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Introduction: IL6 signaling plays an important role in triggering labor and IL6 is an established biomarker of intrauterine infection/inflammation (IUI) driven preterm labor (PTL). The biology of IL6 during IUI at the maternal-fetal interface was investigated in samples from human subjects and non-human primates (NHP). Methods: Pregnant women with histologic chorioamnionitis diagnosed by placenta histology were recruited (n=28 term, n=43 for preterm pregnancies from 26-36 completed weeks of gestation). IUI was induced in Rhesus macaque by intraamniotic injection of lipopolysachharide (LPS, n=23). IL1 signaling was blocked using Anakinra (human IL-1 receptor antagonist, n=13), and Tumor necrosis factor (TNF) signaling was blocked by anti TNF-antibody (Adalimumab n=14). The blockers were given before LPS. All animals including controls (intraamniotic injection of saline n=27), were delivered 16h after LPS/saline exposure at about 80% gestation. Results: IUI induced a robust expression of IL6 mRNAs in the fetal membranes (chorion-amnion-decidua tissue) both in humans (term and preterm) and NHP. The major sources of IL6 mRNA expression were the amnion mesenchymal cells (AMC) and decidua stroma cells. Additionally, during IUI in the NHP, ADAM17 (a protease that cleaves membrane bound IL6 receptor (IL6R) to release a soluble form) and IL6R mRNA increased in the fetal membranes, and the ratio of IL6 and soluble forms of IL6R, gp130 increased in the amniotic fluid signifying upregulation of IL6 trans-signaling. Both IL1 and TNF blockade suppressed LPS-induced IL6 mRNAs in the AMC and variably decreased elements of IL6 trans-signaling. Discussion: These data suggest that IL1 and TNF blockers may be useful anti-inflammatory agents via suppression of IL6 signaling at the maternal-fetal interface.
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Interleucina-6 , Macaca mulatta , Transdução de Sinais , Fator de Necrose Tumoral alfa , Feminino , Gravidez , Humanos , Animais , Interleucina-6/metabolismo , Fator de Necrose Tumoral alfa/metabolismo , Corioamnionite/imunologia , Corioamnionite/metabolismo , Corioamnionite/veterinária , Lipopolissacarídeos/imunologia , Interleucina-1/metabolismo , Adulto , Trabalho de Parto Prematuro/imunologia , Trabalho de Parto Prematuro/metabolismo , Inflamação/imunologia , Inflamação/metabolismo , Proteína Antagonista do Receptor de Interleucina 1/metabolismo , Proteína Antagonista do Receptor de Interleucina 1/farmacologia , Placenta/metabolismo , Placenta/imunologiaRESUMO
The process of human parturition involves inflammation at the interface where fetal chorion trophoblast cells interact with maternal decidual stromal (DS) cells and maternal immune cells in the decidua (endometrium of pregnancy). This study tested the hypothesis that inflammation at the chorion-decidua interface (CDI) induces labor by negating the capacity for progesterone (P4) to block labor and that this is mediated by inactivation of P4 in DS cells by aldo-keto reductase family 1 member C1 (AKR1C1). In human, Rhesus macaque, and mouse CDI, AKR1C1 expression increased in association with term and preterm labor. In a human DS cell line and in explant cultures of term human fetal membranes containing the CDI, the prolabor inflammatory cytokine, interleukin-1ß (IL-1ß), and media conditioned by LPS-stimulated macrophages increased AKR1C1 expression and coordinately reduced nuclear P4 levels and P4 responsiveness. Loss of P4 responsiveness was overcome by inhibition of AKR1C1 activity, inhibition of AKR1C1 expression, and bypassing AKR1C1 activity with a P4 analog that is not metabolized by AKR1C1. Increased P4 activity in response to AKR1C1 inhibition was prevented by the P4 receptor antagonist RU486. Pharmacologic inhibition of AKR1C1 activity prevented parturition in a mouse model of inflammation-induced preterm parturition. The data suggest that inflammatory stimuli at the CDI drive labor by inducing AKR1C1-mediated P4 inactivation in DS cells and that inhibiting and/or bypassing of AKR1C1-mediated P4 inactivation is a plausible therapeutic strategy to mitigate the risk of inflammation-associated preterm birth.
