MIMM Trevor Owens Mon Oct 4 - Lymphoid tissue - organization and development Wed Oct 6 - Lymphocyte traffic and adhesion signaling Fri Oct 8 - B cell tolerance - deletion versus anergy Mon Oct 18 - Transgenic models of B cell tolerance Mid-term Oct 15 - One Owens question, from Lectures 1,2. Kerfoot, S. M.,Kubes, P., Overlapping roles of P-selectin and alpha 4 integrin to recruit leukocytes to the central nervous system in experimental autoimmune encephalomyelitis. J Immunol 2002; 169: Kim, M., Carman, C.V., Springer, T.A Bidirectional transmembrane signaling by cytoplasmic domain separation in integrins. Science 301:

Mice without secreted TNF but with functional normally- regulated and expressed membrane-bound TNF (Mem-TNF∆/∆ mice) were created by knocking-in the uncleavable ∆1-9,K11E TNF allele. In contrast to TNF-deficient mice (TNF-/-), mem-TNF supported many features of lymphoid structure, except generation of primary B cell follicles. Splenic chemokine production was nearly normal in Mem-TNF mice Mem-TNF was suboptimal for development of inflammation. Ruuls et al, 2001, Immunity 15:533 Mem-TNF Tg mice have normal LN development

Ruuls et al, 2001, Immunity 15:533 SPLEEN No primary B cell follicles T:B segregation less than in WT, but improved over TNF-/- Marginal zone metallophilic macrophages restored MadCAM restored PNA + Germinal centres CR1 + FDC networks

(from von Andrian and Mackay, NEJM 343:1020)

Adhesion and trafficking concepts 1. Combinatorial construction of specific homing pathways eg. rolling adhesion model, multiple ‘yes-no’ decisions 2. Bidirectional cross-talk between lymphocytes and microenvironment Adhesion signaling Chemokine receptor expression 3. Competitive niche homing in controlling lymphocyte homeostasis and shaping the immune repertoire Deletion/anergy of autoimmune B cells via PALS exclusion (from Butcher and Picker, Science 1996;272:60-66)

(from Hickey MJ, Clin Sci :1-12) ~4000  m/s ~40  m/s 1-20 s Minutes (reversible) ~10 minutes

L-selectin deficient mice Antigen presentation and effector mechanisms intact. Virtually no antigen-specific T cells within draining peripheral nodes after a contact challenge (skin) Defect resides primarily in the inability of antigen-specific T cells to home to and be activated in these nodes

AdhesionLymphocyte LymphocytesTargetMoleculeCounter-receptor NaïveLymph nodePNAdL-selectin NaïvePeyer’s patchMAdCAM-1L-selectin +  4ß7 Gut-homingLaminaMAdCAM-1  4ß7 cellspropria Skin-homingSkinE-selectinCLA +  4ß1 cellsVCAM-1 (PNAd): a group of endothelial sialomucins — CD34, podocalixin, glycosylation-dependent cell-adhesion molecule 1 (GlyCAM-1), and sialylated glycoprotein of 200 kd (sgp200) — all of which include a sulfated sialyl-LewisX (sLeX)–like motif. (from Butcher and Picker, Science 1996;272:60-66)

(from von Andrian and Mackay, NEJM 343:1020)

Th1 T cells - induce/mediate inflammation in tissues Uniquely capable of transferring Delayed Type Hypersensitivity Preferentially express functional PSGL-1, ligand for P-selectin, and therefore preferentially migrate to skin and other tissues. Experimental autoimmune encephalomyelitis (EAE) is a Th1- mediated autoimmune disease. Myelin protein-specific T cells infiltrate the central nervous system and induce inflammation and demyelination. EAE is a model for MS.  4integrin (eg. VLA-4/VCAM) interactions have been implicated in controlling entry of cells to CNS in EAE. A clinical trial of an integrin-blocking reagent (Antegren®) has shown promising results in MS. Limited selectin expression in CNS vasculature

(from Kerfoot & Kubes, J Immunol 2002; 169: 1000) Intravital microscopy Visualization of fluorescent- labeled cells in cerebral blood vessel in mice without (A) and with (B) EAE. Cells that roll can be distinguished from cells that adhere.

(from Kerfoot & Kubes, J Immunol 2002; 169: 1000) Rolling and adhesion increase in mice with EAE.

(from Kerfoot & Kubes, J Immunol 2002; 169: 1000) Infiltrating cells are T cells (CD3 + ) and macrophages/granulocytes (Mac- 1 + )

(from Kerfoot & Kubes, J Immunol 2002; 169: 1000) Rolling is preferentially inhibited by anti-P- Selectin before disease initiates. As disease progresses, this becomes less effective and more of the rolling is mediated by  4 integrin