Data för: Persistent and Compartmentalized T cell Immunity in ALS Data for: Persistent and Compartmentalized T cell Immunity in ALS 2026-229-1 https://doi.org/10.48723/cypd-qp27 Swedish National Data Service Svensk nationell datatjänst Landing page Data för: Persistent and Compartmentalized T cell Immunity in ALS Data for: Persistent and Compartmentalized T cell Immunity in ALS 2026-229-1 https://doi.org/10.48723/cypd-qp27 Andersson, John Andersson, John 802091 802091 R01TS000348 R01TS000348 2023-02428_VR 2023-02428_VR Swedish National Data Service Svensk nationell datatjänst Landing page Amyotrophic Lateral Sclerosis Amyotrofisk lateralskleros Sequence Analysis, RNA RNA-sekvensanalys Neurodegenerative Diseases Neurodegenerativa sjukdomar Neurogenic Inflammation Neurogen inflammation In this project, we aimed to understand the activation requirements and temporal dynamics of T cells in the peripheral and intrathecal compartments in amyotrophic lateral sclerosis (ALS). The present data is on single-cell sequencing isolated from human cerebrospinal fluid (CSF) cells and blood samples from newly diagnosis ALS patients (n=17) and ALS follow up (n=4). This analysis was conducted as part of a bigger project which is summarized in the section below. Using single-cell RNA and TCR sequencing in combination with imaging flow cytometry of cells derived from cerebrospinal fluid (CSF) and blood, we identified increased amount of antigen-presenting cell:T cell complexes in CSF, suggesting local activation. Clonal T cell expansion in CSF was observed with high inter-patient variability, indicating immunological heterogeneity among ALS patients. Notably, CMV-specific T cells in CSF, but not blood, positively correlated with the rate of disease progression, implicating a chronic viral component in ALS. Longitudinal flow cytometric analysis revealed shifts in T cell composition after ALS diagnosis, with peripheral T cell expansion linked to faster progression, whereas increased CSF T cell levels correlated with better prognosis. These findings provide critical insights into ALS immunopathology and lay the groundwork for developing targeted immunotherapies. The dataset consists of 248 GNU zipped FASTQ files. The total size of the dataset is approximately 1.1 TB. Detta projekt syftade till att förstå aktiveringskraven och den tidsmässiga dynamiken hos T-celler i de perifera och intratekala kompartmenten vid amyotrofisk lateralskleros (ALS). Föreliggande data avser encellssekvensering isolerad från humana cerebrospinalvätska (CSF)-celler och blodprover från nydiagnostiserade ALS-patienter (n=17) och ALS-uppföljning (n=4). Denna analys genomfördes som en del av ett större projekt som sammanfattas i avsnittet nedan. Med hjälp av encells-RNA och TCR-sekvensering i kombination med avbildningsflödescytometri av celler härledda från cerebrospinalvätska (CSF) och blod identifierade vi en ökad mängd antigenpresenterande cell:T-cellskomplex i CSF, vilket tyder på lokal aktivering. Klonal T-cellsexpansion i CSF observerades med hög interpatientvariabilitet, vilket indikerar immunologisk heterogenitet bland ALS-patienter. Det är värt att notera att CMV-specifika T-celler i CSF, men inte blod, korrelerade positivt med sjukdomsprogressionshastigheten, vilket implicerar en kronisk viral komponent i ALS. Longitudinell flödescytometrisk analys visade förändringar i T-cellsammansättningen efter ALS-diagnos, med perifer T-cellsexpansion kopplad till snabbare progression, medan ökade CSF T-cellsnivåer korrelerade med bättre prognos. Dessa fynd ger viktiga insikter i ALS-immunopatologi och lägger grunden för att utveckla riktade immunterapier. Datasetet består av 248 GNU-zippade FASTQ-filer. Datasetets totala storlek uppgår till ca 1,1 TB Sweden Sverige IndividualIndividual IndividIndivid Participants were recruited through the ALS Clinical Research Centre of Karolinska University Hospital in Stockholm, Sweden Deltagarna rekryterades genom ALS Clinical Research Centre på Karolinska Universitetssjukhuset i Stockholm, Sverige. Numeric Text Sample collection and processing Fresh CSF and blood samples were collected from ALS patients at the time of diagnosis for scRNAseq analysis. Blood samples (3.5 mL) were collected in EDTA vacutainers (BD), and CSF (20 mL) was collected via lumbar puncture using two 10 mL plastic tubes (Sarstedt). Samples were transported and processed within 2 hours of collection. CSF samples were centrifuged at 300×g for 10 mins at 4 °C. The supernatant was carefully removed without disturbing the cell pellet, which was resuspended in 50 µL PBS containing 0.04% bovine serum albumin (BSA). Peripheral blood mononuclear cells (PBMCs) were isolated using the prepTM (STEMCELL) protocol, following the manufacturer’s instructions. After centrifugation, PBMCs from the interface layer were collected and washed twice in 15 mL PBS supplemented with 0.04 % BSA. Cell counts were determined using a Fuchs-Rosenthal chamber (Biosigma) with 5 µL of single-cell suspension. CSF samples with visible blood contamination or >200 RBCs/µL were excluded. A total of 10,000 PBMCs and all available CSF cells per patient were used for library preparation. Library preparation and sequencing Single-cell suspensions were loaded onto