Analysis of KRT1, KRT10, KRT19, TP53 and MMP9 expression in pediatric and adult cholesteatoma.

Cholesteatoma is an epidermal cyst with still unknown pathomechanism. The aim of the current study was to investigate molecular differences in the background of the hyperproliferative property and aggressive behavior typical of the cholesteatoma epithelium. The expression of three cytokeratin genes (KRT1, KRT10 and KRT19), the matrix metalloproteinase 9 gene (MMP9) and the tumor suppressor TP53 gene was measured by qRT-PCR in surgical samples of pediatric and adult cholesteatoma cases and their expression level was compared to that of normal skin samples from the retroauricular region of control individuals. Cholesteatoma samples were stratified according to the age of onset and recurrence for more detailed analysis. Our results showed identical expression pattern for KRT1 and KRT10, their expression was higher in pediatric cases than in adults, especially in pediatric recurrent samples. The expression level of KRT19 was inversely proportional to that of KRT1/KRT10, it was lower in the more invasive recurrent cases both in our pediatric and adult groups. As it was expected from the bone destructive behavior of cholesteatoma, a significantly elevated expression of MMP9 was measured in cholesteatoma samples, the highest level was found in adult recurrent cases. Low expression levels characterize the TP53 gene without significant differences in our samples. These findings demonstrate that cytokeratin expression distinguishes between pediatric/adult, nonrecurrent/recurrent cases, suggesting that distinct differentiation state and cell division potential characterize these cholesteatoma cases. KRT19 with a tumor suppressor potential might restrict the recurrence of cholesteatoma. The differences observed in gene expression profiles between cholesteatoma and control samples support the notion that cholesteatoma is a cystic lesion with tumor-like behavior because it is characterized by invasive, destructive growth and high tendency for recurrence.

Comparative analysis of preoperative diagnostic values of HRCT and CBCT in patients with histologically diagnosed otosclerotic stapes footplates.

This prospective case review was performed with the aim to compare and asses the diagnostic values of cone-beam computed tomography (CBCT) and high-resolution computed tomography (HRCT) in the preoperative evaluation of otosclerosis. A total of 43 patients with histologically confirmed stapedial otosclerosis, who underwent unilateral stapedectomies were analyzed. Preoperative temporal bone CBCT and HRCT scans were performed in all cases. Both CBCT and HRCT imaging were characterized by a slice thickness of 0.4-0.625 mm and multiplanar image reconstruction. Histopathologic examination of the removed stapes footplates was performed in all cases. Findings of CBCT and HRCT were categorized according to the modified Marshall's grading system (fenestral or retrofenestral lesions). Histopathologic results were correlated with multiplanar reconstructed CBCT and HRCT scans, respectively. Negative control groups for CBCT (n = 36) and HRCT (n = 27) examinations consisted of patients, who underwent CBCT imaging due to various dental disorders or HRCT analysis due to idiopathic sudden sensorineural hearing loss. Histologically active foci of otosclerosis (n = 31, 72 %) were identified by both CBCT and HRCT in all cases with a sensitivity of 100 %. However, CBCT could not detect histologically inactive otosclerosis (n = 12, 23 %; sensitivity 0 %). In contrast, HRCT showed inactive otosclerosis with a sensitivity of 59.3 %. According to CBCT results, no retrofenestral lesions were found and the overall sensitivity for hypodense lesions was 61.37 %. In conclusion, CBCT is a robust imaging method in the detection of histologically active fenestral hypodense foci of otosclerosis with high sensitivity and radiologic specificity. In the light of these results, HRCT still remains the basic imaging method in the preoperative diagnosis of otosclerosis, since it has much greater sensitivity and specificity in the detection of retrofenestral hypodense lesions and histologically inactive otosclerotic foci in the oval window niche.

The c-MYC protooncogene expression in cholesteatoma.

