ExoCounter Assays Identify Women Who May Develop Early-Onset Preeclampsia From 12.5 μL First-Trimester Serum by Characterizing Placental Small Extracellular Vesicles

Abstract

BACKGROUND: Preeclampsia is a major cause of maternal and perinatal morbidity and mortality worldwide. Identifying women with high risk of developing preeclampsia in early pregnancy remains challenging. Extracellular vesicles released from the placenta offer an attractive biomarker but have been elusive to quantify. METHODS: Here, we tested ExoCounter, a novel device that immunophenotypes size-selected small extracellular vesicles <160 nm, for its ability to perform qualitative and quantitative placental small extracellular vesicles (psEV) analysis. To investigate disease-specific and gestational age-specific changes, we analyzed psEV counts in maternal plasma samples taken at each of the 3 trimesters from women who had (1) normal pregnancy (n=3); (2) women who developed early-onset preeclampsia ([EOPE], n=3); and (3) women who developed late-onset preeclampsia (n=4) using 3 antibody pairs, CD10-placental alkaline phosphatase (PLAP), CD10-CD63, and CD63-PLAP. We further validated the findings in first-trimester serum samples among normal pregnancy (n=9), women who developed EOPE (n=7), and women who developed late-onset preeclampsia (n=8). RESULTS: We confirmed that CD63 was the major tetraspanin molecule coexpressed with PLAP—a known placental extracellular vesicles marker on psEV. Higher psEV counts for all 3 antibody pairs were detected in the plasma of women who developed EOPE than the other 2 groups in the first trimester, which persisted through the second and third trimesters. Significantly higher CD10-PLAP ( P <0.01) and CD63-PLAP ( P <0.01) psEV counts were validated in the serum of the first trimester of women who developed EOPE compared with normal pregnancy. CONCLUSIONS: Application of the ExoCounter assay developed here could identify patients at risk of developing EOPE in the first trimester, thereby providing a window of opportunity for early intervention.

Association between Plasma HLA-DR+ Placental Vesicles and Preeclampsia: A Pilot Longitudinal Cohort Study

“…In this study, we demonstrated that levels of PLAP+ STEVs increase from the first to the second trimester, likely due to the increase in placental size from the first to the second trimester, then decrease from the second to the third trimester. This observation is consistent with a recent study analyzing circulating STEVs from the sixth to twelfth weeks of gestation [14] but does not confirm a previous observation made in a small population of pregnant women suggesting a linear increase in circulating STEVs from the first to third trimester [15]. Such a discrepancy might be related to the different purification methods used to isolate EVs and thus to the possibly different populations of EVs obtained in terms of size and content.…”

“…Such a discrepancy might be related to the different purification methods used to isolate EVs and thus to the possibly different populations of EVs obtained in terms of size and content. Indeed, in our study and in that by Jiang and co-authors, the purification method for EVs was only ultracentrifugation [14], while in other reports, the purification methods were both ultracentrifugation and concentration gradient [15,16]. Further studies are required to assess whether the two different methods of EV isolation are equivalent or add to the collection of different EV populations.…”

Syncytiotrophoblast extracellular vesicles contain functional kynurenine metabolising enzymes: potential implications for preeclampsia

“…STB-EVs were obtained from placentae using a modified ex-vivo dual lobe placental perfusion and differential ultracentrifugation method, as previously detailed by our group [32][33][34][35] . Placentae were subjected to a 3-hour perfusion process, after which the maternal side perfusate was collected and promptly centrifuged twice using a Beckman Coulter Avanti J-20XP centrifuge and a Beckman Coulter JS-5.3 swing-out rotor.…”