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20-Hidroxiesteroide Desidrogenases , Decídua , Inflamação , Macaca mulatta , Parto , Progesterona , Células Estromais , Feminino , Animais , Progesterona/metabolismo , Progesterona/farmacologia , Decídua/metabolismo , Humanos , Camundongos , Células Estromais/metabolismo , Gravidez , Inflamação/metabolismo , 20-Hidroxiesteroide Desidrogenases/metabolismo , 20-Hidroxiesteroide Desidrogenases/genética , Interleucina-1beta/metabolismo , Córion/metabolismoRESUMO
The amnion is a thin layer of fetal origin in contact with the amniotic fluid which plays a key role at the feto-maternal interface during pregnancy. Here, we present a protocol for isolation of human and Rhesusmacaque amnion cells. We describe steps for tissue dissection, cell isolation for flow cytometry analysis, and RNA isolation for RNA sequencing library preparation and analysis. This protocol can provide insights into altered immunological pathways during intrauterine infections to develop new therapeutic strategies. For complete details on the use and execution of this protocol, please refer to Presicce et al.1.
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Âmnio , Separação Celular , Citometria de Fluxo , Placenta , Âmnio/citologia , Humanos , Citometria de Fluxo/métodos , Feminino , Gravidez , Animais , Placenta/citologia , Separação Celular/métodos , Perfilação da Expressão Gênica/métodos , Transcriptoma/genética , Macaca mulattaRESUMO
BACKGROUND: Preterm birth is often associated with chorioamnionitis and leads to increased risk of neurodevelopmental disorders, such as autism. Preterm birth can lead to cerebellar underdevelopment, but the mechanisms of disrupted cerebellar development in preterm infants are not well understood. The cerebellum is consistently affected in people with autism spectrum disorders, showing reduction of Purkinje cells, decreased cerebellar grey matter, and altered connectivity. METHODS: Preterm rhesus macaque fetuses were exposed to intra-amniotic LPS (1 mg, E. coli O55:B5) at 127 days (80%) gestation and delivered by c-section 5 days after injections. Maternal and fetal plasma were sampled for cytokine measurements. Chorio-decidua was analyzed for immune cell populations by flow cytometry. Fetal cerebellum was sampled for histology and molecular analysis by single-nuclei RNA-sequencing (snRNA-seq) on a 10× chromium platform. snRNA-seq data were analyzed for differences in cell populations, cell-type specific gene expression, and inferred cellular communications. RESULTS: We leveraged snRNA-seq of the cerebellum in a clinically relevant rhesus macaque model of chorioamnionitis and preterm birth, to show that chorioamnionitis leads to Purkinje cell loss and disrupted maturation of granule cells and oligodendrocytes in the fetal cerebellum at late gestation. Purkinje cell loss is accompanied by decreased sonic hedgehog signaling from Purkinje cells to granule cells, which show an accelerated maturation, and to oligodendrocytes, which show accelerated maturation from pre-oligodendrocytes into myelinating oligodendrocytes. CONCLUSION: These findings suggest a role of chorioamnionitis on disrupted cerebellar maturation associated with preterm birth and on the pathogenesis of neurodevelopmental disorders among preterm infants.
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Corioamnionite , Nascimento Prematuro , Recém-Nascido , Feminino , Lactente , Animais , Humanos , Gravidez , Proteínas Hedgehog , Macaca mulatta , Escherichia coli , Recém-Nascido Prematuro , Cerebelo , RNA Nuclear PequenoRESUMO
Intrauterine infection/inflammation (IUI) is a frequent complication of pregnancy leading to preterm labor and fetal inflammation. How inflammation is modulated at the maternal-fetal interface is unresolved. We compared transcriptomics of amnion (a fetal tissue in contact with amniotic fluid) in a preterm Rhesus macaque model of IUI induced by lipopolysaccharide with human cohorts of chorioamnionitis. Bulk RNA sequencing (RNA-seq) amnion transcriptomic profiles were remarkably similar in both Rhesus and human subjects and revealed that induction of key labor-mediating genes such as IL1 and IL6 was dependent on nuclear factor κB (NF-κB) signaling and reversed by the anti-tumor necrosis factor (TNF) antibody Adalimumab. Inhibition of collagen biosynthesis by IUI was partially restored by Adalimumab. Interestingly, single-cell transcriptomics, flow cytometry, and immunohistology demonstrated that a subset of amnion mesenchymal cells (AMCs) increase CD14 and other myeloid cell markers during IUI both in the human and Rhesus macaque. Our data suggest that CD14+ AMCs represent activated AMCs at the maternal-fetal interface.
