[Retrocochlear diagnostics for acute hearing loss and successful therapy]. / Retrocochleäre Diagnostik im Rahmen der Hörsturzuntersuchung und erfolgreiche Therapie.

A 41-year-old female patient presented due to acute onset of unilateral hearing loss 3 months previously and persistent since then. Systemic therapy with oral glucocorticoids in decreasing doses had been performed beforehand, but did not lead to any improvement. In the course of audiological diagnostics, based on subjective and objective methods, a retrocochlear hearing disorder was suspected. A meningioma was diagnosed by diagnostic imaging. Subsequent surgical removal achieved a significant hearing improvement.

Multicenter clinical trial on functional evaluation of transoral laser microsurgery for supraglottic laryngeal carcinomas.

BACKGROUND: Transoral laser microsurgery (TLM) is an accepted and effective treatment strategy for supraglottic carcinomas. Data on oncologic and to a lesser extent functional outcomes have been published by mainly European specialized single institutions. TLM for supraglottic carcinomas has never been tested in a multicenter trial on its applicability as surgical standard at every hospital. OBJECTIVES: To test the efficacy of TLM supraglottic laryngectomy (TLM-SGL) in terms of swallowing function, oncologic outcome parameters, morbidity, complications of treatment, and quality of life in a multicenter setting. METHODS: The study is designed as a multicenter (approximately 25 centers), non-randomized, single-arm study with a targeted number of 200 previously untreated patients with squamous cell carcinomas (SCC) of the supraglottic larynx T2/T3 N0-3 M0; UICC stage II-IVa. The surgical treatment consists of TLM-SGL and elective or therapeutic uni- or bilateral selective neck dissection (SND). After pathologic risk stratification adjuvant radio- (RT) or radiochemotherapy (RCT) is indicated. Patients are followed-up for 2 years post surgically. Swallowing function is assessed by fibreoptic endoscopic evaluation of swallowing (FEES). The primary endpoint is aspiration-free swallowing at 12 months as established using FEES and defined as grade < 6 of penetration-aspiration scale (PAS). Secondary endpoints include local control, larynx preservation, overall and disease-free survival, complications and side effects of treatment, prevalence of tracheostomy and percutaneous endoscopic gastrostomy (PEG)-tube-feeding, and dysphagia-specific quality of life (QoL) assessed by the MD Anderson Dysphagia Inventory (MDADI) as well as voice-related QoL assessed by the Voice Handicap Index (VHI).

Lysyl oxidase like-4 monoclonal antibody demonstrates therapeutic effect against head and neck squamous cell carcinoma cells and xenografts.

A new member of the lysyl oxidase (LOX) family, lysyl oxidase-like 4 (LOXL4), is overexpressed in head and neck squamous cell carcinoma (HNSCC) compared to normal squamous epithelium. A monoclonal antibody (mAb) derived from fusion of Balb/c mouse splenocytes immunized with LOXL4 specific peptide was used to evaluate its therapeutic efficacy in 15 HNSCC cell lines associated with LOXL4 overexpression. For xenograft experiments 41 severe combined immunodeficient (SCID) mice were used to analyze LOXL4-mAb mediated tumor regression. Cell viability was analyzed using cytotoxicity-, and clonogenic-assays. Significant suppression of tumor cell growth was observed in 12 out of 15 (80%) tumor cell lines after 48 hr exposure to the mAb (LD50 of 15 µg/ml to 45 µg/ml). The effect induced by the antibody could be blocked by pre-incubation of the antibody with the peptide used for immunization of the mice and antibody generation, indicating that the effect of the antibody is specific. In mice inoculated with HNSCC cells, i.v. injections of the LOXL4-mAb resulted within 70 days in extensive tumor destruction in all treated animals whereas no tumor regression occurred in control animals. In mice pre-immunized i.v. with LOXL4-mAb and subsequently injected with HNSCC cells, tumor development was considerably delayed in contrast to non LOXL4-mAb pre-immunized animals. These results demonstrate that the LOXL4-mAb has potent antitumor activity and suggest its suitability as a therapeutic immune agent applicable to HNSCC exhibiting tumor specific upregulation of LOXL4.

