a comprehensive study of vesicular and non vesicular mirnas from a volume of cerebrospinal fluid compatible with clinical practice

• DOI: 10.5281/zenodo.3605969
• Publication Date: 2019-01-01
• Authors: Juan José Lozano; Monika González; Africa Garcia-Orad; Javier Benito; Juan M. Falcón-Pérez; Juan M. Falcón-Pérez; Ana M. Aransay; Raquel Pérez Garay; Felix Royo; Endika Prieto-Fer...; Esperanza Gonzalez; Borja Santos-Zorr...; Nuria Macias-Camara
• Keywords: 0301 basic medicine; microrna; diagnosis; Exosomes; Real-Time Polymerase Chain Reaction; CSF exosomes; CSF miRNAs; clinical samples; infants; microRNA profiling; differential expression; Central Nervous System Neoplasms; CSF miRNAs; Extracellular Vesicles; 03 medical and health sciences; exosome; Humans; visualization; disease; 0303 health sciences; infants; Sequence Analysis, RNA; clinical samples traumatic brain-injury; microRNA profiling; glioblastoma; Infant, Newborn; Infant; Reproducibility of Results; 3. Good health; MicroRNAs; CSF exosomes; biomarker; extracellular vesicles; Research Paper

Cerebrospinal fluid (CSF) microRNAs (miRNAs) have emerged as potential biomarkers for minimally invasive diagnosis of central nervous system malignancies. However, despite significant advances in recent years, this field still suffers from poor data reproducibility. This is especially true in cases of infants, considered a new subject group. Implementing efficient methods to study miRNAs from clinically realistic CSF volumes is necessary for the identification of new biomarkers. Methods: We compared six protocols for characterizing miRNAs, using 200-µL CSF from infants (aged 0-7). Four of the methods employed extracellular vesicle (EV) enrichment step and the other two obtained the miRNAs directly from cleared CSF. The efficiency of each method was assessed using real-time PCR and small RNA sequencing. We also determined the distribution of miRNAs among different CSF shuttles, using size-exclusion chromatography. Results: We identified 281 CSF miRNAs from infants. We demonstrated that the miRNAs could be efficiently detected using only 200 µL of biofluid in case of at least two of the six methods. In the exosomal fraction, we found 12 miRNAs that might be involved in neurodevelopment. Conclusion: The Norgen and Invitrogen protocols appear suitable for the analysis of a large number of miRNAs using small CSF samples.