Insulin exposed endometrial epithelial cells cultured in a microfluidic device alters transcripts involved in translation that may contribute to reduced implantation capacity of the endometrium

Abstract

ABSTRACTObesity is a rapidly growing public health issue among women of reproductive age. It is also associated with decreased reproductive function including implantation failure. Implantation failure can result from a myriad of factors including impaired gametes and endometrial dysfunction. The mechanisms of how obesity-related hyperinsulinaemia disrupts endometrial function and implantation are poorly understood. Our study aims to investigate potential mechanisms by which insulin alters endometrial transcript expression, which may affect endometrial receptivity. Ishikawa cells mimicking human endometrial epithelium were seeded into a microfluidics organ-on-chip device to produce an in vitro endometrium. Syringe pump was attached to the microfluidics device to deliver three varying treatments into Ishikawa cells: 1) media control 2) vehicle control (PBS acidified to pH3 with acetic acid) 3) Insulin (2mg/mL) at a constant flow rate of 1uL/min for 24 hours to mimic secretion in vivo. Three biological replicates were obtained. Insulin-induced transcriptomic response of the in vitro endometrium was quantified via RNA sequencing, and subsequently analysed using DAVID and Webgestalt to identify Gene Ontology (GO) terms and signalling pathways. A Total of 29 transcripts showed differential expression levels across two comparison groups (control v vehicle control; vehicle control v insulin). There were nine transcripts significantly differentially expressed in vehicle control v insulin group (p<0.05). Functional annotation analysis of transcripts altered by insulin (n=9) identified three significantly enriched GO terms: SRP-dependent cotranslational protein targeting to membrane, poly(A) binding, and RNA binding (p<0.05). Over-representation analysis found three significantly enriched signalling pathways relating to insulin-induced transcriptomic response: protein export, glutathione metabolism, and ribosome pathways (p<0.05). Insulin-induced dysregulation of biological functions and pathways highlight potential mechanisms by which high insulin concentrations within maternal circulation may perturb endometrial receptivity.