Pathophysiological effects of glucocorticoids on nasal polyps: an update

PURPOSE OF REVIEW: The exact mechanisms by which glucocorticoids exert their beneficial effects on nasal polyps are not clearly defined. Nasal polyps, asthma and allergic rhinitis share common features such as mucosal infiltration with eosinophils and mast cells as well as local IgE production. The present review is an update on the pathophysiological mechanisms of glucocorticoids on nasal polyps described during the last 2 years. RECENT FINDINGS: The reduction of leukocyte numbers in nasal polyps following glucocorticoid treatment depends on several mechanisms, for example altered balance between the two isoforms of the human glucocorticoid receptors, GRalpha and GRbeta. Another explanation may be inhibition of CD4+ T by CD8+ T cells. Increased expression of the antiinflammatory cytokine transforming growth factor beta may contribute to this. A DNA microarray study which examined the expression of some 22 000 genes showed increased expression of several antiinflammatory genes in nasal polyps after treatment with glucocorticoids. The antiinflammatory gene that increased most was uteroglobin (also known as Clara cell protein 16) which is abundantly expressed in airway secretions and thought to have an important role in regulating inflammation. SUMMARY: Glucocorticoids affect both pro and antiinflammatory pathways in nasal polyps. Upregulation of antiinflammatory genes such as transforming growth factor beta and uteroglobin may play an important role. Elucidation of these mechanisms may help us to understand not only the effects of glucocorticoids on nasal polyps, but also on related disorders such as allergic rhinitis and asthma.     

Role of prostaglandin D2 and E2 terminal synthases in chronic rhinosinusitis

BACKGROUND: Prostaglandin (PG)D(2) and E(2), two major cyclooxygenase (COX) products, are generated by PGD(2) synthase (PGDS) and PGE(2) synthase (PGES), respectively, and appear to mediate airway inflammation. OBJECTIVE: We sought to determine the role of PGDS and PGES in the pathophysiology of chronic rhinosinusitis (CRS). METHODS: The study examined the expression of PGDS and PGES in nasal polyps of 22 CRS patients. As controls, uncinate process mucosae were obtained from 12 CRS patients not having nasal polyps and five subjects without sinusitis. Immunohistochemistry and quantitative real-time PCR were used to evaluate the expression. RESULTS: Both PGDS and PGES were detected in nasal polyps by immunohistochemistry. Significantly greater levels of PGDS mRNA and lesser levels of PGES mRNA were observed in the nasal polyps as compared with uncinate process mucosae, and an inverse correlation between PGDS and PGES expression was observed. Levels of PGDS mRNA in nasal polyps were positively correlated with degree of infiltration by EG2+ eosinophils, whereas the levels of PGES were inversely correlated. Significantly increased levels of PGDS and conversely decreased levels of PGES were observed in asthmatics as compared with non-asthmatics. In addition, PGDS and PGES levels were positively and inversely correlated with the radiological severity of sinusitis, respectively. CONCLUSIONS: These results suggest that PGDS and PGES display an opposite and important role in the pathophysiology of CRS such as polyp formation, and more specifically, a biased expression of these synthases might contribute to the development of CRS by affecting eosinophilic inflammation.     

Regulation of glucocorticoid receptor in nasal polyps by systemic and intranasal glucocorticoids

Background: Poor response of nasal polyps to glucocorticoids (GCs) may be because of abnormal expression of GC receptors (GR) α and β or to downregulation of GRα. We aimed to evaluate the in vivo regulation of GR isoforms in GC‐treated nasal polyps and to assess the relationship between clinical response to GCs and GR levels. Methods: Patients with nasal polyps were randomly (3:1) treated (n = 51) or not (n = 14) with oral prednisone and intranasal budesonide for 2 weeks, plus intranasal budesonide for 10 additional weeks. Nasal symptoms were evaluated. Biopsies were obtained before (w0) and after 2 (w2) and 12 (w12) weeks of treatment, and analysed for their inflammatory content and GR mRNA (10² cDNA copies/μg total RNA) and protein (% immunoreactive inflammatory cells) expression. Healthy nasal mucosa (n = 11) was also investigated. Data are presented as median and 25–75th percentile. Results: At w0, nasal polyps expressed less GRα mRNA (1343;683–2263; P < 0.05) and GR protein (41;29–54; P < 0.05) than nasal mucosa (2474;1346–2933; 60;51–72, respectively). GRβ immunoreactivity was higher in nasal polyps (11;4–19; P < 0.05) than in nasal mucosa (5;2–5). At w2, increased GRα mRNA (2010;1037–2732; P < 0.01) and GR protein (56;27–71; P = 0.056) were found compared with w0 (1177;759–2058; 37;29–55, respectively). At w12, GRα mRNA and GR protein were similar to w0. GRβ expression was unaltered by treatment. Neither GRα nor GRβ correlated with nasal symptoms. GR immunoreactivity negatively correlated with eosinophils (r = ?0.478; P < 0.001). Conclusions: GRα is downregulated in nasal polyps and upregulated by GC treatment. Neither GRα nor GRβ appear to determine the sensitivity to GCs in nasal polyposis.     

