Genotoxicity of nitroso compounds and sodium dichromate in a model combining organ cultures of human nasal epithelia and the comet assay

Genotoxic effects of xenobiotics are a possible step in tumor initiation in the mucosa of the upper aerodigestive tract. Using the comet assay, detecting genotoxicity in human tissue has been restricted to single incubations in vitro, but in vivo most xenobiotics harm their target in a repetitive or chronic manner. Therefore, we propose a model, which provides repetitive incubations in human upper aerodigestive tract mucosa cultures. Samples of human inferior nasal turbinate mucosa (n = 25) were cultured according to a modified version of a technique originally described by Steinsvag. On day 1 fresh samples and on days 7, 9 and 11 organ cultures were incubated with N-nitrosodiethylamine (NDEA), sodium dichromate (Na2Cr2O7) and N'-methyl-N-nitro-N-nitrosoguanidine(MNNG). Mucosa samples and organ cultures, respectively, underwent a modified comet assay on days 1, 7 and 11. Genotoxicity could be shown for NDEA, Na2Cr2O7 and MNNG on days 1, 7 and 11. Duration of tissue culture and repetitive incubations did not significantly influence the results for NDEA. Nevertheless, Na2Cr2O7 and MNNG caused higher genotoxic effects on cultures subjected to the comet assay on day 11. This model may help to assess genotoxic hazards posed by environ mental pollutants that have a cumulative character in repetitive or chronic exposure in vivo. Copyright (C) 2001 S. Karger AG, Basel

Primary cold atmospheric plasma combined with low dose cisplatin as a possible adjuvant combination therapy for HNSCC cells—an in-vitro study

Background The aim of the present study was to examine the cytostatic effects of cold atmospheric plasma (CAP) on different head and neck squamous carcinoma (HNSCC) cell lines either in isolation or in combination with low dose cisplatin. The effect of CAP treatment was investigated by using three different HNSCC cell lines (chemo-resistant Cal 27, chemo-sensitive FaDu and OSC 19). Materials and method Cell lines were exposed to CAP treatment for 30, 60, 90, 120 and 180 s (s). Cisplatin was added concurrently (cc) or 24 h after CAP application (cs). Cell viability, DNA damage and apoptosis was evaluated by dye exclusion, MTT, alkaline microgel electrophoresis assay and Annexin V-Fit-C/PI respectively. Results In all cell lines, 120 s of CAP exposure resulted in a significant reduction of cell viability. DNA damage significantly increased after 60 s. Combined treatment of cells with CAP and low dose cisplatin showed additive effects. A possible sensitivity to cisplatin could be restored in Cal 27 cells by CAP application. Conclusion CAP shows strong cytostatic effects in HNSCC cell lines that can be increased by concurrent cisplatin treatment, suggesting that CAP may enhance the therapeutic efficacy of low dose cisplatin

30-Day Postoperative Outcomes in Adults with Obstructive Sleep Apnea Undergoing Upper Airway Surgery

Background: Obstructive sleep apnea (OSA) is a chronic disorder of the upper airway. OSA surgery has oftentimes been researched based on the outcomes of single-institutional facilities. We retrospectively analyzed a multi-institutional national database to investigate the outcomes of OSA surgery and identify risk factors for complications. Methods: We reviewed the American College of Surgeons National Surgical Quality Improvement Program (NSQIP) database (2008–2020) to identify patients who underwent OSA surgery. The postoperative outcomes of interest included 30-day surgical and medical complications, reoperation, readmission, and mortality. Additionally, we assessed risk-associated factors for complications, including comorbidities and preoperative blood values. Results: The study population included 4662 patients. Obesity (n = 2909; 63%) and hypertension (n = 1435; 31%) were the most frequent comorbidities. While two (0.04%) deaths were reported within the 30-day postoperative period, the total complication rate was 6.3% (n = 292). Increased BMI (p = 0.01), male sex (p = 0.03), history of diabetes (p = 0.002), hypertension requiring treatment (p = 0.03), inpatient setting (p < 0.0001), and American Society of Anesthesiology (ASA) physical status classification scores ≥ 4 (p < 0.0001) were identified as risk-associated factors for any postoperative complications. Increased alkaline phosphatase (ALP) was identified as a risk-associated factor for the occurrence of any complications (p = 0.02) and medical complications (p = 0.001). Conclusions: OSA surgery outcomes were analyzed at the national level, with complications shown to depend on AP levels, male gender, extreme BMI, and diabetes mellitus. While OSA surgery has demonstrated an overall positive safety profile, the implementation of these novel risk-associated variables into the perioperative workflow may further enhance patient care

