Blocking OLFM4/galectin-3 axis in placental polymorphonuclear myeloid-derived suppressor cells triggers intestinal inflammation in newborns.

Fetal growth restriction (FGR) is a major cause of premature and low-weight births, which increases the risk of necrotizing enterocolitis (NEC); however, the association remains unclear. We report a close correlation between placental polymorphonuclear myeloid-derived suppressor cells (PMN-MDSCs) and NEC. Newborns with previous FGR exhibited intestinal inflammation and more severe NEC symptoms than healthy newborns. Placental PMN-MDSCs are vital regulators of fetal development and neonatal gut inflammation. Placental single-cell transcriptomics revealed that PMN-MDSCs populations and olfactomedin-4 gene (Olfm4) expression levels were significantly increased in PMN-MDSCs in later pregnancy compared to those in early pregnancy and non-pregnant females. Female mice lacking Olfm4 in myeloid cells mated with wild-type males showed FGR during pregnancy, with a decreased placental PMN-MDSCs population and expression of growth-promoting factors (GPFs) from placental PMN-MDSCs. Galectin-3 (Gal-3) stimulated the OLFM4-mediated secretion of GPFs by placental PMN-MDSCs. Moreover, GPF regulation via OLFM4 in placental PMN-MDSCs was mediated via hypoxia inducible factor-1α (HIF-1α). Notably, the offspring of mothers lacking Olfm4 exhibited intestinal inflammation and were susceptible to NEC. Additionally, OLFM4 expression decreased in placental PMN-MDSCs from pregnancies with FGR and was negatively correlated with neonatal morbidity. These results revealed that placental PMN-MDSCs contributed to fetal development and ameliorate newborn intestinal inflammation.

Evaluation of systemic immune-inflammation index for predicting late-onset fetal growth restriction.

INTRODUCTION: To determine a cut-off value for systemic immune-inflammation index (SII) (neutrophil × platelet/lymphocyte) in the prediction of fetal growth restriction (FGR). MATERIALS AND METHODS: This case-control study was conducted retrospectively at the Obstetrics-Gynecology and Perinatology Clinics of Etlik Zubeyde Hanim Women's Health Education and Training Hospital. Singleton pregnant women with late-onset FGR who were followed up in outpatient clinics or hospitalized and whose pregnancy resulted at our hospital were included in the study group (group I). Healthy early and full-term singleton pregnant women with spontaneous labor who were followed up in the same hospital and whose pregnancy resulted at the same hospital were included in the control group (group II). Receiver-operating characteristic curves were used to assess the performance of SII value in predicting FGR. RESULTS: We recruited 79 cases (pregnant with late-onset fetal growth restriction) and 79 controls (healthy pregnant), matched for age, body mass index, and parity. ΔSII was statistically significantly higher in the pregnant with late-onset FGR compared with healthy pregnant (123 vs - 65; p = 0.039). The values in ROC curves with the best balance of sensitivity/specificity were > 152 109/L (49% sensitivity, 70% specificity) and > 586 109/L (27% sensitivity, 90% specificity) for late-onset FGR. DISCUSSION: Higher ΔSII levels in maternal blood indicate an inflammatory process causing FGR. The cut-off value for ΔSII (> 586 109/L) at 90% specificity can be used as a screening test. In the presence of ΔSII levels > 586 109/L (27% sensitivity and 90% specificity), the physicians should be more cautious about risk for FGR. Therefore, pregnant women at risk for FGR should be checked more frequently and monitored closely. However, further studies are needed to confirm our findings.

Cytoplasmic RNA quality control failure engages mTORC1-mediated autoinflammatory disease.

