The aims of the study were (1) to determine whether miRNAs (microRNAs) can be detected in the cerebrospinal fluid (CSF) and blood of patients with ischemic stroke and (2) to compare these miRNA profiles with corresponding profiles from other neurological patients to address whether the miRNA profiles of CSF or blood have potential usefulness as diagnostic biomarkers of ischemic stroke. CSF from patients with acute ischemic stroke (n = 10) and patients with other neurological diseases (n = 10) was collected by lumbar puncture. Blood samples were taken immediately after. Expression profiles in the cell-free fractions of CSF and blood were analyzed by a microarray technique (miRCURY LNA™ microRNA Array, Exiqon A/S, Denmark) using a quantitative PCR (qPCR) platform containing 378 miRNA primers. In total, 183 different miRNAs were detected in the CSF, of which two miRNAs (let-7c and miR-221-3p) were found upregulated in relation to stroke. In the blood, 287 different miRNAs were detected of which two miRNAs (miR-151a-3p and miR-140-5p) were found upregulated and one miRNA (miR-18b-5p) was found downregulated in the stroke group. Some miRNAs occurred exclusively in the CSF including miR-523-3p which was detected in 50 % of the stroke patients, whereas it was completely absent in controls. Our preliminary results demonstrate that it is possible to detect and profile miRNAs in CSF and blood from patients with neurological diseases. Some miRNAs appear differentially expressed in the CSF and others in the blood of stroke patients. Currently, we are validating our results in larger groups of patients.

BackgroundMicroRNAs (miRNAs) are small non-coding RNA molecules that function as posttranscriptional regulators of gene expression. Measurements of miRNAs in cerebrospinal fluid (CSF) and blood have just started gaining attention as a novel diagnostic tool for various neurological conditions. The purpose of this exploratory investigation was to analyze the expression of miRNAs in CSF and blood of patients with Alzheimer’s disease (AD) and other neurodegenerative disorders in order to identify potential miRNA biomarker candidates able to separate AD from other types of dementia.MethodsCSF was collected by lumbar puncture performed on 10 patients diagnosed with AD and 10 patients diagnosed with either vascular dementia, frontotemporal dementia or dementia with Lewy bodies. Blood samples were taken immediately after. Total RNA was extracted from cell free fractions of CSF and plasma, and a screening for 372 known miRNA sequences was carried out by real time quantitative polymerase chain reactions (miRCURY LNA™ Universal RT miRNA PCR, Polyadenylation and cDNA synthesis kit, Exiqon).ResultsFifty-two miRNAs were detected in CSF in at least nine out of ten patients in both groups. Among these, two miRNAs (let-7i-5p and miR-15a-5p) were found significantly up-regulated and one miRNA (miR-29c-3p) was found significantly down-regulated in patients with AD compared to controls. One hundred and sixty-eight miRNAs were frequently detected in the blood, among which miR-590-5p and miR-142-5p were significantly up-regulated and miR-194-5p was significantly down-regulated in AD patients compared to controls.ConclusionsDetection of miRNA expression profiles in blood and in particular CSF of patients diagnosed with different types of dementia is feasible and it seems that several expressional differences between AD and other dementia types do exist when measured in a clinically relevant setup. In this explorative pilot study, the deregulated miRNAs in CSF of AD patients may be associated with relevant target genes related to AD pathology, including APP and BACE1, which suggests that miRNAs are interesting candidates for AD biomarkers in the future.Electronic supplementary materialThe online version of this article (doi:10.1186/s40035-016-0053-5) contains supplementary material, which is available to authorized users.

BackgroundThe purpose of this study was to investigate the potential of cerebrospinal fluid miRNAs as diagnostic biomarkers of acute ischemic stroke using three different profiling techniques in order to identify and bypass any influence from technical variation.MethodsCerebrospinal fluid (CSF) from patients with acute ischemic stroke (n = 21) and controls (n = 21) was collected by lumbar puncture. miRNA analysis was performed with three different methods: 1) Trizol RNA extraction followed by Illumina Next Generation Sequencing (NGS) on all small RNAs, 2) Exiqon RNA extraction protocol and miRNA qPCR assays, and 3) validation of 24 selected miRNAs with Norgen Biotek RNA extraction protocol and Applied Biosystems qPCR assays.ResultsNGS detected 71 frequently expressed miRNAs in CSF of which brain-enriched miR-9-5p and miR-128-3p were significantly higher in CSF of stroke patients compared to controls. When dividing stroke patients into groups according to infarct size several brain-enriched miRNAs (miR-9-5p, miR-9-3p, miR-124-3p, and miR-128-3p) were elevated in patients with infarcts >2 cm3. This trend appeared in data from both NGS, qPCR (Exiqon), and qPCR (Applied Biosystems) but was only statistically significant in some of the measurement platforms.ConclusionsSeveral brain-enriched miRNAs are elevated in the CSF three days after stroke onset, suggesting that these miRNAs reflect the brain damage caused by ischemia. The expression differences seem, however, limited to patients with larger ischemic brain injury, which argues against the use of CSF miRNAs as diagnostic biomarkers of stroke based on current methods.Electronic supplementary materialThe online version of this article (doi:10.1186/s40364-017-0104-9) contains supplementary material, which is available to authorized users.

Comparison of two methods to detect circulating tumor cells (CTC) CytoTrack and CellSearch through recovery of MCF-7 breast cancer cells, spiked into blood collected from healthy donors. Spiking of a fixed number of EpCAM and pan-cytokeratin positive MCF-7 cells into 7.5 mL donor blood was performed by FACSAria flow sorting. The samples were shipped to either CytoTrack or CellSearch research facilities within 48 h, where evaluation of MCF-7 recovery was performed. CytoTrack and CellSearch analyses were performed simultaneously. Recoveries of MCF-7 single cells, cells in clusters, and clusters were determined. The average numbers of MCF-7 cells/cells in clusters/clusters recovered from blood by the CytoTrack and CellSearch methods were 103 ± 5.9/27 ± 7.9/11 ± 3.5 (95 % CI) and 107 ± 4.4/20 ± 7.1/10 ± 3.5, respectively, with no difference between the two methods (p = 0.37/p = 0.23/p = 0.09). Overall, the recovery of CytoTrack and CellSearch was 68.8 ± 3.9 %/71.1 ± 2.9 %, respectively (p = 0.58). In spite of different methodologies, CytoTrack and CellSearch found similar number of CTCs, when spiking was performed with the EpCAM and pan cytokeratin-positive cell line MCF-7. The results suggest that CytoTrack and CellSearch have similar abilities to identify CTC in vitro.