Curcumin and Other Polyphenolic Compounds in Head and Neck Cancer Chemoprevention

Despite clear results of observational studies linking a diet rich in fruits and vegetables to a decreased cancer risk, large interventional trials evaluating the impact of dietary micronutrient supplementation, mostly vitamins, could not show any beneficial effects. Today it has become clear that a single micronutrient, given in supernutritional doses, cannot match cancer preventive effects of whole fruits and vegetables. In this regard polyphenols came into focus, not only because of their antioxidant potential but also because of their ability to interact with molecular targets within the cells. Because polyphenols occur in many foods and beverages in high concentration and evidence for their anticancer activity is best for tissues they can come into direct contact with, field cancerization predestines upper aerodigestive tract epithelium for cancer chemoprevention by polyphenols. In this paper, we summarize cancer chemopreventive attempts with emphasis on head and neck carcinogenesis and discuss some methodological issues. We present data regarding antimutagenic effects of curcumin and epigallocatechin-3-gallate in human oropharyngeal mucosa cultures exposed to cigarette smoke condensate

Graphical modeling can be used to illustrate associations between variables describing functioning in head and neck cancer patients

Objective: To examine the associations between variables of functioning measured by the International Classification of Functioning, Disability and Health (ICF) in head and neck cancer (HNC) patients by means of graphical modeling.Study design and setting: Graphical modeling was used on a data set of a cross-sectional multicentric study of 145 patients with HNC. Functioning was qualified using the extended ICF checklist. Multiple imputation was used to handle missing data. The least absolute shrinkage and selection operator for generalized linear models was used to identify conditional associations between the ICF categories. Bootstrap aggregating was used to enhance the accuracy and validity of model selection.Results: The resulting graph shows largely meaningful associations between the ICF categories. One central point could be visualized consisting of a circular path of d330 Speaking, d350 Conversation, b510 Ingestion functions, s320 Structure of mouth, and b310 Voice functions. Another important structure in the graph were the bow-shaped associations beginning with d335 Producing nonverbal messages to b130 Energy and drive functions.Conclusion: Graphical modeling can be used to describe associations between different areas of functioning in HNC patients. They found associations can be the basis for improved rehabilitation and gives a deeper understanding of functioning in HNC patients

Cytotoxic and genotoxic effects of resin monomers in human salivary gland tissue and lymphocytes as assessed by the single cell microgel electrophoresis (Comet) assay

