Exosomes Represent an Immune Suppressive T Cell Checkpoint in Human Chronic Inflammatory Microenvironments.

Background: T cells present in chronic inflammatory tissues such as nasal polyps (from chronic rhinosinusitis patients) have been demonstrated to be hypo-responsive to activation via the TCR, similar to tumor-specific T cells in multiple different human tumor microenvironments. While immunosuppressive exosomes have been known to contribute to the failure of the tumor-associated T cells to respond optimally to activation stimuli, it is not known whether they play a similar role in chronic inflammatory microenvironments. In the current study, we investigate whether exosomes derived from chronic inflammatory microenvironments contribute to the immune suppression of T cells. Methods: Exosomes were isolated by ultracentrifugation and characterized by size and composition using nanoparticle tracking analysis, scanning electron microscopy, antibody arrays and flow exometry. Immunosuppressive ability of the exosomes was measured by quantifying its effect on activation of T cells, using nuclear translocation of NFκB as an activation endpoint. Results: Exosomes were isolated and characterized from two different types of chronic inflammatory tissues - nasal polyps from chronic rhinosinusitis patients and synovial fluid from rheumatoid arthritis patients. These exosomes arrest the activation of T cells stimulated via the TCR. This immune suppression, like that which is seen in tumor microenvironments, is dependent in part upon a lipid, ganglioside GD3, which is expressed on the exosomal surface. Conclusion: Immunosuppressive exosomes present in non-malignant chronic inflammatory tissues represent a new T cell checkpoint, and potentially represent a novel therapeutic target to enhance the response to current therapies and prevent disease recurrences.

Humanized mouse model used to monitor MUC gene expression in nasal polyps and to preclinically evaluate the efficacy of montelukast in reducing mucus production.

OBJECTIVES: To determine whether MUC gene expression could be down-regulated in nasal polyps by the leukotriene receptor antagonist montelukast, we developed a system in which nondisrupted human nasal polyps could be successfully implanted into severely immunocompromised mice, and in which the histopathology of the original nasal polyp tissue could be preserved for long periods. In addition, the histopathologic changes in the human nasal polyps were carefully examined to determine the origin of the submucosal glands (SMGs) that develop in true nasal polyps found in the anterior third of the nose. METHODS: Small, nondisrupted pieces of human nasal polyp tissues were subcutaneously implanted into NOD-scid IL-2rgamma(null) mice. Xenograft-bearing mice were treated with either montelukast or saline solution. Xenografts at 8 to 12 weeks after implantation were examined histologically, and expression of MUC genes 4, 5AC, and 7 was studied in the polyps before implantation and in the 8-week xenograft. Alzet pumps were inserted into the mice, and montelukast (Singulair) was continuously delivered to determine its effect on goblet cell hyperplasia, mucus production, and the enlargement of nasal polyps over an 8-week period. RESULTS: The xenografts were maintained in a viable and functional state for up to 3 months and retained a histopathology similar to that of the original tissue, but with a noticeable increase in goblet cell hyperplasia and marked mucus accumulation in the SMGs. MUC4 and MUC5AC were significantly increased in the xenograft 8 weeks after implantation, but MUC7 was significantly decreased compared to the preimplantation polyps. Inasmuch as MUC7 is found exclusively in serous glands, the findings suggest that serous glands are not found in polyps in the anterior third of the nose. The histopathologic findings confirm the original findings of Tos et al suggesting that the SMGs are derived from pinching-off of the epithelium of the enlarging polyp following inflammatory changes. These SMGs have the same epithelium as surface epithelium and consist of multiple goblet cells that secrete periodic acid Schiff stain-positive mucin into the interior of the SMGs. A progressive increase in the volume of the xenografts was observed, with little or no evidence of mouse cell infiltration into the human leukocyte antigen-positive human tissue. An average twofold increase in polyp volume was found 2 months after engraftment. Montelukast did not decrease the growth of the xenograft in the 8-week NOD-scid mice, nor did it affect MUC gene expression. CONCLUSIONS: The use of innate and adaptive immunodeficient NOD-scid mice homozygous for targeted mutations in the IL-2 gamma-chain locus NOD-scid IL-2r gamma(null) for establishing engraftment of nondisrupted pieces of human nasal polyp tissues represents a significant advancement in studying chronic inflammation over a long period of time. In the present study, we utilized this humanized mouse model to confirm our prediction that MUC genes 4 and 5AC are highly expressed and significantly increased over those of preimplanted polyps. The overexpression of these 2 MUC genes correlates with both the goblet cell hyperplasia and the excessive mucus production that are found in nasal polyp xenografts. MUC7, which is primarily associated with the submucosa, as opposed to MUC4 and MUC5AC, which are primarily expressed in the epithelium, was significantly decreased in the nasal polyp xenografts. Montelukast had no significant effect on MUC gene expression in the xenografts. In addition to the MUC gene expression patterns, the histology of the xenografts supports the concept that mucinous glands that are characteristic of true nasal polyps are significantly different from those in the mucosa found in the lateral wall of the nose in patients with chronic sinusitis without nasal polyps. The mucinous glands seen in nasal polyps (which appear to be derived from an invagination of hyperplastic epithelial mucosa containing large numbers of goblet cells) are histologically distinct from the seromucinous glands found in the submucosa of hyperplastic middle turbinates. The data presented here establish a humanized mouse model as a viable approach to study nasal polyp growth, to assess the therapeutic efficacy of various drugs in this chronic inflammatory disease, and to contribute to our understanding of the pathogenesis of this disease.