Cholesteatoma is an epidermoid cyst, which is most frequently found in the middle ear. The matrix of cholesteatoma is histologically similar to the matrix of the epidermoid cyst of the skin (atheroma); their epithelium is characterized by hyperproliferation. The c-MYC protooncogene located on chromosome 8q24 encodes a transcription factor involved in the regulation of cell proliferation and differentiation. Previous studies have found aneuploidy of chromosome 8, copy number variation of c-MYC gene, and the presence of elevated level c-MYC protein in cholesteatoma. In this study we have compared the expression of c-MYC gene in samples taken from the matrix of 26 acquired cholesteatomas (15 children and 11 adults), 15 epidermoid cysts of the skin (atheromas; head and neck region) and 5 normal skin samples (retroauricular region) using RT-qPCR, providing the first precise measurement of the expression of c-MYC gene in cholesteatoma. We have found significantly elevated c-MYC gene expression in cholesteatoma compared to atheroma and to normal skin samples. There was no significant difference, however, in c-MYC gene expression between cholesteatoma samples of children and adults. The significant difference in c-MYC gene expression level in cholesteatoma compared to that of atheroma implies a more prominent hyperproliferative phenotype which may explain the clinical behavior typical of cholesteatoma.

Middle ear gas pressure regulation: the relevance of mastoid obliteration.

OBJECTIVES: To establish a mathematical model of middle ear gas pressure regulation and to discuss potential implications for pathophysiology-oriented theoretical approach to middle ear surgery, with particular attention to mastoid obliteration. BACKGROUND: Numerous studies support that small mastoid volume is associated with cholesteatoma. Latest studies show that mastoid obliteration is an effective technique to lower the recurrence rate in these ears. METHODS: A mathematical model was used to predict the development of gas pressure balance in the function of different middle ear volumes (VME), considering normal and dysfunctional Eustachian tube. Published data as gas pressure input values and our 3D CT reconstruction data in healthy and pathologic middle ears of children were applied. RESULTS: The model predicted ≤6.66 daPa pressure fluctuations in VME ≥3 ml, compared to ≥16 daPa of a VME ≤1 ml at perfect ET function, because of the different pressure change rate and pressure buffer effect of the MEs. Substantially larger fluctuations can be expected in a VME <3 ml with malfunctioning ET. Modeling mastoid obliteration predicts similar pressure fluctuations to a VME ≥3 ml resulting from elimination of gas exchange surface. CONCLUSION: Pressure change is faster in smaller MEs than in larger ones. Healthy MEs between 3 and 6 ml are very sensitive to the duration of a potential ET dysfunction to develop ME pathology. In MEs with poor mastoid pneumatization and dysfunctional ET, typical in cholesteatoma cases, mastoid obliteration as surgical reduction of mucosal surface for gas exchange can improve ME gas pressure balance resulting in better long-term outcome.

Primary tuberculosis of the middle ear cleft: diagnostic and therapeutic considerations.

Tuberculosis remains one of the most challenging infectious diseases, which rarely manifests in the middle ear cleft exclusively. Typical symptoms of tuberculosis have become more and more confusing due to the genetic evolution of different Mycobacterium species. In the diagnosis of tuberculous otitis media (TOM), clinical suspicion plays a fundamental role, when topical and/or systemic antibiotic treatment cannot lead to improvement in ear discharge and inflammation. If there is no other reason of persisting otorrhea, microbiological sampling and culturing are the subsequent steps of diagnosis. These investigations, however, have low sensitivity; therefore a canal wall-up mastoidectomy is recommended, which includes the removal of necrotic bone and multiple histological sampling from various locations. Currently, histopathological analysis is the most robust and reliable method in the diagnosis of TOM. Tuberculin skin test, Mycobacterium-specific PCR and interferon-gamma release assay cannot distinguish between active, inactive or post-infective conditions. According to these considerations, these methods may serve as supplementary assays for the final diagnosis. Having the appropriate diagnosis after surgical intervention and laboratory analysis, medical management should be continued by anti-tuberculosis chemotherapy. Hereby, we demonstrate two cases with primary TOM and provide an overview of the literature in the light of diagnostic and therapeutic guidelines in the management of TOM.

Genetic association analysis in a clinically and histologically confirmed otosclerosis population confirms association with the TGFB1 gene but suggests an association of the RELN gene with a clinically indistinguishable otosclerosis-like phenotype.

