Author Of 2 Presentations
P0136 - Profiling of small non-coding RNAs across cellular and biofluid compartments in patients with Multiple Sclerosis (ID 1195)
Abstract
Background
Small non-coding RNAs (sncRNAs) are important regulators of gene expression at the transcriptional and post-transcriptional levels in various biological contexts, such as regulation of immune system functions, homeostasis, and autoimmunity development. While the role of microRNAs (miRNAs) in multiple sclerosis (MS) has attained major interest, studies investigating other sncRNA classes, especially in the target central nervous system compartment, are still scarce.
Objectives
We aimed to perform a comprehensive, comparative analysis of all classes of sncRNAs in matching peripheral blood mononuclear cells (PBMCs), plasma, cerebrospinal fluid (CSF) cells, and cell-free CSF from MS patients and controls.
Methods
23 relapsing-remitting (RRMS, n=12 in relapse, n=11 in remission), 6 secondary progressive (SPMS) MS patients and 16 non-inflammatory and inflammatory neurological disease controls (NINDC, n=11; INDC, n=5) were included in the analysis. We utilized Small-seq (Faridani et al., Nat Biotechnol. 2016) to quantify sncRNA transcripts.
Results
We observed distinct and variable profiles of sncRNA classes across all analyzed cellular and biofluid compartments. While miRNAs were the most abundant class of sncRNAs in PBMCs and plasma, transfer RNAs represented the most abundant class in CSF cells and cell-free CSF. Furthermore, we observed an opposing quantitative pattern of small nuclear, nucleolar, transfer RNAs, and miRNAs changes between the blood and CNS compartments. In CSF cells, 133/133 and 115/117 differentially expressed sncRNAs were increased in RRMS relapse compared to remission and RRMS compared to NINDC, respectively. In contrast, 65/67 differentially expressed PBMC sncRNAs were decreased in RRMS compared to NINDC. These findings underscore the importance of including both the peripheral and intrathecal compartments in studies investigating the role of sncRNAs in MS.
Conclusions
Our findings demonstrate widespread alterations of several classes of sncRNAs, particularly during the relapse phase in CSF cells. Genome-wide small non-coding RNA profiling provides therefore an informative moleculer panel for addressing MS pathogenesis, where further research may lead to the identification of novel biomarkers and possible treatment targets.
P0956 - DNA methylation of cerebrospinal fluid cells in Multiple Sclerosis (ID 1193)
Abstract
Background
Multiple Sclerosis (MS) is a chronic inflammatory disease of a yet unknown cause characterized by autoimmune destruction of myelin and neurons in the central nervous system. Immune cells from the cerebrospinal fluid (CSF) can provide valuable insight into the pathogenic processes occurring in the typically inaccessible target organ; however, they have not been sufficiently utilized due to low numbers. DNA methylation is an epigenetic modification that without altering the genetic code can stably change the expression of genes and thus may play an important role in MS development. Moreover, due to stability, high specificity and the ability to be measured in limited amounts of material genome-wide, methylation changes may assist in studying MS pathogenesis.
Objectives
We aimed to establish and optimize a genome-wide methylation approach to investigate methylation changes that render immune cells pathogenic in MS with the prospect of better understanding disease pathogenesis.
Methods
Cells from the CSF of relapsing-remitting MS (RRMS n=5) and age- and sex-matched non-inflammatory controls (NINDC n=5) were extracted. Whole-genome bisulfite sequencing (WGBS) libraries were generated using the post-bisulfite adaptor tagging (PBAT) protocol, which originally was designed for single-cell and adapted to the limited cell amount as previously described (1). Results were replicated in a larger cohort (RRMS n=18 and NINDC n=7), while the reliability of the methodology was confirmed with technical replication of the original cohort.
Results
We first explored the global landscape of CpG methylation levels, as well as specific genomic features including promoters, enhancers and CTCF binding sites. We conducted statistical analysis based on three distinct methods, which identified differentially methylated CpG (DMCs). Methylation levels calculated in the sequencing data were well correlated with the independent technical replicates where the sequencing depths were comparatively lower which gave more evidence the less sequenced data can reflect the true values to some degree. Besides, in the replicate sample groups, a large proportion of DMCs changed in the same direction when applying a minimal 0.2 difference. Moreover, NFATC1, NFKBID, MAPK1, MAP2K2 and other genes participated in the regulation of the immune response were differentially modified. In addition, the differentially modified genes were significantly enriched in pathways including integrin signaling pathway which is important for cell movement and activation, CD40 signaling pathway which is related with the lymphocyte activation, as well as various cytokine signaling pathways such as IL-1, IL-6, IL-17.
Conclusions
Genome-wide methylation analysis, based on a low cell-number library preparation, will enable us to study MS pathogenesis using samples that are difficult to comprehensively study by many other methods.
Presenter Of 1 Presentation
P0136 - Profiling of small non-coding RNAs across cellular and biofluid compartments in patients with Multiple Sclerosis (ID 1195)
Abstract
Background
Small non-coding RNAs (sncRNAs) are important regulators of gene expression at the transcriptional and post-transcriptional levels in various biological contexts, such as regulation of immune system functions, homeostasis, and autoimmunity development. While the role of microRNAs (miRNAs) in multiple sclerosis (MS) has attained major interest, studies investigating other sncRNA classes, especially in the target central nervous system compartment, are still scarce.
Objectives
We aimed to perform a comprehensive, comparative analysis of all classes of sncRNAs in matching peripheral blood mononuclear cells (PBMCs), plasma, cerebrospinal fluid (CSF) cells, and cell-free CSF from MS patients and controls.
Methods
23 relapsing-remitting (RRMS, n=12 in relapse, n=11 in remission), 6 secondary progressive (SPMS) MS patients and 16 non-inflammatory and inflammatory neurological disease controls (NINDC, n=11; INDC, n=5) were included in the analysis. We utilized Small-seq (Faridani et al., Nat Biotechnol. 2016) to quantify sncRNA transcripts.
Results
We observed distinct and variable profiles of sncRNA classes across all analyzed cellular and biofluid compartments. While miRNAs were the most abundant class of sncRNAs in PBMCs and plasma, transfer RNAs represented the most abundant class in CSF cells and cell-free CSF. Furthermore, we observed an opposing quantitative pattern of small nuclear, nucleolar, transfer RNAs, and miRNAs changes between the blood and CNS compartments. In CSF cells, 133/133 and 115/117 differentially expressed sncRNAs were increased in RRMS relapse compared to remission and RRMS compared to NINDC, respectively. In contrast, 65/67 differentially expressed PBMC sncRNAs were decreased in RRMS compared to NINDC. These findings underscore the importance of including both the peripheral and intrathecal compartments in studies investigating the role of sncRNAs in MS.
Conclusions
Our findings demonstrate widespread alterations of several classes of sncRNAs, particularly during the relapse phase in CSF cells. Genome-wide small non-coding RNA profiling provides therefore an informative moleculer panel for addressing MS pathogenesis, where further research may lead to the identification of novel biomarkers and possible treatment targets.