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Clinical evidence points to a function for B cell-activating factor (BAFF) in pregnancy. However, direct roles for BAFF-axis members in pregnancy have not been examined. Here, via utility of genetically modified mice, we report that BAFF promotes inflammatory responsiveness and increases susceptibility to inflammation-induced preterm birth (PTB). In contrast, we show that the closely related A proliferation-inducing ligand (APRIL) decreases inflammatory responsiveness and susceptibility to PTB. Known BAFF-axis receptors serve a redundant function in signaling BAFF/APRIL presence in pregnancy. Treatment with anti-BAFF/APRIL monoclonal antibodies or BAFF/APRIL recombinant proteins is sufficient to manipulate susceptibility to PTB. Notably, macrophages at the maternal-fetal interface produce BAFF, while BAFF and APRIL presence divergently shape macrophage gene expression and inflammatory function. Overall, our findings demonstrate that BAFF and APRIL play divergent inflammatory roles in pregnancy and provide therapeutic targets for mitigating risk of inflammation-induced PTB.
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Nascimento Prematuro , Animais , Feminino , Camundongos , Gravidez , Fator Ativador de Células B , Inflamação , Transdução de Sinais , Membro 13 da Superfamília de Ligantes de Fatores de Necrose Tumoral/genéticaRESUMO
Perinatal inflammatory stress is associated with early life morbidity and lifelong consequences for pulmonary health. Chorioamnionitis, an inflammatory condition affecting the placenta and fluid surrounding the developing fetus, affects 25 to 40% of preterm births. Severe chorioamnionitis with preterm birth is associated with significantly increased risk of pulmonary disease and secondary infections in childhood, suggesting that fetal inflammation may markedly alter the development of the lung. Here, we used intra-amniotic lipopolysaccharide (LPS) challenge to induce experimental chorioamnionitis in a prenatal rhesus macaque (Macaca mulatta) model that mirrors structural and temporal aspects of human lung development. Inflammatory injury directly disrupted the developing gas exchange surface of the primate lung, with extensive damage to alveolar structure, particularly the close association and coordinated differentiation of alveolar type 1 pneumocytes and specialized alveolar capillary endothelium. Single-cell RNA sequencing analysis defined a multicellular alveolar signaling niche driving alveologenesis that was extensively disrupted by perinatal inflammation, leading to a loss of gas exchange surface and alveolar simplification, with notable resemblance to chronic lung disease in newborns. Blockade of the inflammatory cytokines interleukin-1ß and tumor necrosis factor-α ameliorated LPS-induced inflammatory lung injury by blunting stromal responses to inflammation and modulating innate immune activation in myeloid cells, restoring structural integrity and key signaling networks in the developing alveolus. These data provide new insight into the pathophysiology of developmental lung injury and suggest that modulating inflammation is a promising therapeutic approach to prevent fetal consequences of chorioamnionitis.
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Corioamnionite , Nascimento Prematuro , Animais , Corioamnionite/induzido quimicamente , Corioamnionite/patologia , Feminino , Pulmão/patologia , Macaca mulatta , Gravidez , Nascimento Prematuro/prevenção & controle , Troca Gasosa PulmonarRESUMO
Up to 40% of preterm births are associated with histological chorioamnionitis (HCA), which leads to elevated levels of pro-inflammatory mediators and microbial products in the amniotic fluid, which come in contact with fetal lungs. Yet, fetal pulmonary immune responses to such exposure remain poorly characterized. To address this gap, we used our established HCA model, in which pregnant Rhesus macaques receive intraamniotic (IA) saline or LPS. IA LPS induced a potent and rapid myeloid cell response in fetal lungs, dominated by neutrophils and monocytes/macrophages. Infiltrating and resident myeloid cells exhibited transcriptional profiles consistent with exposure to TLR ligands, as well as cytokines, notably IL-1 and TNFα. Although simultaneous, in vivo blockade of IL-1 and TNFα signaling did not prevent the inflammatory cell recruitment, it blunted the lung overall inflammatory state reducing communication between, and activation of, infiltrating immune cells. Our data indicate that the fetal innate immune system can mount a rapid multi-faceted pulmonary immune response to in utero exposure to inflammation. These data provide mechanistic insights into the association between HCA and the postnatal lung morbidities of the premature infant and highlight therapeutic potential of inflammatory blockade in the fetus.