Human nasal polyp microenvironments maintained in a viable and functional state as xenografts in NOD-scid IL2rgamma(null) mice.

OBJECTIVES: The objective was to develop a model with which to study the cellular and molecular events associated with nasal polyp progression. To accomplish this, we undertook to develop a system in which nondisrupted human nasal polyp tissue could be successfully implanted into severely immunocompromised mice, in which the histopathology of the original nasal polyp tissue, including inflammatory lymphocytes, epithelial and goblet cell hyperplasia, and subepithelial fibrosis, could be preserved for prolonged periods. METHODS: Small, non-disrupted pieces of human nasal polyp tissues were subcutaneously implanted into NOD-scid IL2rgamma(null) mice. Xenografts at 8 to 12 weeks after implantation were examined histologically and immunohistochemically to identify human inflammatory leukocytes and to determine whether the characteristic histopathologic characteristics of the nasal polyps were maintained for a prolonged period. The xenografts, spleen, lung, liver, and kidneys were examined histologically and immunohistochemically and were evaluated for changes in volume. The sera of these mice were assayed for human cytokines and immunoglobulin. RESULTS: Xenografts of human nasal polyp tissues were established after their subcutaneous implantation into NOD-scid IL2rgamma(null) mice. The xenografts were maintained in a viable and functional state for up to 3 months, and retained a histopathologic appearance similar to that of the original tissue, with a noticeable increase in goblet cell hyperplasia and marked mucus accumulation in the submucosal glands compared to the original nasal polyp tissue. Inflammatory lymphocytes present in the polyp microenvironment were predominantly human CD8+ T cells with an effector memory phenotype. Human CD4+ T cells, CD138+ plasma cells, and CD68+ macrophages were also observed in the xenografts. Human immunoglobulin and interferon-gamma were detected in the sera of xenograft-bearing mice. The polyp-associated lymphocytes proliferated and were found to migrate from the xenografts to the spleens of the recipient mice, resulting in a significant splenomegaly. A progressive increase in the volume of the xenografts was observed with little or no evidence of mouse cell infiltration into the human leukocyte antigen-positive human tissue. An average twofold increase in polyp volume was found at 3 months after engraftment. CONCLUSIONS: The use of innate and adaptive immunodeficient NOD-scid mice homozygous for targeted mutations in the interleukin-2 receptor gamma-chain locus NOD-scid IL2rgamma(null) for establishing xenografts of nondisrupted pieces of human nasal polyp tissues represents a significant improvement over the previously reported xenograft model that used partially immunoincompetent CB17-scid mice as tissue recipients. The absence of the interleukin-2 receptor gamma-chain results in complete elimination of natural killer cell development, as well as severe impairments in T and B cell development. These mice, lacking both innate and adaptive immune responses, significantly improve upon the long-term engraftment of human nasal polyp tissues and provide a model with which to study how nasal polyp-associated lymphocytes and their secreted biologically active products contribute to the histopathology and progression of this chronic inflammatory disease.

