C. Patrick Case

Secretions from placenta, after hypoxia/reoxygenation, can damage developing neurones of brain under experimental conditions.

Some psychiatric diseases in children and young adults are thought to originate from adverse exposures during foetal life, including hypoxia and hypoxia/reoxygenation. The mechanism is not understood. Several authors have emphasised that the placenta is likely to play an important role as the key interface between mother and foetus. Here we have explored whether a first trimester human placenta or model barrier of primary human cytotrophoblasts might secrete factors, in response to hypoxia or hypoxia/reoxygenation, that could damage neurones. We find that the secretions in conditioned media caused an increase of [Ca(2+)]i and mitochondrial free radicals and a decrease of dendritic lengths, branching complexity, spine density and synaptic activity in dissociated neurones from embryonic rat cerebral cortex. There was altered staining of glutamate and GABA receptors. We identify glutamate as an active factor within the conditioned media and demonstrate a specific release of glutamate from the placenta/cytotrophoblast barriers invitro after hypoxia or hypoxia/reoxygenation. Injection of conditioned media into developing brains of P4 rats reduced the numerical density of parvalbumin-containing neurones in cortex, hippocampus and reticular nucleus, reduced immunostaining of glutamate receptors and altered cellular turnover. These results show that the placenta is able to release factors, in response to altered oxygen, that can damage developing neurones under experimental conditions.

Treating the placenta to prevent adverse effects of gestational hypoxia on fetal brain development.

Some neuropsychiatric disease, including schizophrenia, may originate during prenatal development, following periods of gestational hypoxia and placental oxidative stress. Here we investigated if gestational hypoxia promotes damaging secretions from the placenta that affect fetal development and whether a mitochondria-targeted antioxidant MitoQ might prevent this. Gestational hypoxia caused low birth-weight and changes in young adult offspring brain, mimicking those in human neuropsychiatric disease. Exposure of cultured neurons to fetal plasma or to secretions from the placenta or from model trophoblast barriers that had been exposed to altered oxygenation caused similar morphological changes. The secretions and plasma contained altered microRNAs whose targets were linked with changes in gene expression in the fetal brain and with human schizophrenia loci. Molecular and morphological changes in vivo and in vitro were prevented by a single dose of MitoQ bound to nanoparticles, which were shown to localise and prevent oxidative stress in the placenta but not in the fetus. We suggest the possibility of developing preventative treatments that target the placenta and not the fetus to reduce risk of psychiatric disease in later life.

Nanoparticle-induced neuronal toxicity across placental barriers is mediated by autophagy and dependent on astrocytes

The potential for maternal nanoparticle (NP) exposures to cause developmental toxicity in the fetus without the direct passage of NPs has previously been shown, but the mechanism remained elusive. We now demonstrate that exposure of cobalt and chromium NPs to BeWo cell barriers, an in vitro model of the human placenta, triggers impairment of the autophagic flux and release of interleukin-6. This contributes to the altered differentiation of human neural progenitor cells and DNA damage in the derived neurons and astrocytes. Crucially, neuronal DNA damage is mediated by astrocytes. Inhibiting the autophagic degradation in the BeWo barrier by overexpression of the dominant-negative human ATG4BC74A significantly reduces the levels of DNA damage in astrocytes. In vivo, indirect NP toxicity in mice results in neurodevelopmental abnormalities with reactive astrogliosis and increased DNA damage in the fetal hippocampus. Our results demonstrate the potential importance of autophagy to elicit NP toxicity and the risk of indirect developmental neurotoxicity after maternal NP exposure.Exposure to nanoparticles induces indirect autophagy-mediated signalling events, leading to neuron damage via astrocytes in a human model of the placenta and to DNA damage in the neonatal hippocampus in vivo.

Maternal treatment with a placental-targeted antioxidant (MitoQ) impacts offspring cardiovascular function in a rat model of prenatal hypoxia