Spatial expression of two anti-inflammatory mediators, annexin 1 and galectin-1, in nasal polyposis

BACKGROUND: There is renewed interest in the role played by specific counter-regulatory mechanisms to control the inflammatory host response, poorly investigated in human pathology. Here, we monitored the expression of two anti-inflammatory mediators, annexin 1 and galectin-1, and assessed their potential link to glucocorticoids' (GCs) effective control of nasal polyposis (NP). METHODS: Total patterns of mRNA and protein expression were analysed by quantitative real-time PCR (qPCR) and Western blotting analyses, whereas ultrastructural immunocytochemistry was used for spatial localization and quantification of each mediator, focusing on mast cells, eosinophils and epithelial cells. RESULTS: Up-regulation of the annexin 1 gene, and down-regulation of galectin-1 gene, was detected in polypoid tissue compared with nasal mucosa. Patient treatment with betamethasone augmented galectin-1 protein expression in polyps. At the cellular level, control mast cells and eosinophils displayed higher annexin 1 expression, whereas marked galectin-1 immunolabelling was detected in the granule matrix of mast cells. Cells of glandular duct epithelium also displayed expression of both annexin 1 and galectin-1, augmented after treatment. CONCLUSION: Mast cells and epithelial cells appeared to be pivotal cell types involved in the expression of both annexin 1 and galectin-1. It is possible that annexin 1 and galectin-1 could be functionally associated with a specific mechanism in NP and that GC exert at least part of their beneficial effects on the airway mucosa by up-regulating, in a specific cell target fashion, these anti-inflammatory agonists.     

The MEK1/2–ERK1/2 Pathway is Activated in Chronic Rhinosinusitis with Nasal Polyps

Chronic rhinosinusitis with nasal polyps (CRSwNP) is a common disease that has a considerable impact on the quality of life. Alterations in signalling pathways may contribute to the ongoing inflammation and proliferation in CRSwNP. The MEK1/2–ERK1/2 pathway transmits signals from many extracellular molecules to regulate cellular processes. We examined tissue samples from nasal polyps and the inferior turbinate of patients with CRSwNP and the inferior turbinate from subjects with healthy mucosa. The expressions of MEK1/2, ERK1/2, and their active phosphorylated forms pMEK1/2 and pERK1/2 were analysed using DNA microarray, quantitative real-time PCR, protein array, Western hybridisation, and immunohistochemistry. We detected increased MEK1/2 protein expression in nasal polyps compared to the inferior turbinates of patients with CRSwNP or healthy mucosa. We also found a higher amount of MEK1/2 in the inferior turbinates of patients with CRSwNP compared to those with healthy mucosa. Most importantly, we observed a significant increase in the phosphorylation of MEK1/2 and ERK1/2 in nasal polyps compared to both types of controls. We observed activation of the MEK1/2–ERK1/2 pathway in nasal polyps. Interestingly, we did not see the same activation pattern in different tiers of the MEK1/2–ERK1/2 signalling cascade. One explanation for this result is that the components enhance the complex MEK–ERK cascade in a distinct manner, enabling a wide variety of functions. The MEK1/2–ERK1/2 pathway appears to play a pivotal role in the pathogenesis of CRSwNP.     