Human nasal polyp microenvironment maintained in viable and functional states as xenografts in SCID mice.

OBJECTIVES: We undertook to maintain human nasal polyp tissue in a viable and functional state in SCID (severe combined immunodeficiency) mice. METHODS: Small, nondisrupted pieces of human nasal polyp tissues were subcutaneously implanted into SCID mice depleted of natural killer cells. The resulting xenografts were examined histologically, and the sera were evaluated for the presence of human protein. RESULTS: The original histologic architecture of the polyp was maintained in the xenografts. The tissues, including pseudostratified columnar epithelial-lined polyps and subepithelial stroma, remained viable, and goblet cells continued to produce mucin for up to 26 weeks after engraftment. Human inflammatory leukocytes, including CD3+ T cells, CD20+ B cells, CD138+ plasma cells, and CD68+ monocytes and/or macrophages, were present. Identification of human immunoglobulin and human interferon-gamma in the sera of xenograft-bearing mice indicated that the B cells or plasma cells and T cells within the xenografts remained functional for 2 weeks after engraftment. CONCLUSIONS: The ability to engraft and maintain nasal polyps provides an in vivo human/mouse chimeric model with which to investigate the role of inflammatory leukocytes and stromal cells in the maintenance and progression of polyposis and to determine how exogenous cytokines may alter the interaction of inflammatory cells, stromal cells, and epithelial cells in the polyp.

Bacterial interference of penicillin-sensitive and -resistant Streptococcus pneumoniae by Streptococcus oralis in an adenoid organ culture: implications for the treatment of recurrent upper respiratory tract infections in children and adults.

OBJECTIVES: The role of the viridans group of streptococci (Streptococcus oralis) in the prevention of colonization with Streptococcus pneumoniae was investigated in an adenoid organ culture system. METHODS: The adenoids from 10 patients who were undergoing adenoidectomy for either hypertrophy or recurrent otitis media were used. RESULTS: Streptococcus oralis Parker and S. oralis Booth (two organisms isolated from the nasopharynges of patients undergoing adenoidectomy only and patients undergoing adenoidectomy and bilateral tympanostomy with tubes, respectively) uniformly inhibited both penicillin-sensitive and penicillin-resistant S. pneumoniae. Although both strains of S. oralis inhibited the growth of both S. pneumoniae strains, strain Parker provided more complete inhibition than did strain Booth. CONCLUSIONS: The results indicate that some strains of S. oralis may inhibit the growth of the most serious pathogens in the nasopharynx. It is therefore possible that colonization of inhibitory strains of viridans streptococci may be used in the nasopharynx as a relatively safe and inexpensive approach to prevention of recurrent otitis media in some children and of recurrent suppurative sinusitis in both children and adults.

Superantigen hypothesis for the early development of chronic hyperplastic sinusitis with massive nasal polyposis.

PURPOSE OF REVIEW: The pathogenesis, pathophysiology, and immunobiology of chronic hyperplastic sinusitis with massive nasal polyposis are starting to become unraveled. Allergy, viral infection, bacterial infection, fungal infection, and environmental pollution have all been suggested as possible initial triggers that may upregulate inflammation of the lateral wall of the nose to develop nasal polyposis. The purpose of this review is to present data from our laboratory that suggest that one of the possible early events in the development of inflammation of the lateral wall of the nose in chronic hyperplastic sinusitis with massive nasal polyposis is the production of exotoxins from Staphylococcus aureus. The exotoxins may act as superantigens and cause activation and clonal expansion of lymphocytes with specific Vbeta regions, resulting in massive cytokine production. RECENT FINDINGS: Recent published studies suggest that S. aureus is the most common organism isolated from the mucus adjacent to massive nasal polyposis. Staphylococci produce exotoxins. These exotoxins, sometimes known as enterotoxins, include SEA, SEB, and TSST-1. These exotoxins are capable of acting as superantigens and therefore, reacting with T lymphocytes with specific Vbetas in the lateral wall of the nose. Thereafter, it is possible that these lymphocytes are stimulated to produce both TH1 and TH2 cytokines, which have also been demonstrated in the nasal polyp. The consequence of these findings may be the upregulation and increased survival of eosinophils in the nasal polyp. SUMMARY: Staphylococcus aureus is present in the mucin adjacent to nasal polyps in about 60 to 70% of cases of massive nasal polyposis. These organism, as studied up to the present, always produce exotoxins, which may act as superantigens, causing activation and clonal expansion of lymphocytes with specific Vbeta region in the lateral wall of the nose. The present review suggests that activation of these lymphocytes produce both TH1 and TH2 cytokines. The potential damage to the nasal mucosa from eosinophils is briefly discussed. Theoretically, topical antibiotics to suppress the colonization of S. aureus may be a logical approach to downregulate the production of superantigen in the lateral wall of the nose after appropriate endoscopic sinus surgery.

