T cell-platelet activation loop
The constitutive surface expression of CD40 on platelets opens the question as to the role that these cells play in immune responses. Danese et al. (p. 2011 ) found a dose-dependent increase in P-selectin expression in platelets exposed to soluble CD40L but not in platelets exposed to thrombin. The response was blocked by mAbs against either CD40 or the CD40 ligand (CD40L). P-selectin expression also was induced by cocultivation of platelets with peripheral blood T cells activated by anti-CD3 plus anti-CD28 stimulation but not by resting T cells; the blocking mAbs decreased the response by 40–50%. RANTES was released by platelets incubated with the soluble CD40L or with paraformaldehyde-fixed cells expressing CD40L. Phosphorylated p38 was detected in platelets treated with soluble CD40L, but it was lacking in resting platelets and in platelets treated with the blocking mAbs. A selective inhibitor of p38 phosphorylation prevented P-selectin up-regulation and the release of RANTES. T cells adhered to the surface of endothelial cells incubated with supernatants from activated platelets; the adherence was prevented by pretreatment of the T cells with a RANTES receptor antagonist. The experiments show that activated T cells stimulate platelets through a CD40-CD40L interaction to release RANTES, which in turn recruits more T cells to stimulate platelets.
Experimental autoimmune encephalitis (EAE) is a neuronal Ag-induced progressive paralytic disorder that may spontaneously resolve. Although regulatory T cells are thought to be involved in the recovery, the specific T cell type has not been identified. Faunce et al. (p. 1991 ) adapted an eye-derived APC model for generating Ag-specific CD8+ T regulatory cells to treatment of murine EAE. Tolerogenic APC were generated by culturing peritoneal exudate cells in serum-free medium containing myelin basic protein (MBP) and TGFβ2. Immunogenic APC were cultured in serum-free medium and TGFβ2 without MBP. Mice immunized with MBP 7 days following injection of tolerogenic cells had a 4-day delay in EAE onset (9 vs 15 days) and had less severe, or no, disease than mice that had received immunogenic APC or no cells before MBP immunization. Transfer of CD4+ T cell-depleted spleen cells from APC-treated animals with low severity of disease into untreated syngeneic recipients 1 wk before MBP immunization resulted in greater suppression of EAE than seen with transfer of total tolerogenic cells. CD8+ T cell depletion resulted in disease incidence and severity similar to control animals that did not receive any cells. The authors show that transferring Ag-pulsed APC induces suppressive CD8+ T regulatory cells and propose that in vitro-generated tolerogenic APC might prevent progression or recurrence of autoimmune disease.
Bradykinin receptors and perfusion injury
Activation of bradykinin B2 receptors following intestinal ischemia and reperfusion injury in rodents reduces local (intestinal), remote (lung), and systemic injury. Administration of a B2 receptor antagonist also reduces reperfusion-induced inflammatory tissue injury. In an attempt to reconcile these contradictory findings, Souza et al. (p. 2542 ) treated mice with the B2 receptor antagonist, HOE 140, before intestinal ischemia and reperfusion. HOE 140 prevented vascular permeability and neutrophil recruitment in the intestines and lungs, prevented intestinal hemorrhage, suppressed the increase in levels of TNF-α, increased the release of IL-1β and IL-10, and delayed lethality compared with control animals that did not receive HOE 140. B1 receptor mRNA was induced in intestine and lungs after reperfusion. Reperfusion of untreated B1R−/− mice resulted in the same resistance to injury seen in HOE 140-treated wild-type mice. In addition, 40% of the B1R−/− animals survived 240 min of reperfusion, twice as long as all wild-type and HOE 140-treated reperfused mice. Surprisingly, B1R−/− mice treated with HOE 140 before reperfusion were not protected and exhibited injury parameters similar to reperfused wild-type mice; lethality was greater than seen for blockage or absence of either receptor alone. The authors conclude that perfusion injury occurs by B1 receptor expression induced by B2 receptor activation.
Self vs almost-self tolerance
Whereas thymic deletion of self-reactive T cells protects against autoimmunity, it has the potential to destroy reactivity toward foreign Ags that are structurally related to self Ags. Kessels et al. (p. 2324 ) looked at T cell responsiveness to single amino acid variants of self-peptides in three systems in which a defined Ag is either self or non-self: B10NP mice that transgenically express a fragment of the nucleoprotein (NP) of influenza A virus; the male-specific HY Ag in male mice; and mice transgenic for the SV40 T Ag. Injection of the NP peptide or wild-type virus into B10NP mice did not elicit a T cell response. However, injection with amino acid substituted NP peptides or viruses encoding mutant NPs did elicit a response in most instances as measured by MHC-tetramer staining and IFN-γ release. T cells from wild-type mice responded to all viruses and peptides. Similar results were found for male mice vaccinated with wild-type and variant HY peptides and for male and female mice transgenic for SV40 T Ag vaccinated with wild-type and variant T Ag peptides. Self-tolerance extended to variant peptides with substitutions at the N-termini in the NP and HY models but not in the T Ag model. Some variant peptides elicited T cell responses to wild-type peptides in all three models. The authors conclude that, after deletion of self-specific T cells, the unfocused polyclonal T cell repertoire retains the capacity to react against most Ags that are closely related to self.