Inborn errors of nucleic acid metabolism often cause aberrant activation of nucleic acid sensing pathways, leading to autoimmune or autoinflammatory diseases. The SKIV2L RNA exosome is cytoplasmic RNA degradation machinery that was thought to be essential for preventing the self-RNA-mediated interferon (IFN) response. Here, we demonstrate the physiological function of SKIV2L in mammals. We found that Skiv2l deficiency in mice disrupted epidermal and T cell homeostasis in a cell-intrinsic manner independently of IFN. Skiv2l-deficient mice developed skin inflammation and hair abnormality, which were also observed in a SKIV2L-deficient patient. Epidermis-specific deletion of Skiv2l caused hyperproliferation of keratinocytes and disrupted epidermal stratification, leading to impaired skin barrier with no appreciable IFN activation. Moreover, Skiv2l-deficient T cells were chronically hyperactivated and these T cells attacked lesional skin as well as hair follicles. Mechanistically, SKIV2L loss activated the mTORC1 pathway in both keratinocytes and T cells. Both systemic and topical rapamycin treatment of Skiv2l-deficient mice ameliorated epidermal hyperplasia and skin inflammation. Together, we demonstrate that mTORC1, a classical nutrient sensor, also senses cytoplasmic RNA quality control failure and drives autoinflammatory disease. We also propose SKIV2L-associated trichohepatoenteric syndrome (THES) as a new mTORopathy for which sirolimus may be a promising therapy.

Prenatal administration of IL-1Ra attenuate the neurodevelopmental impacts following non-pathogenic inflammation during pregnancy.

Prenatal inflammation negatively affects placental function, subsequently altering fetal development. Pathogen-associated molecular patterns (PAMPs) are used to mimics infections in preclinical models but rarely detected during pregnancy. Our group previously developed an animal model of prenatal exposure to uric acid (endogenous mediator), leading to growth restriction alongside IL-1-driven placental inflammation (Brien et al. in J Immunol 198(1):443-451, 2017). Unlike PAMPs, the postnatal impact of prenatal non-pathogenic inflammation is still poorly understood. Therefore, we investigated the effects of prenatal uric acid exposure on postnatal neurodevelopment and the therapeutic potential of the IL-1 receptor antagonist; IL-1Ra. Uric acid induced growth restriction and placental inflammation, which IL-1Ra protected against. Postnatal evaluation of both structural and functional aspects of the brain revealed developmental changes. Both astrogliosis and microgliosis were observed in the hippocampus and white matter at postnatal day (PND)7 with IL-1Ra being protective. Decreased myelin density was observed at PND21, and reduced amount of neuronal precursor cells was observed in the Dentate Gyrus at PND35. Functionally, motor impairments were observed as evaluated with the increased time to fully turn upward (180 degrees) on the inclined plane and the pups were weaker on the grip strength test. Prenatal exposure to sterile inflammation, mimicking most clinical situation, induced growth restriction with negative impact on neurodevelopment. Targeted anti-inflammatory intervention prenatally could offer a strategy to protect brain development during pregnancy.

NF-κB regulation in maternal immunity during normal and IUGR pregnancies.

Intrauterine Growth Restriction (IUGR) is a leading cause of perinatal death with no effective cure, affecting 5-10% pregnancies globally. Suppressed pro-inflammatory Th1/Th17 immunity is necessary for pregnancy success. However, in IUGR, the inflammatory response is enhanced and there is a limited understanding of the mechanisms that lead to this abnormality. Regulation of maternal T-cells during pregnancy is driven by Nuclear Factor Kappa B p65 (NF-κB p65), and we have previously shown that p65 degradation in maternal T-cells is induced by Fas activation. Placental exosomes expressing Fas ligand (FasL) have an immunomodulatory function during pregnancy. The aim of this study is to investigate the mechanism and source of NF-κB regulation required for successful pregnancy, and whether this is abrogated in IUGR. Using flow cytometry, we demonstrate that p65+ Th1/Th17 cells are reduced during normal pregnancy, but not during IUGR, and this phenotype is enforced when non-pregnant T-cells are cultured with normal maternal plasma. We also show that isolated exosomes from IUGR plasma have decreased FasL expression and are reduced in number compared to exosomes from normal pregnancies. In this study, we highlight a potential role for FasL+ exosomes to regulate NF-κB p65 in T-cells during pregnancy, and provide the first evidence that decreased exosome production may contribute to the dysregulation of p65 and inflammation underlying IUGR pathogenesis.

The Acute Hepatic NF-κB-Mediated Proinflammatory Response to Endotoxemia Is Attenuated in Intrauterine Growth-Restricted Newborn Mice.