Manifestation of eosinophilic granulomatosis with polyangiitis in head and neck.

INTRODUCTION: Besides an obvious clinical involvement of the ear, nose and throat (ENT)-region in Eosinophilic Granulomatosis with Polyangiitis (EGPA), systematic data is sparse. Only a few case series and case reports are available that particularly describe rhinological, otological or other manifestations of EGPA in the ENT-region. Therefore, the objective of this study is to systematically describe data on ENT-region involvement in a large series of EGPA patients. METHOD: EGPA patients examined in the Department of Otorhinolaryngology of the Christian-Albrechts-University of Kiel between 1990 and 2010 were included in the study. Criteria for ENT-manifestation were assigned to five subgroups (history, ENT examination, audiological and rhinological diagnostic findings and cranial MRI) and documented cumulatively. EGPA patients were examined in a standardized way based on the validated Ear Nose and Throat Activity Score (ENTAS) or its precursor, including audiological and rhinological diagnostic findings. MRI scans were analysed to further evaluate ENT involvement. RESULTS: A total of 95 EGPA patients were included in the study. In approximately 80% of them, ENT-involvement was documented and the assumption of a frequent rhinological manifestation in patients with EGPA was confirmed. Moreover, the data reveals remarkable evidence for an otological manifestation. A missing correlation between the rhinological and the otological manifestation indicates an independent autoimmune-inflammatory process for this manifestation. CONCLUSION: The data of the largest monocentric study presented here confirms the hypothesis of a frequent ENT involvement in EGPA patients, in whom rhinological and otological manifestations are most common. Therefore, treatment should include long term follow-up and should be managed interdisciplinary.

Head and neck cancer cells and xenografts are very sensitive to palytoxin: decrease of c-jun n-terminale kinase-3 expression enhances palytoxin toxicity.

OBJECTIVES: Palytoxin (PTX), a marine toxin isolated from the Cnidaria (zooanthid) Palythoa caribaeorum is one of the most potent non-protein substances known. It is a very complex molecule that presents both lipophilic and hydrophilic areas. The effect of PTX was investigated in a series of experiments conducted in head and neck squamous cell carcinoma (HNSCC) cell lines and xenografts. MATERIALS AND METHODS: Cell viability, and gene expression of the sodium/potassium-transporting ATPase subumit alpha1 (ATP1AL1) and GAPDH were analyzed in HNSCC cells and normal epithelial cells after treatment with PTX using cytotoxicity-, clonogenic-, and enzyme inhibitor assays as well as RT-PCR and Northern Blotting. For xenograft experiments severe combined immunodeficient (SCID) mice were used to analyze tumor regression. The data were statistically analyzed using One-Way Annova (SPSS vs20). RESULTS: Significant toxic effects were observed in tumor cells treated with PTX (LD50 of 1.5 to 3.5 ng/ml) in contrast to normal cells. In tumor cells PTX affected both the release of LDH and the expression of the sodium/potassium-transporting ATPase subunit alpha1 gene suggesting loss of cellular integrity, primarily of the plasma membrane. Furthermore, strong repression of the c-Jun N-terminal kinase 3 (JNK3) mRNA expression was found in carcinoma cells which correlated with enhanced toxicity of PTX suggesting an essential role of the mitogen activated protein kinase (MAPK)/JNK signalling cascades pathway in the mechanisms of HNSCC cell resistance to PTX. In mice inoculated with carcinoma cells, injections of PTX into the xenografted tumors resulted within 24 days in extensive tumor destruction in 75% of the treated animals (LD50 of 68 ng/kg to 83 ng/kg) while no tumor regression occurred in control animals. CONCLUSIONS: These results clearly provide evidence that PTX possesses preferential toxicity for head and neck carcinoma cells and therefore it is worth further studying its impact which may extend our knowledge of the biology of head and neck cancer.

Facilitation properties in electrically evoked compound action potentials depending on spatial location and on threshold.

