Next generation sequencing analysis of the single-cell transcriptomes of the T cell eclited by fecal phageome

By performing single-cell RNA sequencing (scRNA-seq) analysis on the T cells which stimulated by phageome from normal or severe acute pancreatitis mice, we seek to understand which T cells contribute to the inflammation in pancreatitis. t-distributed stochastic neighbor embedding (tSNE) identified 5 distinct T cell cluster, including effector CD8, naïve CD8, CD4, proliferating CD8 and stem like CD8. Stimulation by different phages did not change the T cell heterogeneity. Comparison of genes revealed a significant enrichment in the cell proliferation after SAP derived phages treatment. Differential gene expression analysis confirmed that SAP derived phages treatment skewed the cell pattern toward more IL-7r expression, while control derived phages treatment induce the IL-22 expression. Overall design: In order to precisely isolate T cells that respond to viral stimulation, an in vitro proximity labeling system FucoID was employed in which DCs were pre-treated with phages isolated from healthy or SAP mice, followed by co-cultured with T cells isolated from the gut of GF mice. Through this interaction-dependent labeling approach, DCs antigen presented T cells can be separated from bystander T cells based on their cell-surface enzymatic fucosyl-biotinylation and next analyzed by single-cell RNA sequencing.