Intrauterine growth restriction (IUGR) is a relevant predictor for higher rates of neonatal sepsis worldwide and is associated with an impaired neonatal immunity and lower immune cell counts. During the perinatal period, the liver is a key immunological organ responsible for the nuclear factor kappa B (NF-κB)-mediated innate immune response to inflammatory stimuli, but whether this role is affected by IUGR is unknown. Herein, we hypothesized that the newborn liver adapts to calorie-restriction IUGR by inducing changes in the NF-κB signaling transcriptome, leading to an attenuated acute proinflammatory response to intraperitoneal lipopolysaccharide (LPS). We first assessed the hepatic gene expression of key NF-κB factors in the IUGR and normally grown (NG) newborn mice. Real-time quantitative PCR (RT-qPCR) analysis revealed an upregulation of both IκB proteins genes (Nfkbia and Nfkbib) and the NF-κB subunit Nfkb1 in IUGR vs. NG. We next measured the LPS-induced hepatic expression of acute proinflammatory genes (Ccl3, Cxcl1, Il1b, Il6, and Tnf) and observed that the IUGR liver produced an attenuated acute proinflammatory cytokine gene response (Il1b and Tnf) to LPS in IUGR vs. unexposed (CTR). Consistent with these results, LPS-exposed hepatic tumor necrosis factor alpha (TNF-α) protein concentrations were lower in IUGR vs. LPS-exposed NG and did not differ from IUGR CTR. Sex differences at the transcriptome level were observed in the IUGR male vs. female. Our results demonstrate that IUGR induces key modifications in the NF-κB transcriptomic machinery in the newborn that compromised the acute proinflammatory cytokine gene and protein response to LPS. Our results bring novel insights in understanding how the IUGR newborn is immunocompromised due to fundamental changes in NF-κB key factors.

Toll-like receptor-4 null mutation causes fetal loss and fetal growth restriction associated with impaired maternal immune tolerance in mice.

Maternal immune adaptation to accommodate pregnancy depends on sufficient availability of regulatory T (Treg) cells to enable embryo implantation. Toll-like receptor 4 is implicated as a key upstream driver of a controlled inflammatory response, elicited by signals in male partner seminal fluid, to initiate expansion of the maternal Treg cell pool after mating. Here, we report that mice with null mutation in Tlr4 (Tlr4-/-) exhibit impaired reproductive outcomes after allogeneic mating, with reduced pregnancy rate, elevated mid-gestation fetal loss, and fetal growth restriction, compared to Tlr4+/+ wild-type controls. To investigate the effects of TLR4 deficiency on early events of maternal immune adaptation, TLR4-regulated cytokines and immune regulatory microRNAs were measured in the uterus at 8 h post-mating by qPCR, and Treg cells in uterus-draining lymph nodes were evaluated by flow cytometry on day 3.5 post-coitum. Ptgs2 encoding prostaglandin-endoperoxide synthase 2, cytokines Csf2, Il6, Lif, and Tnf, chemokines Ccl2, Cxcl1, Cxcl2, and Cxcl10, and microRNAs miR-155, miR-146a, and miR-223 were induced by mating in wild-type mice, but not, or to a lesser extent, in Tlr4-/- mice. CD4+ T cells were expanded after mating in Tlr4+/+ but not Tlr4-/- mice, with failure to expand peripheral CD25+FOXP3+ NRP1- or thymic CD25+FOXP3+ NRP1+ Treg cell populations, and fewer Treg cells expressed Ki67 proliferation marker and suppressive function marker CTLA4. We conclude that TLR4 is an essential mediator of the inflammation-like response in the pre-implantation uterus that induces generation of Treg cells to support robust pregnancy tolerance and ensure optimal fetal growth and survival.

Primipaternities and human birthweights.

