Background: Studies have shown circulating miRNA dysregulation in pre-eclampsia, however difficulties in reproducibility have limited their use as biomarkers. Exosomal miRNA are a promising avenue to identify reproducibly dysregulated miRNA biomarkers for pre-eclampsia.
Aims: To optimise exosome isolation from maternal serum in pregnancy and examine serum-derived exosomal miR-1 and miR-210 dysregulation in pre-eclampsia.
Methods: Exosome yields from maternal serum isolated by ultracentrifugation and ExoQuickTM isolation methods were compared using electron microscopy and BCA assay-measured protein content. Exosomal RNA was extracted using a modified and manufacturer's mirVanaTM PARISTM protocol and TRIzol(R) and SeraMirTM protocols. qRT-PCR was performed on SeraMirTM-extracted RNA for U6 small nuclear RNA, miR-1 and miR-210.
Results: ExoQuickTM isolated the highest exosome yield as indicated by electron microscopy visualisation and protein content; 11215 [mu]g vs ultracentrifugation <= 12 [mu]g. SeraMirTM extracted the highest and only exosomal RNA concentration above the limit of detection (15.7 ng/[mu]L). No amplification products were detected by qRT-PCR.
Discussion: This is the first study to isolate serum-derived exosomes and exosomal RNA in pregnancy and provides a workflow to discover exosomal miRNA biomarkers for pre-eclampsia and other pregnancy complications. Findings suggest more sensitive PCR techniques; pre-amplification or digital PCR, are required to demonstrate exosomal miRNA dysregulation and discover these biomarkers.
(C) 2015 Royal College of Pathologists of Australasia