Metabolomics,Unknown,Transcriptomics,Genomics,Proteomics

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Inflammatory environment and oxidized LDL convert circulating human proangiogenic cells (PACs) into functional antigen presenting cells (APCs)

ABSTRACT: The function of human circulating proangiogenic cells (PACs) has been described extensively. However, little focus has been placed on understanding how these cells differ in their functions in the presence of microenvironments mimicking vascular inflammation. We hypothesized that exposure to proinflammatory cytokines or the oxLDL, an autoantigen abundant in advanced atherosclerotic plaques, converts PACs into immune-modulating/proinflammatory cells. Hence, we examined the effect of oxLDL and inflammatory stimuli on their phenotype by use of a functional genomics model based on secretome and whole genome transcriptome profiling. PACs obtained from culturing a PBMC fraction in angiogenic medium were primed with DC differentiation cytokines for 3 days and then exposed to proinflammatory cytokines or oxLDL for 2 days. Under these conditions, PACs converted into APCs (APCcy and APCox, respectively), expressed maturation markers CD80 and CD83, and showed an up-regulation of CD86. APCcy and APCox induced a robust T cell BrdU incorporation. Despite a similar ability to induce lymphocyte proliferation, APCcy and APCox differed for the secretory pathway and mRNA expression. Analysis of the differentially expressed genes identified 4 gene "clusters", showing reciprocal modulation in APCcy vs. APCox justifying, according to functional genomics analyses, a different putative function of the cells in antigen processing. Together, these data show that treatment with inflammatory cytokines or oxLDL converts human PAC phenotype and function into that of APCs with similar lymphocyte-activating ability but distinct maturation degree and paracrine functions. PBMC-derived PACs were committed with GM-CSF and IL-4 to a dendritic phenotype, and then exposed to proinflammatory cytokines (IL-1β and TNF-α) or to the oxidized form of LDL (oxLDL). Functional assays and whole genome gene expression profiling were performed both on PACs (n=6) and or resulting cytokine- (APCcy, n=6) and oxLDL-induced APCs (APCox, n=6). RNA was reverse transcribed, labeled, and linearly amplified, and then hybridized to HumanHT-12 v.4 Expression BeadChip microarrays. To estimate technical variability, ≈20% of the samples (n=4) were hybridized in duplicate.

ORGANISM(S): Homo sapiens

SUBMITTER: Gualtiero Colombo

PROVIDER: E-GEOD-60208 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:Antigen presenting cells (APCs) regulate the balance of our immune response towards microbes. Whereas immunogenic APCs boost inflammation and activate lymphocytes, the highly plastic cells can switch into a tolerogenic/suppressive phenotype that dampens and resolves the response. Thereby the initially mediated inflammation seems to prime the switch of APCs while the strength of activation determines the grade of the suppressive phenotype. Recently we showed that pathogen recognition receptor-mediated pro-inflammatory cytokines reprogram differentiating human blood monocytes in vitro towards an immunosuppressive phenotype through prolonged activation of signal transducer and activator of transcription (STAT) 3. The TLR7/8 ligand R848 (Resiquimod) triggers the high release of cytokines from GMCSF/IL-4-treated monocytes. These cytokines subsequently upregulate T cell factors, such as Programmed death-ligand 1 (PD-L1) and Indolamin-2,3-Dioxygenase (IDO) suppressive through cytokine receptor-mediated STAT3 activation. Here we reveal an essential role for the micro-RNA (miR) hsa-miR99b/let-7e/miR125a cluster in stabilizing the suppresive phenotype of R848-stimulated APCs on different levels. On the one hand the miR cluster boosts R848-stimulated cytokine production through regulation of MAPkinase inhibitor Tribbles homolog 2 (TRIB2), thereby enhancing cytokine-stimulated activation of STAT3. One the other hand, the STAT3 inhibitor suppressor of cytokine signaling-1 (SOCS-1) is targeted by the miR cluster, stabilizing the STAT3-induced expression of immunosuppressive factors PD-L1 and IDO. Finally hsa-miR-125a/let7e/99b cluster regulates generation of the suppressive tryptophan (Trp) metabolite kynurenine by targeting the tryptophanyl -tRNA synthetase WARS, the direct competitor of IDO in terms of availability of Trp. In summary, our results reveal the hsa-miR99b/let-7e/miR125a cluster as an important player in the concerted combination of mechanisms that stabilizes STAT3 activity and thus regulate R848-stimulated suppressive APCs.

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Inflammatory environment and oxidized LDL convert circulating human proangiogenic cells (PACs) into functional antigen presenting cells (APCs)

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