Cancer stem cells in laryngeal cancer: what we know.

Cancer stem cells (CSCs) are a group of cells with eternal life or infinite self-renewal ability, which have high migrating, infiltrative, and metastatic abilities. Though CSCs only account for a small proportion in tumors, the high resistance to traditional therapy exempts them from therapy killing and thus they can reconstruct tumors. Previous studies found that heterogeneity of cancer cells extensively exists in head and neck cancers. Our current knowledge, about CSCs in the laryngeal cancer (LC), largely depends on head and neck studies. Given the lack of systematic data about CSCs in LC, we propose a review of the literature, reporting the evidences of CSCs in tumorigenesis of LC, with the purpose to provide new insights into the prevention and/or treatment of laryngeal carcinoma.

Severe obstructive sleep apnoea syndrome and erectile dysfunction: a prospective randomised study to compare sildenafil vs. nasal continuous positive airway pressure.

BACKGROUND: A high incidence of erectile dysfunction (ED) among patients with obstructive sleep apnoea syndrome (OSAS) has been reported, with a strong correlation between obstructive sleep apnoea, ED, and quality of life (QOL), and it has been estimated that 10-60% of patients with OSAS suffer from ED. In this prospective randomised controlled trial, we investigated 82 men with ED consecutively who were referred to the outpatient clinic for sleep disorders and had severe OSAS (AHI> 30 events/h) without any other comorbidities as a possible cause of ED. The aim of this study was to evaluate and compare the efficacy of sildenafil vs. continuous positive airway pressure (CPAP) in men with ED and severe OSAS. METHODS: Eighty-two patients were randomised to two main treatment groups: group 1 patients (n = 41) were treated with 100-mg sildenafil 1 h before sexual intercourse without CPAP, and group 2 patients (n = 41 men) were treated with only nasal CPAP during night time sleep. Both groups were evaluated with the same questionnaires (International Index of Erectile Function-EF domain; Sex Encounter Profile; Erectile Dysfunction Inventory Treatment Satisfaction) 12 weeks after treatment. RESULTS: In patients receiving sildenafil treatment, 58.2% of those who attempted sexual intercourses were successful compared to 30.4% in the CPAP group. The mean number of successful attempts per week was significantly higher in the sildenafil group compared with the CPAP group (2.9 vs. 1.7, respectively; p < 0.0001). The mean IIEF-EF domain scores were significantly higher in the sildenafil group compared with the CPAP group (p < 0.0001). The overall satisfaction rate was 68% with sildenafil treatment and 29% with CPAP treatment. CONCLUSIONS: This study confirms that severe OSAS is strongly associated with erectile dysfunction. CPAP and sildenafil (100 mg) are safe and effective therapies for OSAS-related ED patients. In the present study sildenafil was more effective than CPAP in treating ED associated with OSAS, as indicated by a significantly higher rate of successful attempts at intercourse and higher IIEF-EF domain scores. Our study, to date, is the only that has investigated sildenafil in patients with severe OSAS.

Technical note: development and validation of a method using ultra performance liquid chromatography coupled with tandem mass spectrometry for determination of vitamin B12 concentrations in milk and dairy products.

A method using ultra performance liquid chromatography coupled with tandem mass spectrometry was developed to measure cobalamins in naturally enriched raw milk and to evaluate their fate during thermal treatments and along the process of cheese making. After addition of methotrexate as internal standard, samples were submitted to heat treatment in the presence of cyanide, which converts all the less-stable cobalamins into cyanocobalamin; then, purification was performed by a solid-phase extraction step. Reverse-phase ultra performance liquid chromatography separation coupled with tandem mass spectrometry provided a fast and reliable determination. Mass spectrometric analysis was carried out in multiple reaction monitoring mode. The monitored transitions were m/z 678.36 → 147.10 and 678.36 → 359.30 for vitamin B12 and m/z 455.22 → 175.13 and 455.22 → 308.22 for methotrexate (internal standard). The limit of quantification was 2 ng/g. The method showed good linearity from 2 to 20 ng/g (R(2) ≥ 0.98) and intra- and interday precisions were always less than 19%.

Carotid stenosis after adjuvant cervical radiotherapy in patients with head and neck cancers: a prospective controlled study.

