Basil O. Gerber

Functional expression of the eotaxin receptor CCR3 in T lymphocytes co-localizing with eosinophils.

BACKGROUND The chemokine eotaxin is produced at sites of allergic inflammation, binds selectively to the chemokine receptor CCR3 and attracts eosinophil and basophil leukocytes, which express high numbers of this receptor. Responses of T lymphocytes to eotaxin have not been reported so far. We have investigated the expression of CCR3 in T lymphocytes and analysed the properties and in vivo distribution of T lymphocytes expressing this receptor. RESULTS In search of chemokine receptors with selective expression in T lymphocytes, we have isolated multiple complementary DNAs (cDNAs) encoding CCR3 from a human CD4+ T-cell cDNA library. T-lymphocyte clones with selectivities for protein and non-protein antigens were analysed for expression of CCR3 and production of Th1- and Th2-type cytokines. Of 13 clones with surface CCR3, nine secreted enhanced levels of interleukin-4 and/or interleukin-5, indicating that CCR3 predominates in Th2-type lymphocytes. CCR3+ T lymphocytes readily migrated in response to eotaxin, and showed the characteristic changes in cytosolic free calcium. Immunostaining of contact dermatitis, nasal polyp and ulcerative colitis tissue showed that CCR3+ T lymphocytes are recruited together with eosinophils and, as assessed by flow cytometry, a large proportion of CD3+ cells extracted from the inflamed skin tissue were CCR3+. By contrast, CCR3+ T lymphocytes were absent from tissues that lack eosinophils, as demonstrated for normal skin and rheumatoid arthritis synovium. CONCLUSIONS We show that T lymphocytes co-localizing with eosinophils at sites of allergic inflammation express CCR3, suggesting that eotaxin/CCR3 represents a novel mechanism of T-lymphocyte recruitment. These cells are essential in allergic inflammation, as mice lacking mature T lymphocytes were insensitive to allergen challenge. Surface CCR3 may mark a subset of T lymphocytes that induce eosinophil mobilization and activation through local production of Th2-type cytokines.

CD69 upregulation on T cells as an in vitro marker for delayed‐type drug hypersensitivity

Background:  T cells play a key role in delayed‐type drug hypersensitivity reactions. Their reactivity can be assessed by their proliferation in response to the drug in the lymphocyte transformation test (LTT). However, the LTT imposes limitations in terms of practicability, and an alternative method that is easier to implement than the LTT would be desirable.

Stimulation of human T cells with sulfonamides and sulfonamide metabolites

BACKGROUND Exposure to sulfonamides is associated with a high incidence of hypersensitivity reactions. Antigen-specific T cells are involved in the pathogenesis; however, the nature of the antigen interacting with specific T-cell receptors is not fully defined. OBJECTIVE We sought to explore the frequency of sulfamethoxazole (SMX)- and SMX metabolite-specific T cells in hypersensitive patients, delineate the specificity of clones, define mechanisms of presentation, and explore additional reactivity with structurally related sulfonamide metabolites. METHODS SMX- and SMX metabolite-specific T-cell clones were generated from 3 patients. Antigen specificity, mechanisms of antigen presentation, and cross-reactivity of specific clones were then explored. Low-lying energy conformations of drugs (metabolites) were modeled, and the energies available for protein binding was estimated. RESULTS Lymphocytes proliferated with parent drugs (SMX, sulfadiazine, and sulfapyridine) and both hydroxylamine and nitroso metabolites. Three patterns of drug (metabolite) stimulation were seen: 44% were SMX metabolite specific, 43% were stimulated with SMX metabolites and SMX, and 14% were stimulated with SMX alone. Most metabolite-responsive T cells were stimulated with nitroso SMX-modified protein through a hapten mechanism involving processing. In contrast to SMX-responsive clones, which were highly specific, greater than 50% of nitroso SMX-specific clones were stimulated with nitroso metabolites of sulfapyridine and sulfadiazine but not nitrosobenzene. Pharmacophore modeling showed that the summation of available binding energies for protein interactions and the preferred spatial arrangement of atoms in each molecule determine a drugs potential to stimulate specific T cells. CONCLUSIONS Nitroso sulfonamide metabolites form potent antigenic determinants for T cells from hypersensitive patients. T-cell responses against drugs (metabolites) bound directly to MHC or MHC/peptide complexes can occur through cross-reactivity with the haptenic immunogen.

Drug-specific in vitro release of IL-2, IL-5, IL-13 and IFN-γ in patients with delayed-type drug hypersensitivity

Background:  The most prevalent drug hypersensitivity reactions are T‐cell mediated. The only established in vitro test for detecting T‐cell sensitization to drugs is the lymphocyte transformation test, which is of limited practicability. To find an alternative in vitro method to detect drug‐sensitized T cells, we screened the in vitro secretion of 17 cytokines/chemokines by peripheral blood mononuclear cells (PBMC) of patients with well‐documented drug allergies, in order to identify the most promising cytokines/chemokines for detection of T‐cell sensitization to drugs.

