Open-ST: High-resolution spatial transcriptomics in 3D.

Spatial transcriptomics (ST) methods unlock molecular mechanisms underlying tissue development, homeostasis, or disease. However, there is a need for easy-to-use, high-resolution, cost-efficient, and 3D-scalable methods. Here, we report Open-ST, a sequencing-based, open-source experimental and computational resource to address these challenges and to study the molecular organization of tissues in 2D and 3D. In mouse brain, Open-ST captured transcripts at subcellular resolution and reconstructed cell types. In primary head-and-neck tumors and patient-matched healthy/metastatic lymph nodes, Open-ST captured the diversity of immune, stromal, and tumor populations in space, validated by imaging-based ST. Distinct cell states were organized around cell-cell communication hotspots in the tumor but not the metastasis. Strikingly, the 3D reconstruction and multimodal analysis of the metastatic lymph node revealed spatially contiguous structures not visible in 2D and potential biomarkers precisely at the 3D tumor/lymph node boundary. All protocols and software are available at https://rajewsky-lab.github.io/openst.

[Recurrent laryngeal papillomatosis]. / Rezidivierende Larynxpapillomatose.

Human papillomaviruses (HPV) 6 and 11 cause 90% of recurrent laryngeal papillomatosis (RLP). It is unclear whether recurrences are caused by new infections or the spread of infected cells. Symptomatic and sometimes curative treatment is laser surgery or conventional microsurgical removal. RLP surgery aims to relieve shortness of breath and improve the voice. Patients (especially children) are affected by voice problems, repetitive surgeries, pulmonary manifestations, and psychological trauma. Vaccination with Gardasil 9 (Merck & Co., Rahway, NJ, USA) prevents new infections with HPV 6, 11, 16, 18, 31, 33, 45, 52, and 58 and induces vaccine antigen-specific antibodies and CD4+ T helper cells. According to current studies, RLP can be avoided with prophylactic vaccination. The treatment is associated with a general vaccination risk (European Medicines Agency approval: girls and boys from 9 years). Studies also show that the vaccine after removal of HPV-associated neoplasia/papilloma prevents recurrences. Extension of the vaccination recommendation to prevent recurrences of HPV-associated diseases in men may promote applicability and herd immunity. For rare and treatment-refractory cases with laryngotracheal involvement, systemic therapy with bevacizumab (e.g. Avastin; Genentech, San Francisco, CA, USA), a VEGF antibody, is a promising adjuvant treatment option.

Analysis of Severe Acute Respiratory Syndrome 2 Replication in Explant Cultures of the Human Upper Respiratory Tract Reveals Broad Tissue Tropism of Wild-Type and B.1.1.7 Variant Viruses.

BACKGROUND: The upper respiratory tract (URT) is the primary entry site for severe acute respiratory syndrome 2 (SARS-CoV-2) and other respiratory viruses, but its involvement in viral amplification and pathogenesis remains incompletely understood. METHODS: In this study, we investigated primary nasal epithelial cultures, as well as vital explanted tissues, to scrutinize the tropism of wild-type SARS-CoV-2 and the recently emerged B.1.1.7 variant. RESULTS: Our analyses revealed a widespread replication competence of SARS-CoV-2 in polarized nasal epithelium as well as in the examined URT and salivary gland tissues, which was also shared by the B.1.1.7 virus. CONCLUSIONS: In our analyses, we highlighted the active role of these anatomic sites in coronavirus disease 2019.

Safety and effectiveness in explantation and re-implantation of hypoglossal nerve stimulation devices.

PURPOSE: Since 2001, hypoglossal nerve stimulators have been implanted in patients with obstructive sleep apnea around the world, initially in trial situations but more recently also in regular care settings. Medium term data indicate effectiveness and tolerability of treatment. However, when assessing the safety of the procedure, the safe feasibility of explantation or reimplantation must also be considered. PATIENTS AND METHODS: Nine patients with an implanted respiratory-driven hypoglossal nerve stimulator. We have evaluated the feasibility and safety of explantation or re-implantation with another stimulation system. RESULTS: In 2012, nine patients were implanted with a respiratory-driven hypoglossal nerve stimulator as part of the Apnex Medical Pivotal Study. The study was ended in 2013. For a variety of reasons, the system was explanted from all nine patients by the year 2019. Three of these patients were re-implanted with a different system with respiratory sensing during the same session (mean incision to closure time for explantation 88.2 ± 35.01 min., mean incision to closure time for re-implantation 221.75 ± 52.73 min.). Due to extensive scar tissue formation, all procedures were technically challenging. Complication rate was significantly higher when re-implantation was performed or attempted in the same surgical session (0 of 5 patients with explantation versus 3 of 4 patients with attempted re-implantation; p = 0.018). There was no significant difference between the AHI values before and after implantation in patients with re-implantation. CONCLUSION: Explantation and re-implantation are technically challenging though possible procedures. The single-staged equilateral reimplantation of another hypoglossal nerve stimulation system can, but need not, be successful.