OBJECTIVES: To determine the prevalence of radiation-induced carotid stenosis, in patients who were treated for head and neck malignancies, using colour-flow duplex scanning. DESIGN: Prospective controlled study at a single medical centre. PARTICIPANTS: We enrolled two groups of patients. The first (radiotherapy group) consisted of patients who received surgical treatment and adjuvant radiotherapy of the neck. The control group consisted of patients with head and neck malignancies who received only surgical treatment. MAIN OUTCOME MEASURES: All patients were evaluated with carotid artery ecoDoppler imaging 1 week before and 36 months after the surgical procedure. Intima-media thickness was measured bilaterally at the internal carotid artery and at the bifurcation. Carotid obstruction was classified as low (0-30%), moderate (31-49%) or severe (≥50%). RESULTS: The preoperative stenosis grade did not differ between groups. In 15/25 patients (60%) in the radiotherapy group, mild stenosis evolved to moderate stenosis, while only 6/37 (16%) of the controls did (P = 0.004). Additionally, 9/39 (23%) patients in the radiotherapy group progressed to severe stenosis compared with only 3/54 (6%) controls (P = 0.029). The overall evolution showed that stenosis worsened in 24/32 (62%) patients in the radiotherapy group and 9/54 (17%) patients in the control groups (P < 0.0001). CONCLUSIONS: These results highlight the need to study the long-term incidence of cerebrovascular events in these two different populations (radiation treated and surgically treated) to identify increased cerebrovascular morbidity.

Autologous serum skin test reactivity and basophil histamine release test in patients with nasal polyposis: preliminary results.

An eosinophilic inflammatory process is generally observed in patients suffering from nasal polyposis (NP), however its onset has not yet been defined. It has been suggested that immune activation of inflammatory cells may be the cause. The aim of this study is to verify whether autoantibodies and/or histamine-releasing factors are present in the serum of patients suffering from NP. In fact, we assume that autoantibodies and/or histamine-releasing factors, as already demonstrated in chronic idiopathic urticaria and asthma, may be involved in the pathogenesis of NP. In this case-control analytical study 40 patients with NP and 27 control subjects underwent the in vivo autologous serum skin test (ASST). The sera from 6 patients suffering from NP and 9 control group subjects, who had all been previously studied and randomly selected, underwent basophil histamine release assay from normal donor as a pilot study. The ASST showed positive results in 55% of patients suffering from NP versus 8% of the control group (p= .00006), the basophil histamine release test (BHRT) turned out positive in all patients tested and in 11% of the control group. We found a weak positive correlation between the percentage of histamine release and the wheal diameter. ASST reactivity is very frequent in patients suffering from NP, thus suggesting the presence of histamine-releasing factors in the blood stream. The BHRT was positive in the serum of all patients, thus suggesting the presence of anti-FcepsilonRI, anti-IgE autoantibodies and/or other histamine-releasing factors, the presence of which can play a role in triggering and maintaining the eosinophilic inflammatory process in NP.

Effect of exposure to benzo[a]pyrene on SODs, CYP1A1/1A2- and CYP2E1 immunopositive proteins in the blood clam Scapharca inaequivalvis.

The effects of water-borne exposure to benzo[a]pyrene (36 h; celite-bound 0.44 mg L(-1) B[a]P) on cytochrome P450 (CYP) and superoxide dismutases (SODs) were examined in digestive gland of the blood clam, Scapharca inaequivalvis. B[a]P accumulation and elimination were rapid, with maximum whole-body concentrations of 1.78 ng g(-1) wet wt after 12 h of treatment, followed by a progressive decline to 0.89 ng g(-1) at 36 h. The presence of B[a]P resulted in an increase in total CYP of digestive gland microsomes from 54+/-14 to 108+/-21 pmol/mg protein (mean+/-SD; p<0.05, 24 h). Increases were also seen in microsomal CYP1A1/1A2-immunopositive protein (50.5 kDa app. mol. wt; p<0.05), but not CYP2E1-immunopositive protein (49 kDa app. mol. wt.), indicating a specific response of the former isoform. Exposure to B[a]P produced a steady increase in Mn-SOD digestive gland activity (p<0.01; p<0.05) but no significant change in Cu/Zn-SOD activity. The respective proteins, measured by western blotting, were not significant induced after B[a]P exposure. Cu/Zn-SOD and Mn-SOD activities were correlated with total CYP levels (r=0.96 and 0.63, respectively), indicating a role for CYP in reactive oxygen species (ROS) production during exposure. Both 'NADPH-independent' and NADPH-dependent metabolism of B[a]P by digestive gland microsomes was seen, producing mainly 1,6-, 3,6- and 6,12-diones, with some phenols and 7,8-dihydrodiol; putative protein adducts were also formed. Redox cycling of the diones may also have contributed to ROS production, leading to the increased SOD activities.

Sialendoscopic management of autoimmune sialadenitis: a review of literature.

Autoimmune diseases of major salivary glands include Sjögren's syndrome and a complex of disorders classified as immunoglobulin G4-related diseases. These pathologies are characterised by an autoimmune reaction mediated by T-helper lymphocytes that targets the ducts of exocrine glands in Sjögren's syndrome and glandular parenchyma in immunoglobulin G4-related diseases. Immunoglobulin G4-related diseases represent recently introduced multi-organ diseases that also involve the salivary glands. However, the morbid conditions once known as Mikulicz's disease and Kuttner's tumour were recently considered as two variants of immunoglobulin G4-related diseases affecting the major salivary glands ( immunoglobulin G4-related sialadenitis). This review briefly summarises the pathogenesis and clinical features of autoimmune diseases of the major salivary glands, focusing on the diagnostic and therapeutic role of sialendoscopy.

