Replication Data for: Refined single-cell profiling captures a CCR5high CD4+ cytotoxic T-cell precursor in multiple sclerosis

Due to patient privacy reasons, here we provide only processed Seurat objects from the single-cell sequencing data reported in this study.

Project background: T helper cells are deemed to play an important role in the onset of multiple sclerosis (MS). We earlier found that a subset that has features of both Th1 and Th17 cells, named Th17.1 cells, is enriched in the cerebrospinal fluid (CSF) of people with MS (Koetzier et al. Neurol N2 2020) and specifically accumulates in the blood upon treatment with natalizumab (blocking brain homing)(van Langelaar, Brain 2018).

Project aim: As Th17.1 also exist in healthy individuals and also have protective features, we aim to uncover pathogenic traits of Th17.1 cells in MS and how this pathogenic profile is impacted by natalizumab therapy.

Experimental design: For this project, we isolated Th17.1 cells and combined with total lymphocytes from the same person (HC or MS) in one 10x run, in a 1:2 ratio (approximately 5k Th17.1 cells and 10k lymphocytes per sample). We stained and barcoded total lymphocytes and Th17.1 respectively with two hashing antibodies. In addition, only the total lymphocyte populations were labeled with CITEseq antibodies against: CCR6, CXCR3, CCR4, CD20, CD8 and CD4. 5' scRNAseq runs were then performed on all samples using the 10x Genomics platform using Next-GEM v2 reagents, in collaboration with Dr. Eric Bindels from the department of Hematology at Erasmus MC.

We included 15 samples, this concerns 3 HC samples (CMS and HCPB samples), 6 untreated MS samples, and 6 MS post-natalizumab treatment (abbreviation “TYS”). The untreated and post-natalizumab samples are from the same patients (paired samples).