OBJECTIVES: To investigate in singleton multiparous pregnancies the effect of having a new father for an index pregnancy on new-borns' birthweights and intrauterine growth restriction. DESIGN: 20 year-observational cohort study (2001-2020). SETTINGS: Centre Hospitalier Universitaire Hospitalier Sud Reunion's maternity (French overseas department, Indian Ocean). MAIN OUTCOMES AND MEASURES: Comparing the 811 multiparas (cases) who had a new partner with the 49,712 who did not (controls), there were no differences concerning maternal age, education, ovulation induction/IVF, previous miscarriages, exams during pregnancies, pre-pregnancy BMI, gestational diabetes, and chronic hypertension. Cases had more previous pregnancies than controls (gravidity 4.2 vs 2.8, p < 0.001), volunteer abortions (OR1.93, p < 0.001), in vitro fecundations (OR 4.34, p < 0.001), were more likely to be unmarried (OR 2.94, p < 0.001) smoker (OR 2.2, p < 0.0001) and consuming alcohol during pregnancy (OR 2.35, p = 0.001). Cases had a much higher risk of preeclampsia than controls (OR 3.94, p < 0.001), especially early-onset preeclampsia (< 34 weeks) with an OR 4.1 (p < 0.001). Controlling for confounding factors (preeclampsia, smoking, alcohol use, early prematurity < 33 weeks, maternal ethnicity), primipaternity was an independent factor for small for gestational age newborns (OR 1.48, p < 0.001). CONCLUSIONS: It has been known for decades that primiparas have lighter babies than multiparas. Primipaternity represents also a risk for lower birth weights. Human birthweight seems to be linked with a "couple habituation" (to paternal genes) which may be not fully established in the first pregnancy of the couple.

Extrathymic Aire-expressing cells support maternal-fetal tolerance.

Healthy pregnancy requires tolerance to fetal alloantigens as well as syngeneic embryonic and placental antigens. Given the importance of the autoimmune regulator (Aire) gene in self-tolerance, we investigated the role of Aire-expressing cells in maternal-fetal tolerance. We report that maternal ablation of Aire-expressing (Aire +) cells during early mouse pregnancy caused intrauterine growth restriction (IUGR) in both allogeneic and syngeneic pregnancies. This phenotype is immune mediated, as IUGR was rescued in Rag1-deficient mice, and involved a memory response, demonstrated by recurrence of severe IUGR in second pregnancies. Single-cell RNA sequencing demonstrated that Aire + cell depletion in pregnancy results in expansion of activated T cells, particularly T follicular helper cells. Unexpectedly, selective ablation of either Aire-expressing medullary thymic epithelial cells or extrathymic Aire-expressing cells (eTACs) mapped the IUGR phenotype exclusively to eTACs. Thus, we report a previously undescribed mechanism for the maintenance of maternal-fetal immune homeostasis and demonstrate that eTACs protect the conceptus from immune-mediated IUGR.

Human Herpesviruses 6A and 6B in Reproductive Diseases.

Human herpesviruses 6A (HHV-6A) and human herpesvirus 6B (HHV-6B)-collectively, HHV-6A/B-are recently-discovered but ancient human viruses. The vast majority of people acquire one or both viruses, typically very early in life, producing an ineradicable lifelong infection. The viruses have been linked to several neurological, pulmonary and hematological diseases. In early human history, the viruses on multiple occasions infected a germ cell, and integrated their DNA into a human chromosome. As a result, about 1% of humans are born with the full viral genome present in every cell, with uncertain consequences for health. HHV-6A may play a role in 43% of cases of primary unexplained infertility. Both the inherited and acquired viruses may occasionally trigger several of the factors that are important in the pathogenesis of preeclampsia. Transplacental infection occurs in 1-2% of pregnancies, with some evidence suggesting adverse health consequences for the child. While emerging knowledge about these viruses in reproductive diseases is not sufficient to suggest any changes in current practice, we write this review to indicate the need for further research that could prove practice-changing.

Immune dysfunction in developmental programming of type 2 diabetes mellitus.