Spiral ganglion neurons (SGNs) facilitation properties can be recorded utilizing electrically evoked compound action potential (ECAP). While intracochlear variation of the ECAP threshold in relation to its electrode channel is reported, no study investigated its impact on facilitation. In this study, we quantified intracochlear variation of the facilitation properties in cochlear implants (CI) using ECAPs. We hypothesized that the facilitation effect is dependent on the electrode channel and its ECAP threshold. Therefore, ECAPs were recorded in 23 CI subjects. For each subject, five default (channel-derived) and up to two additional (threshold-derived) stimulation sites were defined. Facilitation was quantified by the paradigm introduced by (Hey et al., 2017) with optimized parameter settings. For each channel the maximum facilitated amplitude was determined by a series of ECAP measurements. A linear mixed-effects model was used to investigate the impact of the electrode channel and ECAP threshold on the maximum facilitated amplitude. The maximum facilitated amplitude was found to be dependent on the ECAP threshold and independent on the electrode channel. We conclude that the facilitation paradigm is a useful and feasible tool to gain local information on the SGNs temporal processing patterns.

Effect of postoperative systemic prednisolone on short-term and long-term outcomes in chronic rhinosinusitis with nasal polyps: A multi-centered randomized clinical trial.

Introduction: The objective of this study was to determine whether postoperative additive systemic steroid administration in chronic rhinosinusitis with nasal polyps (CRSwNP) impacted selected endoscopic, subjective and objective outcome measures. Methods: This was a prospective, randomized, double-blind, placebo-controlled, noninferiority multicenter trial of n=106 patients with CRSwNP. All patients underwent primary functional endoscopic sinus surgery (FESS) followed by topical nasal steroids. Patients were randomized to a systemic steroid or placebo for 1 month. Patients were followed up for 2 years over 9 time points. The primary outcome measures were the differences between groups with respect to the nasal polyp score (NPS) and sinonasal quality of life (SNQoL). Secondary outcome measures included interactions with respect to the Lund-Kennedy score (LKS), sinonasal symptoms, general quality of life (GQoL), 16-item odor identification test scores, recurrence rates, need for revision surgery and mucus biomarker levels. Results: 106 patients were randomized to either the placebo or the systemic steroid group (n=53 per group). Postoperative systemic steroids were not superior to placebo with respect to all primary (p= 0.077) and secondary outcome measures (p>0.05 for all). Reported adverse events were similar between the two groups. Conclusion: In conclusion, the addition of postoperative systemic steroids after primary FESS did not confer a benefit over topical steroid nasal spray alone with respect to NPS, SNQOL, LKS, GQOL, sinonasal symptoms, smell scores, recurrence rates, the need for revision surgery or biomarkers over a short-term follow-up of up to 9 months and a long-term follow-up of up to 24 months in CRSwNP patients. Functional endoscopic surgery did, however, show a strong effect on all outcome measures, which remained relatively stable up to the endpoint at 2 years.

Rituximab for refractory granulomatosis with polyangiitis (Wegener's granulomatosis): comparison of efficacy in granulomatous versus vasculitic manifestations.

OBJECTIVE: First, to investigate the overall efficacy and safety of rituximab (RTX) in refractory granulomatosis with polyangiitis (GPA) in a tertiary referral centre. Second, to compare the efficacy of RTX in granulomatous and vasculitic manifestations in GPA. PATIENTS AND METHODS: This study comprised a retrospective, standardised data collection from all patients who received RTX for refractory Wegener's granulomatosis from 2002 to 2010. Patients were assessed by a standardised interdisciplinary diagnostic procedure (including ear, nose and throat and ophthalmology assessment, MRI, immunodiagnostics, B-cell levels and Birmingham Vasculitis Activity Score) and were treated by standardised therapeutic regimens according to available evidence. RESULTS: 59 patients received 75 cycles of RTX. 9.3% achieved complete remission. A response was documented in 61.3% (improvement in 52%, unchanged disease activity in 9.3%), 26.7% had refractory disease. Birmingham Vasculitis Activity Score, disease extent index, erythrocyte sedimentation rate, C-reactive protein and prednisolone demand decreased significantly. All patients achieved B-cell depletion. Granulomatous manifestations such as orbital granuloma and pachymeningitis were more frequently refractory to RTX than vasculitis or other granulomatous manifestations. Thus, for example, complete remission/improvement was found in 89.2% of patients with renal disease and in only 44.4% of those with orbital masses (p=0.003). The relapse rate was 44.4% after a median period of 13.5 months. Adverse events occurred in 29%, pneumonia in 15% and death in 3%. CONCLUSION: The overall response rate of refractory GPA to RTX was high (61.3% complete remission or improvement). Response rates of vasculitic manifestations were excellent; failure of response/progress was mostly due to granulomatous manifestations, especially orbital masses. Relapse rates were high (40%) despite maintenance treatment.

