Analysis of miRNA signatures in CSF identifies upregulation of miR-21 and miR-146a/b in patients with multiple sclerosis and active lesions.

BACKGROUND: MicroRNAs (miRNAs) have been reported as deregulated in active brain lesions derived from patients with multiple sclerosis (MS). In there, these post-transcriptional regulators may elicit very important effects but proper identification of miRNA candidates as potential biomarkers and/or therapeutic targets is scarcely available. OBJECTIVE: The aim of the study was to detect the presence of a set of candidate miRNAs in cell-free cerebrospinal fluid (CSF) and to determine their association with gadolinium-enhancing (Gd+) lesions in order to assess their value as biomarkers of MS activity. METHODS: Assessment of 28 miRNA candidates in cell-free CSF collected from 46 patients with MS (26 Gd+ and 20 Gd- patients) was performed by TaqMan assays and qPCR. Variations in their relative abundance were analyzed by the Mann-Whitney U test and further evaluated by receiver operating characteristic (ROC) analysis. Signaling pathways and biological functions of miRNAs were analyzed using bioinformatic tools (miRTarBase, Enrichr, REVIGO, and Cytoscape softwares). RESULTS: Seven out of 28 miRNA candidates were detected in at least 75% of CSF samples. Consistent increase of miR-21 and miR-146a/b was found in Gd+ MS patients. This increase was in parallel to the number of Gd+ lesions and neurofilament light chain (NF-L) levels. Gene Ontology enrichment analysis revealed that the target genes of these miRNAs are involved in biological processes of key relevance such as apoptosis, cell migration and proliferation, and in cytokine-mediated signaling pathways. CONCLUSION: Levels of miR-21 and miR-146a/b in cell-free CSF may represent valuable biomarkers to identify patients with active MS lesions.

miRNA Signature in CSF From Patients With Primary Progressive Multiple Sclerosis.

BACKGROUND AND OBJECTIVES: Primary progressive multiple sclerosis (PPMS) displays a highly variable disease progression with a characteristic accumulation of disability, what makes difficult its diagnosis and efficient treatment. The identification of microRNAs (miRNAs)-based signature for the early detection in biological fluids could reveal promising biomarkers to provide new insights into defining MS clinical subtypes and potential therapeutic strategies. The objective of this cross-sectional study was to describe PPMS miRNA profiles in CSF and serum samples compared with other neurologic disease individuals (OND) and relapsing-remitting MS (RRMS). METHODS: First, a screening stage analyzing multiple miRNAs in few samples using OpenArray plates was performed. Second, individual quantitative polymerase chain reactions (qPCRs) were used to validate specific miRNAs in a greater number of samples. RESULTS: A specific profile of dysregulated circulating miRNAs (let-7b-5p and miR-143-3p) was found downregulated in PPMS CSF samples compared with OND. In addition, in serum samples, miR-20a-5p and miR-320b were dysregulated in PPMS against RRMS and OND, miR-26a-5p and miR-485-3p were downregulated in PPMS vs RRMS, and miR-142-5p was upregulated in RRMS compared with OND. DISCUSSION: We described a 2-miRNA signature in CSF of PPMS individuals and several dysregulated miRNAs in serum from patients with MS, which could be considered valuable candidates to be further studied to unravel their actual role in MS. CLASSIFICATION OF EVIDENCE: This study provides Class II evidence that specific miRNA profiles accurately distinguish PPMS from RRMS and other neurologic disorders.

Inhibition of pro-inflammatory signaling in human primary macrophages by enhancing arginase-2 via target site blockers.

The modulation of macrophage phenotype from a pro-inflammatory to an anti-inflammatory state holds therapeutic potential in the treatment of inflammatory disease. We have previously shown that arginase-2 (Arg2), a mitochondrial enzyme, is a key regulator of the macrophage anti-inflammatory response. Here, we investigate the therapeutic potential of Arg2 enhancement via target site blockers (TSBs) in human macrophages. TSBs are locked nucleic acid antisense oligonucleotides that were specifically designed to protect specific microRNA recognition elements (MREs) in human ARG2 3' UTR mRNA. TSBs targeting miR-155 (TSB-155) and miR-3202 (TSB-3202) MREs increased ARG2 expression in human monocyte-derived macrophages. This resulted in decreased gene expression and cytokine production of TNF-α and CCL2 and, for TSB-3202, in an increase in the anti-inflammatory macrophage marker, CD206. Proteomic analysis demonstrated that a network of pro-inflammatory responsive proteins was modulated by TSBs. In silico bioinformatic analysis predicted that TSB-3202 suppressed upstream pro-inflammatory regulators including STAT-1 while enhancing anti-inflammatory associated proteins. Proteomic data were validated by confirming increased levels of sequestosome-1 and decreased levels of phosphorylated STAT-1 and STAT-1 upon TSB treatment. In conclusion, upregulation of Arg2 by TSBs inhibits pro-inflammatory signaling and is a promising novel therapeutic strategy to modulate inflammatory signaling in human macrophages.