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Corioamnionite , Pneumonia , Nascimento Prematuro , Líquido Amniótico , Animais , Corioamnionite/patologia , Feminino , Humanos , Inflamação , Interleucina-1 , Lipopolissacarídeos , Pulmão , Macaca mulatta , Gravidez , Nascimento Prematuro/patologia , Fator de Necrose Tumoral alfaRESUMO
Intrauterine infection/inflammation (IUI) is a major contributor to preterm labor (PTL). However, IUI does not invariably cause PTL. We hypothesized that quantitative and qualitative differences in immune response exist in subjects with or without PTL. To define the triggers for PTL, we developed rhesus macaque models of IUI driven by lipopolysaccharide (LPS) or live Escherichia coli. PTL did not occur in LPS challenged rhesus macaques, while E. coli-infected animals frequently delivered preterm. Although LPS and live E. coli both caused immune cell infiltration, E. coli-infected animals showed higher levels of inflammatory mediators, particularly interleukin 6 (IL-6) and prostaglandins, in the chorioamnion-decidua and amniotic fluid (AF). Neutrophil infiltration in the chorio-decidua was a common feature to both LPS and E. coli. However, neutrophilic infiltration and IL6 and PTGS2 expression in the amnion was specifically induced by live E. coli. RNA sequencing (RNA-seq) analysis of fetal membranes revealed that specific pathways involved in augmentation of inflammation including type I interferon (IFN) response, chemotaxis, sumoylation, and iron homeostasis were up-regulated in the E. coli group compared to the LPS group. Our data suggest that the intensity of the host immune response to IUI may determine susceptibility to PTL.
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Imunidade , Trabalho de Parto Prematuro/patologia , Complicações na Gravidez/imunologia , Animais , Modelos Animais de Doenças , Escherichia coli/patogenicidade , Infecções por Escherichia coli/complicações , Infecções por Escherichia coli/imunologia , Feminino , Inflamação , Lipopolissacarídeos/toxicidade , Macaca mulatta , GravidezRESUMO
Background: Granulocyte-macrophage colony-stimulating factor (GM-CSF) is a pro-inflammatory cytokine that is increased in the amniotic fluid in chorioamnionitis and elevated in the fetal lung with endotoxin exposure. Although GM-CSF has a pivotal role in fetal lung development, it stimulates pulmonary macrophages and is associated with the development of bronchopulmonary dysplasia (BPD). How antenatal GM-CSF results in recruitment of lung macrophage leading to BPD needs further elucidation. Hence, we used a transgenic and knock-out mouse model to study the effects of GM-CSF focusing on the fetal lung macrophage. Methods: Using bitransgenic (BTg) mice that conditionally over-expressed pulmonary GM-CSF after doxycycline treatment, and GM-CSF knock-out (KO) mice with no GM-CSF expression, we compared the ontogeny and immunophenotype of lung macrophages in BTg, KO and control mice at various prenatal and postnatal time points using flow cytometry and immunohistology. Results: During fetal life, compared to controls, BTg mice over-expressing pulmonary GM-CSF had increased numbers of lung macrophages that were CD68+ and these were primarily located in the interstitium rather than alveolar spaces. The lung macrophages that accumulated were predominantly CD11b+F4/80+ indicating immature macrophages. Conversely, lung macrophages although markedly reduced, were still present in GM-CSF KO mice. Conclusion: Increased exposure to GM-CSF antenatally, resulted in accumulation of immature macrophages in the fetal lung interstitium. Absence of GM-CSF did not abrogate but delayed the transitioning of interstitial macrophages. Together, these results suggest that other perinatal factors may be involved in modulating the maturation of alveolar macrophages in the developing fetal lung.
