Metalloproteinase function in chronic rhinosinusitis with nasal polyposis

OBJECTIVES: Chronic rhinosinusitis with nasal polyposis (CRSwNP) and asthma share characteristic inflammatory features and histopathologic findings of airway remodeling. Remodeling, which is controlled by matrix metalloproteinases (MMP), is a key event in the pathogenesis of asthma. The MMP functions have rarely been evaluated in CRSwNP. STUDY DESIGN: Prospective and in vivo. METHODS: MMP-7, MMP-8, MMP-9, and tissue inhibitor of metalloproteinase (TIMP)-1 concentrations were analyzed by enzyme-linked immunosorbent assay and their molecular forms by Western immunoblotting and gelatin zymography in 24 patients operated on for CRSwNP and in nasal lavages from 19 healthy controls. MMP function, protective or destructive, was evaluated by comparing MMP/TIMP-1 levels with the disease activity, estimated by tissue eosinophilia and a need for re-operations. RESULTS: Significantly increased levels of MMP-8/TIMP-1 and MMP-9/TIMP-1 were found in patients without tissue eosinophilia relative to eosinophil-positive CRSwNP patients and controls, as well as in patients who did not require re-operation in comparison with re-operated patients. In eosinophil-positive and re-operated patients, these parameters were within the same range than in controls. CONCLUSIONS: Proteolytic spectrum is different in eosinophilic and noneosinophilic CRSwNP, suggesting a new mechanism for eosinophil accumulation in the disease pathogenesis. Enhanced MMP-8 and MMP-9 expression was associated with a better prognosis/clinical outcome, and thus these results may represent a synergic, protective role of MMP-8 and MMP-9 in host response in CRSwNP. Because synthetic MMP inhibitors, capable of equilibrating the unfavorable MMP/TIMP-ratio, may be of potential therapeutic value in chronic respiratory tract diseases, the MMP functions in inflammatory conditions need to be carefully established.     

The importance of follow-up examinations in chronic rhinosinusitis with nasal polyposis

PurposeClinical examinations following functional endoscopic sinus surgery (FESS) for chronic rhinosinusitis with nasal polyps (CRSwNP) are critical for physicians to assess results of the intervention and to early identify recurrences. However, no clear consensus on the frequency and timing of clinical examinations following surgery exists. The aim of this study was to analyze CRSwNP recurrences after FESS with a specific focus on the adherence to follow-up examinations.Materials and methodsSixty patients who underwent FESS for CRSwNP were enrolled. Clinical parameters were recorded. Adherence to follow-up examinations with nasal fiber optic endoscopy and regular administration of nasal steroids were analyzed.ResultsAdherence to periodic clinical examinations and regular treatment with nasal steroids was 25 %. CRSwNP recurrence was observed in 56.7 % of cases at the last follow-up examination. No statistically significant difference concerning nasal symptoms was observed between patients with and without current recurrence (p > 0.05). Subjects who underwent regular examinations and prompt treatment of small recurrences had a lower probability of relapse at their last examination (7.7 % versus 38.2 %, respectively; p < 0.001). Polyp grade > 2 in patients with or without adherence were seen in 15.4 % and 42.9 % of cases, respectively (p < 0.05).ConclusionsAdherence to follow-up examinations with nasal optic fiber endoscopy is crucial to early identify recurrence after surgery and promptly treat it with medical therapy.     

The presence of CD209 expressing dendritic cells correlates with biofilm positivity in chronic rhinosinusitis with nasal polyposis

Biofilm-positive cases of chronic rhinosinusitis with nasal polyposis (CRSwNP) may form a separate clinical entity, which is characterized by high recurrence rates and resistance against different therapeutic strategies. This can be explained by a special immunologic phenotype. Biofilm existence has been supposed to correlate with increased amount of dendritic cells that are responsible for antigen presentation in CRSwNP. A total of 20 patients with CRSwNP undergoing endoscopic sinus surgery (ESS) were analyzed. The negative control group consisted of ten patients undergoing septoplasty without CRSwNP. Three series of individual nasal polyps and control specimens were processed to hematoxylin–eosin (HE) and Gram staining and to CD209-specific immunofluorescent assay, respectively. Biofilm was detected in 13 of 20 patients (65 %) with CRSwNP and in none of the ten negative controls. The subepithelial layer of biofilm-positive nasal polyps displayed a statistically significant (p < 0.001) increase in the numbers of CD209-expressing dendritic cells compared to biofilm-negative specimens. It was found that biofilm detectability showed strong correlation to the architecture of respiratory mucosa and to the dominant inflammatory cell type of the subepithelial layer. Persisting bacterial biofilms may affect the type of antigen presentation and consecutive immune reactions in the subepithelial layer of nasal mucosa. This phenomenon may result in different inflammatory pathways with specific cytokine profile compared to biofilm-negative cases. Co-existence of bacterial biofilms and dominant pattern of dendritic cells suggest a biofilm-associated immunologic phenotype in CRSwNP. This can explain the mucosal changes, functional disorders and therapy resistance featuring CRSwNP.     