Description of a CSF-Enriched miRNA Panel for the Study of Neurological Diseases.

BACKGROUND: The study of circulating miRNAs in CSF has gained tremendous attention during the last years, as these molecules might be promising candidates to be used as biomarkers and provide new insights into the disease pathology of neurological disorders. OBJECTIVE: The main aim of this study was to describe an OpenArray panel of CSF-enriched miRNAs to offer a suitable tool to identify and characterize new molecular signatures in different neurological diseases. METHODS: Two hundred and fifteen human miRNAs were selected to be included in the panel, and their expression and abundance in CSF samples were analyzed. In addition, their stability was studied in order to propose suitable endogenous controls for CSF miRNA studies. RESULTS: miR-143-3p and miR-23a-3p were detected in all CSF samples, while another 80 miRNAs were detected in at least 70% of samples. miR-770-5p was the most abundant miRNA in CSF, presenting the lowest mean Cq value. In addition, miR-26b-5p, miR-335-5p and miR-92b-3p were the most stable miRNAs and could be suitable endogenous normalizers for CSF miRNA studies. CONCLUSIONS: These OpenArray plates might be a suitable and efficient tool to identify and characterize new molecular signatures in different neurological diseases and would improve the yield of miRNA detection in CSF.

Assessing the presence of oligoclonal IgM bands as a prognostic biomarker of cognitive decline in the early stages of multiple sclerosis.

BACKGROUND: An association has been found between the presence of lipid-specific oligoclonal IgM bands (LS-OCMB) in cerebrospinal fluid and a more severe clinical multiple sclerosis course. OBJECTIVE: To investigate lipid-specific oligoclonal IgM bands as a prognostic biomarker of cognitive impairment in the early stages of multiple sclerosis. METHODS: Forty-four patients underwent neuropsychological assessment at baseline and 4 years. Cognitive performance at follow-up was compared adjusting by age, education, anxiety-depression, and baseline performance. RESULTS: LS-OCMB+ patients only performed worse for Long-Term Storage in the Selective Reminding Test (p = .018). CONCLUSION: There are no remarkable cognitive differences between LS-OCMB- and LS-OCMB+ patients in the early stages of MS.

Orphan CpG islands amplify poised enhancer regulatory activity and determine target gene responsiveness.

CpG islands (CGIs) represent a widespread feature of vertebrate genomes, being associated with ~70% of all gene promoters. CGIs control transcription initiation by conferring nearby promoters with unique chromatin properties. In addition, there are thousands of distal or orphan CGIs (oCGIs) whose functional relevance is barely known. Here we show that oCGIs are an essential component of poised enhancers that augment their long-range regulatory activity and control the responsiveness of their target genes. Using a knock-in strategy in mouse embryonic stem cells, we introduced poised enhancers with or without oCGIs within topologically associating domains harboring genes with different types of promoters. Analysis of the resulting cell lines revealed that oCGIs act as tethering elements that promote the physical and functional communication between poised enhancers and distally located genes, particularly those with large CGI clusters in their promoters. Therefore, by acting as genetic determinants of gene-enhancer compatibility, CGIs can contribute to gene expression control under both physiological and potentially pathological conditions.

Radiologically isolated syndrome: targeting miRNAs as prognostic biomarkers.

Aim: Some clinical and biological characteristics have been described as prognostic factors for clinical conversion into clinically definite multiple sclerosis in radiologically isolated syndrome (RIS) population. The aim of this study was to assess signatures of circulating miRNAs in those patients according to their conversion status after 5 years of follow-up. Patients & methods: OpenArray plates assessing 216 miRNA candidates were run in 15 RIS patients, and their relative abundances were analyzed. Results: A specific profile of deregulated circulating miRNAs (miR-144-3p, miR-448 and miR-653-3p in cerebrospinal fluid and miR-142-3p, miR-338-3p, miR-363-3p, miR-374b-5p, miR-424-5p, miR-483-3p in plasma) differentiated individuals who remained as RIS after 5 years of follow-up. Conclusion: Circulating miRNAs might be used as prognostic biomarkers for RIS patients.