Kristina M. Adams Waldorf

Fetal brain lesions after subcutaneous inoculation of Zika virus in a pregnant nonhuman primate

We describe the development of fetal brain lesions after Zika virus (ZIKV) inoculation in a pregnant pigtail macaque. Periventricular lesions developed within 10 d and evolved asymmetrically in the occipital–parietal lobes. Fetal autopsy revealed ZIKV in the brain and significant cerebral white matter hypoplasia, periventricular white matter gliosis, and axonal and ependymal injury. Our observation of ZIKV-associated fetal brain lesions in a nonhuman primate provides a model for therapeutic evaluation.

A hemolytic pigment of Group B Streptococcus allows bacterial penetration of human placenta

Group B Streptococcus invades human amniotic epithelial cells using a hemolytic pigment.

The cervical mucus plug: Structured review of the literature

The cervical mucus plug is a large, complex structure within the cervical canal that is shed shortly before or during labor. We propose that the cervical mucus plug fulfills critical ‘gate‐keeper’ functions based on its physical and immunologic properties, which help prevent ascending infection and preterm labor. The viscoelastic properties of the cervical mucus plug are determined by mucins (large glycoproteins), which can inhibit viral replication and exclude larger molecules and bacteria by preventing their diffusion through the plug. Furthermore, the innate and adaptive immunological properties of the cervical mucus plug are well suited for arresting bacterial infection by stimulating a robust inflammatory response. A possible association between an impaired gate‐keeper function of the cervical mucus plug and preterm birth is discussed.

Choriodecidual inflammation: a harbinger of the preterm labor syndrome.

Causal, cellular, and inflammatory links between choriodecidual infection with group B streptococcus (GBS) and preterm labor were assessed in a nonhuman primate model. Rhesus monkeys received varying doses of a clinical isolate of GBS, type III or saline, via an indwelling catheter placed between the chorion/decidua and myometrium in the lower pole of the uterus. Choriodecidual inoculation of GBS was followed by a graded response in amniotic fluid (AF) leukocytes, proinflammatory cytokines, prostaglandin E2 and F2α, and uterine activity (P < .05). The magnitude of the inflammatory response in AF was related, in part, to the initial inoculum size and whether AF cultures remained negative or became positive for GBS. Microbial invasion of AF was associated with advanced inflammation and preterm labor. We provide experimental evidence that choriodeciduitis is a transitional stage of intrauterine infection, which may be self-limited, remain dormant, or progress to intraamniotic infection. These data, coupled with clinical observations, suggest that choriodecidual inflammation is an antecedent event in the pathogenesis of premature cervical ripening (functional cervical insufficiency), premature rupture of the fetal membranes, or preterm labor.

The hidden maternal-fetal interface: events involving the lymphoid organs in maternal-fetal tolerance.

The genetic disparity between the mother and fetus has long enticed immunologists to search for mechanisms of maternal tolerance to fetal antigens. The study of antigen-specific tolerance in murine and human pregnancy has gained new momentum in recent years through the focus on antigen-presenting cells, uterine lymphatics and fetal antigen-specific maternal T cell responses. In mice, we now know that these responses occur within the secondary lymphoid structures as they can be conveniently tracked through the use of defined, often transgenic fetal antigens and maternal T cell receptors. Although the secondary lymphoid organs are sites of both immunization and tolerization to antigens, the immunological processes that occur in response to fetal antigens during the healthy pregnancy must invariably lead to tolerance. The molecular properties of these maternal-fetal tolerogenic interactions are still being unraveled, and are likely to be greatly influenced by tissue-specific microenvironments and the hormonal milieu of pregnancy. In this article, we discuss the events leading to antigen-specific maternal tolerance, including the trafficking of fetal antigens to secondary lymphoid organs, the properties of the antigen-presenting cells that display them to maternal T lymphocytes, and the nature of the ensuing tolerogenic response. Experimental data generated from human biological specimens as well as murine transgenic models are considered.

