(1146) Menstrual discs for cervicovaginal cellular phenotyping: a novel methodology to define tissue-specific adaptations in pregnancy.

Softdiscs were collected from non-pregnant (NP) non-menstruating and pregnant participants enrolled in an ongoing preterm birth study. Single cell RNA-sequencing (scRNA-seq) was performed on cells isolated from discs. Standard Seurat based workflows identified cell clusters and cell type enrichment (PangaloaDB and EnrichR). Supervised differential expression analysis and pathway analysis (Metascape) were performed.  


Results:

27 discs were collected. scRNA-seq was performed for 5 pregnant and 7 NP samples. Characteristics were similar between groups with respect to age (26.8 years, SD 2.6) and race. Mean gestational age at disc collection was 20.6 weeks (SD 2.6) for pregnant participants. After processing, 24,976 cells were isolated in total. Cell viability was ~40% with no correlation to the sampled population. There was a 10-fold higher cell recovery in pregnant individuals. Two large clusters and seven sub-clusters of cells were identified (Fig. 1). The majority of cells were epithelial with a smaller cluster of myeloid cells. Pathway analysis of the top upregulated genes in pregnant vs NP revealed pathways related to cytokine signaling, leukocyte influx, and bacterial defense (Fig. 2).


Conclusion:

We demonstrate feasibility of a novel noninvasive sampling methodology to define cell signatures at a critical biologic site, with increased immune activation being a hallmark of pregnancy physiology in the cervicovaginal space. Molecular phenotyping of exfoliated epithelial and immune cells in high-risk populations using this approach may be leveraged in the future to unveil cellular regulators of premature cervical remodeling. 1R01HD114611-01 (KDG)