Project description:T cells infiltrate pancreatic islets during the progression of type 1 diabetes (T1D) but their differentiation states have not been completely defined. We used unbiased single-cell RNA sequencing analyses to gain further insight into the phenotypic complexity of islet-infiltrating T cells in non-obese diabetic (NOD) mice.
Project description:In type 1 diabetes (T1D) autoreactive CD8 T cells infiltrate pancreatic islets and destroy insulin-producing β cells. Progression to T1D onset is a chronic process, which suggests that the effector activity of β-cell autoreactive CD8 T cells needs to be maintained throughout the course of disease development. The mechanism that sustains diabetogenic CD8 T cell effectors during the course of T1D progression has not been completely defined. Here we used single-cell RNA sequencing to gain further insight into the phenotypic complexity of islet-infiltrating CD8 T cells in NOD mice. We identified two functionally distinct subsets of activated CD8 T cells, CD44highTCF1+CXCR6- and CD44highTCF1-CXCR6+, in islets of prediabetic NOD mice. Compared to CD44highTCF1+CXCR6- CD8 T cells, the CD44highTCF1-CXCR6+ subset expressed higher levels of inhibitory and cytotoxic molecules and was more prone to apoptosis. Adoptive cell transfer experiments revealed that CD44highTCF1+CXCR6- CD8 T cells, through continuous generation of the CD44highTCF1-CXCR6+ subset, were more capable than the latter population to promote insulitis and the development of T1D. We further showed that direct interleukin-27 (IL-27) signaling in CD8 T cells promoted the generation of terminal effectors from the CD44highTCF1+CXCR6- population. These results indicate that islet CD44highTCF1+CXCR6- CD8 T cells are a progenitor-like subset with self-renewing capacity and under an IL-27 controlled mechanism they differentiate into the CD44highTCF1-CXCR6+ terminal effector population. Our study provides new insight into the sustainability of the CD8 T cell response in the pathogenesis of T1D.
Project description:Pancreas specific deletion of the Haster promoter region results in a variegated phenotype in pancreatic islets with overexpression or silencing of the Hnf1a gene. To determine the transcriptional consequence of the overexpression or silencing of Hnf1a is islet cells from the Haster pKO mice (Haster loxP/loxP;Pdx1-Cre), we performed scRNA-seq of pancreatic islets from control and adult female Haster pKO mice.
Project description:The transcriptome of pancreatic islets from four mouse strains was compared - NODk.RagKO, B10k.RagKO, NODk.RagKO.insHEL and B10k.RagKO.insHEL. This comparison allows the assessment of altered transcriptome due to two factors - genetic background of NOD vs B10 strain, and the effect of the insHEL transgene. The use of the MHC k haplotype and the RagKO background remove contamination from infiltrating immune cells, in order to obtain a pure islet transcriptome. Three mice of each gentoype were analyzed in this study.
Project description:To comprehensively characterize the changes within the TME during TREM1 deficiency and anti-PD-1 immune checkpoint blockade therapy, we performed scRNA-seq analysis of the CD45+ TICs in melanoma-bearing C57BL/6 mice receiving the various treatments. We analyzed approximately 8,249 CD45+ cells from the treatment groups with t-SNE analysis, identifying 10 distinct clusters of tumor-infiltrating immune cells
Project description:We performed an in vitro co-culture of allogeneic PBMCs and SC-islet clusters. SC-islet clusters were enriched for β cells (using CD49A magnetic sorting; SC-α still remain at lower numbers) (Veres et al., 2019), dissociated and reaggregated to obtain a more uniform cell count between wells. SC-islets were co-cultured with human allogeneic PBMCs for 24 or 48 hours. As controls [time(t)=0], SC-islets remained in culture without PBMC addition. These samples, in addition to PBMCs alone (t=0), were used for scRNA-seq (Figure 2A).
Project description:To investigate the effect of miR-203 in type 2 diabetes, target genes of miR-203 need to be investigated. The β cell specific miR-203 transgene (miR-203 TG) mice was constructed, and scRNA-seq was then performed on mouse islets.
Project description:The transcriptome of pancreatic islets from four mouse strains was compared - NODk.RagKO, B10k.RagKO, NODk.RagKO.insHEL and B10k.RagKO.insHEL. This comparison allows the assessment of altered transcriptome due to two factors - genetic background of NOD vs B10 strain, and the effect of the insHEL transgene. The use of the MHC k haplotype and the RagKO background remove contamination from infiltrating immune cells, in order to obtain a pure islet transcriptome.
Project description:We use scRNA-seq to show the differences in tumor-infiltrating immune cells among IgG, anti-PD-1, anti-PSGL-1, and combination anti-PD-1 and anti-PSGL-1 treated mice. We show that anti-PSGL-1 treatment resulted in an increase in neutrophil and T cells, anti-PD-1 treatment resulted in an increase in macrophages, and the combination resulted in an increase in T cells and macrophages when compared to the tumors of IgG treated mice. Additionally, we show that Tregulatory cells are decreased in the tumors of anti-PSGL-1 and combination treated mice. Further, we find that anti-PSGL-1 treated CD8 T cells show upregulation of activation and survival genes, while combination treatment increased effector gene expression in CD8 T cells. Both anti-PSGL-1 treatment and combination treatment increase effector gene expression in CD4 T cells when compared to IgG. This scRNA-seq study shows the impact of IgG, anti-PD-1, anti-PSGL-1, and combination anti-PSGL-1 and anti-PD-1 antibody tteatment on tumor-infiltrating immune cells in B16-GP33 melnoma tumor bearing mice.