Human Kupffer Cell Subsets Defined by CD32
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ABSTRACT:
PROVIDER: PRJNA645956 | ENA |
REPOSITORIES: ENA
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Project description:Human liver myeloid cells are imperfectly defined, but it is broadly agreed that cells of stellate appearance in situ, expressing the markers CD11b and CD68, are the liver’s resident macrophages, termed Kupffer cells. Recent investigations using single cell RNA sequencing and unsupervised clustering algorithms suggest there are two populations of cells with the characteristics of tissue macrophages in human liver. We therefore analyzed dissociated human liver tissue using the markers CD11b and CD68 to define Kupffer cells and found within this population two subsets that differ in their expression of multiple surface markers. These subsets were FACS-sorted based on CD32 expression, and gene expression analysis identified them with human liver myeloid cell subsets defined by two independent single cell RNA sequencing studies. These two subsets differed in the expression of genes associated with T cell activation and immunosuppression, suggesting distinct roles in T cell tolerance.
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Project description:Human liver myeloid cells are imperfectly defined, but it is broadly agreed that cells of stellate appearance in situ, expressing the markers CD11b and CD68, are the liver's resident macrophages, classically termed Kupffer cells. Recent investigations using single cell RNA sequencing and unsupervised clustering algorithms suggest there are two populations of cells with the characteristics of tissue macrophages in human liver. We therefore analyzed dissociated human liver tissue using the markers CD11b and CD68 to define macrophage-like cells and found within this population two subsets that differ in their expression of multiple surface markers. These subsets were FACS-sorted based on CD32 expression, and gene expression analysis identified them with human liver myeloid cell subsets that were previously defined by two independent single cell RNA sequencing studies. Using qRT-PCR we found that the two subsets differed in the expression of genes associated with T cell activation and immunosuppression, suggesting distinct roles in T cell tolerance. In addition, one subset expressed two markers, CD1C and CD11c, more often seen on classical dendritic cells. Criteria used to distinguish macrophages from dendritic cells in other tissues may need to be revised in the human liver.
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Project description:HIV reservoirs in tissues form the major hurdle to an HIV cure. Despite major progress made on the understanding of the establishment and persistence of viral reservoirs, the characterization of composition and dynamics of the viral reservoir is still uncomplete. In addition, most studies in HIV infection are limited to blood. Here we take advantage of non-human primate models to provide a longitudinal analysis on potential viral target cells in distinct body compartments : blood, lymph nodes (LN), spleen ileum, jejunum and liver. We observed an increase in CD32+CD4+ T cells in secondary lymphoid tissues and intestine during primary and chronic pathogenic SIVmac infection. In the natural host (African green monkey, AGM), increase of the CD4+CD32+T cell levels was observed in tissues with higher replication and immune activation. CD32+CD4+ T cells expressed more often markers associated with HIV infected and/or reservoir cells (PD-1, CXCR5 for TFH cells in SLT, and CXCR3 for Th1 cells) than CD32- cells. The tissue CD32+CD4+T cells displayed higher levels of actively transcribed SIV RNA than CD32-CD4+T cells. The genome-wide transcriptome of CD32+CD4+ T cells in spleen from SIV-infected animals indicated that the CD32+CD4+ T cells shared B cell markers. The CD20+ expressing CD32+CD4+ T cells were increased in the tissues but not in the blood during SIV infection. Altogether, the study showed that SIV infection increases the frequencies of CD32+CD4+ T cells in tissues more than in blood. These cells might represent a not well-described subpopulation of activated CD4+ T cells.
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Project description:During embryonic development, blood cells emerge from a subset of specialized endothelial cells, named hemogenic endothelial cells (HECs), via a process known as endothelial-to-hematopoietic transition (EHT). A thorough characterization of HECs and their EHT is essential to guide the efforts to derive this population from human pluripotent stem cells (hPSCs), a critical step to generate therapeutic blood products in vitro. However, current known markers used to isolate HECs are insufficient as they also enrich for arterial endothelial cells that are associated with HECs. To identify specific human HEC markers, we performed transcriptomic analysis of 28-32-day human embryos, a developmental stage characterized by active EHT. We observed that the expression of FCGR2B, encoding for the Fc receptor CD32 previously associated with other specialized endothelia, is highly enriched in the ACE+CD34+ endothelial cell population that contains HECs. Functional ex vivo analyses confirmed that multilineage hematopoietic potential is highly enriched in CD32+ endothelial cells isolated from the aorta-gonad-mesonephros region and yolk sac of human embryos. In addition, CD32 emerged as selective marker for hPSC-derived HECs across different hematopoietic programs. Remarkably, our analyses showed that CD32 expression identifies HECs that are irreversibly bound to undergo EHT. As such, CD32 expression enriches for cells with hemogenic potential with a higher specificity for hPSC-derived HECs than other known HEC markers. These findings provide a simple method for isolating HECs from human embryos and hPSC cultures, allowing the efficient generation of hematopoietic cells in vitro.
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