Malignant tumors of the three major pairs and the numerous minor salivary glands in humans are rare, and little is known about their various etiologies. Considering the fact that resin monomers from dental restorative materials are released into the saliva and diffuse into the tooth pulp or gingiva, mucosa, and salivary glands, this may potentially contribute to tumorigenesis. Resin monomers may also be reabsorbed and reach the circulating blood as well. Whereas the cytotoxic potential of some components has been clearly documented, data on genotoxicity in human target cells require further investigation. In the present study, genotoxic and cytotoxic effects of three common methacrylates are investigated in human samples of salivary glands and peripheral lymphocytes. The Comet assay was used to quantify DNA single strand breaks, alkali labile and incomplete excision repair sites in salivary gland probes and lymphocytes of 10 volunteers. The xenobiotics investigated were triethyleneglycoldimethacrylate (TEGDMA), urethanedimethacrylate (UDMA), and 2-hydroxyethylmethacrylate (HEMA), with N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) and dimethyl sulfoxide (DMSO) as controls. DNA migration was analyzed using the tail moment according to Olive (OTM). Cytotoxicity was monitored using trypan blue staining. With TEGDMA concentrations at 10(-5)m (10(-3)m), UDMA at 10(-7)m (10(-7)m), and HEMA at 10(-3)m (10(-5)m) significant enhancements of DNA migration were achieved in tissue cells (lymphocytes) as compared to the negative controls. At higher concentrations of up to 2.5x10(-2)m, induced DNA migration was expressed by OTM at 10.7 for TEGDMA in tissue cells (8.7 in lymphocytes), 10.5 for UDMA (6.4), and 9.7 for HEMA (6.1). The viability of the cell systems was not affected as concerns the threshold level for the assay of 75% viable cells except for the highest concentration tested for TEGDMA and UDMA in tissue cells. At higher concentration levels, all tested substances induced significant enhancement of DNA migration in the Comet assay as a possible sign for genotoxic effects in human salivary glands and lymphocytes. These data add to the results of prior studies in human peripheral lymphocytes and give evidence of a possible risk factor for tumor initiation in human salivary glands

Rapid and Non-Enzymatic <em>In Vitro</em> Retrieval of Tumour Cells from Surgical Specimens

<div><p>The study of tumourigenesis commonly involves the use of established cell lines or single cell suspensions of primary tumours. Standard methods for the generation of short-term tumour cell cultures include the disintegration of tissue based on enzymatic and mechanical stress. Here, we describe a simple and rapid method for the preparation of single cells from primary carcinomas, which is independent of enzymatic treatment and feeder cells. Tumour biopsies are processed to 1 mm³ cubes termed explants, which are cultured 1–3 days on agarose-coated well plates in specified medium. Through incisions generated in the explants, single cells are retrieved and collected from the culture supernatant and can be used for further analysis including <em>in vitro</em> and <em>in vivo</em> studies. Collected cells retain tumour-forming capacity in xenotransplantation assays, mimic the phenotype of the primary tumour, and facilitate the generation of cell lines.</p> </div

Single cells from head and neck specimens recapitulate primary tumours.

<p>(<b>A</b>) Biopsy of a hypopharynx carcinoma was processed to explants of approx. 1 mm³ and transferred into agarose-coated 24-well plates. Shown is the primary tumour biopsy and explants in wells. (<b>B</b>) Upon incision and cultivation in specified medium, cells migrate out of the explant and settle in the well. Shown is one representative example at day one. (<b>C</b>) Upper panels: Explants were collected after one day, cryo-preserved and processed to sequential sections before staining with EpCAM-, CD133-specific or an isotype control antibody as indicated. The structure of explants begins to disintegrate and cells migrate away from the major tumour area and are released into the supernatant. Cells express EpCAM but no CD133. Lower panels: Single cells migrating out of explants were subjected to an EpCAM-, CD133 or isotype control antibody staining. The majority of cells expressed EpCAM and only rarely expressed CD133. (<b>D</b>) Matched primary carcinoma was stained for the expression of EpCAM and Ki-67. Shown are 40× (upper panel) and 200× magnifications (middle and lower panels) of the indicated stainings. (<b>E</b>) Single cells from explants of primary specimen shown in (D) were stained in cytospins with EpCAM-specific (upper panel) or an isotype control antibody (lower panel). (<b>F</b>) Cell surface expression of EpCAM on single cells from explants of primary specimen shown in (D) was analysed upon flow cytometry with specific antibodies. Left dot plot represents isotype control, while right dot plot represents EpCAM staining. Dot plot statistics according to the marker set for both graphs are given for isotype control and EpCAM staining. (<b>G</b>) Single cells (1–5×10⁶) from explants of primary specimen shown in (D) were xenotransplanted subcutaneously in immunocompromised NOD-SCID mice. After 28 days tumours were surgically removed, cryo-preserved, and stained for the expression of EpCAM and Ki-67. Shown are 100× (upper panel) and 200× magnifications (middle and lower panels) of the indicated stainings.</p