Transcriptional profiling of mucus production in rhesus macaque endocervical cells under hormonal regulation

Abstract Objective Endocervical mucus production is a key regulator of fertility throughout the menstrual cycle. With cycle-dependent variability in mucus quality and quantity, cervical mucus can either facilitate or block sperm ascension into the upper female reproductive tract. This study seeks to identify genes involved in the hormonal regulation of mucus production, modification, and regulation through profiling the transcriptome of endocervical cells from the non-human primate, the rhesus macaque (Macaca mulatta). Intervention We treated differentiated primary endocervical cultures with estradiol (E2) and progesterone (P4) to mimic peri-ovulatory and luteal-phase hormonal changes. Using RNA-sequencing, we identified differential expression of gene pathways and mucus-producing and mucus-modifying genes in cells treated with E2 compared to hormone-free conditions and E2 compared to E2-primed cells treated with P4. Main Outcome Measures We pursued differential gene expression analysis on RNA-sequenced cells. Sequence validation was done using quantitative PCR (qPCR). Results Our study identified 158 genes that show significant differential expression in E2-only conditions compared to hormone-free control and 250 genes that show significant differential expression in P4-treated conditions compared to E2-only conditions. From this list, we found hormone-induced changes in transcriptional profiles for genes across several classes of mucus production, including ion channels and enzymes involved in post-translational mucin modification that have not previously been described as hormonally regulated. Conclusion Our study is the first to use an in vitro culture system to create an epithelial cell–specific transcriptome of the endocervix. As a result, our study identifies new genes and pathways altered by sex steroids in cervical mucus production. Summary Sentence In vitro hormonal regulation of mucus production, modification, and secretion was profiled using primary epithelial endocervical cells.