicles of first trimester trophoblast cell line overexpressing VPAC2 induce anti-inflammatory signals in HB cell and maternal monocytes

Conditioning of maternal monocytes to myeloid regulatory cells by endometrial stromal cells

Conditioning of maternal monocytes to myeloid regulatory cells by endometrial stromal cells

Immune Cell and Adipose Gene Expression Reprogramming in Juvenile Monkeys Born to Health and Weight-Diverse Mothers

Over 93 million Americans are obese and 66 million suffer from metabolic disease. Roughly 40% of obese people do not have metabolic abnormalities (metabolically health obese [MHO]), while approximately 15% of lean do (metabolically unhealthy lean [MUL]). African green monkeys (AGMs) demonstrate naturally occurring obesity and metabolic syndrome (MetS) without diet manipulation, and MetS criteria are heritable. Age-matched maternal AGMs were classified by adjusted MetS criteria ([n=44]; waist >40cm, fasting glucose (FG) >100 mg/dL, SBP/DBP >135/85mmHg, and HDL-c <50mg/dL) and classified as metabolically healthy lean (MHL), MHO, MUL or metabolically unhealthy obese (MUO). Age, weight and sex-matched pre-pubertal juvenile offspring from these mothers were additionally selected (n=9-11/group; ages=1.1-3.4years) for evaluation. We assessed monocyte subtypes by flow cytometry, and subcutaneous adipose gene expression patterns by RNAseq. Non-classical monocytes were increased in obese and unhealthy mothers (MHO p=0.02, MUL p=0.003, MUO p=0.00002) compared to MHL. MUL and MUO juvenile offspring also had more non-classical monocytes compared to MHL (p=0.05 and p=0.07). Monocyte chemoattractant protein-1 (MCP)-1 was measured in plasma and found to be elevated in MUO juveniles (p=0.02). Patterns of increased cytokine and extracellular matrix gene expression were seen in MUL and MUO juveniles’ adipose (6-7/group), mirroring obese and unhealthy mothers’ adipose gene expression patterns. Maternal health and obesity influence offspring immune cells and adipose gene expression prior to weight gain and metabolic disease onset. Our data underscore maternal monocyte and adipose profiles as inherited phenotypes that present prior to adipose expansion and may be targets to improve intergenerational health trajectories.

Influence of Covid-19 infection on fetal thymus size after recovery

To investigate the long-term effects of the SARS-CoV-2 infection on the fetal immune system by fetal thymus size measurements with ultrasound (USG). This prospective study was conducted in the Turkish Ministry of Health Ankara City Hospital between November 1, 2020 and April 1, 2021, with recovered, pregnant women, four weeks after they had been confirmed for the SARS-CoV-2 infection by real-time polymerase-chain-reaction (RT-PCR). COVID-19 recovered (CR) pregnant women compared with age-matched pregnant controls in terms of demographic features, fetal thymic-thoracic ratio (TTR), and laboratory parameters. There was no difference in demographic features between the two groups. TTR found significantly lower in the CR group than the control group (p=0.001). The fetal TTR showed a significant and moderate correlation with maternal monocyte counts, monocyte to lymphocyte ratio (MLR), and red cell distribution width (RDW); while it did not correlate with lymphocyte counts, c-reactive protein (CRP), and procalcitonin levels. The 2019 novel coronavirus disease (COVID-19) reduces fetal thymus size in pregnant women with mild or moderate symptoms after recovery from the infection.

Maternal Monocytes Respond to Cell-Free Fetal DNA and Initiate Key Processes of Human Parturition.

Throughout gestation, the maternal immune system is tightly modulated to allow growth of a semiallogeneic fetus. During the third trimester, the maternal immune system shifts to a proinflammatory phenotype in preparation for labor. What induces this shift remains unclear. Cell-free fetal DNA (cffDNA) is shed by the placenta and enters maternal circulation throughout pregnancy. Levels of cffDNA are increased as gestation progresses and peak before labor, coinciding with a shift to proinflammatory maternal immunity. Furthermore, cffDNA is abnormally elevated in plasma from women with complications of pregnancy, including preterm labor. Given the changes in maternal immunity at the end of pregnancy and the role of sterile inflammation in the pathophysiology of spontaneous preterm birth, we hypothesized that cffDNA can act as a damage-associated molecular pattern inducing an inflammatory cytokine response that promotes hallmarks of parturition. To test this hypothesis, we stimulated human maternal leukocytes with cffDNA from primary term cytotrophoblasts or maternal plasma and observed significant IL-1β and CXCL10 secretion, which coincides with phosphorylation of IFN regulatory factor 3 and caspase-1 cleavage. We then show that human maternal monocytes are crucial for the immune response to cffDNA and can activate bystander T cells to secrete proinflammatory IFN-γ and granzyme B. Lastly, we find that the monocyte response to cffDNA leads to vascular endothelium activation, induction of myometrial contractility, and PGE2 release in vitro. Our results suggest that the immune response to cffDNA can promote key features of the parturition cascade, which has physiologic consequences relevant to the timing of labor.