Dynamics of COX-2 in nasal mucosa and nasal polyps from aspirin-tolerant and aspirin-intolerant patients with asthma

BACKGROUND: Only dynamic studies can elucidate the discrepancies concerning the expression of the inducible COX-2 gene in inflammatory airway diseases. OBJECTIVES: To quantify the expression and spontaneous regulation of COX-1 and COX-2 mRNAs in nasal polyps and nasal mucosa by real-time PCR. METHODS: Nasal polyps were obtained from 16 aspirin-tolerant patients with asthma/rhinitis (ATAR) and 18 aspirin-intolerant patients with asthma/rhinitis (AIAR) undergoing nasal polypectomy. Nasal mucosa was obtained from 12 subjects undergoing nasal corrective surgery. All specimens were cut into 3 pieces. One was immediately snap-frozen in liquid nitrogen, and the remaining 2 were left at room temperature for 30 or 60 minutes before freezing. Data are presented as medians and 25th to 75th percentiles of 10 6 cDNA molecules/microg total RNA. RESULTS: Baseline COX-2 mRNA levels were significantly lower in both ATAR (0.45; 0.13-1.20; P <.05) and AIAR (0.24; 0.12-0.41; P <.001) nasal polyps than in nasal mucosa (1.35; 0.52-3.90). COX-2 mRNA expression did not change over time in nasal mucosa but increased significantly in ATAR nasal polyps ( P <.05), reaching similar levels to nasal mucosa after 60 minutes. In contrast, COX-2 mRNA showed no significant change over time in AIAR nasal polyps. COX-1 mRNA was higher in nasal polyps than in nasal mucosa, and its expression was not modified over time in any group of patients. CONCLUSION: These results suggest differential kinetics of COX-2 mRNA between nasal mucosa and nasal polyps. AIAR nasal polyps appear to have a greater abnormality of the COX-2 pathway than ATAR.     

变形链球菌gcp基因敲除菌株表达谱基因芯片

背景：前期研究中经证实变形链球菌内部存在单磷酸鸟苷环二聚体信号通路,构建了变形链球菌gcp基因敲除菌株。 目的：比较变形链球菌野生菌种和gcp基因突变菌株基因表达的差异情况,筛选与生物膜相关的基因,进入后续研究。 方法：提取两种细菌的总RNA,反转录后分别用cy3和cy5染色。与基因芯片杂交后,扫描结果,进行数据分析,获取差异基因信息,对筛选的基因进行Real-Time PCR验证。 结果与结论：差异基因主要与糖代谢、生物膜形成有关,选择了2个基因进行验证,PCR结果与芯片结果相符合。变形链球菌gcp基因敲除后,突变菌株ahpC基因表达上调,磷酸转移酶系统基因表达下调,说明这2个基因与c-di-GMP信号通路的下游途径相关。     

Specific immunoglobulin E for staphylococcal enterotoxins in nasal polyps from patients with aspirin-intolerant asthma

BACKGROUND: Nasal polyps infiltrated with eosinophils are commonly found in chronic asthmatic patients, more frequently in those with aspirin-intolerant asthma (AIA) than aspirin-tolerant asthma (ATA). Some studies have suggested a contribution of superantigens derived from Staphylococcus sp to nasal polyposis and eosinophilia, but their relative importance in AIA and ATA subjects is unknown. OBJECTIVE: We investigated whether local production of specific IgE to staphylococcal enterotoxins A and B (SEA and SEB) and relationships with markers of eosinophilic inflammation differ in the nasal polyps of AIA and ATA subjects. METHODS: Fifteen AIA subjects with positive responses to lysine-aspirin bronchoprovocation and 15 ATA subjects underwent polypectomy. Immunoassays were used to quantify eosinophil cationic protein (ECP), IL-5, mast cell tryptase, soluble IL-2 receptors (sIL-2R), total IgE, and specific IgE for SEA and SEB. RESULTS: ECP levels in nasal polyp homogenates were higher in AIA subjects than in ATA subjects (P < 0.02), with no significant differences in tryptase, IL-5 or sIL-2R. Total IgE, and specific IgE to both SEA and SEB, were detectable in some nasal polyps from both subject groups, but median levels were markedly higher in AIA subjects than in ATA subjects (P = 0.04, 0.01, 0.05, respectively). Levels of specific IgE to SEA and SEB correlated significantly with levels of ECP and IL-5, but not those of tryptase or sIL-2R. CONCLUSION: These findings suggest that staphylococcal superantigens may drive local eosinophilic inflammation in nasal polyp tissue, and that this is exacerbated in subjects with AIA.     