Incidence and survival of HNSCC patients living with HIV compared with HIV-negative HNSCC patients.

PURPOSE: The aim was to analyze the incidence and survival of patients living with HIV (PLWH) with head and neck squamous cell carcinoma (HNSCC) and to compare with a control group of HIV-negative HNSCC patients. METHODS: Clinicopathological data and predictors for overall survival (OS) and disease-free survival (DFS) were investigated (2009-2019). RESULTS: 50 of 5151 HNSCC patients (0.97%) were PLWH, and 76% were smokers. Age ≤ 60 years, HIV-PCR ≤ 50 copies, CD4 cells ≤ 200/mm3, cART treatment, T and UICC classification, oral cavity and nasal/paranasal sinuses, and therapy were significantly associated with OS in univariate analysis. In the multivariate analysis, only age and HIV-PCR independently predicted OS. The OS of the 50 PLWH was not significantly altered compared with the 5101 HIV-negative controls. However, OS and DFS were significantly inferior in advanced tumor stages of PLWH compared with an age-matched control group of 150 HIV-negative patients. CONCLUSIONS: PLWH were diagnosed with HNSCC at a significantly younger age compared to HIV-negative patients. Taking into account patient age at initial diagnosis, both OS and DFS rates in PLWH are significantly worse compared with a matched control group of HIV-negative patients in advanced tumor stages UICC III/IV. The prognosis (OS) is improved when taking cART treatment, the HIV viral load is undetectable and CD4 count is high.

Effects of surgical treatment of hypertrophic turbinates on the nasal obstruction and the quality of life.

PURPOSE: Chronic hyperplasia of the inferior nasal concha is accompanied by a nasal obstruction; however, there is no standardised surgical treatment for this condition. Here, we compared the outcome of three surgical techniques frequently used to treat the hyperplasia of inferior turbinates: turbinectomy with lateralization, submucosal electrocautery and laser cautery additional to septoplasty. MATERIALS AND METHODS: One hundred and nine patients participated in this prospective randomized study upon signing written consent. The subjects were randomly assigned to one of three intervention groups: 1) submucosal turbinectomy with lateralization, 2) submucosal electrocautery or 3) laser cautery. All groups were followed-up for up to 6months after surgical intervention. During the four follow-up appointments, the outcomes were measured with the modified German version of Sino-Nasal Outcome Test 20 questionnaire. In addition, the nasal breathing and the absolute nasal flow rates and respective mucosal component were determined by the anterior rhinomanometry. RESULTS: Following surgery, the subjective and objective nasal obstruction decreased significantly in all three groups. Moreover, the subjective symptoms measured by modified Sino-Nasal Outcome Test 20 improved significantly, although there were some temporal differences between groups regarding subjective nasal obstruction, ear pressure, nasal discomfort, daytime fatigue, cough and dry mouth. The mucosal component of nasal congestion decreased significantly after surgery. CONCLUSIONS: All surgical techniques used to reduce the conchae mucosa led to a significant improvement in the objective and subjective nasal breathing and the quality of life. Septoplastic reduction proved to be of additional benefit.

Nodal yield of neck dissections and influence on outcome in laryngectomized patients.

The appropriate extent of neck dissection (ND) in head and neck squamous cell carcinoma (SCC) continues to be investigated. This study aimed to determine whether the extent of ND as measured by nodal yield (NY) is a prognostic factor in patients undergoing primary total laryngectomy (TL) for advanced laryngeal SCC. A retrospective review at a tertiary referral centre identified 54 patients who underwent TL with elective (n = 39, cN0) or therapeutic (n = 15, cN+) ND with curative intent between 2002 and 2014. Survival analysis was obtained via regression analysis, calculated for overall (OS) and disease-free survival (DFS). Stage-independent 5-year OS was 32.1 %. Mean NY was 18.67 (standard deviation 9.898; range 0-45). The rate of cervical lymph node metastasis on pathology (pN+) was 27.8 %. Uni- and multivariate regression analysis detected no association between NY and OS or DFS (P ≥ 0.05). Advantages in OS and DFS were seen among patients without cervical lymph node metastasis (pN0) in comparison to those with pN+ necks (OS: hazard ratio [HR] 0.179; 95 % confidence interval [CI] 0.046-0.693; P = 0.013, and DFS: HR 0.190; 95 % CI 0.061-0.590; P = 0.004). ND resulted in pathologic upstaging of the neck in seven patients and downgrading in eight; increasing NY was not associated with an increased probability of a change in nodal stage. No significant association was found between NY and OS or DFS in patients undergoing TL with ND as primary therapy for advanced laryngeal SCC.