IL-17C expression in nasal epithelial cells of chronic rhinosinusitis with nasal polyposis

Interleukin 17C (IL-17C) is a functionally distinct member of the IL-17 family that is selectively induced in epithelia by bacterial challenge and inflammatory stimuli. The goal of this study was to explore the expression of IL-17C in nasal epithelial cells and their role in the pathogenesis of chronic rhinosinusitis with nasal polyposis (CRSwNPs). IL-17C expression was detected using immunohistochemistry (IHC) of the epithelial cell layers and using the western blot assay on whole tissue homogenates from control subjects (n = 10) and CRSwNP patients [10 non-eosinophilic polyps and 10 eosinophilic polyps (EPs)]. Expression of IL-17C and P47-phox were evaluated in the human nasal epithelial cells (RPMI-2650 cells) after treatment with staphylococcal enterotoxin B (SEB) and pretreatment with reactive oxygen species (ROS) scavenger, N-acetyl l-cysteine (NAC). Finally, IL-17C expression was demonstrated in eosinophilic rhinosinusitis murine model using IHC. Epithelial expression of IL-17C was higher in nasal polyps (especially in EPs) compared to control mucosa. SEB increased the expression of IL-17C and P47-phox in RPMI-2650 cells. SEB-induced expressions of both IL-17C and P47-phox were significantly decreased in NAC-pretreated cells. Epithelial expression of IL-17C was significantly higher in experimental mice compared to control mice. SEB-induced IL-17C expression in nasal epithelial cells is mediated by ROS production. This pathway may be associated with the pathogenesis of CRSwNP, especially eosinophilic nasal polyps.     

Current concepts in therapy of chronic rhinosinusitis and nasal polyposis

The exact pathophysiological mechanisms leading to chronic rhinosinusitis (CRS) still to a large extent remain obscure. However, recently there has been some progress in elucidating the etiology of nasal polyposis, especially regarding tissue eosinophilia as well as the role of aspirin intolerance and eicosanoid mediators. Endonasal sinus surgery has evolved to be the treatment of choice in CRS and nasal polyposis in all cases where conservative treatment has failed or resulted in only a partial or temporary relief. Today, state of the art in surgical technique includes the ability to combine microscopic and endoscopic procedures. Regardless of technical advances like powered instrumentation or computer-aided surgery, in a modern protocol, surgical therapy can offer only one option within a complex and individually tailored therapeutical concept. This review discusses current concepts and new developments in the diagnosis and treatment of CRS and nasal polyposis.     

慢性鼻-鼻窦炎鼻息肉患者细菌生物膜的观察

目的 观察慢性鼻-鼻窦炎鼻息肉患者手术黏膜组织中细菌生物膜的形态特征,并分析其与临床因素、术后疗效的关系.方法 试验组获取72例慢性鼻-鼻窦炎鼻息肉患者手术中筛窦黏膜组织,对照组获取15例鼻中隔偏曲患者、10例鼻骨骨折患者钩突黏膜组织(患者均知情同意),所有组织标本均行扫描电镜检查.对两组患者进行术后随访,分别应用Lund-Kennedy鼻内镜检查、慢性鼻窦炎疗效评定标准(1997年,海口)评估患者病情改善程度.采用t检验分析细菌生物膜对术后Lund-Kennedy评分的影响;采用卡方检验分析细菌生物膜与内镜手术疗效的相关性,以P＜0.05为差异有统计学意义.结果 试验组72例慢性鼻-鼻窦炎鼻息肉患者中有3例患者失访,失访率为4.2%.69例慢性鼻-鼻窦炎鼻息肉患者的细菌生物膜阳性率71.0%(49/69),并且可以观察到黏膜组织纤毛不同程度的损伤.对照组未发现细菌生物膜,可以观察到黏膜纤毛较浓密,排列整齐.卡方检验结果显示,性别、慢性鼻-鼻窦炎鼻息肉类型、病程等因素在试验组细菌生物膜阴性组与阳性组间差异无统计学意义(P值均＞0.05).细菌生物膜阳性组患者术后6个月和12个月的Lund-Kennedy 评分(4.78 ±1.67;4.55 ±1.61)分别高于细菌生物膜阴性组患者(3.65 ±1.39;3.65 ±1.18),差异均有统计学意义(t=-2.654,P＜0.01;t=-2.264,P＜0.05);细菌生物膜阴性组术后6个月和12个月的疗效明显优于细菌生物膜阳性组,差异有统计学意义(x2值分别为18.014、22.063,P值均＜0.001).结论 慢性鼻.鼻窦炎鼻息肉患者手术黏膜组织中存在细菌生物膜的生命周期不同形态,细菌生物膜的存在与否,与慢性鼻-鼻窦炎鼻息肉的类型、性别、病程无明显相关.细菌生物膜可能对患者术后疗效产生不良的影响.