Intra- and inter-rater reliability of endonasal activity estimation in granulomatosis with polyangiitis (Wegener´s).

OBJECTIVES: Granulomatosis with polyangiitis (GPA) frequently starts with an affection of the nasal and paranasal mucosa. Localised GPA of the nasal mucosa or persistent disease activity ('grumbling disease') is often encountered even under immunosuppressive therapy. Necessity for reconstructive surgery is common and careful scheduling to prevent failure and minimise revision rates is crucial. Therefore, reliable estimation of GPA activity in the upper airways using a score is mandatory for diagnosis, follow-up and scheduling reconstructive surgery. METHODS: Fifty endoscopic, endonasal images of 45 patients with GPA were used. Twelve (4 German, 8 Mexican) experienced (n=7) and inexperienced (n=5) physicians assessed GPA-activity at two times (T1/T2) in dichotomy and in a grading approach (none, mild, moderate and high activity) using the novel ENT Activity Score (ENTAS). All documents were written in English. RESULTS: Estimation of activity in dichotomy (none vs. mild/moderate/high): Cohen's Kappa (κ) for intra-rater reliability T1/T2 in inexperienced and experienced physicians was κ=0.58 (agreement 85%) and κ=0.72 (agreement 91%). The inter-rater reliability (Fleiss's κ) T1/T2 for inexperienced and experienced physicians was κ=0.62/κ=0.59 and κ=0.50/κ=0.58 respectively. Estimation of activity in grading approach (none, mild, moderate, high): for inexperienced physicians the intra-rater reliability T1/T2 was κ=0.67 (agreement 56%) and the inter-rater reliability at T1/T2 was κ=0.29 (intraclass correlation coefficient, ICC=0.69) and κ=0.27 (ICC=0.59). For experienced physicians the intra-rater reliability T1/T2 was κ=0.80 (agreement 67%) and the inter-rater reliability at T1 and T2 was κ=0.41 (ICC=0.77) and κ=0.39 (ICC=0.75) respectively. CONCLUSIONS: Intra-rater reliability is high in decision in dichotomy and even in grading activity. There is no difference for experienced or inexperienced physicians. Inter-rater reliability is high in dichotomy, but low for activity grading. Thus, the ENTAS provides a reliable instrument for assessing, documenting and following GPA-related disease activity in the upper respiratory tract. The relationship of activity and following damage needs to be investigated in further studies.

Improved outcome in 445 patients with Wegener's granulomatosis in a German vasculitis center over four decades.

OBJECTIVE: To determine the long-term outcome in patients with Wegener's granulomatosis (WG) over 4 decades in an academic hospital unit specializing in rheumatology. METHODS: We included 290 patients, divided them into 2 cohorts, and compared them with the historical cohort of 155 patients. Comparisons were retrospective regarding disease manifestations, therapy, mortality, and incidence of malignancies. The historical cohort (cohort 1) included 155 patients diagnosed between 1966 and 1993, cohort 2 included 123 patients diagnosed between 1994 and 1998, and cohort 3 included 167 patients diagnosed between 1999 and 2002. RESULTS: Over time, the interval between first symptoms and diagnosis was reduced by half (from 8 months to 4 months). Organ manifestations were similar in the 3 cohorts, and more than 80% of patients still required cyclophosphamide (CYC); however, the median cumulative dose was reduced significantly (from 67 gm in cohort 1 to 36 gm in cohort 2 and to 24 gm in cohort 3). The standardized mortality ratios (SMRs) declined (from 2.1 in cohort 1 to 1.41 in cohort 2 and to 1.03 in cohort 3), with fewer deaths related to WG and/or therapy (86.4% in cohort 1, 76.9% in cohort 2, 50% in cohort 3), decreasing relapse rates (63.9% in cohort 1, 51.2% in cohort 2, 35.3% in cohort 3), and no increased rate of malignancies. Compared with young females, young males had a considerably higher SMR (8.87 [95% confidence interval 4.05-16.8]) and more frequent renal manifestations (54.4% versus 33.8%). CONCLUSION: Mortality of WG patients declined over the last 4 decades, probably due to improved diagnostic and therapeutic procedures and increased awareness of WG, which led to earlier diagnosis and therapy, reduction in relapse rates, and lower cumulative CYC dose with fewer deaths related to therapy.