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Respiratory complicËations are the major cause of morbidity and mortality among preterm infants, which is partially prevented by the administration of antenatal corticosteroids (ACS). Most very preterm infants are exposed to chorioamnionitis, but short- and long-term effects of ACS treatment in this setting are not well defined. In low-resource settings, ACS increased neonatal mortality by perhaps increasing infection. We report that treatment with low-dose ACS in the setting of inflammation induced by intraamniotic lipopolysaccharide (LPS) in rhesus macaques improves lung compliance and increases surfactant production relative to either exposure alone. RNA sequencing shows that these changes are mediated by suppression of proliferation and induction of mesenchymal cellular death via TP53. The combined exposure results in a mature-like transcriptomic profile with inhibition of extracellular matrix development by suppression of collagen genes COL1A1, COL1A2, and COL3A1 and regulators of lung development FGF9 and FGF10. ACS and inflammation also suppressed signature genes associated with proliferative mesenchymal progenitors similar to the term gestation lung. Treatment with ACS in the setting of inflammation may result in early respiratory advantage to preterm infants, but this advantage may come at a risk of abnormal extracellular matrix development, which may be associated with increased risk of chronic lung disease.
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Corticosteroides/farmacologia , Pulmão/efeitos dos fármacos , Nascimento Prematuro/tratamento farmacológico , Corticosteroides/efeitos adversos , Animais , Animais Recém-Nascidos , Corioamnionite/tratamento farmacológico , Corioamnionite/genética , Dexametasona/farmacologia , Modelos Animais de Doenças , Feminino , Glucocorticoides/farmacologia , Inflamação/tratamento farmacológico , Inflamação/genética , Macaca mulatta , Masculino , Gravidez , Surfactantes Pulmonares/farmacologiaRESUMO
Intra-amniotic (IA) inflammation is associated with significant morbidities for both the mother and the fetus. Prior studies have illustrated many of the effects of IA inflammation on the uterine lining (decidua) and membranous layers of the placenta at the fetal-maternal interface. However, much less is known about the immunological response occurring within the villous placenta. Using a rhesus macaque model of lipopolysaccharide (LPS)-induced IA inflammation, we showed that pregnancy-matched choriodecidua and villi have distinct immunological profiles in rhesus pregnancies. In the choriodecidua, we show that the abundance of neutrophils, multiple populations of antigen-presenting cells, and two populations of natural killer (NK) cells changes with prenatal IA LPS exposure. In contrast, in immune cells within the villous placenta we observed alterations in the abundance of B cells, monocytes, and CD8 T cells. Prior work has illustrated that IA inflammation leads to an increase in tumor necrosis factor alpha (TNFα) at the fetal-maternal interface. In this study, pretreatment with a TNFα blockade partially reversed inflammation in the placental villi. Furthermore, we report that immune cells in the villous placenta sensed LPS during our experimental window, and subsequently activated T cells to produce proinflammatory cytokines. Moreover, this study is the first report of memory T cells in third-trimester non-human primate placental villi and provides evidence that manipulation of immune cells in the villi at the fetal-maternal interface should be considered as a potential therapeutic target for IA inflammation.
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Corioamnionite/imunologia , Vilosidades Coriônicas/imunologia , Decídua/imunologia , Leucócitos/imunologia , Ativação Linfocitária , Animais , Biomarcadores/metabolismo , Corioamnionite/induzido quimicamente , Corioamnionite/tratamento farmacológico , Corioamnionite/metabolismo , Vilosidades Coriônicas/efeitos dos fármacos , Vilosidades Coriônicas/metabolismo , Decídua/efeitos dos fármacos , Decídua/metabolismo , Modelos Animais de Doenças , Feminino , Imunofenotipagem , Leucócitos/efeitos dos fármacos , Leucócitos/metabolismo , Lipopolissacarídeos , Macaca mulatta , Gravidez , Transdução de Sinais , Inibidores do Fator de Necrose Tumoral/farmacologia , Fator de Necrose Tumoral alfa/metabolismoRESUMO
Acute chorioamnionitis is characterized by neutrophilic infiltration and inflammation at the maternal fetal interface. It is a relatively common complication of pregnancy and can have devastating consequences including preterm labor, maternal infections, fetal infection/inflammation, fetal lung, brain, and gastrointestinal tract injury. In this review, we will discuss current understanding of the pathogenesis, immunobiology, and mechanisms of this condition. Most commonly, acute chorioamnionitis is a result of ascending infection with relatively low-virulence organisms such as the Ureaplasma species. Furthermore, recent vaginal microbiome studies suggest that there is a link between vaginal dysbiosis, vaginal inflammation, and ascending infection. Although less common, microorganisms invading the maternal-fetal interface via hematogenous route (e.g., Zika virus, Cytomegalovirus, and Listeria) can cause placental villitis and severe fetal inflammation and injury. We will provide an overview of the knowledge gleaned from different animal models of acute chorioamnionitis and the role of different immune cells in different maternal-fetal compartments. Lastly, we will discuss how infectious agents can break the maternal tolerance of fetal allograft during pregnancy and highlight the novel future therapeutic approaches.