CCL22-induced responses are powerfully enhanced by synergy inducing chemokines via CCR4: evidence for the involvement of first β-strand of chemokine

In an attempt to clarify how cells integrate the signals provided by multiple chemokines expressed during inflammation, we have uncovered a novel mechanism regulating leukocyte trafficking. Our data indicate that the concomitant exposure to CCR4 agonists and CXCL10/IP‐10 strongly enhances the chemotactic response of human T lymphocytes. This enhancement is synergistic rather than additive and occurs via CCR4 since it persists after CXCR3 blockade. Besides chemotaxis, other cellular responses are enhanced upon stimulation of CCR4‐transfected cells with CCL22/MDC plus CXCL10. Several other chemokines in addition to CXCL10 were able to increase CCL22‐mediated chemotaxis. The first β‐strand of the chemokine structure is highly and specifically implicated in this phenomenon, as established using synergy‐inducing and non‐synergy‐inducing chimeric chemokines. As shown in situ for skin from atopic and allergic contact dermatitis patients, this organ becomes the ideal environment in which skin‐homing CCR4+ T lymphocytes can accumulate under the stimulus offered by CCR4 agonists, together with the synergistic chemokines that are concomitantly expressed. Overall, our results indicate that chemokine‐induced synergism strengthens leukocyte recruitment towards tissues co‐expressing several chemokines.

I-TAC/CXCL11 is a natural antagonist for CCR5

The selective CXC chemokine receptor 3 (CXCR3) agonists, monokine induced by interferon‐γ (IFN‐ γ)/CXC chemokine ligand 9 (CXCL9), IFN‐inducible protein 10/CXCL10, and IFN‐inducible T cell α chemoattractant (I‐TAC)/CXCL11, attract CXCR3+ cells such as CD45RO+ T lymphocytes, B cells, and natural killer cells. Further, all three chemokines are potent, natural antagonists for chemokine receptor 3 (CCR3) and feature defensin‐like, antimirobial activities. In this study, we show that I‐TAC, in addition to these effects, acts as an antagonist for CCR5. I‐TAC inhibited the binding of macrophage‐inflammatory protein‐1α (MIP‐1α)/CC chemokine ligand 3 (CCL3) to cells transfected with CCR5 and to monocytes. Furthermore, cell migration evoked by regulated on activation, normal T expressed and secreted (RANTES)/CCL5 and MIP‐1β/CCL4, the selective agonist of CCR5, was inhibited in transfected cells and monocytes, respectively. In two other functional assays, namely the release of free intracellular calcium and actin polymerization, I‐TAC reduced CCR5 activities to minimal levels. Sequence and structure analyses indicate a potential role for K17, K49, and Q51 of I‐TAC in CCR5 binding. Our results expand on the potential role of I‐TAC as a negative modulator in leukocyte migration and activation, as I‐TAC would specifically counteract the responses mediated by many “classical,” inflammatory chemokines that act not only via CCR3 but via CCR5 as well.

Noncovalent Interactions of Drugs With Immune Receptors May Mediate Drug-induced Hypersensitivity Reactions

Drug-induced hypersensitivity reactions are instructive examples of immune reactions against low molecular weight compounds. Classically, such reactions have been explained by the hapten concept, according to which the small antigen covalently modifies an endogenous protein; recent studies show strong associations of several HLA molecules with hypersensitivity. In recent years, however, evidence has become stronger that not all drugs need to bind covalently to the major histocompatibility complex (MHC)-peptide complex in order to trigger an immune response. Rather, some drugs may bind reversibly to the MHC or possibly to the T-cell receptor (TCR), eliciting immune reactions akin to the pharmacological activation of other receptors. While the exact mechanism is still a matter of debate, noncovalent drug presentation clearly leads to the activation of drug-specific T cells. In some patients with hypersensitivity, such a response may occur within hours of even the first exposure to the drug. Thus, the reaction to the drug may not be the result of a classical, primary response but rather be mediated by existing, preactivated T cells that display cross-reactivity for the drug and have additional (peptide) specificity as well. In this way, certain drugs may circumvent the checkpoints for immune activation imposed by the classical antigen processing and presentation mechanisms, which may help to explain the idiosyncratic nature of many drug hypersensitivity reactions.

Synergy‐inducing chemokines enhance CCR2 ligand activities on monocytes

The migration of monocytes to sites of inflammation is largely determined by their response to chemokines. Although the chemokine specificities and expression patterns of chemokine receptors are well defined, it is still a matter of debate how cells integrate the messages provided by different chemokines that are concomitantly produced in physiological or pathological situations in vivo. We present evidence for one regulatory mechanism of human monocyte trafficking. Monocytes can integrate stimuli provided by inflammatory chemokines in the presence of homeostatic chemokines. In particular, migration and cell responses could occur at much lower concentrations of the CCR2 agonists, in the presence of chemokines (CCL19 and CCL21) that per se do not act on monocytes. Binding studies on CCR2+ cells showed that CCL19 and CCL21 do not compete with the CCR2 agonist CCL2. Furthermore, the presence of CCL19 or CCL21 could influence the degradation of CCL2 and CCL7 on cells expressing the decoy receptor D6. These findings disclose a new scenario to further comprehend the complexity of chemokine‐based monocyte trafficking in a vast variety of human inflammatory disorders.

Processing-dependent and -independent pathways for recognition of iodinated contrast media by specific human T cells.

Background One to three percent of patients exposed to intravenously injected iodinated contrast media (CM) develop delayed hypersensitivity reactions. Positive patch test reactions, immunohistological findings, and CM‐specific proliferation of T cells in vitro suggest a pathogenetic role for T cells. We have previously demonstrated that CM‐specific T cell clones (TCCs) show a broad range of cross‐reactivity to different CM. However, the mechanism of specific CM recognition by T cell receptors (TCRs) has not been analysed so far.

The p-i Concept: Evidence and Implications

For decades, the hapten concept has served as the paradigm which explains how low molecular weight antigens are capable of triggering an immune response. In accordance with this, many drugs have be