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Expression data from human choroidal melanocyte cultures

ABSTRACT: Purpose: To determine to which extend inflammatory cytokines affect chemokine secretion by primary human choroidal melanocytes (HCMs), their capacity to attract monocytes and if HCMs are able to influence the proliferation of activated T cells. Methods: Primary cultures of HCMs were established from eyes of thirteen donors. HCMs were stimulated with IFN-γ and TNF-α or with supernatant from activated T-cells (T-cell conditioned media (TCM)). Gene expression analysis was performed using microarrays. Protein levels were quantified with ELISA or cytometric bead array. Supernatants of HCMs were assessed for the capability to attract monocytes in a trans-well plate. Proliferation of activated T-cells was assessed in a direct co-culture with HCMs by a [3H]-thymidine incorporation assay. Results: Stimulation of HCMs with TCM or IFN-γ and TNF-α resulted in increased expression and secretion of CXCL8, CXCL9, CXCL10, CXCL11, CCL2, CCL5 and intercellular adhesion molecule 1. Vascular endothelial growth factor and monocyte migration inhibitory factor were constitutively expressed without changes in response to pro-inflammatory cytokines. Supernatants derived from unstimulated cultures of ten HCM donors induced a high initial level of monocyte migration, which decreased upon stimulation with either TCM or IFN-γ and TNF-α. The supernatants from three HCM donors initially showed a low level of monocyte attraction which increased following exposure to pro-inflammatory cytokines. Direct co-culture of HCMs with T-cells resulted in inhibition of T-cell proliferation. Conclusion: These results show that normal and activated HCMs are immunologically active by secreting chemokines, and that HCMs are able to attract monocytes in addition to inhibit T-cell proliferation.

ORGANISM(S): Homo sapiens

PROVIDER: GSE70762 | GEO | 2016/12/21

SECONDARY ACCESSION(S): PRJNA289500

REPOSITORIES: GEO

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Project description:A major population of placenta macrophages represented throughout the pregnancy consists of CD14+ macrophages, but their characteristics remain badly understood. Here we purified from placentas at term CD14+ macrophages using positive selection. The phenotyping of CD14+ macrophages performed using flow cytometry revealed that placenta CD14+ macrophages expressed a series of markers distinct of those of circulating monocytes monocyte-derived macrophages. Placenta CD14+ macrophages spontaneously matured in multinucleated giant cells (MGCs) as demonstrated by size, number of nuclei display and specific cytoskeleton organization. Placenta CD14+ macrophages and MGCs were phagocytic cells but the potential of MGCs to mount an inflammatory response was lower than that of their precursors. Placenta CD14+ macrophages and MGCs stimulated with interferon and interleukin-4 were not polarized into typical M1 or M2 profiles. Placenta macrophages exhibited specific activation transcriptional programs. Indeed, principal component analysis and hierarchical clustering show that placental macrophages formed a distinct group from circulating monocytes and monocyte-derived macrophages. Among placenta macrophages, it was also possible to distinguish CD14+ macrophages and MGCs. In addition, networks based on gene interactions were clearly different in CD14+ macrophages and MGCs. Finally, the microenvironment of placenta CD14+ macrophages governs their differentiation into MGCs because CD14+ macrophages incubated with trophoblasts exhibited exarcerbated characteristics of MGCs and because the co-incubation of circulating monocytes from working women with trophoblast supernatants resulted into the formation of MGCs whereas monocytes from non-pregnant women incubated with trophoblast supernatants did not differentiate into MGCs. Taken together, these results clearly demonstrated specific feaures of placenta CD14+ macrophages. Three replicates of each of the following: 1. Placental macrophages just after isolation (CD14+ macrophages) 2. Placental macrophages after 9 days in culture (MGCs) 3. CD14+ cells isolated from PBMC which are extracted from the whole human blood of healthy donors (Monocytes) 4. Macrophages derived from monocytes (MDMs)

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Expression data from human choroidal melanocyte cultures

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