BACKGROUND/HYPOTHESIS: Otosclerosis is a frequent cause of hearing impairment characterized by abnormal resorption and deposition of bone in the human otic capsule. It is a disease of complex etiopathogenesis that is caused by both environmental and genetic factors. The goal of this study is to replicate association for genes that were previously reported to be associated with otosclerosis. However, in this study, patients were used in which the presence of otosclerotic foci was confirmed by histologic investigation, in contrast to previous studies, that did not use histologic confirmation. METHODS: Case-control association study using 153 cases and 300 controls. Thirteen single nucleotide polymorphisms (SNPs) in 6 genes (COL1A1, TGFB1, BMP2, BMP4, AGT, and RELN) were genotyped. RESULTS: An association between TGFB1 (rs1800472) and otosclerosis was detected, confirming several previous reports. It is surprising that no association was found between RELN and otosclerosis because the current analysis had very reasonable power and the RELN association has been published before in different articles using several independent populations. CONCLUSION: Our findings strengthen the association of TGFB1 (rs1800472) with otosclerosis. The fact that other genes did not replicate could be due to different reasons like lack of power (BMP2 and BMP4) and possible false-positive initial association (COL1A1 and AGT). A plausible explanation for the lack of association for RELN is that RELN could be associated with a specific otosclerosis-like phenotype that is different from the histologically confirmed phenotype of the patients in this study, and that is clinically not distinguishable.

Effects of intranasal steroid treatment on the presence of biofilms in non-allergic patients with chronic rhinosinusitis with nasal polyposis.

Microbial biofilms have been implicated in the pathogenesis of chronic rhinosinusitis with nasal polyposis (CRSwNP). Intranasal application of corticosteroids (INCS) is a reliable option in the management of CRSwNP. INCS medication has been suspected to influence the presence and thickness of microbial biofilms and inflammatory cell patterns in CRSwNP. Two series of identical nasal polyps obtained from non-allergic patients with CRSwNP (n = 56), who underwent endoscopic sinus surgery (ESS), were processed to hematoxylin-eosin (H.E.) and Gram staining, respectively. Patients were recruited into three groups. Group A (n = 21) consisted of patients with continuous preoperative INCS treatment. In group B (n = 17), patients were never treated by INCS, while in group C (n = 18) INCS medication was stopped at least 6 months before ESS. Biofilm positivity varied from 76.4 to 88.8% in different subject groups. These values and average thickness of biofilms did not reach statistically significant levels (Mann-Whitney's U probe, p > 0.05) in different patient groups. In contrast, microscopic pattern and numbers of predominant inflammatory cell populations displayed obvious differences according to INCS treatment (Mann-Whitney's U probe, p < 0.001). According to these observations, INCS treatment does not affect the presence and thickness of microbial biofilms in CRSwNP. In contrast, it has significant effects on the pattern of inflammatory cells infiltrating the subepithelial layer, which might result in beneficially altered extracellular matrix production and cytokine release.

Diagnostic value of cone-beam CT in histologically confirmed otosclerosis.

This retrospective case review was performed with the aim to asses the value of cone-beam computed tomography (CBCT) in the preoperative diagnosis of otosclerosis. A total of 32 patients with histologically confirmed stapedial otosclerosis, who underwent unilateral stapedectomies were analyzed. Preoperative temporal bone CBCT scans were performed in all cases. CBCT imaging was characterized by a slice thickness of 0.3 mm and multiplanar image reconstruction. Histopathologic examination of the removed stapes footplates was performed in all cases. Findings of CBCT were categorized according to Marshall's grading system (from grade 0 to grade 3). Histopathologic results were correlated to multiplanar reconstructed CBCT scans, respectively. Histologically active foci of otosclerosis (n = 21) were identified by CBCT in all cases with a sensitivity of 100 %. However, CBCT was unable to detect histologically inactive otosclerosis (n = 11, sensitivity = 0 %). According to CBCT scans, no retrofenestral lesions were found and all positive cases were recruited into the grade 1 group indicating solely fenestral lesions at the anterior pole of stapes footplates. In conclusion, CBCT is a reliable imaging method with considerably lower radiation dose than high-resolution CT (HRCT) in the preoperative diagnosis of otosclerosis. These results indicate that CBCT has high sensitivity and specificity in the detection of hypodense lesions due to histologically active otosclerosis.

Microbiological profile of adenoid hypertrophy correlates to clinical diagnosis in children.