Longitudinal evaluation of azithromycin and cytokine concentrations in amniotic fluid following one-time oral dosing in pregnancy.

To utilize noninvasive collection of amniotic fluid in the setting of preterm premature rupture of membranes (PPROMs) to report the time concentration profile of azithromycin in amniotic fluid over 7 days from a single dose, and evaluate the correlation between azithromycin concentration and inflammatory markers in amniotic fluid. Prospective cohort study of five pregnant patients admitted with PPROMs and treated with a single 1 g oral azithromycin dose. Amniotic fluid was collected from pads and used to quantify azithromycin concentration as well as TNFa, IL‐1a, IL‐1b, IL‐6, IL‐8, and IL‐10 concentrations. Primary outcome was time/concentration profile of azithromycin in amniotic fluid. Secondary outcome included correlation between azithromycin concentration and cytokine concentrations. Five patients were enrolled. Mean gestational age on admission with PPROM was 27.5 ± 2.3 weeks with a median latency of 7 days (interquartile range [IQR] = 4–13). A median of two samples/day (IQR = 1–3) were collected per participant. Azithromycin was quantified in duplicate; intra‐assay coefficient of variation was 17%. Azithromycin concentration was less than 60 ng/ml after day 3. Azithromycin concentration was positively correlated with IL‐8 (r = 0.38, p = 0.03), IL1a (r = 0.39, p = 0.03), and IL‐1b (r = 0.36, p = 0.04) in amniotic fluid. Azithromycin is detectable in amniotic fluid over 7 days from a single 1 g maternal dose, however, it is not sustained over the range of minimum inhibitory concentration for common genitourinary flora. Based on correlation with specific cytokines, azithromycin penetration in amniotic fluid may relate to maternal monocyte concentration in amniotic fluid in the setting of PPROM.

OR08: Decidualization program conditions maternal monocytes to a DC‐10 profile inducing regulatory T‐cell response

OR08: Decidualization program conditions maternal monocytes to a DC‐10 profile inducing regulatory T‐cell response

P92: Innate responses to cell‐free fetal DNA by maternal monocytes promote bystander cell activation and inflammation

P92: Innate responses to cell‐free fetal DNA by maternal monocytes promote bystander cell activation and inflammation

720 Comparing transcriptional signatures of fetal hofbauer cells derived from bulk versus single-Cell RNA sequencing approaches

Fetal Hofbauer Cells (HBC) play an instrumental role in the maintenance of a healthy pregnancy and in response to infection. Single-cell RNA sequencing (scRNAseq) has recently identified potential markers of HBC (Vento-Tormo, Nature, 2018). However, it is unknown whether these transcriptional signatures are congruent with profiles obtained from bulk RNA sequencing (bRNAseq) of purified cell populations. Here we compared the results from these two different approaches to transcriptomic studies. This is a comparative study of results from a publicly available scRNAseq database that analyzed first trimester placentas and bRNAseq data generated by our group analyzing term placentas from normal pregnancies. In the bRNAseq analysis, maternal intervillous monocytes (MIM) and HBC were purified using targeted isolation techniques. Differentially expressed genes in paired samples were identified using LIMMA and compared to a list of 30 proposed HBC markers. Transcript levels were normalized against the mean expression in MIM for each gene to determine relative gene expression; normalized expression values were compared with the 30 markers. HBC:MIM expression ratios were also calculated in paired samples. 19 of 30 (63%) HBC markers defined by scRNAseq were also identified by bRNAseq (adjusted p-value<0.01). Hierarchical clustering demonstrated that HBC and MIM samples clustered separately and that all markers were more highly expressed in HBC, with the exception of DAB2 (Fig 1). Analysis of HBC:MIM expression ratios in paired samples, also demonstrated low relative expression of DAB2 in HBC. ATP1B1, LGMN, NINJ1, and EGFL7 showed higher expression in HBC which was consistent between patient samples (Fig 2). Results from scRNAseq and bRNAseq approaches showed >60% concordance in the identification of HBC markers, despite differences in gestational age and approach. Future studies are needed to validate the use of these potential biomarkers of HBCs, which will serve as important tools in understanding the role of these cells in placental disease.View Large Image Figure ViewerDownload Hi-res image Download (PPT)

Isolation of First-Trimester and Full-term Human Placental Hofbauer Cells.

