Settings and mechanisms for trans-cellular diapedesis

Abstract

Immune system functions require blood leukocytes to continuously traffic throughout the body and repeatedly cross endothelial barriers (i.e., diapedese) as they enter (intravasation) and exit (extravasation) the circulation. The earliest studies to directly characterize diapedesis in vivo suggested co-existence of two distinct migratory pathways: between (para-cellular) and through (trans-cellular) individual endothelial cells. However, in the absence of conclusive in vitro observations, the latter pathway remained poorly accepted. The recent emergence of unambiguous in vitro reports of trans-cellular diapedesis has begun to illuminate mechanisms for this pathway and has renewed interest in its physiological roles. A thorough reevaluation of the existing literature reveals a large number of studies documenting significant use of the trans-cellular pathway in diverse in vivo settings. These include constitutive trafficking in bone marrow and lymphoid organs as well as upregulated extravasation in peripheral tissues during inflammation. Here we collectively summarize these in vivo observations alongside the emerging in vitro data in order to provide a framework for understanding the settings, mechanisms and roles for the trans-cellular route of diapedesis.

Figure 1

Mechanisms for trans-cellular diapedesis. The schematic summarizing basic morphologic features broadly observed in vivo and in vitro as a leukocyte (tan) progressively migrates across the endothelium through a trans-cellular pore. A small segment of endothelium is depicted in which two individual endothelial cells are distinguished by green and blue coloring. Locations where specific junctional adhesion complexes (i.e., tight, adherens and gap junctions) form are indicated (orange). A. Podosome Probing. The schematic depicts a ‘snapshot’ of a lymphocyte laterally migrating toward an intact inter-endothelial junction. During migration dozens of actin-dependent podosome-like protrusions dynamically form (downward pointing arrows) and retract (upward pointing arrows), concomitantly forcing endothelial invaginations termed ‘podo-prints’. This dynamic protrusion behavior is thought serve in migratory pathfinding as a means of ‘probing’ the endothelial surface for sites permissive to trans-cellular diapedesis. I. Shows a trailing edge podosome retracting. II. Shows a podosome protruding into, and being frustrated by, the rigid nuclear lamina. III. Shows a podosome and its cross-section (inset), highlighting the peripheral LFA-1 integrin/talin/vinculin zone and the actin-rich core. IV. Highlights a specific podosome that progressively extends to become an ‘invasive podosome’ and facilitate trans-cellular pore formation in panels B and C. Endothelial vesicles, VVO and caveolae (‘vesicles’) are seen enriched near or directly fused to podo-prints. B. Transmigratory Cup Formation. Overlapping temporally with podosome probing (A), endothelial cells proactively protrude actin-dependent, ICAM-1/VCAM-1-enriched protrusions (*) that embrace adherent leukocytes, forming ‘transmigratory cups’ that are thought to facilitate transition from lateral to trans-endothelial migration. Note many podosomes-like protrusions have or are retracting while one continues to protrude. C. Trans-cellular pore formation. At permissive sites a podosome-like protrusions progressively extend, transitioning to ‘invasive podosomes’, which forces the endothelial apical and basal plasma membrane into close opposition thereby facilitating initial trans-cellular pore formation for diapedesis. Active SNARE complex-dependent fusion of endothelial vesicle at the site of protrusion may facilitate this process. D. Pore expansion. The leukocyte progressively pushes across the trans-cellular pore causing expansion of its diameter to as much as 5 microns. E. Pore contraction. As the leukocyte completes diapedesis the pore contracts, maintaining close endothelial cell-leukocyte contacts. F. Pore closure/resolution. The leukocyte finally, exits the pore completely. Substantial in vitro and in vivo data support the existence of rapid resealing of the vacated pore. However, no details currently exist on the mechanisms of this important process.

Figure 2

Potential factors influencing route of diapedesis. The schematic emphasizes three main factors (endothelial cells, leukocyte and stimulus) and some of the key variables related to each of these that likely influence cell phenotype and behavior and, therefore, the route of diapedesis. Thus, route of diapedesis may be a highly context-specific issue.