Salvage total laryngectomy after conservation laryngeal surgery for recurrent laryngeal squamous cell carcinoma.

The aim of the present study was to evaluate the oncological efficacy of salvage total laryngectomy in patients who had previously undergone supracricoid partial laryngectomy or transoral laser microsurgery for treatment of laryngeal squamous cell carcinoma. We retrospectively reviewed the medical, surgical and pathological records of 35 patients who underwent salvage total laryngectomy after recurrence of laryngeal cancer (following supracricoid partial laryngectomy or transoral laser microsurgery). Kaplan-Meier survival curves as well as univariate and multivariate analyses of prognostic factors were performed. No statistically significant differences were seen comparing the supracricoid partial laryngectomy group with the transoral laser microsurgery group for overall survival and disease-specific survival at 3 years (OS = 38% vs. 52%, p = 0.16; DSS = 40% vs. 61%, p = 0.057) or locoregional control at 2 years (LRC = 40% vs. 54%, p = 0.056). A trend indicating worse survival and locoregional control for supracricoid partial laryngectomy patients emerged. Preservation of the osteocartilaginous frame in transoral laser microsurgery could hypothetically result in better salvageability of anterior recurrences with extralaryngeal spread.

Outcomes of interventional sialendoscopy for obstructive salivary gland disorders: an Italian multicentre study.

Interventional sialendoscopy has become the predominant therapeutic procedure for the management of obstructive salivary disorders, but only a few multicentre studies of large series of patients with a long-term follow-up have been published. This Italian multicentre study involved 1152 patients (553 females; mean age 50 years) who, after at least a clinical and ultrasonographic evaluation, underwent a total of 1342 diagnostic and interventional sialendoscopies, 44.6% of which involved the parotid gland. 12% (n = 138) of patients underwent multiple treatments. The procedure was successful in 1309 cases. In 33 cases (2.4%) the procedure could not be concluded mainly because of complete duct stenosis (21 cases). Salivary stones were the main cause of obstruction (55%), followed by ductal stenosis and anomalies (16%), mucous plugs (14.5%) and sialodochitis (4.7%). Complete therapeutic success was obtained in 92.5% of patients after one or more procedures, and was ineffective in < 8%. Untoward effects (peri and postoperative complications) were observed in 5.4% of cases. Sialendoscopy proved to be an effective, valid and safe procedure in the diagnostic and therapeutic management of non-neoplastic obstructive salivary gland diseases.

Extended parotidectomy.

Malignant tumours of the parotid gland represent a group of relatively rare lesions. The medical records of 363 patients with parotid swelling treated between 1974 and 2003 at the "G. Ferreri" Department of Otorhinolaryngology, "La Sapienza" University in Rome were retrospectively analysed. Clinical presentation, pre-operative investigations, surgical procedure, histopathology report, post-operative complications, and the oncological results of 19 patients who underwent extended radical parotidectomy for malignant neoplasm of the parotid gland are discussed. Extended radical parotidectomy, reserved for neoplasms in an advanced stage, involves the removal of the entire parotid gland, with sacrifice of the facial nerve and the resection en bloc of the adjacent structures affected by neoplastic infiltration, such as the temporal bone, the mandibular bone, the skin, blood vessels and nerves. In addition to this surgical treatment, a cycle of adjuvant radiotherapy is also necessary. The overall rate of survival at 10 years depends mainly on the histological characteristics of the tumour, and, in this series, is reported to be approximately 58%. These data indicate that total extended radical parotidectomy combined with post-operative radiotherapy, represents the best therapeutic approach with regard both to quality of life and life expectancy, in patients with an advanced stage of malignant neoplasm of the parotid gland.

New High-performance Liquid Chromatography-DAD Method for Analytical Determination of Arbutin and Hydroquinone in Rat Plasma.

Natural substances present in herbal preparations should be carefully used because they can give toxic or therapeutic effects despite of their amount or the way of administration. The safety of products of vegetable origin must be assessed before commercialisation by monitoring the active ingredients and their metabolites. This study was therefore designed to identify and quantify arbutin and its metabolite hydroquinone, naturally present in Arctostaphylos uva-ursi (L.) Spreng plant in rat plasma, after an acute and subacute administration of aqueous arbutin solution in Wistar rats. For this purpose a reversed-phase high-performance liquid chromatography coupled with photodiode array detection was developed to assess the pharmacokinetic of arbutin and hydroquinone in plasma of female rats treated with aqueous arbutin solutions. The detection (arbutin: 0.0617 µg/ml and hydroquinone 0.0120 µg/ml) and quantification (arbutin: 0.2060 µg/ml and hydroquinone: 0.0400 µg/ml) limits were determined. At the arbutin concentration level of 10.7 µg/ml repeatability was 13.33% and its recovery 93.4±6.93%, while at the hydroquinone concentration level of 10.6 µg/ml repeatability was 11.66% and its recovery 92.9±7.75%. Furthermore the method was fully validated and the obtained data indicate that the new method provides good performances.