Polydimethylsiloxane for injection laryngoplasty: two cases necessitating tracheotomy.

The surgical treatment of glottic insufficiency due to lesions of the recurrent laryngeal nerve has become a routine procedure in the last few decades. In particular, injection laryngoplasty with polydimethylsiloxane (PDMS) has proved to be an easy, effective and safe method for vocal fold medialization. It is a biologically inert substance having almost ideal properties as a filler; complications related to its intralaryngeal use such as migration, or granuloma formation are extremely rare and allergic reactions have not been reported as yet. We discuss two cases representing the first description of acute severe complications after injection laryngoplasty with PDMS.

Laryngeal injuries following endotracheal intubation in ENT surgery: predictive value of anatomical scores.

Endotracheal intubation has been associated with a threefold higher incidence of laryngopharyngeal complaints following anesthesia in comparison to laryngeal mask airway. Such complaints, including hoarseness and sore throat, have been reported in up to 90% of patients within 24 h of extubation. The purpose of this study was to determine which preoperatively documented clinical and anatomic parameters are predictive of laryngo-pharyngeal trauma resulting from elective endotracheal intubation. Fifty-three patients undergoing ENT procedures requiring general anesthesia with endotracheal intubation were recruited. Pre and postoperative laryngostroboscopic examination was performed and findings correlated to preoperative clinical and anatomic parameters. Readily assessed anatomic parameters including height (>180 cm) and weight (>80 kg) correlated significantly to the Eckerbom grade of intubation-associated acute laryngeal injury (rs = 0.374; p = 0.006 and rs = 0.278; p = 0.044, respectively). The mandibular protrusion test also correlated significantly to the Eckerbom grade (rs = 0.462, p = 0.001) while the upper-lip-bite test showed significant correlation to impaired vocal fold oscillation (rs = 0.288, p = 0.036), with injury prediction sensitivities of 37.5 and 39.4%, respectively. No parameters correlated to subjective complaints (n = 5, 9.2%). This study provides suggestions on how to improve the classification of intubation-associated laryngeal injuries as well as providing the basis for larger clinical trials in other surgical subspecialties.

The Diagnostic Yield of Excisional Biopsy in Cervical Lymphadenopathy: A Retrospective Analysis of 158 Biopsies in Adults.

OBJECTIVES: Cervical lymph nodes are the most common site of peripheral lymphadenopathy. The underlying etiologies are usually benign and self-limiting but may include malignancies or other severe life-threatening diseases. The aim of the current study was to investigate the various underlying pathologies of cervical lymphadenopathy as assessed by the diagnostic yield of excisional lymph node biopsies of the neck in a tertiary adult practice. The evaluation was performed in light of previous literature and regional epidemiological patterns. METHODS: Retrospective analysis of hospital charts of 158 adult patients who underwent an excisional biopsy for suspected cervical lymphadenopathy at a tertiary referral head and neck service between January 2017 and December 2019. RESULTS: The most common underlying pathology was unspecific and/or reactive lymphadenitis in 44.5% of specimens, followed by malignant disease in 38.6% of cases. An age above 40 years was significantly correlated with an increased likelihood of malignant disease. Lower jugular and posterior triangle lymph nodes showed higher malignancy rates than other groups (100% and 66.7%, respectively). The overall surgical complication rate was 2.5%. CONCLUSIONS: The results of the current study serve as an indicator of the variety of etiologies causing cervical lymphadenopathy. In particular, given the increasing incidence of malignant diseases in recent decades, the findings should alert physicians to the importance of lymph node biopsy for excluding malignancy in persistent cervical lymphadenopathy especially in older adults. The findings emphasize the value of excisional lymph node biopsy of the neck as a useful diagnostic tool in adult patients with peripheral lymphadenopathy.

Clinical Applicability of Ultrasound Shear Wave Elastography in Patients under Hypoglossal Nerve Stimulation Therapy.