Abstract：

Objective To explore the presence of bacterial biofilms(BF) in patients with CRS and the effect of BF on clinical symptoms and postoperative outcomes.Methods Seventy-two patients with chronic sinusitis were enrolled in this study.The control group included 15 patients with deviation of the nasal septum and 10 patients had a fracture of the nasal bone.Mucosa of the uncinate process or ethmoid near the ostium of the maxillary sinus was obtained during endoscopic sinus surgery.The specimens were subjected to scanning electron microscopy.Patients were followed for 1 year and observed by the LundKennedy endoscopy,and the Haikou standard classification (ESS-1997).Statistical analysis was performed by t-test or chi-square test Results Three patients were lost to follow-up.The scanning electron microscopy analysis showed bacterial biofilms in 49 of the 69 patients with chronic sinusitis.A marked destruction of the epithelium and cilia was observed in samples positive for bacterial biofilms.No bacterial biofilms were detected in the control group,and scanning electron microscopy showed normal epithelium and cilia in those specimens.There was no significant difference in gender,classification or duration of disease between the BF(-) and BF( + ) groups.At six months and one year postoperative,the Lund-Kennedy endoscopy scores for CRS patients with BF(4.78 ± 1.67 ;4.55 ± 1.61) were significantly higher than those without BF(3.65 ±1.39;3.65 ±1.18) (t =-2.654,P＜0.01;l =-2.264,P＜0.05).Based on the Haikou standard classification,there was a significantly difference between patients with BF and those without BF( x2 = 18.014,22.063 ,P ＜ 0.001,respectively).Conclusions Different life stages of bacterial biofilms were demonstrated to be present in CRS.Gender,classification or duration of disease did not affect the presence of bacterial biofilms in patients with CRS.There is a correlation between bacterial biofilms and an unfavorable outcome in patients with CRS after ESS.     

Bacterial colonization of nasal steroid inhalers in chronic rhinosinusitis

BACKGROUND: The aim of this study was to determine if nasal steroid inhalers harbor bacteria. METHODS: Nasal steroid inhalers were randomly selected from an adult patient population with chronic rhinosinusitis. Swab cultures of the tip of the nasal inhaler were obtained and submitted for microbiological analysis. Contemporaneous control cultures were obtained from freshly opened nasal steroid inhalers. Comparisons were conducted between bacterial recovery rates and types of organisms recovered from the patient and control groups. RESULTS: Among 31 nasal inhalers in use, 14 inhalers (45%) were found to harbor bacteria. The most common organisms were coagulase negative Staphylococci (11 inhalers) followed by oral flora (2 inhalers) and bacillus species (1 inhaler). None of the 10 control cultures were found to harbor bacteria. Nasal steroid inhalers in use were more likely to have bacterial colonization than new inhalers (p = 0.008, chi-square). CONCLUSIONS: Nasal steroid inhalers may harbor pathogenic bacteria. Therefore, they may serve as a vehicle for subsequent reinfection.     

Optical coherence tomography for biofilm detection in chronic rhinosinusitis with nasal polyposis

Chronic rhinosinusitis with nasal polyposis (CRSwNP) is a multifactorial disease that seems to be associated with the presence of microbial biofilms and corresponding subepithelial inflammatory reactions. Optical coherence tomography (OCT) might be applied to detect bacterial and fungal biofilms in patients with CRSwNP. A total of 27 patients with CRSwNP undergoing endoscopic sinus surgery (ESS) were analyzed. The negative control group consisted of six patients undergoing septoplasty for nasal obstruction without CRSwNP. The nasal polyps and inferior turbinate mucosa specimens applied as negative controls were processed to OCT analysis and H.E. and Gram staining. Biofilm was detected in 22 of 27 patients (81.5 %) with CRSwNP and in none of six negative controls. In our series, OCT scan showed an obvious association with the findings of H.E. and Gram staining and was allocated to be a good predictor of biofilm existence. On OCT images, biofilms were displayed as distinct superficial layers with high optical density. It was found that microscopic architecture of biofilms was strongly associated with the integrity of nasal mucosa and to the cellular pattern of subepithelial inflammatory reaction. This study confirmed the presence of microbial biofilms in patients with CRSwNP according to OCT scans and histological analysis. Since biofilms may affect the severity and recurrence rate of CRS treated by ESS they should be detected preoperatively. In conclusion, single application of OCT analysis or combination with conventional histological protocols provides a robust and reliable method for the detection of bacterial and fungal biofilms in CRSwNP. Level of evidence 3b, individual case–control study.     