Cortical habituation deficit in tinnitus sufferers: contingent negative variation as an indicator of duration of the disease.

Cortical attention and habituation parameters are altered in patients suffering from tinnitus. The aim of the study was to quantify cortical attention and habituation parameters in tinnitus patients by recording the contingent negative variation (CNV) response and to correlate amplitudes of different CNV parameters with duration of disease. Twenty patients suffering from tinnitus (median: 44 years) and twenty age- and sex-matched healthy controls (median: 41 years) were tested by a CNV paradigm. We recorded overall CNV, initial CNV, and terminal CNV and calculated habituation slopes. All CNV parameters were Spearman-correlated with individual duration of disease. Highly significant between groups differences emerged in total (tinnitus: -8.4 uV vs. controls: -3.8 uV), initial (-11.2 vs. -6.0 uV), and terminal CNV (-11.9 vs. -6.5 uV) demonstrating higher negative amplitudes in tinnitus patients. Habituation differed in total and terminal CNV, indicating missing habituation in tinnitus patients. Overall CNV (ϱ = -.365) and initial CNV (ϱ = -.529) showed a medium Spearman correlation with duration of disease. We conclude that the correlation between duration of tinnitus and the initial CNV amplitudes indicates an altered state of cortical excitability that can also be observed in more negative CNV-amplitudes in tinnitus patients. We assume that this state indicates a chronicity process in tinnitus disease.

Tonsillar swabs and sputum predict SLPI- and AnxA2 expression in tonsils: A prospective study on smoking dependent SLPI- and AnxA2-expression, and tonsillar HPV infection.

Previous retrospective studies have elucidated a correlation between secretory leucocyte protease inhibitor (SLPI) and Annexin A2 (AnxA2), patient smoking status and tonsillar human papilloma virus (HPV) status. The current study assessed these parameters prospectively and to the best of our knowledge, analyzed SLPI-/AnxA2-expression for the first time in tonsillar swabs and sputum. Samples were obtained from 52 patients with tonsillar squamous cell carcinoma and 163 patients with tonsillar hyperplasia (H; n=56) and chronic or recurrent tonsillitis (CRT; n=107). HPV-DNA, SLPI and AnxA2 gene expression was analyzed in sputum, tonsillar swabs and tissue by performing reverse transcription-quantitative PCR. Results were compared with smoking status, revealing that smoking resulted in significantly increased SLPI gene expression in all biomaterials of all cases. SLPI-gene expression was significantly decreased in all HPV-DNA-positive samples (tissue/swab/sputum), while AnxA2 was significantly increased in all HPV-DNA-positive samples. Results from swabs and sputum were able to predict SLPI- and AnxA2 gene expression of the corresponding tonsil. The current prospective study confirmed previous retrospective results underlining this hypothesis: Smoking enhances SLPI-expression, preventing HPV-binding to AnxA2. HPV-binding to AnxA2 appears essential for successful cell-entry. SLPI/AnxA2-gene expression in swabs and sputum reflect their expression in tonsillar tissue. Accordingly, a positive AnxA2/SLPI-ratio in sputum/swabs could possibly be used to reduce HPV-associated carcinogenesis, by performing tonsillectomy or HPV-vaccination in patients with positive AnxA2/SLPI-ratios.

Antimicrobial peptides in nasal secretion and mucosa with respect to S. aureus colonisation in Wegener´s granulomatosis.