Update on the molecular biology of nasal polyposis.

The definition of CRS with and without nasal polyposis continues to evolve. It may require an understanding of a broader range of etiologies and pathogenesis than bacterial or viral infection. One must know whether the inflammation is of infectious or noninfectious origin. Therapeutic options will include pharmacotherapies and surgery. The pharmacotherapeutic approach will include antibiotics, systemic and topical steroids, possibly antifungals, novel anti-inflammatory therapies such as the use of antibodies directed against inflammatory cytokines and antileukotrienes, and perhaps low-dose macrolide therapy. In the case of massive nasal polyposis, modern surgical techniques will have to be performed before these therapeutic options will be possible. Finally, the use of topical diuretics such as amiloride and furosemide has been studied and the initial responses seem to be encouraging.

Molecular analysis of bacterial flora associated with chronically inflamed maxillary sinuses.

Chronic maxillary sinusitis is a chronic inflammatory condition in which the role of microbial infection remains undefined. Bacteria have been isolated from chronically inflamed sinuses; however, their role in the chronicity of inflammation is unknown. The objective of this study was to determine whether bacteria are present in clinical samples from chronic maxillary sinusitis and to assess the diversity of the flora present. Washes and/or tissue samples from endoscopic sinus surgery on 11 patients with chronic maxillary sinusitis were subjected to PCR amplification of bacterial 16S rDNA using three universal primer pairs, followed by cloning and sequencing. The samples were also assessed for the presence of bacteria and fungi by conventional culture methods. Viable bacteria and/or bacterial 16S rDNA were detected from maxillary sinus samples of five of the 11 patients examined (45 %). Three sinus samples were positive by both PCR and culture methods, one was positive only by PCR, and one only by culture. Thirteen bacterial species were identified: Abiotrophia defectiva, Enterococcus avium, Eubacterium sp., Granulicatella elegans, Neisseria sp., Prevotella sp., Pseudomonas aeruginosa, Serratia marcescens, Staphylococcus aureus, Stenotrophomonas maltophilia, Streptococcus gordonii, Streptococcus mitis/Streptococcus oralis and Streptococcus sp. Fungi were not detected. In one patient Streptococcus mitis/Streptococcus oralis, and in another patient Pseudomonas aeruginosa, were detected from both the sinus and the oral cavity using species-specific PCR primers. These results suggest that both aerobic and anaerobic bacteria can be detected in nearly half of chronic maxillary sinusitis cases.

Treatment of recurrent chronic hyperplastic sinusitis with nasal polyposis.

OBJECTIVE: To demonstrate the long-term efficacy of intranasal furosemide, an inhibitor of the sodium chloride cotransporter channel at the basolateral surface of the respiratory epithelial cell, vs no therapeutic intervention vs intranasal mometasone furoate, a corticosteroid, in preventing relapses of chronic hyperplastic sinusitis with nasal polyposis. DESIGN: Randomized prospective controlled study. Patients were examined every 6 months during follow-up (range, 1-9 years). PATIENTS: One hundred seventy patients with bilateral obstructive or minimally obstructive chronic hyperplastic sinusitis with nasal polyposis. INTERVENTION: All patients were surgically treated in the ENT Department, University of Siena Medical School. One month after surgery, group 1 patients (n = 97) started treatment with intranasal furosemide, group 2 (n = 40) received no therapeutic treatment, and group 3 (n = 33) were treated with mometasone. MAIN OUTCOME MEASURES: Clinical and instrumental evaluation of postoperative outcomes. RESULTS: Seventeen (17.5%) of 97 patients in group 1, 12 (30.0%) of 40 patients in group 2, and 8 (24.2%) of 33 patients in group 3 experienced nasal polyposis relapses. We noted a prevalence of early-stage relapse in patients treated with furosemide or mometasone, whereas patients who did not receive any treatment experienced more severe grades of chronic hyperplastic sinusitis with nasal polyposis (P<.005). CONCLUSION: Use of intranasal furosemide represents a valid therapeutic treatment in the prevention of chronic hyperplastic sinusitis with nasal polyposis.