Graphical abstract Figure. No caption available. &NA; Intrauterine growth restriction, a common consequence of prenatal hypoxia, is a leading cause of fetal morbidity and mortality with a significant impact on population health. Hypoxia may increase placental oxidative stress and lead to an abnormal release of placental‐derived factors, which are emerging as potential contributors to developmental programming. Nanoparticle‐linked drugs are emerging as a novel method to deliver therapeutics targeted to the placenta and avoid risking direct exposure to the fetus. We hypothesize that placental treatment with antioxidant MitoQ loaded onto nanoparticles (nMitoQ) will prevent the development of cardiovascular disease in offspring exposed to prenatal hypoxia. Pregnant rats were intravenously injected with saline or nMitoQ (125 &mgr;M) on gestational day (GD) 15 and exposed to either normoxia (21% O2) or hypoxia (11% O2) from GD15‐21 (term: 22 days). In one set of animals, rats were euthanized on GD 21 to assess fetal body weight, placental weight and placental oxidative stress. In another set of animals, dams were allowed to give birth under normal atmospheric conditions (term: GD 22) and male and female offspring were assessed at 7 and 13 months of age for in vivo cardiac function (echocardiography) and vascular function (wire myography, mesenteric artery). Hypoxia increased oxidative stress in placentas of male and female fetuses, which was prevented by nMitoQ. 7‐month‐old male and female offspring exposed to prenatal hypoxia demonstrated cardiac diastolic dysfunction, of which nMitoQ improved only in 7‐month‐old female offspring. Vascular sensitivity to methacholine was reduced in 13‐month‐old female offspring exposed to prenatal hypoxia, while nMitoQ treatment improved vasorelaxation in both control and hypoxia exposed female offspring. Male 13‐month‐old offspring exposed to hypoxia showed an age‐related decrease in vascular sensitivity to phenylephrine, which was prevented by nMitoQ. In summary, placental‐targeted MitoQ treatment in utero has beneficial sex‐ and age‐dependent effects on adult offspring cardiovascular function.

Evidence for bystander signalling between human trophoblast cells and human embryonic stem cells

Maternal exposure during pregnancy to toxins can occasionally lead to miscarriage and malformation. It is currently thought that toxins pass through the placental barrier, albeit bi-layered in the first trimester, and damage the fetus directly, albeit at low concentration. Here we examined the responses of human embryonic stem (hES) cells in tissue culture to two metals at low concentration. We compared direct exposures with indirect exposures across a bi-layered model of the placenta cell barrier. Direct exposure caused increased DNA damage without apoptosis or a loss of cell number but with some evidence of altered differentiation. Indirect exposure caused increased DNA damage and apoptosis but without loss of pluripotency. This was not caused by metal ions passing through the barrier. Instead the hES cells responded to signalling molecules (including TNF-α) secreted by the barrier cells. This mechanism was dependent on connexin 43 mediated intercellular ‘bystander signalling’ both within and between the trophoblast barrier and the hES colonies. These results highlight key differences between direct and indirect exposure of hES cells across a trophoblast barrier to metal toxins. It offers a theoretical possibility that an indirectly mediated toxicity of hES cells might have biological relevance to fetal development.

Preeclamptic placentae release factors that damage neurons: implications for foetal programming of disease

Abstract Prenatal development is a critical period for programming of neurological disease. Preeclampsia, a pregnancy complication involving oxidative stress in the placenta, has been associated with long-term health implications for the child, including an increased risk of developing schizophrenia and autism spectrum disorders in later life. To investigate if molecules released by the placenta may be important mediators in foetal programming of the brain, we analysed if placental tissue delivered from patients with preeclampsia secreted molecules that could affect cortical cells in culture. Application of culture medium conditioned by preeclamptic placentae to mixed cortical cultures caused changes in neurons and astrocytes that were related to key changes observed in brains of patients with schizophrenia and autism, including effects on dendrite lengths, astrocyte number as well as on levels of glutamate and γ-aminobutyric acid receptors. Treatment of the placental explants with an antioxidant prevented neuronal abnormalities. Furthermore, we identified that bidirectional communication between neurons and astrocytes, potentially via glutamate, is required to produce the effects of preeclamptic placenta medium on cortical cells. Analysis of possible signalling molecules in the placenta-conditioned medium showed that the secretion profile of extracellular microRNAs, small post-transcriptional regulators, was altered in preeclampsia and partially rescued by antioxidant treatment of the placental explants. Predicted targets of these differentially abundant microRNAs were linked to neurodevelopment and the placenta. The present study provides further evidence that the diseased placenta may release factors that damage cortical cells and suggests the possibility of targeted antioxidant treatment of the placenta to prevent neurodevelopmental disorders.

In vitro placenta barrier model using primary human trophoblasts, underlying connective tissue and vascular endothelium

Fetal development may be compromised by adverse events at the placental interface between mother and fetus. However, it is still unclear how the communication between mother and fetus occurs through the placenta. In vitro - models of the human placental barrier, which could help our understanding and which recreate three-dimensional (3D) structures with biological functionalities and vasculatures, have not been reported yet. Here we present a 3D-vascularized human primary placental barrier model which can be constructed in 1 day. We illustrate the similarity of our model to first trimester human placenta, both in its structure and in its ability to respond to altered oxygen and to secrete factors that cause damage cells across the barrier including embryonic cortical neurons. We use this model to highlight the possibility that both the trophoblast and the endothelium within the placenta might play a role in the fetomaternal dialogue.