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Corioamnionite/imunologia , Infecções/imunologia , Vagina/imunologia , Animais , Disbiose , Feminino , Humanos , Microbiota , Gravidez , Complicações Infecciosas na Gravidez , Vagina/microbiologiaRESUMO
Accumulation of activated neutrophils at the feto-maternal interface is a defining hallmark of intrauterine inflammation (IUI) that might trigger an excessive immune response during pregnancy. Mechanisms responsible of this massive neutrophil recruitment are poorly investigated. We have previously showed that intraamniotic injection of LPS in rhesus macaques induced a neutrophil predominant inflammatory response similar to that seen in human IUI. Here, we demonstrate that anti-TNF antibody (Adalimumab) inhibited ~80% of genes induced by LPS involved in inflammatory signaling and innate immunity in chorio-decidua neutrophils. Consistent with the gene expression data, TNF-blockade decreased LPS-induced neutrophil accumulation and activation at the feto-maternal interface. We also observed a reduction in IL-6 and other pro-inflammatory cytokines but not prostaglandins concentrations in the amniotic fluid. Moreover, TNF-blockade decreased mRNA expression of inflammatory cytokines in the chorio-decidua but not in the uterus, suggesting that inhibition of TNF-signaling decreased the inflammation in a tissue-specific manner within the uterine compartment. Taken together, our results demonstrate a predominant role for TNF-signaling in modulating the neutrophilic infiltration at the feto-maternal interface during IUI and suggest that blockade of TNF-signaling could be considered as a therapeutic approach for IUI, the major leading cause of preterm birth.
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Corioamnionite/imunologia , Neutrófilos/imunologia , Fator de Necrose Tumoral alfa/imunologia , Adalimumab/farmacologia , Animais , Anti-Inflamatórios/farmacologia , Corioamnionite/induzido quimicamente , Feminino , Lipopolissacarídeos/toxicidade , Macaca mulatta , Infiltração de Neutrófilos/efeitos dos fármacos , Infiltração de Neutrófilos/imunologia , Gravidez , Fator de Necrose Tumoral alfa/antagonistas & inibidoresRESUMO
Preterm birth (PTB) is a major cause of neonatal mortality and morbidity, often triggered by chorioamnionitis or intrauterine inflammation (IUI) with or without infection. Recently, there has been a strong association of IL-1 with PTB. We hypothesized that IL-1R-associated kinase 1 (IRAK1), a key signaling mediator in the TLR/IL-1 pathway, plays a critical role in PTB. In human fetal membranes (FM) collected immediately after birth from women delivering preterm, p-IRAK1 was significantly increased in all the layers of FM with chorioamnionitis, compared with no-chorioamnionitis subjects. In a preterm rhesus macaque model of IUI given intra-amniotic LPS, induction of p-IRAK1 and downstream proinflammatory signaling mediators were seen in the FM. In a C57BL/6J wild-type PTB mouse model of IUI given intrauterine LPS, an IRAK1 inhibitor significantly decreased PTB and increased live birth in a dose-dependent manner. Furthermore, IRAK1 knockout mice were protected from LPS-induced PTB, which was seen in wild-type controls. Activation of IRAK1 was maintained by K63-mediated ubiquitination in preterm FM of humans with chorioamnionitis and rhesus and mouse IUI models. Mechanistically, IRAK1 induced PTB in the mouse model of IUI by upregulating expression of COX-2. Thus, our data from human, rhesus, and mouse demonstrates a critical role IRAK1 in IUI and inflammation-associated PTB and suggest it as potential therapeutic target in IUI-induced PTB.