Bacterial Biofilms are Associated with Inflammatory Cells Infiltration and the Innate Immunity in Chronic Rhinosinusitis With or Without Nasal Polyps

To investigate the role of bacterial biofilms in the pathogenesis of chronic rhinosinusitis, we applied scanning electron microscopy to detect bacterial biofilms, immunohistochemical and hemotoxylin–eosin staining to identify the types of inflammatory cells infiltration, and real-time PCR array analysis to evaluate the innate immune responses to bacteria biofilms. Biofilms were found in 14 of 19 (73.7 %) of CRSwNP (chronic rhinosisusitis with nasal polyps), 11 of 15 (73.3 %) of CRSsNP (chronic rhinosisusitis without nasal polyps), and none of biofilms were found in 13 normal controls. T helper lymphocytes, cytotoxic T lymphocytes, and neutrophils were the most frequently observed immune cells. The inflammatory cells did not reveal any significant differences in CRSwNP between with biofilms and without biofilms. CRSsNP with biofilms had significantly more neutrophils than CRSsNP without biofilms, and it was inclined to Th1 inflammatory response. Fifty and fifty-one genes were upregulated respectively in CRSwNP and CRSsNP with biofilms. Twenty genes were upregulated separately when comparing biofilms-positive CRSwNP with biofilms-positive CRSsNP. CRSwNP and CRSsNP with biofilms had different types of inflammatory cells infiltration and characteristic changes of the innate immunity. Further research about the direct role of bacterial biofilms in the pathogenesis of CRS will provide a new target for CRS.     

Differential Expression of Toll-Like Receptor Signaling Pathway Is Associated with Microscopic Polyangiitis in Peripheral Blood Neutrophils

Constitutive or excessive activation of Toll-like receptor (TLR) signaling pathway can disrupt the body’s immune tolerance to autoantigen, thus promoting the development of autoimmune disease. However, the expression profile of TLR signaling pathway in peripheral blood neutrophils in the pathogenesis of microscopic polyangiitis (MPA) remains unclear. Thus, improved understanding of the pathobiology of this disease may aid in the development of therapeutic targets for patients with MPA. In the present study, we assessed the expression of TLR signaling pathway-related genes in peripheral blood neutrophils in patients with MPA. PCR array analysis was performed on 20 patients with MPA and 12 healthy controls. Gene expression pro?le was performed using the human TLR for autoimmunity and inflammation PCR array of Genecopoeia, containing 84 genes related to TLR signaling pathway and six house-keeping genes. We then used quantitative real-time PCR to validate the array test. The array results identified 13 upregulated genes and 5 genes which were downregulated. The resulting qRT-PCR was consistent with the findings by PCR array. Our results suggest that peripheral blood neutrophils display changes in the expression of TLR signaling pathway-related genes associated with the pathogenesis of microscopic polyangiitis.     

Upregulation of nasal mucosal eotaxin in patients with allergic rhinitis during grass pollen season: effect of a local glucocorticoid

BACKGROUND: Allergic rhinitis is a common disease characterized by infiltration of eosinophils into the nasal mucosa during the periods of symptoms. Among chemokines, which attract cells to the site of inflammation, eotaxin is relatively specific for eosinophils. OBJECTIVE: We examined the influence of grass pollen season on nasal eotaxin expression in patients with seasonal allergic rhinitis, as well as the effect of a nasal glucocorticoid on this eotaxin expression. METHODS: Nineteen patients with allergic rhinitis received treatment with either nasal beclomethasone (400 microgram/day) or placebo over a grass pollen season. In these patients, nasal biopsies were taken prior to and during the peak of the pollen season and stained immunohistochemically for eotaxin and EG2 + eosinophils. Five healthy subjects served as controls and gave nasal biopsies once prior to the pollen season. RESULTS: Prior to pollen season, there was no significant difference in nasal eotaxin expression between patients with allergic rhinitis and healthy subjects. Grass pollen season induced significant increase in eotaxin expression in placebo-treated (P = 0.04; n = 9) but not in beclomethasone-treated rhinitis patients (P = 0.8; n = 10). During peak grass pollen season, the eotaxin expression in placebo-treated patients was significantly higher compared with healthy subjects outside season (P = 0.03). There was no significant correlation between the expression of eotaxin and the number of EG2 + eosinophils in nasal mucosa. The serum levels of eotaxin in rhinitis patients remained stable over the pollen season. CONCLUSION: Expression of eotaxin in nasal mucosa of grass-pollen allergic rhinitis patients is upregulated during pollen season and treatment with a nasal glucocorticoid protects against this upregulation.     