OBJECTIVE: Adenoid hypertrophy is a common condition in childhood, which may be associated with recurring acute otitis media (RAOM), otitis media with effusion (OME), and obstructive sleep apnea syndrome (OSAS). These different clinical characteristics have some clinical overlap; however, they might be explained by distinct immunologic and infectious profiles and result in various histopathologic findings of adenoid specimens. METHODS: A total of 59 children with adenoid hypertrophy undergoing adenoidectomy were studied. Three series of identical adenoid specimens were processed to hematoxylin-eosin (H.E.) and Gram staining and to respiratory virus specific real-time PCR, respectively. RESULTS: According to the clinical characteristics, patients were recruited into three groups: RAOM (n = 25), OME (n = 19), and OSAS (n = 15). Bacterial biofilms were detected in 21 cases, while at least one of the studied respiratory viruses was detected in 52 specimens. RAOM cases were significantly associated with biofilm existence (n = 20, P < 0.001). In contrast, OME group was characterized by the absence of bacterial biofilm and by normal mucosa. Showing a statistically significant correlation, all OME cases were positive for human bocavirus (HBoV, P < 0.001). CONCLUSIONS: Bacterial biofilms might contribute to the damage of respiratory epithelium and recurring acute infections resulting in RAOM. In OME cases persisting respiratory viruses, mainly HBoV, can cause subsequent lymphoid hyperplasia leading to ventilation disorders and impaired immunoreactivity of the middle ear cleft.

DPOAE Intensity Increase at Individual Dominant Frequency after Short-Term Auditory Exposure.

Previous experiments suggested the possibility of a short-term sound stimulus-evoked and transient increase in DPOAE amplitudes. This phenomenon is possibly due to the complexity of the outer hair cells and their efferent control system and the different time scales of regulatory processes. A total of 100 healthy subjects ranging from 18 to 40 years of age with normal hearing and normal DPOAE values in the range of 781-4000 Hz were recruited in the study. Diagnostic DPOAE measurements were performed after short-term sound exposure. We proposed a 10 sec, 50 dB sound impulse as the most effective stimulus for clinical practice between 40 and 60 sec poststimulus time to detect the aforementioned transient DPOAE increase. We developed a procedure for detection of this transient increase in DPOAE by the application of a short-term sound exposure. The phenomenon was consistent and well detectable. Based on our findings, a new aspect of cochlear adaptation can be established that might be introduced as a routine clinical diagnostic tool. A mathematical model was provided that summarizes various factors that determine electromotility of OHCs and serves as a possible clinical application using this phenomenon for the prediction of individual noise susceptibility.

No evidence for the expression of renin-angiotensin-aldosterone system in otosclerotic stapes footplates.

INTRODUCTION: Recent studies have reported genetic associations between with single nucleotide polymorphism (SNP) of the several genes of the renin-angiotensin-aldosterone (RAA) system in otosclerosis without the confirmation of RAA system expression in human stapes footplates. There are conflicting results. These results are conflicting because RAA system expression has been attributed exclusively to neural, vascular, and renal tissues, exclusively. MATERIALS AND METHODS: Ankylotic stapes footplates (n = 20), cortical bone fragments (n = 10), and human kidney tissue specimens (n = 10) were processed to hematoxylin-eosin (HE) staining and RAA system-specific immunofluorescent assay (IFA), respectively. RESULTS: Histologic diagnosis of otosclerosis was established in all ankylotic stapes footplates. Histologically active- (n = 13) and inactive (n = 7) foci of otosclerosis were consequently characterized by negative immunoreactions for renin, angiotensin converting enzyme (ACE), angiotensin-II (AT-II), and angiotensin-II receptor (AT-IIR), consequently. In cortical bones, a considerable RAA system expression was observed confirmed in the perivascular bone marrow progenitor cells. Kidney specimens, applied as positive controls, showed intense RAA system-specific immunoreactions. CONCLUSION: Concerning current observations, the 4 studied members of RAA system that did not display active expression were not expressed at protein level in otosclerotic stapes footplates. This phenomenon was independent from the histologic activity of otosclerosis. Between these conditions, the etiologic role of RAA system is questionable in the pathogenesis of otosclerosis.

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.

Low-frequency ultrasound for biofilm disruption in chronic rhinosinusitis with nasal polyposis: in vitro pilot study.

OBJECTIVES/HYPOTHESIS: Microbial biofilms have been implicated in the pathogenesis of chronic rhinosinusitis with nasal polyposis (CRSwNP). Although biofilms are characterized by an extremely high resistance against chemical and physical agents, low-frequency ultrasound (LFU) treatment has been suspected to be an efficient and safe method for biofilm disruption. STUDY DESIGN: Basic science experimental study. METHODS: A total of 10 patients with CRSwNP undergoing endoscopic sinus surgery were analyzed. Two series of identical nasal polyps (n = 20) were processed to hematoxylin-eosin (HE) and Gram staining and to continuous-wave LFU treatment (5 minutes, 0.4 MHz, 37°C), respectively. RESULTS: Presence of microbial biofilms was confirmed in all patients with CRSwNP. HE staining showed a strong correlation with the results of Gram protocol in biofilm detection. In the LFU-treated group (n = 10), a significantly decreased inflammatory cell count was found in the subepithelial layer of nasal polyps (P < .001). In addition, bacterial biofilms were completely removed from the surface of the epithelial layer. Microscopic tissue injuries or significant temperature changes were not detected due to LFU treatment. CONCLUSIONS: Between in vitro conditions, LFU treatment appeared to be a reliable and microscopically safe method for the disruption of microbial biofilms in CRSwNP. These results may provide a basis for a prospective human study investigating the efficacy and safety of this therapeutic modality alone or in combination with antibiotics or topical steroids in biofilm-positive cases of CRSwNP.