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Membranas Extraembrionárias/metabolismo , Quinases Associadas a Receptores de Interleucina-1/metabolismo , Nascimento Prematuro/metabolismo , Útero/imunologia , Adulto , Animais , Corioamnionite , Modelos Animais de Doenças , Membranas Extraembrionárias/patologia , Feminino , Humanos , Quinases Associadas a Receptores de Interleucina-1/genética , Lipopolissacarídeos/imunologia , Macaca mulatta , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Gravidez , Nascimento Prematuro/imunologia , Adulto JovemRESUMO
Adequate iron supply during pregnancy is essential for fetal development. However, how fetal or amniotic fluid iron levels are regulated during healthy pregnancy, or pregnancies complicated by intraamniotic infection or inflammation (IAI), is unknown. We evaluated amniotic fluid and fetal iron homeostasis in normal and complicated murine, macaque, and human pregnancy. In mice, fetal iron endowment was affected by maternal iron status, but amniotic fluid iron concentrations changed little during maternal iron deficiency or excess. In murine and macaque models of inflamed pregnancy, the fetus responded to maternal systemic inflammation or IAI by rapidly upregulating hepcidin and lowering iron in fetal blood, without altering amniotic fluid iron. In humans, elevated cord blood hepcidin with accompanying hypoferremia was observed in pregnancies with antenatal exposure to IAI compared with those that were nonexposed. Hepcidin was also elevated in human amniotic fluid from pregnancies with IAI compared with those without IAI, but amniotic fluid iron levels did not differ between the groups. Our studies in mice, macaques, and humans demonstrate that amniotic fluid iron is largely unregulated but that the rapid induction of fetal hepcidin by inflammation and consequent fetal hypoferremia are conserved mechanisms that may be important in fetal host defense.
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Líquido Amniótico/metabolismo , Homeostase , Ferro/metabolismo , Complicações na Gravidez/metabolismo , Animais , Estudos de Casos e Controles , Feminino , Sangue Fetal/metabolismo , Feto/metabolismo , Humanos , Ferro/sangue , Macaca mulatta , Camundongos , Camundongos Endogâmicos C57BL , GravidezRESUMO
Zika virus (ZIKV) infection of pregnant women is associated with congenital Zika syndrome (CZS) and no vaccine is available, although several are being tested in clinical trials. We tested the efficacy of ZIKV DNA vaccine VRC5283 in a rhesus macaque model of congenital ZIKV infection. Most animal vaccine experiments have a set pathogen exposure several weeks or months after vaccination. In the real world, people encounter pathogens years or decades after vaccination, or may be repeatedly exposed if the virus is endemic. To more accurately mimic how this vaccine would be used, we immunized macaques before conception and then exposed them repeatedly to ZIKV during early and mid-gestation. In comparison to unimmunized animals, vaccinated animals had a significant reduction in peak magnitude and duration of maternal viremia, early fetal loss, fetal infection, and placental and fetal brain pathology. Vaccine-induced neutralizing antibody titers on the day of first ZIKV exposure were negatively associated with the magnitude of maternal viremia, and the absence of prolonged viremia was associated with better fetal outcomes. These data support further clinical development of ZIKV vaccine strategies to protect against negative fetal outcomes.
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Vacinação/métodos , Vacinas de DNA/uso terapêutico , Infecção por Zika virus/prevenção & controle , Animais , Anticorpos Neutralizantes/metabolismo , Feminino , Macaca mulatta , Gravidez , Complicações Infecciosas na Gravidez/imunologia , Complicações Infecciosas na Gravidez/prevenção & controle , Viremia/imunologia , Viremia/prevenção & controle , Zika virus/imunologia , Zika virus/patogenicidadeRESUMO
Genetic manipulation of NOD/SCID (NS) mice has yielded numerous sub-strains with specific traits useful for the study of human hematopoietic xenografts, each with unique characteristics. Here, we have compared the engraftment and output of umbilical cord blood (UCB) CD34+ cells in four immune-deficient strains: NS, NS with additional IL2RG knockout (NSG), NS with transgenic expression of human myeloid promoting cytokines SCF, GM-CSF, and IL-3 (NSS), and NS with both IL2RG knockout and transgenic cytokine expression (NSGS). Overall engraftment of human hematopoietic cells was highest in the IL2RG knockout strains (NSG and NSGS), while myeloid cell output was notably enhanced in the two strains with transgenic cytokine expression (NSS and NSGS). In further comparisons of NSG and NSGS mice, several additional differences were noted. NSGS mice were found to have a more rapid reconstitution of T cells, improved B cell differentiation, increased levels of NK cells, reduced platelets, and reduced maintenance of primitive CD34+ cells in the bone marrow. NSGS were superior hosts for secondary engraftment and both strains were equally suitable for experiments of graft versus host disease. Increased levels of human cytokines as well as human IgG and IgM were detected in the serum of humanized NSGS mice. Furthermore, immunization of humanized NSGS mice provided evidence of a functional response to repeated antigen exposure, implying a more complete hematopoietic graft was generated in these mice. These results highlight the important role that myeloid cells and myeloid-supportive cytokines play in the formation of a more functional xenograft immune system in humanized mice.