Intrauterine growth restriction (IUGR) is a common complication of pregnancy and increases the risk of the offspring developing type 2 diabetes mellitus (T2DM) later in life. Alterations in the immune system are implicated in the pathogenesis of IUGR-induced T2DM. The development of the fetal immune system is a delicate balance as it must remain tolerant of maternal antigens whilst also preparing for the post-birth environment. In addition, the fetal immune system is susceptible to an altered intrauterine milieu caused by maternal and placental inflammatory mediators or secondary to nutrient and oxygen deprivation. Pancreatic-resident macrophages populate the pancreas during fetal development, and their phenotype is dynamic through the neonatal period. Furthermore, macrophages in the islets are instrumental in islet development as they influence ß-cell proliferation and islet neogenesis. In addition, cytokines, derived from ß-cells and macrophages, are important to islet homeostasis in the fetus and adult and, when perturbed, can cause islet dysfunction. Several activated immune pathways have been identified in the islets of people who experienced IUGR, with alternations in the levels of IL-1ß and IL-4 as well as changes in TGFß signalling. Leptin levels are also altered. Immunomodulation has shown therapeutic benefit in T2DM and might be particularly useful in IUGR-induced T2DM.

Male fetal sex affects uteroplacental angiogenesis in growth restriction mouse model†.

Abnormally increased angiotensin II activity related to maternal angiotensinogen (AGT) genetic variants, or aberrant receptor activation, is associated with small-for-gestational-age babies and abnormal uterine spiral artery remodeling in humans. Our group studies a murine AGT gene titration transgenic (TG; 3-copies of the AGT gene) model, which has a 20% increase in AGT expression mimicking a common human AGT genetic variant (A[-6]G) associated with intrauterine growth restriction (IUGR) and spiral artery pathology. We hypothesized that aberrant maternal AGT expression impacts pregnancy-induced uterine spiral artery angiogenesis in this mouse model leading to IUGR. We controlled for fetal sex and fetal genotype (e.g., only 2-copy wild-type [WT] progeny from WT and TG dams were included). Uteroplacental samples from WT and TG dams from early (days 6.5 and 8.5), mid (d12.5), and late (d16.5) gestation were studied to assess uterine natural killer (uNK) cell phenotypes, decidual metrial triangle angiogenic factors, placental growth and capillary density, placental transcriptomics, and placental nutrient transport. Spiral artery architecture was evaluated at day 16.5 by contrast-perfused three-dimensional microcomputed tomography (3D microCT). Our results suggest that uteroplacental angiogenesis is significantly reduced in TG dams at day 16.5. Males from TG dams are associated with significantly reduced uteroplacental angiogenesis from early to late gestation compared with their female littermates and WT controls. Angiogenesis was not different between fetal sexes from WT dams. We conclude that male fetal sex compounds the pathologic impact of maternal genotype in this mouse model of growth restriction.

Identification and Characterization of Extrachromosomal Circular DNA in Human Placentas With Fetal Growth Restriction.

Many studies have confirmed that extrachromosomal circular DNAs (eccDNAs/ecDNAs) exist in tumor and normal cells independently of the chromosome and are essential for oncogene plasticity and drug resistance. Studies have confirmed that there are many eccDNAs/ecDNAs in maternal plasma derived from the fetus. Fetal growth restriction (FGR) is a pregnancy-related disease associated with high newborn morbidity and mortality. However, the characteristics and nature of eccDNAs/ecDNAs in FGR are poorly understood. This study aims to deconstruct the properties and potential functions of eccDNAs/ecDNAs in FGR. We performed circle-seq to identify the expression profile of eccDNAs/ecDNAs, analyzed by bioinformatics, and verified by real-time Polymerase Chain Reaction (PCR) combined with southern blot in FGR compared with the normal groups. A total of 45,131 eccDNAs/ecDNAs (including 2,118 unique ones) were identified, which had significantly higher abundance in FRG group than in normal group, and was bimodal in length, peaking at ~146bp and ~340bp, respectively. Gestational age may be one independent factor affecting the production of eccDNAs/ecDNAs, most of which come from genomic regions with high gene density, with a 4~12bp repeat around the junction, and their origin had a certain genetic preference. In addition, some of the host-genes overlapped with non-coding RNAs (ncRNAs) partially or even completely. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis revealed that host-genes on the differentially expressed eccDNAs/ecDNAs (DEEECs/DEECs) were mainly enriched in immune-related functions and pathways. The presence of some ecDNAs were verified, and whose variability were consistent with the circle-seq results. We identified and characterized eccDNAs/ecDNAs in placentas with FGR, and elucidated the formation mechanisms and the networks with ncRNAs, which provide a new vision for the screening of new biomarkers and therapeutic targets for FGR.

