CITE-seq reveals inhibition of NF-kB pathway in B cells from vitamin D-treated multiple sclerosis patients

Vitamin D deficiency is a recognized risk factor for multiple sclerosis (MS) and has been associated with disease activity and progression. Vitamin D treatment has emerged as potentially protective, despite conflicting results from randomized controlled trials. Here, we used single-cell RNA-sequencing (scRNA-seq) combined with barcoded antibodies targeting surface markers (CITE-seq) to uncover candidate genes and pathways regulated in PBMC subpopulations from MS patients receiving high-dose vitamin D (n=5) or placebo (n=5). Best candidates were combined with genes involved in immune function and vitamin D metabolism for validation in a new cohort (n=8 in each group) by high-throughput quantitative polymerase chain reaction (HT-qPCR) in FACS-sorted naive CD4, Th1, Th17, Treg, naive CD8, memory and naive B cells, and MAIT cells. CITE-seq revealed no significant changes in the proportions of these subpopulations in response to vitamin D treatment. Out of the 92 candidate genes identified by CITE-seq, we validated differential expression of five genes (UXT, SNRPN, SUB1, GNLY and KLF6) using HT-qPCR. Furthermore, CITE-seq uncovered vitamin D-induced regulation of several pathways in naive and memory B cells, including MAPK, TLR and interleukin pathways, that may contribute to counteract Epstein-Barr virus (EBV)-induced resistance to apoptosis, notably through inhibition of the NF-{kappa}B pathway. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=119 SRC="FIGDIR/small/559400v1_ufig1.gif" ALT="Figure 1"> View larger version (30K): org.highwire.dtl.DTLVardef@1ad28c1org.highwire.dtl.DTLVardef@14ed135org.highwire.dtl.DTLVardef@188fea1org.highwire.dtl.DTLVardef@105353c_HPS_FORMAT_FIGEXP M_FIG C_FIG