Cutaneous lymphocyte antigen and α4ß7 T-lymphocyte responses are associated with peanut allergy and tolerance in children.

BACKGROUND: It is unclear whether the initial route of allergen exposure in early life could influence the subsequent development of allergy, with cutaneous sensitization leading to peanut allergy (PA), and tolerance induced by oral exposure. The skin- and gastrointestinal (GI)-homing markers, cutaneous lymphocyte antigen (CLA) and α4ß7 integrin, are used to determine whether the state of PA correlates with peanut-specific CLA responses, with tolerance associated with predominant α4ß7 responses. METHODS: CLA+ and α4ß7+ memory T cells were isolated and cultured with peanut extract to assess their proliferation. Stimulation indices were compared in peanut allergic and non-allergic (NA) groups, and peanut-specific cytokine production was measured. RESULTS: In peanut allergic patients, peanut-specific proliferation predominates in the skin-homing CLA+ subset, whilst peanut-tolerant groups have a mixed CLA/α4ß7 response (P = 0.008). Comparison with a control food antigen (ovalbumin) showed that these differences are allergen specific. Cytokine responses showed trends towards Th1 skewing in the GI-homing α4ß7+ cells of peanut-tolerant groups and Th2 skewing in the skin-homing CLA+ cells of peanut allergic patients. CONCLUSION: The predominance of the CLA+ response to peanut in peanut allergic patients is consistent with the hypothesis that allergic sensitization occurs through the skin. The predominant α4ß7+ response in peanut-tolerant groups suggests that allergen exposure through the GI tract induces tolerance.

IgE-mediated facilitated antigen presentation underlies higher immune responses in peanut allergy.

BACKGROUND: Peanut allergy poses significant healthcare problems, because its prevalence is increasing in many countries, and it is rarely outgrown. To explore the immunological mechanisms that underlie peanut allergy and tolerance, we compared the peanut-specific responses of peanut-allergic (PA) and nonallergic (NA) individuals. METHODS: We measured peanut-specific peripheral blood mononuclear cells (PBMC) proliferation using tritiated thymidine. The frequency of peanut-specific T cells amongst PBMC was determined by carboxyfluorescein succinimidyl ester labelling. The role of IgE-dependent facilitated antigen presentation (FAP) in modulating proliferation was investigated by depleting IgE from plasma with anti-IgE-coated beads and then assessing PBMC proliferation in the presence of IgE-depleted or nondepleted plasma. RESULTS: We found that peanut-specific PBMC proliferation is higher and peaks earlier in PA than in NA donors. We investigated the immunological mechanisms that could underlie these differences. We found that both PA and NA have memory responses to peanut, but the frequency of peanut-specific T cells is higher in PA than in NA. Facilitated antigen presentation could cause both the higher proliferation and precursor frequency in PA. Facilitated antigen presentation activity in vitro was confirmed by showing that IgE depletion decreases proliferation, while adding IgE back restores it. CONCLUSION: Our results identify FAP as a mechanism that underlies higher responses to peanut in PA. In these individuals, high levels of peanut-specific IgE could furthermore maintain long-term allergic T-cell responses. We raise the question whether, in the future, therapies targeting IgE such as anti-IgE antibodies may be used to suppress these T-cell responses.

Tissue factor pathway inhibitor (TFPI) levels in the plasma and urine of children with meningococcal disease.

Tissue factor pathway inhibitor (TFPI) is a potent inhibitor of the TF-dependent coagulation system. In meningococcal disease, up-regulation of tissue factor expression on blood monocytes and possibly on endothelial cells has the potential to trigger the activation of the TF-dependent pathway of coagulation. Intravascular coagulation is considered to be a major pathogenic factor in meningococcal disease. We postulated that imbalance between TF expression and TFPI concentration might lead to uncontrolled coagulation in meningococcal disease. The aim of this study was to assess the levels of total TFPI in the plasma of patients with meningococcal disease and assess whether increased leaking of the TFPI was occurring. TFPI antigen levels and activity were measured in the plasma of 54 patients with meningococcal disease, and 13 healthy control children. TFPI antigen level were also determined in the urines of 14 of the 54 and 9 healthy control children. Plasma TFPI activity was reduced in the meningococcal diseased patients (mean of 0.503 +/- 0.341 U/ml; control, 1.010 +/- 0.199 U/ml: p <0.0001), as was the TFPI antigen levels (mean of 54.85 +/- 35.05 ng/ml; Control, 94.51 +/- 11.44 ng/ml; p <0.0001). In contrast, TFPI antigen levels were increased in the urine of these patients when compared to the levels found in the urine of the healthy control children (mean of 12.96 +/- 5.392 ng/mmol creatinine; Control, 0.239 +/- 0.191 ng/mmol creatinine; p <0.035). A lack of correlation between TFPI-activity and TFPI-antigen plasma levels was observed (r = 0.002, p = 0.85). This data is consistent with the hypothesis that in meningococcal disease there is increased inactivation of plasma TFPI by the up regulation of tissue factor expression but in addition increased clearance of TFPI in urine is occurring.

Suppression of human monocyte tissue factor synthesis by antisense oligodeoxynucleotide.

Tissue factor (TF) is a cellular receptor and cofactor for factor V11/V11a which initiates the blood coagulation cascade. An understanding of TF cell biology is therefore of critical importance to the pathophysiology of many disorders. We have utilized an antisense oligodeoxynucleotide (aODN), directed against human blood monocyte TF mRNA, to investigate the feasibility of inhibiting the synthesis of TF. CD14 receptor-mediated endocytosis was used as a means of delivering TF aODN to monocytes. This DNA carrier system consists of the fab portion of anti-CD14 antibody covalently linked to poly (L-lysine). Co-treatment of monocytes with TF aODN and endotoxin resulted in 80.4 +/- 2.2% suppression of TF activity when compared with control endotoxin stimulated cells. Control experiments with TF sense ODN, mismatched aODN, and an irrelevant aODN were performed to exclude nonspecific inhibitory effects. The cytotoxicity of the DNA carrier complex was also evaluated. These results demonstrate that this TF mRNA antisense ODN specifically suppressed the synthesis of biologically active monocyte TF and that antisense ODNs might therefore represent a useful tool in the investigation of monocyte/macrophage TF function.

Co-localization of tissue factor protein and its mRNA in human blood monocytes.

Triggering of both the extrinsic and intrinsic coagulation pathways is mediated by the cell surface receptor Tissue Factor (TF). The ability to observe the cell surface TF protein and its mRNA at the single cell level would facilitate our understanding of the cellular biology of TF in health and diseased states. Employing the methods of immuno-gold silver staining and in situ hybridization using non-isotopically labelled oligoprobes, tissue factor antigen and mRNA were detected simultaneously in endotoxin stimulated human peripheral blood monocytes.

Production of tissue factor by monocyte progenitor cells.

Tissue factor (TF) is known to be produced by monocytes from human peripheral blood. However the production of this factor by haematopoietic progenitor cells is not yet known. We thus studied human monocyte progenitor cells isolated from bone marrow of normal and diseased individuals. These cells were non-adherent, monocytic and able to phagocytose particles ranging from 0.3-1 microns. Unactivated partial thromboplastin time clotting assay demonstrated procoagulant activity consistent with TF function, which was blocked by a neutralizing anti-TF monoclonal antibody, G12. The production of TF messenger RNA was demonstrated on dot blot and northern blot analysis utilizing an oligonucleotide probe.