Study of the fetal and maternal microbiota in pregnant women with intrauterine growth restriction and its relationship with inflammatory biomarkers: A case-control study protocol (SPIRIT compliant).

In general terms, fetal growth restriction (FGR) is considered the impossibility of achieving the genetically determined potential size. In the vast majority of cases, it is related to uteroplacental insufficiency. Although its origin remains unknown and causes are only known in 30% of cases, it is believed to be related to an interaction of environmental and genetic factors with either a fetal or maternal origin. One hypothesis is that alterations in the gastrointestinal microbiota composition, and thus alteration in the immune response, could play a role in FGR development. We performed an observational, prospective study in a subpopulation affected with FGR to elucidate the implications of this microbiota on the FGR condition.A total of 63 fetuses with FGR diagnosed in the third trimester as defined by the Delphi consensus, and 63 fetuses with fetal growth appropriate for gestational age will be recruited. Obstetric and nutritional information will be registered by means of specific questionnaires. We will collect maternal fecal samples between 30 to 36 weeks, intrapartum samples (maternal feces, maternal and cord blood) and postpartum samples (meconium and new-born feces at 6 weeks of life). Samples will be analyzed in the Department of Biochemistry and Molecular Biology II, Nutrition and Food Technology Institute of the University of Granada (UGR), for the determination of the gastrointestinal microbiota composition and its relationship with inflammatory biomarkers.This study will contribute to a better understanding of the influence of gastrointestinal microbiota and related inflammatory biomarkers in the development of FGR.Trial registration: NCT04047966. Registered August 7, 2019, during the recruitment stage. Retrospectively registered. Ongoing research.

Altered Levels of Decidual Immune Cell Subsets in Fetal Growth Restriction, Stillbirth, and Placental Pathology.

Immune cells are critically involved in placental development and functioning, and inadequate regulation of the maternal immune system is associated with placental pathology and pregnancy complications. This study aimed to explore numbers of decidual immune cells in pregnancies complicated with fetal growth restriction (FGR) and stillbirth (SB), and in placentas with histopathological lesions: maternal vascular malperfusion (MVM), fetal vascular malperfusion (FVM), delayed villous maturation (DVM), chorioamnionitis (CA), and villitis of unknown etiology (VUE). Placental tissue from FGR (n = 250), SB (n = 64), and healthy pregnancies (n = 42) was included. Histopathological lesions were classified according to criteria developed by the Amsterdam Placental Workshop Group. Tissue slides were stained for CD68 (macrophages), CD206 (M2-like macrophages), CD3 (T cells), FOXP3 [regulatory T (Treg) cells], and CD56 [natural killer (NK) cells]. Cell numbers were analyzed in the decidua basalis using computerized morphometry. The Mann-Whitney U-test and Kruskal Wallis test with the Dunn's as post-hoc test were used for statistical analysis. Numbers of CD68+ macrophages were higher in FGR compared to healthy pregnancies (p < 0.001), accompanied by lower CD206+/CD68+ ratios (p < 0.01). In addition, in FGR higher numbers of FOXP3+ Treg cells were seen (p < 0.01) with elevated FOXP3+/CD3+ ratios (p < 0.01). Similarly, in SB elevated FOXP3+ Treg cells were found (p < 0.05) with a higher FOXP3+/CD3+ ratio (p < 0.01). Furthermore, a trend toward higher numbers of CD68+ macrophages was found (p < 0.1) in SB. Numbers of CD3+ and FOXP3+ cells were higher in placentas with VUE compared to placentas without lesions (p < 0.01 and p < 0.001), accompanied by higher FOXP3+/CD3+ ratios (p < 0.01). Elevated numbers of macrophages with a lower M2/total macrophage ratio in FGR suggest a role for a macrophage surplus in its pathogenesis and could specifically indicate involvement of inflammatory macrophages. Higher numbers of FOXP3+ Treg cells with higher Treg/total T cell ratios in VUE may be associated with impaired maternal-fetal tolerance and a compensatory response of Treg cells. The abundant presence of placental lesions in the FGR and SB cohorts might explain the increase of Treg/total T cell ratios in these groups. More functionality studies of the observed altered immune cell subsets are needed.

