Oral Presentation ESA-SRB 2023 in conjunction with ENSA

Human seminal fluid extracellular vesicles induce immune responses in female cervical cells in vitro (#41)

Cottrell T Tamessar 1 2 , Chishan Burch 1 2 , Tegan Bryde 1 2 , Shanu Parameswaran 1 2 , Judith Weidenhofer 3 , Hui-ming Zhang 4 , Sarah A Robertson 5 , Geoffry N De Iuliis 1 2 , Elizabeth G Bromfield 1 2 , David J Sharkey 5 , Brett Nixon 1 2 , John E Schjenken 1 2
  1. Priority Research Centre for Reproductive Science and School of Environmental and Life Sciences, The University of Newcastle, Newcastle, NSW, Australia
  2. Infertility and Reproduction Research Program, Hunter Medical Research Institute, New Lambton Heights, NSW, Australia
  3. School of Biomedical Sciences and Pharmacy, College of Health, Medicine and Wellbeing, University of Newcastle, Ourimbah, NSW, Australia
  4. Central Analytical Facility, Research and Innovation Division, The University of Newcastle, Newcastle, NSW, Australia
  5. Robinson Research Institute and Adelaide Medical School, University of Adelaide, Adelaide, South Australia, Australia

Seminal fluid (SF) interacts with epithelial cells lining the female reproductive tract to activate proinflammatory responses that facilitate pregnancy. Several SF factors, including transforming growth factor-β (TGF-β), have been identified as signalling agents but do not fully explain the female response. Seminal fluid extracellular vesicles (SFEVs) likely have signalling potential given their suggested roles in immune regulation; however, their signalling capacity remains elusive. To investigate the impact of SFEVs on female reproductive tract signalling, we utilised a well-established human ectocervical epithelial (Ect1) cell culture model. Human SFEVs were isolated from SF using density-gradient ultracentrifugation and incubated with Ect1 cells for 8 h prior to assessing gene expression profiles by transcriptomics (n=4/group) and qPCR (n=8-9/group). Untreated Ect1 cells were used as a control. Transcriptomic data were analysed using Ingenuity Pathway Analysis. Following 8 h co-incubation, SFEVs altered gene expression profiles in Ect1 cells (>1.5FC, 216 genes induced, 211 suppressed, FDR<0.05). Gene expression changes and signalling pathways were predominantly associated with the inflammatory response and were confirmed by qPCR. These included IL1A (p≤0.01, 2.6FC), IL6 (p≤0.01, 2.8FC) and CXCL2 (p≤0.01, 2.1FC), which were induced in Ect1 cells following SFEV treatment. Additional comparative analysis between SF and SFEV treated Ect1 cells indicated IL1A (SFEV:2.6FC, SF:2.1FC) and IL6 (SFEV:2.8FC, SF:3.0FC) were induced to equivalent levels in SFEV and SF treated cells. In contrast, genes such as CXCL2 (SFEV:2.1FC, SF:5.8FC) appeared primarily regulated by other components of SF. Bioinformatic prediction of SFEV signalling agents identified TGF-β (Z-score=4.8, p<0.05) and NFkB (Z-score=5.78, p<0.05) as potential SFEV cargo responsible for inducing changes in Ect1 gene expression. Our ongoing research delves into the potential delivery mechanisms of these signalling agents from SFEVs to Ect1 cells. This study highlights that SFEVs serve an important role in shaping the immune environment of the female reproductive tract to promote reproductive success.