Expression and modulation of atypical chemokine receptors by epithelial cells

[摘要] The immune system relies on the correct spatial and temporal positioning of cells in order to function; cells need to be able to move throughout the circulatory system to survey for pathogens, to migrate from their resident sites in tissues when they sense infection or injury to alert other cells, or to migrate to the site of damage or infection to help mobilise a response. These functions often involve chemokines, small cytokines that signal through chemokine receptors, which are G-protein coupled receptors expressed on the cell membrane. Different chemokines are regulated differentially and can be associated with certain tissues or developmental processes, meaning the suite of receptors expressed by each cell type determines which tissues it is capable of entering, and the precise location it takes up once inside the tissue.Atypical chemokine receptors are a class of chemokine receptors that do not initiate the downstream signalling pathways typical of a G-protein coupled receptor, as they do not recruit intracellular G-proteins, and generally don't induce cell migration. Instead, these receptors are thought to function mainly as chemokine scavenging receptors, internalising and destroying their ligands before rapidly recycling to the cell surface. In this way, the levels of chemokines in the body are prevented from becoming oversaturated thus dampening the ability of cells to respond to signals. ACKR3 and ACKR4 are two atypical chemokine receptors that are expressed on endothelial cells and keratinocytes in the skin. Here, I have studied their expression on cultured human lymphatic endothelial cells and keratinocytes, and modulation in response to immune stimuli on these cells using a combination of qPCR and immunofluorescent staining. These strategies revealed that ACKR3 and ACKR4 are expressed on cultured LECs and keratinocytes and may be differentially regulated by both cell types in response to inflammatory stimuli including bacterial (LPS) and viral (Unmethylated CpG DNA) signatures. Although chemokine scavenging activity could not be detected on these cells, these findings suggested a role for ACKRs 3 and 4 in the inflammatory response.Further experiments in vivo explored the expression and modulation of ACKR3/CXCR4 and ACKR4 on epithelial cells including lymphatic endothelial cells, keratinocytes and vascular endothelial cells in the spleen. Flow cytometry was used to examine tissues both at rest and after inflammation (Aldara-mediated psoriasis model, or TPA-mediated contact hypersensitivity model) and investigate the regulation of ACKR3/CXCR4 or ACKR4 in response to these stimuli. Key findings included the strong overlap and differential regulation of ACKR3 and CXCR4 in response to TPA in the infundibulum subset of keratinocytes. Additionally, lymph nodes of Ackr4-/- mice were significantly enlarged after repeated treatments with Aldara. This appeared to be due to CCL19 dysregulation, but adoptive transfer suggested that there was no defect in leukocyte homing in these mice. This suggested an as yet undetermined response to enhanced CCL19 bioavailability that did not prevent the correct migration of leukocytes to secondary lymphoid organs.Overall, these experiments suggested that ACKR3 and ACKR4 are modulated in response to several inflammatory stimuli both in vivo and in vitro, and that the modulation of homeostatic chemokines can play a role in the response to inflammatory events. This was particularly important in the context of skin inflammation, where inflammatory chemokines, CXCR4 and ACKR2 have all been implicated in severity and duration of inflammatory events, but few studies have yet described the potential contributions of ACKR3 or ACKR4.

[发布机构] University:University of Glasgow;Department:Institute of Infection Immunity and Inflammation

[关键词] Chemokine, CCRL1, CXCR7, ACKR, CCL19, CCL21, CXCL12.