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Single-cell RNA-sequencing of CD14+ monocyte differentiation with M-CSF stimulus

ABSTRACT: We obtained single-cell RNA-sequencing (scRNA-seq) profiles of CD14+ monocytes isolated from human peripheral blood at 0, 3 and 6 days after M-CSF stimulation (to differentiate the cells into macrophages) across multiple donors. Integration of single-cell RNA sequencing (scRNA-seq) data from multiple experiments, laboratories, and technologies can uncover biological insights, but current methods for scRNA-seq data integration are limited by a requirement for datasets to derive from functionally similar cells. We use a novel algorithm, Scanorama, to identify and merge the shared cell types among all pairs of datasets and to accurately integrate heterogeneous scRNA-seq datasets. Scanorama is sensitive to subtle temporal changes within the same cell lineage, successfully integrating functionally similar cells across time series data of CD14+ monocytes at different stages of differentiation into macrophages. Scanorama is not only able to differentiate between completely disparate cell types but is also sensitive to subtler transcriptional changes within a cell type due to processes like stimulation.

ORGANISM(S): Homo sapiens

PROVIDER: GSE126085 | GEO | 2019/02/06

REPOSITORIES: GEO

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Project description:BACKGROUND: Combination antiretroviral therapy (cART) is able to control HIV-1 viral replication, however long-lived latent infection in resting memory CD4+ T-cells persist. The mechanisms for establishment and maintenance of latent infection in resting memory CD4+ T-cells remain unclear. Previously we have shown that HIV-1 infection of resting CD4+ T-cells co-cultured with CD11c+ myeloid dendritic cells (mDC) produced a population of non-proliferating T-cells with latent infection. Here we asked whether different antigen presenting cells (APC), including subpopulations of DC and monocytes, were able to induce post-integration latent infection in resting CD4+ T-cells, and examined potential cell interactions that may be involved using RNA-seq. RESULTS: mDC (CD1c+), SLAN+ DC and CD14+ monocytes were most efficient in stimulating proliferation of CD4+ T-cells during syngeneic culture and in generating post-integration latent infection in non-proliferating CD4+ T-cells following HIV-1 infection of APC-T-cell co-cultures. In comparison, plasmacytoid DC (pDC) and B-cells did not induce latent infection in APC-T-cell co-cultures. We compared the RNA expression profiles of APC subpopulations that could and could not induce latency in non-proliferating CD4+ T-cells. Gene expression analysis, comparing the mDC, SLAN+ DC and CD14+ monocyte subpopulations to pDC identified 53 upregulated genes that encode proteins expressed on the plasma membrane that could signal to CD4+ T-cells via cell-cell interactions (32 genes), immune checkpoints (IC) (5 genes), T-cell activation (9 genes), regulation of apoptosis (5 genes), antigen presentation (1 gene) and through unknown ligands (1 gene). CONCLUSIONS: APC subpopulations from the myeloid lineage, specifically mDC subpopulations and CD14+ monocytes, were able to efficiently induce post-integration HIV-1 latency in non-proliferating CD4+ T-cells in vitro. Inhibition of key pathways involved in mDC-T-cell interactions and HIV-1 latency may provide novel targets to eliminate HIV latency. mRNA profiles of sorted, pure antigen presenting cells including, CD1c+ myleoid dendirtic cells (mDC), SLAN+ mDC, CD14+ monocytes and plasmacytoid DC (pDC), were generated using next generation sequencing in triplicate, using Illumina Illumina Hiseq 2000.

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Single-cell RNA-sequencing of CD14+ monocyte differentiation with M-CSF stimulus

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