Choriodecidual Group B Streptococcal Inoculation Induces Fetal Lung Injury without Intra-Amniotic Infection and Preterm Labor in Macaca nemestrina

Background Early events leading to intrauterine infection and fetal lung injury remain poorly defined, but may hold the key to preventing neonatal and adult chronic lung disease. Our objective was to establish a nonhuman primate model of an early stage of chorioamnionitis in order to determine the time course and mechanisms of fetal lung injury in utero. Methodology/Principal Findings Ten chronically catheterized pregnant monkeys (Macaca nemestrina) at 118–125 days gestation (term = 172 days) received one of two treatments: 1) choriodecidual and intra-amniotic saline (n = 5), or 2) choriodecidual inoculation of Group B Streptococcus (GBS) 1×106 colony forming units (n = 5). Cesarean section was performed regardless of labor 4 days after GBS or 7 days after saline infusion to collect fetal and placental tissues. Only two GBS animals developed early labor with no cervical change in the remaining animals. Despite uterine quiescence in most cases, blinded review found histopathological evidence of fetal lung injury in four GBS animals characterized by intra-alveolar neutrophils and interstitial thickening, which was absent in controls. Significant elevations of cytokines in amniotic fluid (TNF-α, IL-8, IL-1β, IL-6) and fetal plasma (IL-8) were detected in GBS animals and correlated with lung injury (p<0.05). Lung injury was not directly caused by GBS, because GBS was undetectable in amniotic fluid (∼10 samples tested/animal), maternal and fetal blood by culture and polymerase chain reaction. In only two cases was GBS cultured from the inoculation site in low numbers. Chorioamnionitis occurred in two GBS animals with lung injury, but two others with lung injury had normal placental histology. Conclusions/Significance A transient choriodecidual infection can induce cytokine production, which is associated with fetal lung injury without overt infection of amniotic fluid, chorioamnionitis or preterm labor. Fetal lung injury may, thus, occur silently without symptoms and before the onset of the fetal systemic inflammatory response syndrome.

Synergy and Interactions Among Biological Pathways Leading to Preterm Premature Rupture of Membranes

Preterm premature rupture of membranes (PPROM) occurs in 1% to 2% of births. Impact of PPROM is greatest in low- and middle-income countries where prematurity-related deaths are most common. Recent investigations identify cytokine and matrix metalloproteinase activation, oxidative stress, and apoptosis as primary pathways to PPROM. These biological processes are initiated by heterogeneous etiologies including infection/inflammation, placental bleeding, uterine overdistention, and genetic polymorphisms. We hypothesize that pathways to PPROM overlap and act synergistically to weaken membranes. We focus our discussion on membrane composition and strength, pathways linking risk factors to membrane weakening, and future research directions to reduce the global burden of PPROM.

Group B Streptococcal Infection of the Choriodecidua Induces Dysfunction of the Cytokeratin Network in Amniotic Epithelium: A Pathway to Membrane Weakening

Early events leading to intrauterine infection remain poorly defined, but may hold the key to preventing preterm delivery. To determine molecular pathways within fetal membranes (chorioamnion) associated with early choriodecidual infection that may progress to preterm premature rupture of membranes (PPROM), we examined the effects of a Group B Streptococcus (GBS) choriodecidual infection on chorioamnion in a nonhuman primate model. Ten chronically catheterized pregnant monkeys (Macaca nemestrina) at 118–125 days gestation (term = 172 days) received choriodecidual inoculation of either GBS (n = 5) or saline (n = 5). Cesarean section was performed in the first week after GBS or saline inoculation. RNA extracted from chorioamnion (inoculation site) was profiled by microarray. Single gene, Gene Set, and Ingenuity Pathway Analysis results were validated using qRT-PCR (chorioamnion), Luminex (amniotic fluid, AF), immunohistochemistry, and transmission electron microscopy (TEM). Despite uterine quiescence in most cases, significant elevations of AF cytokines (TNF-α, IL-8, IL-1β, IL-6) were detected in GBS versus controls (p<0.05). Choriodecidual infection resolved by the time of cesarean section in 3 of 5 cases and GBS was undetectable by culture and PCR in the AF. A total of 331 genes were differentially expressed (>2-fold change, p<0.05). Remarkably, GBS exposure was associated with significantly downregulated expression of multiple cytokeratin (CK) and other cytoskeletal genes critical for maintenance of tissue tensile strength. Immunofluorescence revealed highly significant changes in the CK network within amniocytes with dense CK aggregates and retraction from the cell periphery (all p = 0.006). In human pregnancies affected by PPROM, there was further evidence of CK network retraction with significantly shorter amniocyte foot processes (p = 0.002). These results suggest early choriodecidual infection results in decreased cellular membrane integrity and tensile strength via dysfunction of CK networks. Downregulation of CK expression and perturbations in the amniotic epithelial cell intermediate filament network occur after GBS choriodecidual infection, which may contribute to PPROM.