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Hematopoese , Subunidade gama Comum de Receptores de Interleucina/genética , Animais , Linfócitos B/citologia , Linfócitos B/metabolismo , Células da Medula Óssea/citologia , Células da Medula Óssea/metabolismo , Diferenciação Celular , Sangue Fetal/citologia , Sangue Fetal/metabolismo , Doença Enxerto-Hospedeiro/patologia , Fator Estimulador de Colônias de Granulócitos e Macrófagos/genética , Fator Estimulador de Colônias de Granulócitos e Macrófagos/metabolismo , Humanos , Subunidade gama Comum de Receptores de Interleucina/deficiência , Interleucina-3/genética , Interleucina-3/metabolismo , Células Matadoras Naturais/citologia , Células Matadoras Naturais/metabolismo , Camundongos , Camundongos Endogâmicos NOD , Camundongos Knockout , Camundongos SCID , Camundongos Transgênicos , Receptor ErbB-2/metabolismo , Fator de Células-Tronco/genética , Fator de Células-Tronco/metabolismo , Linfócitos T/citologia , Linfócitos T/metabolismoRESUMO
Zika virus (ZIKV) infection of pregnant women can cause fetal microcephaly and other neurologic defects. We describe the development of a non-human primate model to better understand fetal pathogenesis. To reliably induce fetal infection at defined times, four pregnant rhesus macaques are inoculated intravenously and intraamniotically with ZIKV at gestational day (GD) 41, 50, 64, or 90, corresponding to first and second trimester of gestation. The GD41-inoculated animal, experiencing fetal death 7 days later, has high virus levels in fetal and placental tissues, implicating ZIKV as cause of death. The other three fetuses are carried to near term and euthanized; while none display gross microcephaly, all show ZIKV RNA in many tissues, especially in the brain, which exhibits calcifications and reduced neural precursor cells. Given that this model consistently recapitulates neurologic defects of human congenital Zika syndrome, it is highly relevant to unravel determinants of fetal neuropathogenesis and to explore interventions.
Assuntos
Modelos Animais de Doenças , Doenças Fetais/patologia , Macaca mulatta , Doenças do Sistema Nervoso/patologia , Complicações Infecciosas na Gravidez/patologia , Infecção por Zika virus/patologia , Zika virus/patogenicidade , Animais , Encéfalo/patologia , Encéfalo/virologia , Feminino , Doenças Fetais/virologia , Feto/patologia , Feto/virologia , Humanos , Masculino , Doenças do Sistema Nervoso/virologia , Gravidez , Complicações Infecciosas na Gravidez/virologia , RNA Viral/isolamento & purificação , Zika virus/genética , Zika virus/isolamento & purificação , Infecção por Zika virus/virologiaRESUMO
Neutrophil infiltration of the chorioamnion-decidua tissue at the maternal-fetal interface (chorioamnionitis) is a leading cause of prematurity, fetal inflammation, and perinatal mortality. We induced chorioamnionitis in preterm rhesus macaques by intraamniotic injection of LPS. Here, we show that, during chorioamnionitis, the amnion upregulated phospho-IRAK1-expressed neutrophil chemoattractants CXCL8 and CSF3 in an IL-1-dependent manner. IL-1R blockade decreased chorio-decidua neutrophil accumulation, neutrophil activation, and IL-6 and prostaglandin E2 concentrations in the amniotic fluid. Neutrophils accumulating in the chorio-decidua had increased survival mediated by BCL2A1, and IL-1R blockade also decreased BCL2A1+ chorio-decidua neutrophils. Readouts for inflammation in a cohort of women with preterm delivery and chorioamnionitis were similar to findings in the rhesus macaques. IL-1 is a potential therapeutic target for chorioamnionitis and associated morbidities.