Background: During pregnancy, the mother undergoes immunologic adaptations to accommodate the semi-allogeneic fetus and there is growing evidence that placental extracellular vesicles contribute to the maternal adaptations in normal pregnancy. Many studies have shown that placental extracellular vesicles interact with immune cells in vitro but in our previous in vivo study, placental microvesicles did not localise to the spleen or thymus. So we examined whether placental microvesicles interact with circulating immune cells in vivo.
Methods: CellTracker™ Red Dye-labelled placental microvesicles were prepared by differential centrifugation and injected into non-pregnant (n=5) or pregnant CD1 female mice (n=7). After 30 minutes or 24 hours of exposure, mice were culled to harvest leukocytes to see whether they had interacted with microvesicles in vivo. For in vitro exposure, leukocytes were prepared from normal CD1 female mice and incubated with equivalent amount of the same microvesicles at 37˚C for the same time points (n=5). Leukocyte samples from in vivo and in vitro experiments were analysed by flow cytometry.
Results: In agreement with other studies, mouse leukocytes significantly interacted with placental microvesicles in vitro. The mean percentage of vesicle-containing cells was 0.07% and 0.76% for 30 minutes and 24 hours, respectively compared to 0.002% and 0.07% in the control groups (p=0.008). In contrast to in vitro incubation, we observed that only 0.002% of leukocytes interacted with placental microvesicles in non-pregnant mice in vivo at 30 minutes (p=0.0079) and only 0.004% of leucocytes interacted with microvesicles at 24 hours (p=0.008). However, in pregnant mice, 0.76% of leukocytes interacted with placental microvesicles after 24 hours of exposure compared to 0.004% in non-pregnant mice (p=0.0043). Preliminary analysis suggests that the leukocyte populations interacting with microvesicles were macrophages and lymphocytes (both B cells and T cells).
Conclusions: Mouse leukocytes significantly interacted with placental microvesicles in vitro but only minimal number of leukocytes interacted in vivo indicating the interactions are exaggerated in vitro. However, in pregnant mice, there was a stronger interaction between leukocytes and placental microvesicles compared to non-pregnant mice, suggesting that altered physiology in pregnancy increases the interaction of placental vesicles with immune cells.