Mechanisms underlying the trafficking of dendritic cells from tissues to draining lymph nodes via the lymphatic receptor LYVE-1

Fundamental to generating a host immune response to infection is the mobilisation of antigen loaded dendritic cells (DCs) in affected tissues and their migration via afferent lymph to draining nodes where they prime and activate T cells (reviewed in Jackson 2009, "Lymphatic regulation of cell trafficking" http://www.omicsonline.org/open-access/lymphatic-regulation-of-cellular-trafficking-2155-9899-5-258.php?aid=31116) 10.4172/2161-0681.1000241 ). The trafficking of DCs and other leucocyte populations in lymph is vital to such immunity, and involves a series of co-ordinated events driven initially by chemokines, including CCL21 and CX3CL1 secreted from initial lymphatic capillaries (Johnson and Jackson 2010, 2013; reviewed in Johnson et al. 2014), but also by many adhesion receptors in lymphatic endothelium (Johnson et al 2006 and see Fig 1). However it is currently unclear how directional guidance of migrating leucocytes towards lymphatic vessels is integrated with subsequent endothelial adhesion and transmigration, and which endothelial receptors are key for vessel entry.

Most leucocytes are thought to access lymphatic capillaries at specialised overlapping junctions, distinct from tight junctions of blood capillaries. Here, the flap-like edges of oakleaf shaped endothelial cells interdigitate to form button-like portals, whose tips are decorated with the lymphatic vessel specific hyaluronan (HA) receptor LYVE-1 (Banerji et al, 1999; Baluk et al 2007 and reviewed in Jackson 2014), and whose sides are pinned by adherens and tight junction receptors VE-cadherin, claudins and JAMs (Fig 2). The implication is that engagement of migrating cells with these portals leads to loosening of the junctions and entry to the vessel lumen (Fig 3).

Recent work in my laboratory using a combination of in vitro and in vivo approaches has produced compelling evidence that DCs engage LYVE-1, through reversible avidity-dependent binding to HA (Lawrance et al 2016) which decorates the DC surface as a form of glycocalyx (Johnson et al submitted). Interestingly, these LYVE-1:HA adhesive interactions create transmigratory cups and we speculate they form conveyor belts in vivo that guide the cells through lymphatic entry portals, likely aided by CCL21 that forms discrete puncta in their vicinity (Tal et al 2011). Given that LYVE-1 is selectively expressed in lymphatic but not blood vessels, and that its HA ligand can form chains of several microns, we speculate this mechanism has evolved to direct initial contacts between DCs and the basolateral surface of lymphatics, temporally overriding underlying conventional adhesion molecules on the leucocyte surface. Further credence to such a mechanism has been provided by our recent finding that pathogenic Group A Streptococci which have a dense hyaluronan capsule, exploit LYVE-1 to gain access to the lymphatic compartment during host infection (Lynskey et al. 2015)

The project on offer will explore the in vivo anatomy LYVE-1 mediated leucocyte interactions during leucocyte trafficking, using confocal/intravital imaging as well as in vitro analytical approaches. In particular it will ask - How does the LYVE-1:HA adhesion axis integrate with chemotaxis for vessel entry ? Does LYVE-1 mediated entry involve concerted interactions with other receptors such as ICAMs that are upregulated in inflamed lymphatics ? Are LYVE-1:HA interactions involved only in DC entry or do they also assist in DC crawling within the vessel lumen ? Do they also mediate DC migration within downstream lymph nodes, where LYVE-1 is an abundant component of subcapsular sinuses

Lastly, there may also be an option for the candidate to explore the role of the LYVE-1:HA axis for leucocyte adhesion and trafficking in other contexts such as the hepatic sinuses, where LYVE-1 is abundant and where HA deposits are known to recruit neutrophils during liver inflammation (Mc Donald et al 2008).

The successful candidate will have the benefit of supervision both from the PI, and from a dedicated senior research scientist, as well as ongoing Unit and international collaborations with other leading scientists in the field. We anticipate the new insights gained in these preclinical studies to seed future development of novel therapeutic strategies for inflammatory diseases.

The work will involve the use of animal models of inflammation, ex vivo tissue explants and in vitro culture models to visualise characterise, manipulate and quantify leucocyte trafficking, hyaluronan glycocalyx/complex formation and tissue vasculature by confocal and intravital imaging. Experiments will take advantage of LYVE-1-/- mice, LYVE-1 function blocking mAbs and appropriate fluorescent reporter mice that will be available in the host laboratory.