OBJECTIVES: Nasal S. aureus carrier rates are significantly higher in patients with Wegener's granulomatosis (WG) compared to healthy controls (HC), and nasal colonisation is a risk-factor for relapse. Antimicrobial peptides (AMP) are important defence molecules maintaining an intact barrier function. It is the aim of this study to see if there is a possible link between the nasal AMP pattern and S. aureus colonisation, a link which has not been investigated so far. METHODS: ELISA was applied to quantify LL-37 and hBD-3 concentrations in nasal secretions (14 WG patients, 13 HC) with and without nasal S. aureus colonisation. Immunohistochemistry was used to detect the cellular sources of AMP in the nasal mucosa. Functional analyses of primary nasal epithelial cell cultures (NEC) of these groups stimulated with S. aureus were performed. RESULTS: LL-37 was found in significantly higher concentrations in colonised individuals (WG: p=0.001; HC: p=0.014).Using immunohistochemistry, local cellular sources for AMP could be demonstrated. After stimulation with S. aureus, significantly higher concentrations of LL-37 and hBD-3 could be detected in the supernatant of NEC of WG patients (LL-37: p=0.001; hBD-3: p=0.001) and HC (LL-37: p=0.019; hBD-3: p=0.001). HBD-3 concentrations were significantly lower in the supernatant of stimulated NEC of WG patients compared to the NEC of HC (p=0.032), and the dynamic range of the hBD-3 answer was significantly smaller in WG compared to HC (p=0.016). CONCLUSIONS: The dynamic response towards challenges with microbes is dysregulated in WG, and this might be one reason for higher S. aureus colonisation rates in WG.

Antimicrobial peptides in nasal secretion and mucosa with respect to Staphylococcus aureus colonization in chronic rhinosinusitis with nasal polyps.

OBJECTIVE: Nasal carriage of Staphylococcus aureus in patients with chronic rhinosinusitis with nasal polyps (NP) is hypothesized to have pathophysiological impact on the disease. Antimicrobial peptides (AMP), especially human beta-defensin-3 (hBD-3) and LL-37, are an important part of the multifactorial defence against microorganisms in barrier organs like the nasal mucosa. The interaction of S. aureus colonization and AMP in nasal secretions and mucosa of NP were investigated in this study. PATIENTS AND METHODS: AMP were quantified in nasal secretions of 13 normal controls (NC) and 12 NP patients, each with and without S. aureus colonization, by ELISA. Immunohistochemistry was used to investigate the cellular sources of AMP in the nasal mucosa. To explore the AMP response of primary nasal epithelial cell cultures (NEC) towards S. aureus stimulation, a functional assay was established. RESULTS: AMP could be demonstrated in nasal secretions of all groups without differences in hBD-3 concentrations comparing S. aureus carriers vs. non-carriers. In NC, higher LL-37 concentrations were observed in S. aureus colonized as compared to non-colonized patients. This effect was not detectable in NP patients. Epithelial cells, submucosal glands and cells of the connective tissue could be identified as sources of AMP by immunohistochemistry. An AMP response of NEC towards S. aureus stimulation was detected in all groups. CONCLUSION: In NP patients, LL-37 response towards S. aureus colonization is disturbed while the ability of NEC to respond on S. aureus challenge is preserved. This deregulation of the nasal barrier could be involved in the multifactorial pathophysiology of NP.

Total nasal reconstruction: a 6-year experience with the three-stage forehead flap combined with the septal pivot flap.

Several different methods can be applied for repairing total nasal defects. Most of them are based on some common principles and techniques widely accepted and adopted by experienced surgeons. We have been using most of these techniques during the past two decades, however modifying and refining them several times. Our observations and sometimes disappointing experiences led to a concept that has remained unchanged for the past 6 years. It comprises three regular operative steps and sometimes a fourth surgical intervention for repair and refinement. First stage: Reconstruction of the septum using a bipedicled composite septal pivot flap (SPF), of the intranasal lining (INL), and the cover being established by elevating a full-thickness paramedian forehead flap (PMFF). Second stage: Re-elevation of the PMFF, thinning of its layers, and reconstruction of the subsurface framework using autogenous rib cartilage. Third stage: Division of the pedicle and minor corrections. We have been using this technique presented here since 2004 in nine consecutive patients with subtotal to supratotal nasal defects. Seven cases have been repaired completely by now and can be evaluated carefully. With this technique, results have significantly improved and have been stable to date.