Chromium Next GEM Chip K, and gel beads-in-emulsion (GEMs) were generated using the Chromium iX instrument (10x Genomics) according to the Chromium Next GEM Single Cell 5’ Reagent Kits v2 (Dual Index) protocol. cDNA amplification and library preparation were performed according to the manufacturer’s instructions, generating 5’ primer gene expression libraries and V(D)J libraries. Gene expression (GEX) libraries were pooled and sequenced at a target depth of ~ 50,000 reads per cell, while V(D)J libraries were sequenced at ~ 5,000 reads per cell using the Illumina NovaSeq6000 platform at Eukaryotic Single Cell Genomics (ESCG), SciLifeLab, Karolinska Institutet. The data set contains information from 21 individuals (17 ALS + 4 follow ups) using single cell RNA sequencing in combination with TCR V(D)J sequencing to study the immune profile of the central nervous compartment (CSF) and blood. Sequencing was done using 10x Genomics platform (5’ scRNAseq & V(D)J Reagent Kits v 2). 5’ and TCR libraries were then pooled and for sequencing on the NovaSeq sequencer. Provided files are in .fastq.gz format and per individual four files are available (Read 1&2 and Lane 1&2).Sample collection and processing Fresh CSF and blood samples were collected from ALS patients at the time of diagnosis for scRNAseq analysis. Blood samples (3.5 mL) were collected in EDTA vacutainers (BD), and CSF (20 mL) was collected via lumbar puncture using two 10 mL plastic tubes (Sarstedt). Samples were transported and processed within 2 hours of collection. CSF samples were centrifuged at 300×g for 10 mins at 4 °C. The supernatant was carefully removed without disturbing the cell pellet, which was resuspended in 50 µL PBS containing 0.04% bovine serum albumin (BSA). Peripheral blood mononuclear cells (PBMCs) were isolated using the prepTM (STEMCELL) protocol, following the manufacturer’s instructions. After centrifugation, PBMCs from the interface layer were collected and washed twice in 15 mL PBS supplemented with 0.04 % BSA. Cell counts were determined using a Fuchs-Rosenthal chamber (Biosigma) with 5 µL of single-cell suspension. CSF samples with visible blood contamination or >200 RBCs/µL were excluded. A total of 10,000 PBMCs and all available CSF cells per patient were used for library preparation. Library preparation and sequencing Single-cell suspensions were loaded onto Chromium Next GEM Chip K, and gel beads-in-emulsion (GEMs) were generated using the Chromium iX instrument (10x Genomics) according to the Chromium Next GEM Single Cell 5’ Reagent Kits v2 (Dual Index) protocol. cDNA amplification and library preparation were performed according to the manufacturer’s instructions, generating 5’ primer gene expression libraries and V(D)J libraries. Gene expression (GEX) libraries were pooled and sequenced at a target depth of ~ 50,000 reads per cell, while V(D)J libraries were sequenced at ~ 5,000 reads per cell using the Illumina NovaSeq6000 platform at Eukaryotic Single Cell Genomics (ESCG), SciLifeLab, Karolinska Institutet. The data set contains information from 21 individuals (17 ALS + 4 follow ups) using single cell RNA sequencing in combination with TCR V(D)J sequencing to study the immune profile of the central nervous compartment (CSF) and blood. Sequencing was done using 10x Genomics platform (5’ scRNAseq & V(D)J Reagent Kits v 2). 5’ and TCR libraries were then pooled and for sequencing on the NovaSeq sequencer. Provided files are in .fastq.gz format and per individual four files are available (Read 1&2 and Lane 1&2). Access to data through SND. Access to data is restricted. Åtkomst till data via SND. Tillgång till data är begränsad. restrictedAccess Zhenzhen Chen, Jannika Mansikkaviita, Christina Seitz, Solmaz Yazdani, Joonas Lehikoinen, Elie Deeba, Anikó Lovik, Marton Keszei, Hans Grönlund, Hannu Laaksovirta, Fang Fang, Pentti J. Tienari, Caroline Ingre, John Andersson. 1. Unit of Integrative Epidemiology, Institute of Environmental Medicine, Karolinska Institutet, Stockholm, Sweden 2. Translational Immunology Research Program, University of Helsinki, Helsinki, Finland. 3. Neurology Clinic, Karolinska University Hospital, Stockholm, Sweden 4. Department of Clinical Neuroscience, Karolinska Institutet, Stockholm, Sweden 5. Methodology & Statistics Unit, Leiden University, Netherlands 6. The Public Health Agency of Sweden, Stockholm, Sweden 7. Department of Neurology, Brain Center, Helsinki University Hospital, Helsinki, Finland. Zhenzhen Chen, Jannika Mansikkaviita, Christina Seitz, Solmaz Yazdani, Joonas Lehikoinen, Elie Deeba, Anikó Lovik, Marton Keszei, Hans Grönlund, Hannu Laaksovirta, Fang Fang, Pentti J. Tienari, Caroline Ingre, John Andersson. 1. Unit of Integrative Epidemiology, Institute of Environmental Medicine, Karolinska Institutet, Stockholm, Sweden 2. Translational Immunology Research Program, University of Helsinki, Helsinki, Finland. 3. Neurology Clinic, Karolinska University Hospital, Stockholm, Sweden 4. Department of Clinical Neuroscience, Karolinska Institutet, Stockholm, Sweden 5. Methodology & Statistics Unit, Leiden University, Netherlands 6. The Public Health Agency of Sweden, Stockholm, Sweden 7. Department of Neurology, Brain Center, Helsinki University Hospital, Helsinki, Finland.