The effectiveness of steroid treatment in nasal polyposis

OBJECTIVE: The objectives of the management of nasal polyposis are to eliminate or reduce the size of polyps, reestablish nasal breathing, reduce symptoms of rhinitis, restore the sense of smell, and prevent the recurrence of nasal polyps. Local or systemic steroids have been used in the treatment of nasal polyps, but efficacy of combined (local and systemic) steroids in nasal polyposis has been little investigated. The aim of this study was to evaluate the influence of combined steroid therapy on the symptoms and extent of the disease in patients with nasal polyposis. METHODS: Seventeen patients with nasal polyps were treated with combined steroids. Before and after the therapy, polyp size, nasal symptoms, sense of smell, and headache or facial pain were assessed by an established scoring system. RESULTS: After the therapy, symptom scores of all the patients improved. Of the patients, 12% showed a polyp-free nasal cavity, 76% a clear involution of polyps, and 12% no response to the therapy. There were statistically significant differences (P<0.001) for symptom scores and polyp size. Medical ablation of polyps using steroids was not achieved in 88% patients. CONCLUSION: Steroids can reduce polyp sizes and improve the symptoms, but are inadequate to eradicate the polyps. Surgery still plays a major part in the treatment of the nasal polyposis, but steroids can delay the necessity for surgical intervention.     

A characteristic protein in nasal discharge differentiating non-allergic chronic rhinosinusitis from allergic rhinitis

To differentiate non-allergic chronic rhinosinusitis (vasomotor rhinitis) from allergic rhinitis, a characteristic protein in the nasal discharge was studied. The subjects consisted of 10 patients with perennial allergic rhinitis to house dust, 10 patients without perennial rhinitis without antigen (clinically defined as non-allergic chronic rhinosinusitis) and 5 normal volunteers without nasal disease as a control group. The total protein in the nasal discharge was determined by Lowry's method and analysis of the protein components was made by SDS-PAGE. It was found that the nasal discharge obtained from the case with perennial allergic rhinitis contained a high concentration of albumin (25.9 micrograms/ml) and a characteristic protein band with an estimated molecular weight of 26 kilo-Daltons (kD) on a SDS-PAGE, in a concentration of 15.8 micrograms/ml. In contrast, the nasal discharge from non-allergic chronic rhinosonusitis patients contained a lower concentration of albumin (12.9 micrograms/ml) than that of the allergic rhinitis patients, and the concentration of the characteristic protein 26 kD was only 2.3 micrograms/ml. The 26 kD protein was considered to originate from the nasal glands, since its secretion could be provoked by stimulation of the nasal glands of the normal volunteers with a 1% pilocarpine spray. The low concentration of albumin and the characteristic protein 26 kD in the nasal discharge thus appeared to differentiate patients with non-allergic chronic rhinosinusitis from those with perennial allergic rhinitis.     