CD1b-glycolipid crystal structure
Recognition of bacterial glycolipid Ags is critical in the T cell response to intracellular bacterial infection. CD1 proteins, MHC class I-like molecules, present lipid Ags to T cells. Knowledge of the architecture of CD1 molecules can be helpful in manipulating the T cell response to bacteria. Batuwangala et al. (p. 2382 ) refolded the extracellular α1-α3 domains of human CD1b in the presence of glucose monomycolate (GMM) and analyzed crystallized CD1b-GMM complexes by x-ray diffraction. The mero-mycolate chain of GMM was found to occupy two hydrophobic channels (A′ and F′) as well as tunnel T′ which connects A′ and F′ below the α1 and α2 helices; the chain protruded out of the F′ channel into the TCR recognition surface. The α-chain of GMM occupied a third channel, C′. Mass spectrometry of mycolic acids freed from CD1b-GMM crystals indicated that there was enrichment for the largest forms of GMM (C58 and C60) compared with C54-GMM, the predominant species of mycolic acids extracted from Nocardia farcinica. Model structures using mycobacterial mycolic acids with substitutions along the mero-mycolate chain showed that the CD1b binding groove had a high degree of flexibility to accommodate the substitutions. This was due, in part, to a complex pattern of hydrophobicity in CD1b that distinguished it from other CD1 molecules (CD1a, CD1c, CD1d) and allowed it to bind lipids with longer acyl chains. Comparisons of CD1b-GMM with crystal structures of MHC-TCR complexes suggested that the CDR3 loops of a TCR would contact parts of GMM. In addition, the authors identified two distinct regions of CD1b and the protruding acyl tail as possible TCR contact points.
Glycoconjugate vaccines, consisting of Hemophilus influenzae type b (Hib) bacterial polysaccharides (PS) covalently linked to carrier proteins, have significantly reduced Hib disease in infants but require multiple doses to be effective. Latz et al. (p. 2431 ) studied the protection from a single dose of an Hib vaccine currently in use in which the PS is conjugated to a complex of outer membrane proteins from meningococcus (OPMC). TLR-2-transfected human embryonic kidney cells produced IL-8 after incubation with Hib-OPMC or with heat-killed Listeria monocytogenes but not with Hib conjugated to other carriers or with unconjugated Hib PS. No IL-8 response was seen in control cells lacking TLR2. Bone-marrow-derived dendritic cells from wild-type, but not from MyD88−/−, mice released TNF-α after stimulation with OPMC and Hib-OPMC. After three doses of Hib-OMP vaccine, wild-type mice produced higher levels of IgM, IgG, and total Igs against Hib PS by 4–6 wk, and against the carrier protein OPMC by 1 wk, compared with immunized TLR2−/− mice. Splenocytes from immunized wild-type mice produced higher levels of IL-6 and TNF-α after stimulation with OMPC ex vivo than stimulated splenocytes from immunized TLR2−/− mice. The authors propose that the enhanced immunogenicity of the Hib-OMPC vaccine is due to engagement of TLR2 and suggest the use of TLR2 agonists to enhance Hib vaccines.
Defining neutrophil-epithelial interactions
Neutrophil migration is regulated through interactions between its signal regulatory protein-α (SIRPα) and tissue-expressed CD47, an Ig superfamily transmembrane glycoprotein. Although the most membrane-distal Ig loop of the extracellular domain of SIRPα is known to bind CD47, the specific binding peptide in either molecule is unknown. Liu et al. (p. 2578 ) panned phage display hexa- and deca-peptide libraries over an anti-CD47 mAb, C5D5, conjugated to Sepharose. From 80 positive C5D5-binding phages, they identified a 12-amino acid consensus peptide, CERVIGTGWVRC, characterized by a conserved glycine-tryptophan doublet, that competitively blocked binding of other phages to mAb C5D5 and inhibited C5D5 Fab binding to CD47. The function-blocking peptide, designated C5D5.1, reduced neutrophil transmigration across intestinal epithelial monolayers by 60–70% compared with a non-peptide treated control; a control peptide had a minimal effect. Immobilized peptide C5D5.1 selectively bound purified SIRPα, but not purified CD47, protein. Ig domain specific SIRPα fusion proteins bound to the most distal extracellular Ig loop of SIRPα. The experiments define the functional epitope on CD47 involved both in binding to SIRPα and in regulating neutrophil transmigration. The authors suggest that this peptide could be useful in the design of anti-inflammatory agents.
Summaries written by Dorothy L. Buchhagen, Ph.D.