Choriodecidual Infection Downregulates Angiogenesis and Morphogenesis Pathways in Fetal Lungs from Macaca Nemestrina

Background Intrauterine exposure to amniotic fluid (AF) cytokines is thought to predispose to bronchopulmonary dysplasia (BPD). We evaluated the effects of GBS exposure on RNA expression in fetal lung tissue to determine early molecular pathways associated with fetal lung injury that may progress to BPD. Methods Ten chronically catheterized pregnant monkeys (Macaca nemestrina) at 118–125 days gestation (term = 172 days) received choriodecidual inoculation of either: 1) Group B Streptococcus (n = 5) or 2) saline (n = 5). Cesarean section and fetal necropsy was performed in the first week after GBS or saline inoculation regardless of labor. RNA was extracted from fetal lungs and profiled by microarray. Results were analyzed using single gene, Gene Set, and Ingenuity Pathway Analysis. Validation was by RT-PCR and immunohistochemistry. Results Despite uterine quiescence in most cases, fetal lung injury occurred in four GBS cases (intra-alveolar neutrophils, interstitial thickening) and one control (peri-mortem hemorrhage). Significant elevations of AF cytokines (TNF-α, IL-8, IL-1β, IL-6) were detected in GBS versus controls (p<0.05). Lung injury was not directly caused by GBS, because GBS was undetectable by culture and PCR in the AF and fetal lungs. A total of 335 genes were differentially expressed greater than 1.5 fold (p<0.05) with GBS exposure associated with a striking upregulation of genes in innate and adaptive immunity and downregulation of pathways for angiogenesis, morphogenesis, and cellular growth and development. Conclusions A transient choriodecidual infection may induce fetal lung injury with profound alterations in the genetic program of the fetal lung before signs of preterm labor. Our results provide a window for the first time into early molecular pathways disrupting fetal lung angiogenesis and morphogenesis before preterm labor occurs, which may set the stage for BPD. A strategy to prevent BPD should target the fetus in utero to attenuate alterations in the fetal lung genetic program.

Congenital Zika virus infection as a silent pathology with loss of neurogenic output in the fetal brain

Zika virus (ZIKV) is a flavivirus with teratogenic effects on fetal brain, but the spectrum of ZIKV-induced brain injury is unknown, particularly when ultrasound imaging is normal. In a pregnant pigtail macaque (Macaca nemestrina) model of ZIKV infection, we demonstrate that ZIKV-induced injury to fetal brain is substantial, even in the absence of microcephaly, and may be challenging to detect in a clinical setting. A common and subtle injury pattern was identified, including (i) periventricular T2-hyperintense foci and loss of fetal noncortical brain volume, (ii) injury to the ependymal epithelium with underlying gliosis and (iii) loss of late fetal neuronal progenitor cells in the subventricular zone (temporal cortex) and subgranular zone (dentate gyrus, hippocampus) with dysmorphic granule neuron patterning. Attenuation of fetal neurogenic output demonstrates potentially considerable teratogenic effects of congenital ZIKV infection even without microcephaly. Our findings suggest that all children exposed to ZIKV in utero should receive long-term monitoring for neurocognitive deficits, regardless of head size at birth.