Impaired Neonatal Immunity and Infection Resistance Following Fetal Growth Restriction in Preterm Pigs.

Background: Infants born preterm or small for gestational age (SGA, due to fetal growth restriction) both show an increased risk of neonatal infection. However, it remains unclear how the co-occurrence of preterm birth and SGA may affect neonatal immunity and infection risk. We hypothesized that fetal growth restricted (FGR) preterm newborns possess impaired immune competence and increased susceptibility to systemic infection and sepsis, relative to corresponding normal birth weight (NBW) newborns. Methods: Using preterm pigs as a model for preterm infants, gene expression in lipopolysaccharide (LPS) stimulated cord blood was compared between NBW and FGR (lowest 25% birth weight percentile) preterm pigs. Next, clinical responses to a systemic Staphylococcus epidermidis (SE) challenge were investigated in newborn FGR and NBW preterm pigs. Finally, occurrence of spontaneous infections were investigated in 9 d-old FGR and NBW preterm pigs, with or without neonatal antibiotics treatment. Results: At birth, preterm FGR piglets showed diminished ex vivo cord blood responses to LPS for genes related to both innate and adaptive immunity, and also more severe septic responses following SE infection (e.g., higher blood lactate, decreased blood pH, neutrophil and platelet counts, relative to NBW pigs). After 9 d, FGR pigs had higher incidence and severity of spontaneous infections (e.g., higher bacterial densities in the bone marrow), increased regulatory T cell numbers, reduced neutrophil phagocytosis capacity, and impaired ex vivo blood gene responses to LPS, especially when receiving neonatal antibiotics. Conclusion: FGR at preterm birth is associated with poor immune competence, impaired infection resistance, and greater sepsis susceptibility in the immediate postnatal period. Our results may explain the increased morbidity and mortality of SGA preterm infants and highlight the need for clinical vigilance for this highly sensitive subgroup of preterm neonates.

Specific inflammatory profile in each pregnancy complication: A comparative study.

PROBLEM: Pre-eclampsia (PE), preterm birth (PTB) and intra-uterine growth restriction (IUGR) affect 5%-12% of pregnancies. They have been associated with placental inflammation, although the detection of inflammatory mediators in the maternal circulation is still controversial. Our goal was to determine the inflammatory changes occurring in the second part of pregnancy to identify profiles distinguishing pathological pregnancies from each other. METHOD OF STUDY: We performed a nested case-control study of 200 women randomly selected from a cohort recruited at the CHU de Quebec-Universite Laval, Quebec, Canada. Women with uncomplicated term pregnancy (CTRL); PE (severe or not); PTB or IUGR (N = 50/each) were included. Plasma samples, obtained from the late second trimester and at delivery, were analysed for over 30 selected mediators (including cytokines/alarmins), by multiplex, ELISA or specific assays. Demographic and obstetrical information were obtained for classification. RESULTS: In CTRL, we observed significant differences between 2nd trimester and delivery, with increased levels of inflammatory mediators (ex. MCP-1, IL-6), supporting an inflammatory profile towards term. Increased levels of IL-6, CXCL10 and CRP were observed in PE as compared to CTRL. In PTB, we observed increased CXCL9 in 2nd trimester and decreased progesterone at delivery. In IUGR, increased HMGB1 and IL-1α were observed only in the 2nd trimester. CONCLUSIONS: Our work showed significant inflammatory changes in uncomplicated pregnancies towards delivery, supporting that normal delivery is pro-inflammatory, although not to the same extent as in pathological pregnancies. Inflammatory profiles are specific to each pregnancy complication which may help to understand the contribution of inflammation to the clinical presentation of these conditions.

Increased placental macrophages and a pro-inflammatory profile in placentas and maternal serum in infants with a decreased growth rate in the third trimester of pregnancy.

