Single nucleotide polymorphisms (SNPs) in key cytokines may modulate food allergy phenotypes.

Single nucleotide polymorphisms (SNPs) can play a direct or indirect role in phenotypic expression in food allergy pathogenesis. Our goal was to quantitate the expression of SNPs in relevant cytokines that were expressed in food allergic patients. SNPs in cytokine genes IL-4 and IL-10 are known to be important in IgE generation and regulation. We examined IL-4 (C-590T), IL-4Rα (1652A/G) and IL-10 (C-627A) SNPs using real-time PCR followed by restriction fragment length polymorphism (RFLP) analysis. Our results show that the AA, AG and GG genotypes for IL-4Rα (1652A/G) polymorphisms were statistically different in radioallergosorbent test (RAST) positive versus negative patients, and although no statistically significant differences were observed between genotypes in the IL-4 (C-590T) and IL-10 (C-627A) SNPs, we observed a significant decrease in IL-4 (C-590T) gene expression and increase in IL-4Rα (1652A/G) and IL-10 (C-627A) gene expression between RAST(+) versus RAST(-) patients, respectively. We also observed significant modulation in the protein expression of IL-4 and IL-10 in the serum samples of the RAST(+) patients as compared to the RAST(-) patients indicating that changes in SNP expression resulted in altered phenotypic response in these patients.

Further observations on the role of Staphylococcus aureus exotoxins and IgE in the pathogenesis of nasal polyposis.

OBJECTIVES/HYPOTHESIS: Recent studies have suggested that Staphylococcus aureus secrete exotoxins that may act as superantigens and upregulate the variable beta region of lymphocytes in chronic hyperplasticsinusitis with nasal polyposis (CHSwNP). The aim of this study was to add further information for correlating the presence of staphylococcal species and the upregulation of the V(ß) region of both nasal polyp lymphocytes and peripheral blood lymphocytes. Furthermore, IgE-mediated hypersensitivity directed against these exotoxins produces an additional independent immunologic mechanism in upregulating the inflammatory response in the lateral wall of the nose in nasal polyposis. STUDY DESIGN: Prospective study. METHODS: Nasal polyps were harvested from 38 patients with CHSwNP. Eleven patients were studied for the correlation of exotoxin from staphylococcal species and the variable beta region of lymphocytes in both the nasal polyp lymphocytes and corresponding peripheral blood lymphocytes. Eight additional patients with CHSwNP were studied for local and systemic IgE-mediated immunity directed against S aureus exotoxins. Flow cytometry was used to analyze the V(ß) repertoire of polyp-derived CD3-positive lymphocytes from 11 patients. S aureus was isolated from nine patients, and coagulase-negative Staphylococcus was isolated from two patients in whom at least 1 × 10(6) T cells could be isolated from their nasal polyps. A quantitative assay for IgE was developed to study the levels of this immunoglobulin directed against S aureus exotoxins in both the nasal polyp and the peripheral blood lymphocytes of 11 patients and in the nasal mucus and serum of eight additional patients. RESULTS: Eleven patients had T-cell V(ß) clonal expansion. S aureus exotoxin upregulated the corresponding V(ß) region of lymphocytes in both the nasal polyp T cells as well as the T cells from the peripheral blood in nine patients, and two patients with coagulase-negative Staphylococcus also demonstrated upregulation of the V(ß) region in the nasal polyps in the absence of exotoxins. In one patient, in vivo exotoxin was isolated, which correlated with the in vitro isolation from the organism itself. IgE directed against staphylococcal enterotoxin B (SEB) and toxic shock syndrome toxin was significantly elevated in the sera of patients with CHSwNP (P < .0001) as compared with the sera of age-related healthy control subjects; IgE directed against staphylococcal enterotoxin A and SEB (P = .0047) was significantly elevated in the mucus of eight patients with CHSwNP as compared with the nasal mucus of healthy controls. CONCLUSIONS: This study augments our understanding of the potential role of S aureus exotoxins behaving as superantigens in the lateral wall of the nose in CHSwNP. Furthermore, local nasal IgE directed against these exotoxins may create a local allergic inflammation in the lateral wall of the nose. These two immunologic mechanisms are independent but may be additive in the inflammatory process in CHSwNP.

In vivo effects of bifidobacteria and lactoferrin on gut endotoxin concentration and mucosal immunity in Balb/c mice.

The aim of the present study was to examine the effects of oral supplementation of newborn Balb/c mice with bifidobacteria (B. infantis, B. bifidum) and iron-free apo-lactoferrin (bovine, human) on gut endotoxin concentration and mucosal immunity. Endotoxin concentration was measured in ileocecal filtrates at 7, 14, 21, and 28 days postdelivery by a quantitative limulus amebocyte lysate test. While endotoxin levels in bifidobacteria-fed mice showed a steady rise over time, they were consistently lower than that observed in control animals. Results of lactoferrin supplementation varied depending on the specific time point, but overall by day 28, all treatment groups showed lower intestinal endotoxin concentrations compared to saline fed animals. Neither bifidobacteria nor lactoferrin stimulated an increase in B or T cells, or in cytokine production (IL-6, TNF-alpha, INF-gamma), in Peyer's patches as measured by flow cytometry. Bifidobacteria and lactoferrin were well tolerated as dietary supplements and showed promising potential to reduce gut endotoxin levels.

Lymphocyte subpopulations and cytokines in nasal polyps: is there a local immune system in the nasal polyp?

