Poster Presentation ESA-SRB 2023 in conjunction with ENSA

Ingenuity Pathway Analysis reveals novel pathways in adult mouse Leydig cells that are altered by in vivo LHCGR activation. (#395)

Liza O'Donnell 1 , Anne-Louise Gannon 2 , Ben Lawrence 2 , Isabelle Stevant 3 , Pauline Sararols 3 , Serge Nef 3 , Diane Rebourcet 2 , Lee B Smith 1
  1. Griffith University, Southport, Queensland, Australia
  2. The University of Newcastle, Callaghan, NSW, Australia
  3. University of Geneva, Geneva, Switzerland

Steroidogenesis in adult Leydig cells (ALC) is stimulated by luteinising hormone receptor (LHCGR) activation. Many studies have investigated LHCGR activation in ALC but most use models that do not recapitulate the in vivo interstitial ALC niche which includes direct contacts with macrophages. To address this, we analysed ALC transcription during acute LHCGR activation in vivo and identified the transcriptional pathways that are regulated. Highly purified mouse ALC were isolated1 after in vivo administration of vehicle or LHCGR agonist (human chorionic gonadotrophin, hCG) for 2 or 6 hours (h). RNASeq identified differentially expressed genes (DEG) altered by hCG and bioinformatic analyses including Ingenuity Pathway Analysis® (IPA) mapped the functional responses of ALC. Most known pathways associated with LHCGR activation2 were altered by 2h. IPA revealed previously unrecognised regulators and pathways responsive to LHCGR activation. hCG acutely altered the production of, or response to, various cytokines. For example, IL6 was highly stimulated, and IL6-responsive DEGs were enriched in extracellular proteins that likely mediate interactions with interstitial macrophages. Novel LHCGR transcriptional targets were identified, such as activation of HIF1A-mediated transcription involving repression of its negative regulator, CITED2. Crosstalk between LHCGR and estrogen receptor signalling was also identified. hCG induced acute changes in genes encoding TGFβ family receptors, integrins and focal adhesion complexes, suggesting cell-cell interactions within the niche are integrally linked to ALC steroidogenic function. Acute stimulation of LHCGR downregulated the glucocorticoid receptor (GR) which mediates transcription of many genes important for ALC function, suggesting that steroidogenesis requires repression of a subset of GR-dependent transcription, possibly explaining why both excess and reduced GR signalling suppress steroidogenesis. This study reveals novel pathways in steroidogenically-active ALC that could be exploited for the design of therapeutic strategies to support ALC androgen production.

1Sararols et al. 2021, Front Cell Dev Biol, 9:695546        2Tremblay. 2015, Steroids, 103:3