PROBLEM: There is growing evidence for the role of placental inflammation in the pathophysiology of pregnancy complications including fetal growth restriction (FGR). This study aimed to characterize the inflammatory profile in the maternal circulation and the placenta of infants who were growth restricted and those that were small for gestational age (SGA). METHOD OF STUDY: Placental villous tissue and maternal serum were obtained from pregnancies where infants were SGA at birth or who had a decreasing growth rate (≥25 centiles) across the third trimester. Immunohistochemical and histological analyses of placental samples were conducted for macrophage number, alongside vascular and cell turnover analysis. Inflammatory profile was analyzed in maternal and placental compartments via ELISAs and multiplex assays. RESULTS: There were significantly more CD163+ macrophages in placentas of infants with a decreased growth rate compared to controls, but not in SGA infants (median 8.6/ nuclei vs 3.8 and 2.9, P = .008 and P = .003, respectively). Uric acid (P = .0007) and IL-8 (P = .0008) were increased in placentas, and S100A8 (P < .0002) was increased in maternal serum of infants with decreased growth rate. No changes in the maternal serum or placental lysates of SGA infants were observed. CONCLUSION: The evidence of an altered inflammatory profile in infants with a decreasing growth rate, but not in those that were born SGA, provides further evidence that inflammation plays a role in true FGR. It remains unclear whether the increased placental macrophages occur as a direct result, or as a consequence of the pro-inflammatory environment observed in fetal growth restriction.

Water-soluble N-2-Hydroxypropyl trimethyl ammonium chloride chitosan enhanced the immunogenicity of inactivated porcine parvovirus vaccine vaccination on sows against porcine parvovirus infection.

Porcine parvovirus (PPV) is one of the most common and important virus causes of infectious infertility in swine throughout the world. Inactivated PPV vaccine is broadly used, however, there is no appropriate immunomodulatory adjuvant for enhancing present vaccines and developing new ones. Therefore, in this study, the water-soluble N-2-Hydroxypropyl trimethyl ammonium chloride chitosan (N-2-HACC) was synthesized, the adjuvant potential of chitosan derivative was evaluated in inactivated PPV vaccine. Twenty adult healthy sows were assigned to four groups and vaccinated with synthesized PPV/N-2-HACC, commercial inactivated vaccine, N-2-HACC adjuvant and PBS. After insemination, all sows were challenged with the homologous PPV-H strain. In vivo immunization showed that sows immunized with the PPV/N-2-HACC induced more long-lasting HI antibodies and strong immune responses. Importantly, immunization of PPV/N-2-HACC significantly protected sows from homologous PPV-H strain infection. However, immunization of PPV/N-2-HACC didn't change the level of IL-2, IL-4 and IFN-γ and the production of CD4+, CD8 + T lymphocyte. The results indicated that PPV/N-2-HACC protect PPV infection mainly through enhancing the humoral immunity rather than cellular immunity. In addition, the mummified fetuses were observed from the control groups, but neither of the two vaccine groups. In conclusion, these results suggest that N-2-HACC can be exploited as an effective adjuvant for vaccine development, and the PPV/N-2-HACC are potent immunization candidates against PPV infection.

Dysfunction of Tregs contributes to FGR pathogenesis via regulating Smads signalling pathway.

Fetal growth restriction (FGR) is ranked number two of most common complication of abnormal pregnancy worldwide. The pathogenesis of FGR is complicated due to multiple aetiologies and the exact mechanism for FGR development is currently unknown. T regulatory cells (Tregs) are proven to play central roles in the maintenance of normal pregnancy. Peripheral blood samples of 102 pregnant human were collected analysed using flow cytometry to identify Tregs. We found that reduced Tregs and down-regulation of Foxp3 were observed in peripheral blood of FGR patients. In FGR mouse model, we have found that Tregs were not only reduced in spleen but also in placenta. In vitro, Foxp3 and its transcription regulatory signalling molecules, including P-Smad2, P-Smad3 and Smad4, were diminished as well. Inhibition on Foxp3 expression was partially reversed by overexpression of Smad2 and Smad4. In FGR patients, Western blot results revealed that Foxp3, P-Smad2, P-Smad3 and Smad4 expression was inhibited in placenta. Our preliminary result suggests that maternal-foetal immune tolerance mediated by Tregs plays an essential role in the development of FGR. The inhibited expression of Foxp3 and down-regulated Smad2/Smad3/Smad4 signalling pathway were involved in the FGR pathogenesis. Targeting maternal-foetal immune tolerance through Tregs might represent a novel therapeutic option for FGR.