Mechanistic study of human placental infection by Listeria monocytogenes

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DESCRIPTION (provided by applicant): A key function of the placenta is to protect the developing fetus. The barrier functions provided at the interface with maternal tissues by extravillous cytotrophoblasts and the syncytiotrophoblast, and at the interface with fetal blood by placental endothelial cells, are crucial for this protection. This protection is so efficient that nly a few pathogens can circumvent it and infect the fetus. One of these pathogens is Listeria monocytogenes, the etiologic agent of the life-threatening foodborne illness listeriosis. Pregnant women are highly susceptible to listeriosis and can transmit this pathogen to the placenta and fetus, leading to fetal and neonatal death (25%) and long-term neurologic sequelae (13%). Two studies convincingly argued that L. monocytogenes enters the human placenta by invading extravillous cytotrophoblasts and the syncytiotrophoblast. However, the fate of L. monocytogenes within and beyond those trophoblastic cells is unsettled. Our preliminary data led to the observation that L. monocytogenes displays an atypical intracellular lifecycle in primary human syncytiotrophoblasts. Indeed, L. monocytogenes is internalized into those cells and initially proliferates in the cytosol. However, the bacterial burden severely decreases 10-20 hours post-infection. These data support the central hypothesis of this proposal that the syncytiotrophoblast can establish an antimicrobial response. This response is not sufficient to completely halt some pathogens, such as L. monocytogenes, which proliferates for enough time to pass on infection to other placental cells and reaches the fetus. To expand on this finding, Aim 1 will establish the susceptibility of primary human placental cells (syncytiotrophoblasts, cytotrophoblasts, fibroblasts, macrophages (Hofbauer cells), and endothelial cells) to L. monocytogenes during the transplacental progression of infection. Culture of primary human placental cells will be used for detailed study of the L. monocytogenes intracellular lifecycle (Ai 1A). Term placental villous explants will be used for study of the successive stages of the infection of the tissue (Aim 1B). Aim 2 will identify innate defense mechanisms activated in syncytiotrophoblastic cells in response to L. monocytogenes infection. This will involve next- generation sequencing platforms to perform comparative transcriptional profiling of cells exposed, or not, to wild type L. monocytogenes. After confirmation of the results, the most pertinent pathways will be selected in order to establish their antimicrobial functions. This proposal focuses on establishing the relative susceptibility to infection of the various primary cells that compose the human placenta, and on identifying the innate immune responses of the human syncytiotrophoblast. In the long term, the data acquired in the course of the proposal will identify virulence mechanisms and host pathways that can be targeted by therapeutic agents to decrease the morbidity and mortality of pregnancy-associated infections.