Background and aim: Through interaction with other cells, extracellular vesicles (EVs) released during malaria infection can transfer their cargo contributing to disease pathogenesis and cellular communication. Recent studies have suggested that EVs are an important source of biomarkers. MicroRNA are small non-coding RNAs that can control cellular metabolism, differentiation and development. EVs isolated from human plasma carry highly stable miRNAs.
We hypothesise that miRNA signature in EVs vary based on disease severity and is able of differentiating patients with complications, including neurocognitive impairment (NCI) observed in both severe malarial anaemia (SMA) and cerebral malaria (CM).
Methods: MicroRNAs present in EVs were analysed using next generation sequencing. Samples were prepared from 6 groups: controls (n=3), CM with full recovery (n=4), CM with fatal outcome (n=4), CM with NCI (n=4), SMA with full recovery (n=4) and SMA with NCI (n=4).
EVs were purified and RNA extracted from ~200µl of plasma (Uganda) using QIAgen® exoEasy and RNeasy kits. The miRNA library was prepared with a QIAseq™ miRNA Library Kit and sequenced on the Illumina miSeq. The data were analysed using Qiagen® CLC Genomics Workbench and differentially expressed miRNA identified.
Results and conclusion: In the various comparisons we found the following miR differentially expressed: (i) CM patients with full recovery versus CM with NCI and CM with fatal outcome: hsa-miR-206, hsa-miR-370-3p, hsa-miR-11400 and hsa-miR-93-3p; (ii) CM with fatal outcome versus CM with full recovery and CM with NCI: hsa-miR-3960; (iii) CM patients with NCI versus CM with full recovery and CM with fatal outcome: hsa-miR-19a-3p, hsa-miR-590-3p, and hsa-miR-1-3p.
We are currently performing pathway analysis to identify the most significant gene networks, and canonical pathways. This study shows for the first time the miRNA content of EVs in malaria patients and our results suggest that they could be used as potential markers of disease severity.