PURPOSE: The pathogenesis of chronic hyperplastic rhinosinusitis with massive nasal polyposis is still not entirely known. The present study evaluates the lymphocyte subpopulations and their production of cytokines using a technique for detection of intracytoplasmic cytokines by flow cytometry. This information may allow us to determine whether the source of these lymphocytes is from peripheral blood, the common mucosal immune system, or both. METHODS: Detection of intracytoplasmic cytokines by flow cytometry was performed using a fluoresceinated monoclonal antibody directed against CD4+ and CD8+ lymphocytes and a rhodamine-labeled intracytoplasmic monoclonal antibody directed against four cytokines. In this way, the percentage of lymphocytes synthesizing TH1 and TH2 cytokines were identified in nasal polyp lymphocytes and the corresponding peripheral blood lymphocytes of 13 patients. RESULTS: Lymphocytes producing interferon-gamma and IL-2, as well as IL-4 and IL-5, were found in the nasal polyps, suggesting that the nasal polyp possesses both TH1 and TH2 cytokine expression. There are also significant differences between the percentage of lymphocytes producing these cytokines between nasal polyps and peripheral blood, suggesting that nasal polyp lymphocytes derive from at least another source than only peripheral blood lymphocytes. Statistical analysis of four groups of patients demonstrated that no statistically significant difference in the lymphocyte subpopulations in atopic versus non-atopic patients, nor aspirin-intolerant versus aspirin-tolerant patients. In general, CD8 cells always produce more interferon-gamma than IL-2 in both peripheral blood and nasal polyps. In contrast with this data, CD4 cells produce more IL-2 in the peripheral blood than in nasal polyps. CONCLUSIONS: Data support the concept that nasal polyp lymphocyte subpopulations may be derived from both the local mucosal immune system as well as from random migration of peripheral blood lymphocytes secondary to adhesion molecules and chemokines, which are known to be present in nasal polyps.

A superantigen hypothesis for the pathogenesis of chronic hyperplastic sinusitis with massive nasal polyposis.

BACKGROUND: The pathogenesis of chronic hyperplastic sinusitis with massive nasal polyposis is still an enigma; however, the molecular biology of this disease is beginning to become unraveled and the proinflammatory cytokines and the message and the product of these cytokines have all been identified in nasal polyps. However, the initial trigger that causes inflammation of the lateral wall of the nose to up-regulate lymphocytes and eosinophils is still unknown. METHODS: Thirteen patients with massive polyposis were studied. The mucus of the nasal cavities surrounding the nasal polyps was studied for both bacterial and fungal species. The lymphocytes of the nasal polyps were extracted and evaluated for the T-cell receptor, particularly, the variable beta region of this receptor. Enterotoxins (superantigens) of the bacteria were studied. Finally, the histopathology of nasal polyps was studied. RESULTS: Fifty-five percent of the patients had toxin-producing Staphylococcus aureus in the nasal mucus adjacent to the polyps. Three different enterotoxins were isolated, including Staphylococcus enterotoxin A, Staphylococcus enterotoxin B, and toxic shock syndrome toxin 1. The variable B specificity for these superantigens was identified also in the polyp lymphocyte T-cell receptor. CONCLUSION: A superantigen hypothesis for massive polyposis is suggested because the most common bacterial species found in the nasal mucus is Staphylococcus aureus. These bacteria produce enterotoxins in all of the cases studied and the corresponding variable beta region of the T-cell receptor also was up-regulated in the polyp lymphocytes in cases studied thus far. These data taken together suggest that the initial injury to the lateral wall of the nose may be the result of toxin-producing Staphylococci. Superantigens (enterotoxins) may up-regulate lymphocytes to produce cytokines that are responsible for the massive up-regulation of lymphocytes, eosinophils, and macrophages, the three most common inflammatory cells found in massive nasal polyposis.

Immune responses in rhesus rotavirus-challenged BALB/c mice treated with bifidobacteria and prebiotic supplements.

Bifidobacterium species (B. bifidum and B. infantis), with or without prebiotic compounds (arabino-galactan, short-chain fructo-oligosaccharide, iso-malto-dextrins), were orally fed to Balb/c pups (n = 192) to evaluate their potential synergistic effects on modulating the course of rhesus rotavirus (RRV) infection, as well as their ability to mediate the associated mucosal and humoral immune responses. Rotavirus-specific IgA and IgG in serum, rotavirus antigen, and specific IgA in feces were measured by ELISA. Mucosal total IgA and IgG levels were determined in Peyer's patches by flow cytometry. Significantly delayed onset (p = 0.001) and early resolution (p < 0.001) of diarrhea were observed in bifidobacteria-treated, RRV-infected mice compared with RRV-infected control mice. Supplementation with prebiotic compounds did not shorten the clinical diarrhea course more than that observed with bifidobacteria treatment alone. Rotavirus-specific IgA in feces was 16-fold elevated on d 5 postinfection in bifidobacteria-treated, RRV-infected mice compared with the RRV-infected alone group. In addition, the level of rotavirus-specific IgA in serum was four-fold higher in bifidobacteria-treated, RRV-infected litters versus mice challenged with RRV alone on 28 and 42 d postinfection. No enhancement of the immune response was found in RRV-infected mice that were treated with both bifidobacteria and prebiotic compounds over those treated with bifidobacteria only. The findings suggest that bifidobacteria may act as an adjuvant by modulating early mucosal and strong humoral rotavirus-specific immune responses, and mitigate severity of rotavirus-induced diarrhea.