The placenta is the crucial organ that regulates the health of both mother and fetus during pregnancy. The human placenta is composed of villous tree-like structures that embed into the maternal decidua. Within the stroma of the villi resides a population of fetally-derived macrophages, the Hofbauer cells (HBC). HBC are the only fetal immune cells found within the placenta in the steady-state and are thought to play a crucial role in placental function. From the 10th week of gestation, maternal blood flow into the intervillous space begins, resulting in the placental villi becoming bathed in maternal blood. To study HBC it is necessary to develop techniques that allow for their specific isolation and distinction from maternal blood monocytes and decidual macrophages. Here, we describe a protocol that explains step-by-step the strategy we have developed that allows the specific isolation of HBC.

Maternal serum leptin, adiponectin, resistin and monocyte chemoattractant protein-1 levels in different types of diabetes mellitus

ObjectiveEvaluation of serum concentration of leptin, adiponectin, resistin, and MCP-1 in pregnant patients with different types of diabetes mellitus (DM) considering preconception planning and method of DM correction in 11–14th and 30–34th weeks of pregnancy. Study designLongitudinal, prospective study included 130 pregnant women divided into the following comparison groups: type 1 DM (T1DM, n = 40), type 2 DM (T2DM, n = 35), GDM (n = 40), and the control group (n = 15). The ELISA method defined the levels of leptin, resistin, adiponectin, and MCP-1 concentration in serum, which was assessed in 11–14th and 30–34th weeks of pregnancy. Statistical analysis was accomplished using SPSS 23.0 and "Prism 8-GraphPad" software. ResultsThe leptin level in the 1st trimester was the highest in T2DM insulin group compared to the control due to gestational age, hence in the 3rd trimester in all groups its serum concentrations appeared higher than in healthy patients (p = 0.0001). In the 1st trimester leptin levels directly correlated with women's BMI, newborns’ weight and macrosomia rate, in the 3rd trimester – with OGTT levels, HbA1c, gestational hypertension, and preeclampsia rates. Resistin levels in the 1st and 3rd trimesters were increased in almost all DM groups compared to the control group (p = 0.0001). The study established direct positive correlation between resistin and HbA1c, birth weight, and preeclampsia. In the 1st trimester, adiponectin demonstrated the lowest levels in T2DM insulin compared to T1DM and the control group (p = 0.0001) while in the 3rd trimester, adiponectin levels declined alongside gestational age in DM patients and all the groups compared to the control group (p < 0.05). Adiponectin negatively correlated with BMI, OGTT levels, and preeclampsia rate. MCP-1 levels in T2DM appeared higher than in T1DM patients and the control group in the 1st trimester, whereas in the 3rd trimester MCP-1 declined, correlating with BMI, preeclampsia and OGTT levels. ConclusionHigh rate of adverse perinatal outcomes in diabetic pregnancy might be developed due to more severe metabolic failures and further disturbances of adipokines expression.

Circulating Monocytes, Tissue Macrophages, and Malaria.

Malaria is a significant cause of global morbidity and mortality. The Plasmodium parasite has a complex life cycle with mosquito, liver, and blood stages. The blood stages can preferentially affect organs such as the brain and placenta. In each of these stages and organs, the parasite will encounter monocytes and tissue-specific macrophages—key cell types in the innate immune response. Interactions between the Plasmodium parasite and monocytes/macrophages lead to several changes at both cellular and molecular levels, such as cytokine release and receptor expression. In this review, we summarize current knowledge on the relationship between malaria and blood intervillous monocytes and tissue-specific macrophages of the liver (Kupffer cells), central nervous system (microglia), and placenta (maternal intervillous monocytes and fetal Hofbauer cells). We describe their potential roles in modulating outcomes from infection and areas for future investigation.

Effect of pregnancy hormone mixtures on cytokine production and surface marker expression in naïve and LPS-activated THP-1 differentiated monocytes/macrophages

In pregnancy, maternal monocytes and macrophages acquire a specific phenotype that enables them to maintain immune tolerance and facilitate hormone–immune cell interactions, which are necessary for gestational progression. The aim of this study was to determine the effect of pregnancy hormone mixtures of the first and third trimesters on both resting and activated monocytes and macrophages. Pregnancy hormone levels (cortisol, estradiol, progesterone, and prolactin) were quantified at the first and third trimesters. The average of the levels obtained was used to prepare two mixtures of synthetic hormones: low and high. These mixtures were then used to stimulate THP-1 monocytes and macrophages, resting or activated with LPS. Cytokine production in the culture supernatants and surface marker expression (CD14, CD86, and CD163) were evaluated by ELISA and flow cytometry, respectively. We found that the hormones modulated the pro-inflammatory response of THP-1 cells, LPS-activated monocytes, and macrophages, inducing high levels of IL-10 and low levels of IL-8, IL-1-β, and IL-6. All hormone stimulation increased the CD163 receptor in both resting and LPS-activated monocytes and macrophages in a dose-independent manner, unlike CD14 and CD86. Pregnancy hormones promote the expression of the markers associated with the M2-like phenotype, modulating their pro-inflammatory response. This phenotype regulation by hormones could be a determinant in pregnancy.