Burt, Claire Louise
The role of D6 in neutrophil migration, melanoma growth and metastasis.
PhD thesis, University of Glasgow.
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Chemokines are members of a family of small structurally related proteins, which can be classified according to the positioning of four conserved cysteine residues, into four families – CC, CXC, CX3C and XC. They have a wide range of functions, from being involved in embryonic development, disease processes and cell migration. Broadly speaking, chemokines can be divided into homeostatic, those that are involved in development, and inflammatory chemokines, which are produced at high levels during infection.
In order to exert their function, chemokines bind to seven transmembrane G protein coupled receptors; this causes the activation of numerous different signalling pathways which can lead to cell activation, migration, survival or apoptosis. The chemokine system is further complicated by the existence of a family of atypical receptors. Four of these receptors have been identified to date; CXCR7, CCX-CKR, DARC and D6, and they all share the apparent inability to signal after ligand binding.
D6 has been identified as a member of this atypical receptor family with the ability to bind and internalise 12 inflammatory CC chemokines. D6 expression has been demonstrated in barrier tissues, such as the skin, gut and lung, as well as being expressed by numerous leukocyte subsets. Studies looking at the function of D6 have demonstrated its involvement in the resolution phase of inflammation, and D6 may be involved in cell migration and inflammation driven tumourigenesis. Here the analysis of the role of D6 in neutrophil migration has been carried out, in the well-characterised TPA model of skin inflammation. This work also shows that D6 plays a crucial role in melanoma metastasis.
In the TPA model of skin inflammation, 129/Bl6 D6KO mice display an exaggerated inflammatory response, which causes the development of a psoriasis-like pathology. This pathology is dependent on T cells and mast cells, and is associated with an alteration in neutrophil positioning. In the inflamed skin of D6KO mice, neutrophils are found at the dermal/epidermal junction, whilst in WT mice, neutrophils are restricted to the dermis. In this model, D6 is required to prevent neutrophils migrating to the dermal/epidermal junction. D6 expression has been demonstrated both in the skin and on neutrophils, so there are two hypotheses to describe how D6 is influencing neutrophil movement in this model. First, D6 on neutrophils is required to limit their migration within the skin, keeping them away from the dermal/epidermal junction, alternatively D6 in the skin may be responsible for chemokine clearance during inflammation, and the localisation of neutrophils is influenced by these chemokines. This work aimed to investigate the role that D6 played on neutrophil localisation in inflamed skin, carried out using a model of neutrophil adoptive transfer. These results demonstrate that expression of D6 by neutrophils does not alter their positioning in inflamed skin, suggesting that the role for D6 in this model is in regulating chemokines within the skin, a hypothesis which requires further investigation.
The second part of this work aimed to investigate the role of D6 in melanoma growth and metastasis. D6 has been shown to be involved in murine models of inflammation driven skin tumourigenesis and colon cancer. These data presented here show that D6 does not influence the primary melanoma growth, but has a profound effect on the development of metastases in the lungs, in the murine B16 model. D6 is required for the development of lung metastases, but the precise mechanism is yet unknown. These data suggest that D6 can alter macrophage numbers within the lungs, which permits the development of metastases, as well as pointing towards a role for D6 in lymphangiogenesis within the lungs, which affects metastasis development, perhaps by altering the development of the pre-metastatic niche. Overall, these studies contribute to the investigation of the role of D6 in skin inflammation, and show, for the first time, that D6 is involved in the pulmonary metastasis of melanoma, suggesting D6 may be a potential therapeutic target in pulmonary metastasis.
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