Methods br Results br Discussion It is
Discussion It is well established that CRSwNP is characterized by TH2 inflammation and eosinophilia.22, 27 Yet it is becoming increasingly apparent that B cells might play an important role in the inflammatory response within the sinus tissue of patients with CRSwNP.8, 9 The lungs and upper airways represent an important mucosal immune site that is in constant contact with airborne antigens and microbial organisms. Many studies have focused on elucidating the mechanisms involved in the induction and maintenance of B-cell responses within the gut mucosa in animal models and human subjects, but there is a paucity of data regarding these mechanisms in the airway mucosa, especially in human subjects (Kato et al, unpublished data). CRS provides us with a unique opportunity to investigate an ongoing inflammatory response in human tissues, in part because the affected tissue is relatively easily accessible, even in healthy control subjects. In the current study we have expanded on our earlier work and found striking evidence of B-cell inflammation and local antibody production in NPs from patients with CRSwNP.8, 9 NP tissue from patients with CRS not only contained increased numbers of inflammatory cells (Fig 1) but also contained significantly more B cells, plasma cells, and Latrunculin B (Fig 2, Fig 3, Fig 4) compared with UT. Whether these B cells enter the tissue as naive cells and become activated or if they enter as memory cells primed to respond within the tissue is not yet clear. Elucidating where B cells and plasma cells are activated in this disease can provide valuable insight and potential new avenues of investigation for therapeutic interventions. Because we know that levels of B cell–activating factor of the TNF family are highly increased in polyp tissue and correlate with expression of CD20, it is tempting to speculate that B cells that do traffic through the polyp tissue will find a favorable microenvironment for activation and differentiation within the tissue itself. Preliminary experiments from our laboratory indicate that NP tissue supports the survival of plasma cells in vitro (data not shown), but the mechanisms involved are presently under investigation. Further studies are needed to determine whether B cells can be activated within the tissue itself and what mechanisms might be involved. We have previously shown increased levels of autoantigen-specific antibodies in NPs, and others have found increased IgE antibody levels directed against staphylococcal enterotoxins in NPs and evidence for local production of IgE, strengthening the argument that enhanced local antibody production might be an important component of CRSwNP pathogenesis. We have extended these findings in this study by demonstrating that the levels of many different antibody isotypes were increased in NPs of patients with CRSwNP but not in sera (Figs 3 and 4 and see Fig E3). This broad array of antibodies could have multiple activating effects on cells known to reside in NPs, including mast cells, eosinophils, and neutrophils. Notably, one of the most highly increased isotypes in NPs was IgG4 (Fig 3), which is often thought to play a protective role in allergic disease. However, it is also well established that IgE can be generated by a 2-step process that involves an IgG4 intermediate. Because levels of both IgE and IgG4 were highly increased in NPs, future studies will investigate whether sequential class-switch recombination through an IgG4 intermediate plays a role in the generation of IgE in NPs. Interestingly, we found reduced expression of pIgR in NPs (Fig 3), despite the fact that pIgR expression can be induced by local immunoglobulin production. Because pIgR is known to be expressed in glandular epithelium, its reduced expression in NPs might be due to the recently documented reduction of glands in NPs. Moreover, it is likely that antibody secretion from NPs constitutes only a small fraction of the total antibodies recovered in nasal lavage fluid, and thus it is difficult to say with certainty that this is a good representation of NP-specific secretion. Nevertheless, the reduced pIgR expression fits well with the fact that we did not find significantly increased IgA or IgM levels in the nasal lavage fluid of patients with CRSwNP (Fig 4 and see Fig E3). In fact, the increased IgA and IgM levels in NP extracts might be a reflection of an inability of the pIgR system to upregulate its function and adequately accommodate the increased antibody production within the NP microenvironment. Accumulation of IgA in the tissue might further exacerbate inflammation through degranulation of eosinophils, and this could also be an important component of CRS pathogenesis. Although the antigen specificity of NP IgA is unknown, our previous studies have shown that at least some of the IgA in NPs is directed against autoantigens. Future studies will investigate whether antibodies derived from NPs are capable of activating innate effector cells, as well as assessing their specificities and the extent of clonal expansion within the tissues. In addition, although we have focused on antibody production as a key effector function of B cells in this work, it is possible that B cells and plasma cells within NPs play multiple roles during chronic inflammation (eg, T-cell costimulation, chemokine expression, cytokine expression, and regulatory roles), and further studies are needed to elucidate other mechanisms used by B cells during CRS pathogenesis.