Ronen Alon Lab

Investigating leukocyte trafficking & metastasis

Research

Circulating immune cells must exit blood vessels near specific sites of injury, inflammation, or tissue repair. At these locations, the vessel wall displays distinct combinations of trafficking signals that operate sequentially to recruit only specific subsets of circulating cells expressing the proper receptors to these signals. We employ adoptive transfer approaches as well as intravital microscopy in genetically modified mice, together with state-of-the-art 3D imaging techniques, to dissect how endothelial trafficking molecules regulate context- and tissue-selective immune cell recruitment and exit (extravasation) through distinct vascular beds nearby sites of infection, inflammation and malignancy.

Another major focus of our lab is understanding how chemotactic and antigenic signals promote critical lymphocyte interactions with dendritic cells during infection, cancer, and vaccination. Specifically, we investigate how defined lymphocyte subsets utilize adhesion molecules, primarily the integrin ligand ICAM-1 to undergo stimulation and differentiation into effective effector cells, and how they subsequently acquire immunological memory. These insights are critical for the development of improved vaccines as well as of novel therapeutic strategies that interfere with immune cell functions underlying inflammatory disorders. We also study the role of dendritic cell-expressed ICAM-1 in the differentiation and education of tumor antigen–specific T cells, and follow how these immune cells acquire specific tissue homing signatures that enable them to infiltrate immune hubs within tumors via subsets of tumor-associated blood vessels.

Another research direction in our lab focuses on lung metastasis, particularly the interactions between circulating cancer cells entering the pulmonary capillaries and immune cells such as neutrophils and T cells circulating inside these blood vessels. Our recent findings suggest that subsets of cancer cells trapped in lung capillaries evade cytotoxic T cell-mediated killing by downregulating their ability to process and present tumor expressed peptides on their surface MHC complexes. Subsets of these cells entrapped inside lung capillaries are also targeted and eliminated by specialized ROS high neutrophils generated specifically in tumor bearing mice.

Our lab has recently become interested in the role of key integrin ligands, such as ICAM-1 and VCAM-1, in thymocyte differentiation. This multistep process involves both positive and negative thymocyte selection through recognition of self-peptides presented on MHC complexes across distinct thymic compartments. To address this standing question, we have generated novel mice models with conditional loss of ICAM-1 and VCAM-1 expression selectively in thymic epithelial cells (TECs). These models will allow us to dissect how the absence of one or both adhesion molecules selectively in TECs affects thymocyte differentiation and selection in the cortex and medulla.

Research page

The role of DC ICAM-1 in immune synapses in vivo

Targeting killer T cells to metastatic lesions

Lymphocyte education in tumor draining lymph nodes

The roles of thymic epithelial CAMs in thymocyte differentiation

Selected Publications

Dendritic cell ICAM-1 strengthens synapses with CD8 T cells but is not required for their early differentiation

Sapoznikov A., Kozlovski S., Levi N., Feigelson S. W., Regev O., Davidzohn N., Ben-Dor S., Haffner-Krausz R., Feldmesser E., Wigoda N., Petrovich-Kopitman E., Biton M. & Alon R. (2023) Cell Reports. 42, 8, 112864.

Lymphocyte priming in lymph nodes (LNs) was postulated to depend on the formation of stable T cell receptor (TCR)-specific immune synapses (ISs) with antigen (Ag)-presenting dendritic cells (DCs). The high-affinity LFA-1 ligand ICAM-1 was implicated in different ISs studied in vitro. We dissect the in vivo roles of endogenous DC ICAM-1 in Ag-stimulated T cell proliferation and differentiation and find that under type 1 polarizing conditions in vaccinated or vaccinia virus-infected skin-draining LNs, Ag-presenting DCs engage in ICAM-1-dependent stable conjugates with a subset of Ag-specific CD8 blasts. Nevertheless, in the absence of these conjugates, CD8 lymphocyte proliferation and differentiation into functional cytotoxic T cells (CTLs) and skin homing effector lymphocytes takes place normally. Our results suggest that although CD8 T cell blasts engage in tight ICAM-1-dependent DC-T ISs, firm ISs are dispensable for TCR-triggered proliferation and differentiation into productive effector lymphocytes.

Transendothelial migration of lymphocytes mediated by intraendothelial vesicle stores rather than by extracellular chemokine depots

Shulman Z., Cohen S. J., Roediger B., Kalchenko V., Jain R., Grabovsky V., Klein E., Shinder V., Stoler-Barak L., Feigelson S., Meshel T., Nurmi S. M., Goldstein I., Hartley O., Gahmberg C. G., Etzioni A., Weninger W., Ben-Baruch A. & Alon R. (2012) Nature Immunology. 13, 1, p. 6776

Chemokines presented by the endothelium are critical for integrin-dependent adhesion and transendothelial migration of naive and memory lymphocytes. Here we found that effector lymphocytes of the type 1 helper T cell (T _H1 cell) and type 1 cytotoxic T cell (T _C1 cell) subtypes expressed adhesive integrins that bypassed chemokine signals and established firm arrests on variably inflamed endothelial barriers. Nevertheless, the transendothelial migration of these lymphocytes strictly depended on signals from guanine nucleotide-binding proteins of the G _i type and was promoted by multiple endothelium-derived inflammatory chemokines, even without outer endothelial surface exposure. Instead, transendothelial migration-promoting endothelial chemokines were stored in vesicles docked on actin fibers beneath the plasma membranes and were locally released within tight lymphocyte- endothelial synapses. Thus, effector T lymphocytes can cross inflamed barriers through contact-guided consumption of intraendothelial chemokines without surface-deposited chemokines or extraendothelial chemokine gradients.

Lymph node chemokines promote sustained T lymphocyte motility without triggering stable integrin adhesiveness in the absence of shear forces

Woolf E., Grigorova I., Sagiv A., Grabovsky V., Feigelson S., Shulman Z., Hartmann T., Sixt M., Cyster J. G. & Alon R. (2007) Nature Immunology. 8, 10, p. 1076-1085

Lymphocyte motility in lymph nodes is regulated by chemokines, but the contribution of integrins to this motility remains obscure. Here we examined lymphocyte migration over CCR7-binding chemokines that 'decorate' lymph node stroma. In a shear-free environment, surface-bound lymph node chemokines but not their soluble counterparts promoted robust and sustained T lymphocyte motility. The chemokine CCL21 induced compartmentalized clustering of the integrins LFA-1 and VLA-4 in motile lymphocytes, but both integrins remained nonadhesive to ligands on lymphocytes, dendritic cells and stroma. The application of shear stress to lymphocytes interacting with CCL21 and integrin ligands promoted robust integrin-mediated adhesion. Thus, lymph node chemokines that promote motility and strongly activate lymphocyte integrins under shear forces fail to stimulate stable integrin adhesiveness in extravascular shear-free environments.

All Publications