Exploring T. pallidum Pathogenesis in the Placenta Using Novel In Vitro Models of Syphilis During Pregnancy

PROJECT SUMMARY A rapid rise in syphilis cases has caused cases of congenital syphilis, the second leading cause of stillbirth worldwide, to skyrocket. Despite its prevalence, very little is known about the mechanisms through which Treponema pallidum (Tp), the causative agent of syphilis, is able to invade the placenta, impact placental function, and cause adverse pregnancy outcomes. This is largely due to a complete lack of viable in vitro models to study pathogenesis of Tp in the placenta. The overall objective of this proposal is to use novel in vitro models to investigate mechanisms through which Tp invades and impacts the placenta. By combining expertise from the Mysorekar and Edmondson/Norris laboratories, Tp has successfully been cultured in vitro in trophoblast cells and this system is being adapted to work with 3D stem cell-derived trophoblast organoids (SC-TOs). This project investigates the hypothesis that pairing these new in vitro models with an integrative, multi-omic approach will allow identification of key pathways and targets underlying placental invasion and replication of Tp at the maternal-fetal interface. Aim 1 will further define the optimal conditions for co-culturing Tp with Jeg-3 cells and SC-TOs. Advanced microscopy techniques will be used to characterize Tp attachment, growth, and possible internalization and migration of various Tp strains in our models. Aim 2 will elucidate the metabolic alterations and nutrient siphoning mechanisms that occur during Tp infection. Unbiased metabolomics will be conducted on trophoblasts infected with Tp to identify changes in trophoblast nutrient and metabolite levels due to host nutrient siphoning by Tp. Western blotting, immunofluorescence, and confocal imaging will then be used to examine changes in the expression and localization of nutrient transporters which regulate materno-fetal nutrient allocation across the placenta. Aim 3 will use integrated multi-omics to explore mechanisms underlying and driving Tp infection of trophoblasts by identifying changes in chromatin accessibility, gene expression, and protein expression elicited by infection. Bioinformatics will be used to integrate these -omic outputs and identify overarching pathways altered in trophoblast cells by Tp. Together, these studies will begin to identify key mechanisms through which Tp infects the placenta and impacts trophoblast function. This will be integral to increasing basic understanding of Tp pathogenesis in the placenta and for identifying novel biomarkers and treatment targets for syphilis-associated pregnancy outcomes.