Relationship between stiffness of genioglossi (GG) and geniohyoidei (GH) muscles under electric hypoglossal nerve stimulation therapy (HNS) in relation to success of therapy was investigated with additional special focus on tongue movement. PATIENTS AND METHODS: Clinical and sleep laboratory parameters of a cohort of 18 patients with known shear wave velocity (SWV) data of the ipsilateral and contralateral musculi GG and GH (sGG, sGH and nGG, nGH) before and under HNS therapy were analyzed. The SWV was already determined using the ultrasonic shear wave elastography (US-SWE) technique. RESULTS: Median Epworth Sleepiness Scale (ESS) was 8 (IQR 12), median baseline Apnoe-Hypopnoe Index (AHI) 31.65 (IQR 25.1), median AHI under HNS therapy 16.3 (IQR 20.03). Therapy success: 9/18 patients (AHI during therapy < 15/h). There was no significant difference in SWV (sGG, sGH, nGG and nGH) between therapy responders and non-responders during therapy. Also, no difference could be seen with respect to the difference and increase in SWV values without and with stimulation. Examination of SWV values (sGG, sGH, nGG, nGH during stimulation, difference of SWV values stimulation - no stimulation, increase factor of SWV) revealed a significant negative correlation between the AHI under therapy and the measured SWV of the musculus GH of the contralateral side during stimulation (-0.622, p = 0.006). Patients with bilateral protrusion of the tongue differed regarding to therapy success in increase in SWV in sGG (p = 0.032). Tongue protrusion to contralateral: A significant difference between patients with AHI during therapy < 15/h in SWV values at sGG without stimulation (p = 0.021) was seen, with also a correlation to the current AHI under therapy (p = 0.047) and the change factor of the AHI (p = 0.015). CONCLUSION: Stiffness of the target muscle does not appear to be an isolated measure of the success of HNS therapy. This observation may have implications for future decision-making processes in the process of titrating electrical therapy parameters. But the technique of US-SWE may be useful for future research of the neurophysiology of the tongue and OSA phenotyping.

The Diagnostic Performance of Multiparametric Ultrasound in the Qualitative Assessment of Inconclusive Cervical Lymph Nodes.

BACKGROUND: Enlarged cervical lymph nodes (CLNs) can result from infection or malignancies, and a definitive diagnosis requires histological examination. Ultrasound (US) remains the first-line imaging modality for detection, and new US techniques may improve characterization. The aim of our study was to investigate whether the qualitative assessment of multiparametric US (mpUS) can improve diagnostic performance in the differentiation of benign and malignant CLNs. METHODS: 107 CLNs in 105 patients were examined by preoperative mpUS consisting of B-mode US, color-coded duplex sonography (CCDS), shear wave elastography (SWE) and contrast-enhanced US (CEUS). US images were evaluated in consensus by two experienced US operators. Histopathological examination was used as reference standard. RESULTS: SWE and CEUS combined showed the highest overall diagnostic performance (91% sensitivity, 77% specificity, 87% positive predictive value (PPV), 83% negative predictive value (NPV), 90% accuracy, χ2 (1) = 51.485, p < 0.001) compared to B-mode US and CCDS (87% sensitivity, 44% specificity, 73% PPV, 65% NPV, 73% accuracy χ2 (1) = 12.415, p < 0.001). In terms of individual techniques, SWE had higher specificity than B-mode and CCDS (71% sensitivity, 90% specificity, 92% PPV, 64% NPV, 78% accuracy, χ2 (1) = 36.115, p < 0.001), while qualitative CEUS showed the best diagnostic performance of all investigated US techniques (93% sensitivity, 85% specificity, 91% PPV, 87% NPV, 90% accuracy, χ2 (1) = 13.219, p < 0.001). Perfusion patterns, homogeneity, presence of necrosis, and malignancy differed significantly between malignant and benign CLNs (p < 0.001). CONCLUSIONS: SWE and CEUS can facilitate the differentiation of inconclusive CLNs when performed to supplement B-mode US and CCDS. MpUS may thus aid the decision between surgery and a watch-and-scan strategy in enlarged CLNs.

Subgroup Analysis of Overall Survival among Smoking and Non-Smoking Elderly Patients with HNSCC.

Smoking is a leading cause of head and neck squamous cell carcinoma (HNSCC). However, non-smokers are also affected by HNSCC, and the prognostic factors applicable to older non-smokers with HNSCC are largely unknown. The aim of this study was to determine predictors of overall survival (OS) in patients both with and without a smoking history aged 70 and over at initial diagnosis. Retrospective data of patients aged ≥70 (initial diagnoses 2004-2018) were examined. Evaluated predictors included tumour stage, biological age, health and therapy. A total of 688 patients (520 smokers, 168 non-smokers) were included with a median age of 74. The 5-year OS was 39.6%. Non-smokers had significantly improved OS compared to smokers (52.0% versus 36.0%, p < 0.001). Disease-free survival (DFS) differed significantly between both groups (hazard ratio = 1.3; 95%CI 1.04-1.626). TNM stage and the recommended therapies (curative versus palliative) were comparable. The proportion of p16-positive oropharyngeal carcinomas was significantly higher in non-smokers (76.7% versus 43.8%, p < 0.001). Smokers were significantly more likely to be men (p < 0.001), drinkers (p < 0.001), and have poorer health status (Karnofsky performance status, KPS, p = 0.023). They were also more likely to have additional tumours (p = 0.012) and lower treatment adherence (p = 0.038). Important predictors of OS identified in both groups, were, among others, alcohol abuse, KPS, Charlson comorbidity index, site of primary tumour, UICC stage and treatment received. Elderly non-smokers are also affected by HNSCC, however, both OS and DFS are increased compared to smokers.