Interleukin 17-producing T helper cells and interleukin 17 orchestrate autoreactive germinal center development in autoimmune BXD2 mice

Interleukin 17 (IL-17) is a cytokine associated with inflammation, autoimmunity and defense against some bacteria. Here we show that IL-17 can promote autoimmune disease through a mechanism distinct from its proinflammatory effects. As compared with wild-type mice, autoimmune BXD2 mice express more IL-17 and show spontaneous development of germinal centers (GCs) before they increase production of pathogenic autoantibodies. We show that blocking IL-17 signaling disrupts CD4+ T cell and B cell interactions required for the formation of GCs and that mice lacking the IL-17 receptor have reduced GC B cell development and humoral responses. Production of IL-17 correlates with upregulated expression of the genes Rgs13 and Rgs16, which encode regulators of G-protein signaling, and results in suppression of the B cell chemotactic response to the chemokine CXCL12. These findings suggest a mechanism by which IL-17 drives autoimmune responses by promoting the formation of spontaneous GCs.     

LPS 刺激诱导大鼠原代枯否细胞基因表达谱变化分析

目的:研究脂多糖(Lipopolysaccharide,LPS)刺激原代大鼠枯否细胞(Kupffer cell,KC)基因表达谱的变化情况。方法:胶原酶灌注消化和不连续密度梯度离心分离培养大鼠原代肝脏KC,采用LPS 刺激细胞。OneArray 基因表达谱芯片检测LPS 刺激后,细胞内基因表达谱的变化。采用实时荧光定量PCR 法对表达上调最显著的基因进行验证。结果:基因芯片结果显示LPS 刺激后,原代KC 内基因表达谱发生了明显变化。与正常对照组相比,LPS 刺激组基因表达上调27 个(包括Ces1f、Slc17a3、Slc21a4、Hsd17b2、Sorbs2、Ccdc116、Mgam、Myo5b、Etl4、Fabp1、Kif4b、Fosl1、Cyp4a1、Penk、Tmem221、Rpl5、Nr2f1、Hoxb1、Gpr165、Fam90a13p、Kpna6、Irak1bp1、Kcnh1 和4 个尚未命名基因),表达下调4 个(包括Oc90、Tagln、Arxes2 和Olr830)。其中Ces1f 为上调最显著的基因。实时荧光定量PCR 验证结果显示,LPS 刺激诱导KC 对Ces1f 基因表达水平上调达23.88倍。结论:LPS 刺激可诱导大鼠原代KC 基因表达谱发生变化。其中,Ces1f 基因的上调表达最为显著。     

Effect of seasonal allergen exposure on mucosal IL-16 and CD4+ cells in patients with allergic rhinitis

BACKGROUND: CD4+ T cells constitute a major source of cytokines in allergic diseases such as allergic rhinitis. Interleukin (IL)-16 selectively recruits CD4+ cells. METHODS: We evaluated the effect of natural allergen exposure during a grass-pollen season on IL-16 expression and number of CD4+ cells in nasal mucosa. Patients with allergic rhinitis (n=16) were treated with either a nasal glucocorticoid beclomethasone (BDP; 400 microg/day) or placebo, and gave nasal biopsies prior to and during the grass-pollen season. The evaluated markers in allergic rhinitis patients were also compared to those in healthy control subjects (n=5). RESULTS: Prior to the pollen season, the expression of IL-16, but not the number of CD4+ cells, was significantly higher in patients with allergic rhinitis than in healthy control subjects. The grass-pollen season further increased IL-16 expression and also increased the number of CD4+ cells in placebo-treated, but not in BDP-treated, allergic rhinitis patients. The pollen-season-induced change in IL-16 expression and in CD4+ cells was significantly more pronounced in placebo- than in BDP-treated patients. There was a significant correlation between the change in IL-16 expression and the number of CD4+ cells. CONCLUSIONS: These data suggest that local upregulation of IL-16 expression contributes to the inflammation observed in seasonal allergic rhinitis. Hypothetically, inhibition of IL-16 expression can be one of several mechanisms by which nasal glucocorticoids achieve their anti-inflammatory effect in allergic rhinitis.