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.

Interference of nontypeable Haemophilus influenzae and Moraxella catarrhalis by Streptococcus oralis in adenoid organ culture: a possible strategy for the treatment of the otitis-prone child.

The role of viridans group streptococci (Streptococcus oralis) in the prevention of colonization with nontypeable Haemophilus influenzae and Moraxella catarrhalis was investigated in an adenoid organ culture system. The adenoids from 100 patients who were undergoing adenoidectomy for either hypertrophy or recurrent otitis media were used. Streptococcus oralis Parker uniformly inhibited colonization with nontypeable H. influenzae or M. catarrhalis over a 24-hour period of incubation in adenoid organ culture. Streptococcus oralis Booth, a noninhibitory strain, did not significantly reduce colonization with nontypeable H. influenzae and M. catarrhalis. The results indicate that some strains of S. oralis may inhibit colonization with potential pathogens in the nasopharynx. It is therefore possible that colonization with inhibitory strains of viridans streptococci may be used in the nasopharynx as a relatively safe and inexpensive approach to prevention of recurrent otitis media in some children.

Memory T cells in the chronic inflammatory microenvironment of nasal polyposis are hyporesponsive to signaling through the T cell receptor.

A majority of T cells from chronic inflammatory tissues derived from patients with nasal polyposis were found to express an effector memory phenotype. We report here that these memory T cells failed to activate NF-κB in response to TCR stimulation but responded normally when the proximal TCR signaling molecules were bypassed with PMA and ionomycin. The dysfunction of these cells was associated with a decrease in the phosphorylation of several TCR proximal signaling molecules including ZAP70, Lck and SLP-76. In addition to the disruption in the TCR signaling pathway, the nasal polyp-associated T cells were shown to have a defect in their ability to translocate LAMP-1 to the cell surface. The results presented here establish that the phenotype and anergy of the T cells in the nasal polyp are similar to those which is seen in memory T cells derived from human tumors and other sites of chronic inflammation.

Bacterial interference in upper respiratory tract infections: a systematic review.

BACKGROUND: Published definitions of bacterial interference (BI) differ, some focusing on changes in the normal flora and others on changes in subsequent infection. A need for consensus was identified at a roundtable discussion of BI in upper respiratory tract infections (URTI). We conducted a systematic review of the available data to justify a consensus definition of BI specific to URTI as "a dynamic, antagonistic interaction between at least 2 organisms that affects the life cycle of each, changes the microenvironment, and alters the organisms' colonization, invasiveness, and ability to affect the health of the host." METHODS: Continued communication among the faculty postroundtable was used to identify and refine the search criteria to (1) in vitro and in vivo studies assessing bacterial URTI, (2) BI evaluated by response to treatment of URTI with antimicrobial agents, and (3) bacterial function in relation to interactions between normal (nonpathogenic) and pathological flora. The criteria were applied to systematic searches of MEDLINE (1950 onward), EMBASE (1974 onward), and the Cochrane Library (2007). RESULTS: Twenty-nine studies met the inclusion criteria, most focused on children with recurrent infections. Qualitative analysis supports the consensus definition. Interfering organisms affected the life cycle of test pathogens and inhibited their colonization, invasiveness, and health outcomes. Data were insufficient for statistical analysis. CONCLUSION: Interactions between interfering organisms and potential pathogens isolated from the same host can alter response to infection and treatment. More studies are needed, particularly in adults, to understand the role of interfering organisms, the influence of antibiotics, and the potential for recolonization posttreatment.

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.

Genetic polymorphisms in chronic hyperplastic sinusitis with nasal polyposis.

