Project description:We compared CD8 T cells specific for a CMV epitope (pp65495-503) and an influenza A virus (IAV) epitope (M158-66) of the same healthy adults and identified genes whose expression are altered in CMV-specific compared to IAV-specific TCM cells
Project description:We compared CD8 TCM cells specific for a CMV epitope (pp65495-503) and an influenza A virus (IAV) epitope (M158-66) of the same healthy adults and identified genes whose expression are altered in CMV-specific compared to IAV-specific TCM cells.
Project description:Reconstitution of cytomegalovirus (CMV)-specific immunity following transplant remains a primary clinical objective to prevent CMV disease, and adoptive immunotherapy of CMV-specific T cells can be an effective therapeutic approach. Due to the persistence of CMV, most CMV-specific CD8+ T cells become terminally differentiated effector cells (TEFF). However, a minor subset retains a memory phenotype (TM). Interestingly, recent studies suggest that CMV-specific CD8+ T cells with different phenotypes may have different abilities to reconstitute sustained immunity following transfer. The immunology of human CMV (HCMV) infections is reflected in the mouse model of MCMV infection. We found that HCMV- and MCMV-specific T cells displayed shared genetic programs, validating the MCMV model for studies of CMV-specific T cells in vivo. After transfer, the proliferative capacity of MCMV-specific TM cells was vastly superior to TEFF cells. Strikingly, TM cells expanded and established sustained and diverse T cell populations even after multiple challenges. Although both TEFF and TM cells could protect Rag-/- mice, only TM cells could consistently survive after transfer into immune replete, latently infected recipients and respond if recipient immunity was lost. These data show that CMV-specific TM cells retain memory function during persistent infection and can re-establish CMV immunity when necessary. C57BL/6 mice were infected intraperitoneally (i.p.) with MCMV strain MW97.01 between 6-16 weeks of age. Splenocytes were isolated from chronically-infected mice and co-stained with three PE-conjugated tetramers loaded with the antigenic peptides from M38, m139 and IE3, all of which promote memory inflation. Cells were then stained with fluorescently conjugated antibodies and sorted on a MoFlo cell sorter. Naïve CD8+ cells were identified as CD44lo. MCMV-specific T cells were identified as CD8+, CD44hi and tetramer binding and then further segregated into memory and effector cells subsets by their expression of KLRG1 and CD127.
Project description:Reconstitution of cytomegalovirus (CMV)-specific immunity following transplant remains a primary clinical objective to prevent CMV disease, and adoptive immunotherapy of CMV-specific T cells can be an effective therapeutic approach. Due to the persistence of CMV, most CMV-specific CD8+ T cells become terminally differentiated effector cells (TEFF). However, a minor subset retains a memory phenotype (TM). Interestingly, recent studies suggest that CMV-specific CD8+ T cells with different phenotypes may have different abilities to reconstitute sustained immunity following transfer. The immunology of human CMV (HCMV) infections is reflected in the mouse model of MCMV infection. We found that HCMV- and MCMV-specific T cells displayed shared genetic programs, validating the MCMV model for studies of CMV-specific T cells in vivo. After transfer, the proliferative capacity of MCMV-specific TM cells was vastly superior to TEFF cells. Strikingly, TM cells expanded and established sustained and diverse T cell populations even after multiple challenges. Although both TEFF and TM cells could protect Rag-/- mice, only TM cells could consistently survive after transfer into immune replete, latently infected recipients and respond if recipient immunity was lost. These data show that CMV-specific TM cells retain memory function during persistent infection and can re-establish CMV immunity when necessary.
Project description:Here we studied the transcriptional profile of virus specific CD8 T cells to gain molecular insights in CD8 T cell functionality in HIV infection. HIV- and CMV-specific CD8 T cells were isolated from HIV infected individuals participating in the Amsterdam Cohort Studies (HIV progressors, long-term non-progressors (LTNP; HLA-B*57 and non-HLA-B*57) and individuals carrying the MAVS minor genotype) and CMV seropositive blood donors (BD). The transcription profile of HIV-specific CD8 T cells of LTNP groups was associated with increased protein/RNA metabolism pathways, indicating that immune control of HIV infection in these individuals is associated with increased functionality. In contrast, CMV-specific CD8 T cells from progressors showed increased expression of genes related to effector functions and suggests recent CMV reactivation. Our study provides novel insights into molecular mechanisms involved in HIV and CMV control in chronic HIV infection.
Project description:Zinc finger E-box binding protein 2 (ZEB2) is a key factor in the differentiation of naive CD8+ T cells into effector and memory T cells. However, the precise regulatory role of ZEB2 in cytotoxic CD8+ T cells remains unknown. Our recent DNA methylation analysis of cytomegalovirus (CMV)-specific human CD8+ T cells revealed two differentially methylated regions (DMRs) within the ZEB2 locus. We showed that these ZEB2 DMRs undergo pronounced demethylation during T cell differentiation. In particular, terminally differentiated CD8+ T cells and cytotoxic CD4+ T cells show almost complete demethylation. A ZEB2 knockout approach in CD8+ effector T cells followed by RNA-seq analysis revealed an altered gene expression profile, affecting genes related to cell-cell adhesion and impairing cytotoxicity in CMV-specific killing assays. Our data show that ZEB2 expression contributes to the differentiation of naive CD8+ T cells into effector and memory T cells and regulates the functional properties of virus-specific cytotoxic CD8+ T cells.
Project description:We introduced single-chain trimer (SCT) technologies into a high throughput platform for pMHC library generation that can be used for screening antigen specific CD8+ T cells. We compared the diversity of T cell receptor repertoire captured by SCT and folded peptide-MHC multimer presenting HLA-A02:01 restricted CMV peptide. We then constructed SCT libraries designed to capture SARS-CoV-2 spike specific CD8+ T cells from COVID-19 participants and healthy donors. TCR sequencing with antigen specificity was analyzed. The immunogenicity of these epitopes was validated by functional assays of T cells with cloned TCRs captured using SCT libraries.
Project description:The liver is an immunologically unique organ in terms of the regulation of immuno-tolerance and immune responses. Recently, non-recirculating tissue-resident memory T (TRM) cells expressing CD69 were found in various peripheral organs. In humans, a large population of CD69+CD8+ memory T cells residing in liver sinusoids shows reduced cytotoxicity. However, the trascriptional characteristics of human liver CD8+ T cells are still unknown. Therefore, we performed a microarray analysis to screen the gene expression profile in liver sinusoidal CD8+ T cells using CMV-specific CD8+ T cells from human peripheral blood and liver sinusoid.