The interaction of smoking habit, SLPI and AnxA2 in HPV associated head and neck and other cancers.

Six own studies confirm a correlation between smoking, expression of the secretory leukocyte protease inhibitor (SLPI, an antileukoproteinase) and expression of Annexin A2 (AnxA2), and their influence on human papilloma virus (HPV)-infections. SLPI and HPV are ligands of AnxA2. This correlation was tested on 928 tissue samples from 892 patients in six independent studies [squamous cell carcinoma of the head and neck (HNSCC), n = 522; non-neoplastic tonsils n = 214; clinically normal mucosa, n = 93 (of these n = 57 were obtained from patients treated for non-malignant diseases and n = 36 were obtained from HNSCC-patients) and vulvar squamous cell carcinoma (VSCC) n = 99]. HPV-DNA-status was determined by GP5+/GP6+-PCR, followed in case of HPV-positivity by Sanger sequencing and RT-PCR using HPV-type specific primers. SLPI- and AnxA2-gene-expression was determined by RT-q-PCR; SLPI-protein-expression was additionally determined by immunohistochemistry (IHC); the data were correlated with each other and with patient characteristics. Smoking results in increased SLPI-gene- and protein- and AnxA2-gene-expression with significantly higher SLPI- than AnxA2-gene-expression. SLPI is decreased in non-smokers with a continuous AnxA2-surplus. HPV-status correlates with smoking habit, with smokers being mostly HPV-negative and non-smokers HPV-positive. We hypothesize that smoking leads to SLPI-overexpression with SLPI-binding to AnxA2. Thus, HPV cannot bind to AnxA2 but this seems pivotal for HPV-cell-entry. Smoking favors SLPI-expression resulting in HPV-negative carcinomas, while HPV-positive carcinomas are more common in non-smokers possibly due to a surplus of unbound AnxA2. In addition, the hypothesis may contribute to understand why smokers show increased oral HPV-prevalence in natural history studies but do not necessarily develop HPV-associated lesions.

HPV DNA/RNA detection in various oral and oropharyngeal biomaterials identifies active HPV infections also in non-neoplastic tonsils.

Previous studies describe a correlation between HPV-positivity and non-smoking in TSCC; p16INK4A-expression as surrogate-marker for HPV-DNA/RNA-positivity is discussed controversially. In the present study, these parameters are assessed prospectively. HPV-status of sputum and tonsillar-swabs was analyzed to determine their validity as surrogate-marker for tissue-HPV-status. TSCC- (n = 52) and non-neoplastic tonsillar tissue (n = 163) were analyzed. HPV-DNA- and HPV-RNA-status of total sputum, cellular fraction and supernatants, tonsillar-swabs and -tissue was determined by (RT)-PCR. Immunohistochemistry determined p16INK4A-expression. 23/163 (14.2%) non-neoplastic tonsils were HPV-DNA-positive; five patients (3 HPV16, 2 HPV11) had active HPV-infections (HPV-RNA-positive), in all biomaterials. 140/163 (85.9%) patients were either HPV-DNA-positive or HPV-DNA-negative in all samples. 21/52 (40.4%) TSCC-tonsils were HPV-DNA-positive; 17 patients were HPV-RNA-positive (14 HPV16; 4 HPV18). 40/52 (76.9%) TSCC-patients were congruent in all biomaterials. p16INK4A-expression alone would have misclassified the HPV-status of 14/52 (26.2%) TSCC-patients. This prospective study confirms the discrepancy between HPV-status and p16INK4A-expression and the significant correlation between non-smoking and HPV-DNA-positivity. HPV-sputum- and/or swab-results do not consistently match tissue-results, possibly having (detrimental) consequences if those were used to assess tissue-HPV-status. In the 5 patients with active HPV infection in the non-neoplasitic tonsils, tonsillectomy likely prevented subsequent development of TSCC.