OBJECTIVES/HYPOTHESIS: Although many proinflammatory cytokines have been identified in nasal polyp tissue, the initial trigger that causes this inflammation characterized by edema, lymphocytosis, and eosinophilia, is still unknown. The purpose of the present study is to identify the presence of genetic polymorphisms in proinflammatory, anti-inflammatory, and chemokine genes that might contribute to genetic susceptibility to chronic hyperplastic sinusitis with nasal polyposis (CHSwNP). STUDY DESIGN: Case control study. METHODS: Buccal swabs were taken from the left and right oral mucosal surfaces from 179 patients with CHSwNP and 153 nonpolyposis controls with the Purgene DNA purification protocol (Gentra). Genotyping assays for cytokine gene loci were performed on 14 cytokine genes using the iPlex Gold and the Mass Array Compact system (Sequenom, San Diego, CA). Tests of Hardy-Weinberg equilibrium proportions were performed separately in the cases and controls. Tests for evidence of association between alleles at each single-nucleotide polymorphism (SNP) and case-control status were performed using unconditional logistic regression. RESULTS: The frequency of the A allele in a SNP located in tumor necrosis factor (TNF)-alpha (rs1800629) is significantly different in patients with nasal polyposis versus controls without nasal polyposis, 18.6% and 11.5%, respectively with an individuals' odds of susceptibility to nasal polyps increasing almost two-fold (odds ratio, 1.86; confidence interval, 1.4-3.09) given at least one copy of the A allele at this SNP. All other cytokine gene polymorphisms of both inflammatory, anti-inflammatory, and chemokine genes were not statistically different between the two groups. CONCLUSIONS: TNF-alpha-308, a SNP in the promoter region of this cytokine gene is associated with increased odds of developing nasal polyposis. TNF-alpha is a potent immuno-mediator and proinflammatory cytokine that has been implicated in the pathogenesis of a large number of human diseases. The location of this gene on the short arm of chromosome 6, with the major histocompatibility complex genes and complement, has raised the probability that polymorphism within this locus may contribute to a genetic association of this region of the genome with a wide variety of infectious and autoimmune diseases.

IL-12 reverses anergy to T cell receptor triggering in human lung tumor-associated memory T cells.

Memory T cells in human non-small cell lung cancer are unresponsive to progressing tumors. T cells were evaluated at the single cell level by imaging the nuclear translocation of NF-kappaB and NFAT via immunofluorescence confocal microscopy as an early measure of responsiveness to T cell receptor triggering. Little translocation of NF-kappaB or NFAT occurred in tumor-associated T cells in response to CD3+CD28 cross-linking under conditions which led to maximal translocation in normal donor peripheral blood T cells. TNF-alpha induced maximal NF-kappaB translocation in these T cells, indicating that they remain receptive to alternative signaling pathways, and pulsing with IL-12 prior to TCR triggering reversed their apparent anergy. T cells from additional chronic inflammatory microenvironments demonstrated a similar refractoriness to TCR activation, suggesting either that a common regulatory mechanism present within the microenvironment controls these cells or that with continuous antigen exposure, they remain refractory to activation via the TCR.

Immunologic aspects of otitis media.

The middle ear cleft is a modified gas pocket which functions normally when the gas contents are regulated by a normal eustachian tube, resulting in equalization of middle ear pressure to that of the environment. The most important regulator of this middle ear pressure is the eustachian tube, a critical passageway from the nasopharynx into the middle ear. Any alteration of eustachian tube mucociliary function caused by virus, allergy, pollutants, or alteration of the normal homeostasis of the nasopharynx will result in eustachian tube obstruction. This, in turn, leads to underventilation of the middle ear, and transudation of fluid. If bacteria or virus or viral-bacterial interaction leads to infectious disease of the middle ear, an immune response is produced as a result of the inflammatory response, allowing lymphocytes and antigen-presenting cells to enter into the middle-ear mucosa. This article summarizes the immunologic reactivity in the middle ear following a viral-bacterial inflammatory reaction in the middle-ear mucosa. Although secretory IgA is critical for protection of the nasopharynx, its function in the middle ear has still not been resolved. The evidence strongly suggests that IgG1 and IgG3 subclasses are responsible for eradication of middle ear pathogens. Finally, a review of alternative approaches to the prevention of otitis media is briefly discussed in this critical period of emergence of resistant bacteria to available antibiotics.

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.