Exosome-like vesicles facilitate intercellular communication between uterine artery smooth muscle cells and perivascular adipose tissue

Introduction Perivascular adipose tissue (PVAT) regulates uterine artery tone during pregnancy. However, the mechanisms underlying its functional role in uterine arteries is unknown. Exosomes are cargo carrying membrane-bound extracellular vesicles used in intercellular communication. It is unknown whether PVAT secretes exosomes. We hypothesized that uterine PVAT sheds exosomes (Exo-PVAT) that are uptaken by neighboring uterine vascular smooth muscle cells (USMCs). Methods Exo-PVAT were isolated and purified with tissue culture and ultracentrifugation, and primary USMCs were isolated using enzymatic digestion from pregnant and non-pregnant rats. Exosome protein content, size and molecular weight were determined via western blot, Malvern Zetasizer and fast protein liquid chromatography (FPLC), respectively. To determine USMC uptake of Exo-PVAT, Exo-PVAT were labelled with a membrane-labeling dye and co-cultured with USMCs for 3 hours. Results Exo-PVAT expressed TSG101, Alix, and CD9. Pregnancy did not affect Exo-PVAT size [Median(IQR) (nm), Non-pregnant: 99.4 (80.5) vs. Pregnant: 46.7 (21.2), p=0.5]. Using FPLC, we identified exosomes of 40-200 kDa in samples from both pregnant and non-pregnant rats. PVAT from pregnant rats secreted a relatively high amount of exosomes of 2000 kDa compared to non-pregnat rats. Immunocytochemical assessments revealed that USMCs took up exosomes derived from their adjacent PVAT. Conclusion Uterine PVAT sheds exosome-like vesicles which are uptaken by adjacent USMCs. The interaction between Exo-PVAT and USMCs is a novel type of intercellular communication that may have important implications in uterine artery function and blood flow in pregnancy.