Perspectives of pharmacological treatment in otosclerosis.

To review our current knowledge of the pathologic bone metabolism in otosclerosis and to discuss the possibilities of non-surgical, pharmacological intervention. Otosclerosis has been suspected to be associated with defective measles virus infection, local inflammation and consecutive bone deterioration in the human otic capsule. In the early stages of otosclerosis, different pharmacological agents may delay the progression or prevent further deterioration of the disease and consecutive hearing loss. Although effective anti-osteoporotic drugs have become available, the use of sodium fluoride and bisphosphonates in otosclerosis has not yet been successful. Bioflavonoids may relieve tinnitus due to otosclerosis, but there is no data available on long-term application and effects on sensorineural hearing loss. In the initial inflammatory phase, corticosteroids or non-steroidal anti-inflammatory drugs may be effective; however, extended systemic application may lead to serious side effects. Vitamin D administration may have effects on the pathological bone loss, as well as on inflammation. No information has been reported on the use of immunosuppressive drugs. Anti-cytokine targeted biological therapy, however, may be feasible. Indeed, one study on the local administration of infliximab has been reported. Potential targets of future therapy may include osteoprotegerin, RANK ligand, cathepsins and also the Wnt-ß-catenin pathway. Finally, anti-measles vaccination may delay the progression of the disease and potentially decrease the number of new cases. In conclusion, stapes surgery remains to be widely accepted treatment of conductive hearing loss due to otosclerosis. Due to lack of solid evidence, the place of pharmacological treatment targeting inflammation and bone metabolism needs to be determined by future studies.

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 value of HRCT in stapes fixations corresponding to hearing thresholds and histologic findings.

OBJECTIVE: To estimate the correlations between high-resolution computed tomography (HRCT) scans, preoperative audiometric findings and histopathologic results in stapes ankylosis. STUDY DESIGN: Retrospective case review. SETTING: Tertiary referral center. PATIENTS: A total of 57 patients with stapes ankylosis, who underwent unilateral stapedectomies were analyzed. INTERVENTIONS: Diagnostic and therapeutic. MAIN OUTCOME MEASURES: Preoperative HRCT examinations were performed in all cases. Findings of HRCT were categorized according to Marshall's grading system. Preoperative air-bone gaps (ABGs) and bone conduction (BC) thresholds were determined. Stapes footplates removed surgically were processed using hematoxylin and eosin staining. RESULTS: Active otosclerosis (n = 29) was demonstrated by HRCT with a sensitivity of 76.31%, whereas its sensitivity to establish inactive otosclerosis (n = 13, 61.9%) and nonotosclerotic stapes fixations (n = 15, 51.7%) was much lower. Nonotosclerotic stapes fixations were characterized by pure conductive hearing loss that was not associated with HRCT findings. HRCT grades showed statistically significant association with BC levels at the averages of 0.5-1-2 kHz frequencies in the group of ears with inactive otosclerosis, exclusively (p < 0.05). CONCLUSION: HRCT is a reliable imaging method in the preoperative diagnosis of different types of stapes fixations. Imaging findings should be evaluated together with clinical history and audiometric data for obtaining as precise diagnosis for stapes fixation as possible.

Expression of bone morphogenetic protein 2, 4, 5, and 7 correlates with histological activity of otosclerotic foci.