鼻用糖皮质激素治疗慢性鼻窦炎的鼻窦CT观察

目的采用影像学方法评价鼻用糖皮质激素治疗慢性鼻窦炎（不合并鼻息肉）的疗效。方法慢性鼻窦炎患者24例（42侧），其中青少年患者11例（20侧），平均（x^-±s，下同）年龄（11．8±3．2）岁；成年患者13例（22侧），平均（36．7±11．0）岁。患者接受鼻用糖皮质激素（布地奈德鼻喷雾剂）治疗，成年和青少年患者的药物剂量分别为256μg／d和128μg／d，平均疗程分别为（7．5±3．2）周和（7．0±3．4）周。采用Lund-Mackay评分方法，在治疗前后分别对患者鼻窦CT进行评分（Lund得分），同时记录患者主诉鼻部症状改善情况。结果①治疗后，成年患者的平均Lund得分为6．5±7．5，显著低于治疗前（12．2±5．7，t=3．82，P〈0．01）；青少年患者的平均Lund得分治疗后为2．6±3．7，显著低于治疗前（10．3±5．7，t=5．08，P〈0．01）。成年患者中5例（38％）达到影像学治愈标准，好转7例（54％），无效1例（8％）；青少年患者中8例（73％）达到影像学治愈标准，好转3例（27％），两组疗效差异无统计学意义（P〉0．05）。②鼻用糖皮质激素对成年慢性前组筛窦炎治愈率最高（47％），对慢性后组筛窦炎治愈率最低（38％）；药物对青少年慢性前、后组筛窦炎和慢性蝶窦炎的疗效优于成年患者。③成年患者的主观疗效与治疗前Lund得分呈正相关关系（r=0．676．P〈0．05），即治疗前Lund得分越高，主观疗效越差。青少年患者的主观疗效，与治疗前Lund得分无相关关系（P〉0．05）。结论鼻用糖皮质激素可有效治疗慢性鼻窦炎，部分患者可以达到影像学上的治愈标准。     

Bacterial biofilms in surgical specimens of patients with chronic rhinosinusitis

OBJECTIVES: Biofilms are bacterial pathogens that organize in several chronic and recalcitrant infectious processes. We hypothesize that biofilms play a role in chronic rhinosinusitis (CRS). Our goal is to demonstrate biofilms in mucosal specimens of patients undergoing surgery for CRS. STUDY DESIGN: A prospective study of the presence of biofilms in patients undergoing endoscopic sinus surgery for CRS compared with control patients without CRS. METHODS: There were a total of 30 subjects and 4 controls enrolled. The samples of 24 subjects and 4 controls were cultured and then prepared using standard methods for scanning electron microscopy (SEM). The remaining six subjects' samples were treated using advanced cryofixation methods as preparation to preserve structure for SEM and transmission electron microscopy (TEM). RESULTS: Using strict SEM morphologic criteria, 24 (80%) of the 30 patients were found to have micrographic evidence of biofilms. All controls had healthy appearing cilia and goblet cells without biofilms. The six cryofixation samples showed biofilm structures on SEM micrographs that were correlated with bacterial structures seen at the mucosal surface on the corresponding TEM cross sections. Bacterial cultures were positive on all patients. CONCLUSIONS: Biofilms were demonstrated to be present in patients undergoing surgery for CRS; none of the patients without CRS had any evidence of biofilms. Although SEM is capable of demonstrating the biofilms' three-dimensional structure, glycocalyx, and water channels, it cannot clearly demonstrate the presence of bacteria within the biofilm. We were able to demonstrate evidence of bacteria in the biofilms on the subjects tested using TEM.     

Increased expression of nucleophosmin is associated with the pathophysiology of chronic rhinosinusitis with nasal polyposis

ObjectiveNucleophosmin (NPM1) has been suggested to be involved in the pathophysiologic mechanism of inflammatory disorders. We measured the expression level of NPM1 in nasal polyp (NP) tissues of patients with chronic rhinosinusitis with nasal polyposis (CRSwNP). We also assessed the correlation between NPM1 expression and other parameters such as eosinophilic infiltration, inflammatory cytokines, and clinical indicators such as Lund-Mackay computed tomography (CT) score.MethodsThirty patients with CRSwNP were included. We performed pre-operative CT scan to determine Lund-Mackay CT scores. During endoscopic sinus surgery, we harvested NP tissues from patients with CRSwNP. We performed Sirius red staining to evaluate eosinophilia and conducted immunohistochemical staining for NPM1 and real-time PCR for cytokines including interleukin (IL)-5, IL-17A, and IL-32.ResultsThe mRNA expression of NPM1 was significantly up-regulated in eosinophilic NP tissues (RQ 0.58 ± 0.06), compared to non-eosinophilic NP tissues (RQ 0.38 ± 0.08, p < 0.05). In the epithelium of NP tissue, a significant positive correlation was observed between eosinophilic infiltration and NPM1 expression. The expression of NPM1 was significantly correlated with that of IL-5 (r = 0.6229, p = 0.0004), IL-17A (r = 0.5971, p = 0.001), and IL-32 (r = −0.5985, p = 0.0068). There was no significant correlation between the mRNA expression of NPM1 and the Lund-Mackay CT score (Spearman r = −0.2563, p = 0.1879).ConclusionExpression of NPM1 was significantly increased in eosinophilic NP tissues from patients with CRSwNP. We observed an association between NPM1 expression and various pro-inflammatory cytokines such as IL-5, IL-17, and IL-32 and eosinophilic infiltration, which is thought to contribute to the pathophysiology of NP.