CONCLUSION: This study is the first to establish that bone morphogenetic protein 5 (BMP5) plays a role in the pathogenesis of otosclerosis. These results confirm that elevated expression levels of BMPs, members of the transforming growth factor (TGF)-ß superfamily, contribute to the pathologically increased bone turnover in early, active stages of otosclerosis. OBJECTIVES: Otosclerosis is a complex bone remodeling disorder of the otic capsule, which might be characterized by increased expression of different types of BMPs. TGF-ß and BMP are both members of the TGF-ß superfamily and play a critical role in bone resorption and new bone formation. It has been suggested that BMP and its receptors may be involved in the pathologically increased bone turnover observed in otosclerosis. METHODS: Fifty-one otosclerotic and 16 non-otosclerotic ankylotic stapes footplates were histologically analyzed: conventional hematoxylin-eosin staining and BMP2, 4, 5, and 7specific immunofluorescent assays were performed. Cortical bone fragments (n = 35) and incus specimens (n = 6) were used as negative controls. RESULTS: Active otosclerosis (n = 39) was characterized by increased expression of BMP2, 4, 5, and 7. Inactive cases of otosclerosis (n = 12) were characterized by negative immunoreaction for BMPs. Non-otosclerotic stapes specimens (n = 16) and negative controls (n = 41) showed negligible BMP expression. The BMP expression pattern showed a strong correlation with the histological activity of otosclerosis.

Tumor necrosis factor-α receptor expression correlates with mucosal changes and biofilm presence in chronic rhinosinusitis with nasal polyposis.

OBJECTIVES/HYPOTHESIS: Biofilms might play a potential role in the pathogenesis and high recurrence rate of chronic rhinosinusitis with nasal polyposis (CRSwNP). Biofilm persistence has been thought to correlate with epithelial damage, subepithelial inflammatory cell infiltration, and tumor necrosis factor-α receptor (TNFR) expression in CRSwNP. STUDY DESIGN: Case-control experimental study. METHODS: A total of 36 patients with CRSwNP undergoing endoscopic sinus surgery were analyzed. The negative control group consisted of eight patients undergoing septoplasty for nasal obstruction without CRSwNP. The nasal polyps and inferior turbinate mucosa samples applied as negative controls were processed by hematoxylin-eosin (HE) and Gram staining and TNFR-I and TNFR-II-specific immunofluorescent assay. RESULTS: Biofilm was detected in 29 of 36 patients with CRSwNP and in none of the eight negative controls. Staining by HE showed strong correlation with the results of Gram staining protocol. In the biofilm-positive cases, TNFR-I and TNFR-II displayed homogeneous pattern of significantly increased epithelial expression compared to the biofilm-negative nasal polyps. In cases of biofilm absence, the expression pattern of TNF-α receptors was characterized by increased TNFR-II-specific immunoreaction. It was found that biofilm detectability corresponded to the integrity of nasal epithelium and to the dominant inflammatory cell type of the subepithelial layer. CONCLUSIONS: Persisting biofilms might increase the epithelial sensitivity against TNF-α that result in epithelium destruction. Coexistence of biofilms and increased TNFR expression might explain the inflammatory mucosal changes, functional disorders, and therapy resistance featuring CRSwNP.

No evidence for disturbed COL1A1 and A2 expression in otosclerosis.

Otosclerosis is a complex bone remodeling disorder of the human otic capsule that might be associated with various mutations of A1 and A2 alleles of type-I collagen. The study herein presented, investigates the possibilty of the genetic involvement of type-I collagen in the pathogenesis of histologically confirmed otosclerosis. A total of 55 ankylotic stapes footplates were analyzed. Cortical bone fragments (n = 30), incus (n = 3) and malleus (n = 2) specimens were employed as negative controls. Specimens were divided into two groups. The first group was processed using conventional H.E. hematoxylin-eosin (H.E.) staining and type-I collagen-specific immunofluorescent assay (IFA), while the second group was examined by COL1A1 and A2-specific RT-PCR. Otosclerotic- (n = 31) and non-otosclerotic stapes footplates (n = 9) as well as cortical bones (n = 20), incus (n = 2) and malleus specimens (n = 1) showed normal and quite similar A1 and A2 allele expression confirmed by IFA. RT-PCR analysis revealed normal and consistent mRNA expression of both alleles in each specimen. Expression levels and patterns of COL1A1/A2 alleles did not show significant correlation with the histological diagnosis of otosclerosis. Type-I collagen is a highly conserved structure protein, which plays a fundamental role in the integritiy of various connective tissues. Mutations of A1 and A2 alleles result in serious systemic disorders of the skeleton, tendons and skin. Since otosclerosis is an organ-specific disease, it is difficult to explain its genetic association with type-I collagen. In conclusion, we found no evidence supporting the putative link of COL1A1 and COL1A2 alleles with otosclerosis.