Systematic isolation and identification of membrane lipid oxidation products.

This report discusses the analytical procedure by which it is possible to isolate and identify the oxidation products of cellular and subcellular membrane lipids. The key point of this procedure is the method used for the transmethylation of the lipid material isolated from the tissues. In effect, both the conversion of the glycerides into methyl esters and the reduction of the hydroperoxyl groups into the corresponding hydroxyl groups is performed in one step, without breaking any oxirane rings that may be present. The methyl esters containing functional groups introduced by oxidative processes are separated from the non-modified ones by preparative TLC and are identified by GLC and GC-MS.

Lipid peroxidation in cancer cells: chemical and physical studies.

Our studies on the biochemical composition and the structural organization of smooth and rough endoplasmic reticulum isolated from Morris hepatomas 9618A and 3924A confirm the results obtained employing the total microsomal fraction. We have definitely established the following facts: (1) Tumor subcellular organelles exhibit the very low degree of peroxidizability that has been shown to be related to the growth rate of the tumor. (2) Associated with such a low susceptibility to peroxidation are (a) changed lipid composition of cellular membranes, whose content in polyunsaturated fatty acid is markedly decreased, and (b) changed static and dynamic properties of the membrane. Previously it was also found that cellular oxy-radical scavenging enzymes are markedly reduced. From these data, it is possible to infer that tumor membranes are altered structurally and functionally in part as the result of an oxy-radical-induced damage that occurs in vivo under conditions of oxygen toxicity. This seems to be supported by recent findings that the spontaneous increase in growth rate of the originally very slow-growing Morris hepatoma 9618A results also in the loss of cytochrome P-450 (an important intramembraneous propagator of lipid peroxidation) as well as of C20:4 and C22:6. Studies performed by GLC and GC-MS on the fatty acid residues of phospholipids of rat liver microsomes show the presence of C20:3-OH and C18:1-OH, but no hydroxyl derivatives of low molecular weight aldehydes. The hydroxyl derivatives of arachidonic acid and linoleic acid are present in much smaller amounts in the microsomes isolated from H9618A and H3924A.

Synthesis of highly pure dolichyl and cholesteryl esters.

Highly pure dolichyl and/or cholesteryl esters can be synthesised at room temperature by reacting a defined pure dolichol and/or pure cholesterol with an acyl chloride in the presence of pyridine. During the reaction some acyl anhydride originates as a secondary product. It can be hydrolysed by methanolic KOH and removed by a rapid percolation of the reaction mixture through a small alumina column. A final purification of the raw product obtained by silicic-acid/celite column chromatography makes it possible to obtain highly pure esters with a satisfactory yield.

A quick LC-MS-MS method for the determination of flunixin in bovine muscle.

A simple, fast and cost-effective liquid chromatographic/tandem mass spectrometry (LC-MS-MS) method for the quantitative determination of flunixin (FLU) in bovine muscle was developed and validated. The sample preparation procedure involved an extraction with acetonitrile, followed by evaporation and reconstitution. Chromatographic separation was achieved on a reverse-phase column under programmed conditions. FLU detection was performed with positive electrospray ionization in selected reaction monitoringmode, monitoring one precursor and two products ions. For quantification purposes, FLU-d3 was used as an internal standard. The matrix effect on the analysis of FLU in bovine muscle was evaluated by comparison between calibration curves prepared with standard solution and in blank matrix extracts. The equivalent responses obtained confirmed the absence of signal suppression or/and enhancement. The method was extensively validated according to the parameters requested by European Commission Decision 2002/657/EC in terms of specificity, limit of detection, linearity, trueness, precision, decision limit (CCα) and detection capability (CCß). FLU stability was also investigated in matrix and in sample extracts at different times and storage conditions.

Recurrent respiratory papillomatosis by HPV: review of the literature and update on the use of cidofovir.

Recurrent respiratory papillomatosis is a viral induced disease characterised by exophytic epithelial lesions affecting the larynx. The problem with its treatment is the high recurrence of papilloma growth after surgical removal. The aim of our review is to analyse the actual use of cidofovir, an agent used in adjuvant therapy. We have reviewed 6 manuscripts that included a total of 118 patients. The parameters taken into account were: concentration of infiltrated cidofovir (mg/ml), therapeutic response, relapse-free time (months), side effects, genotypes (HPV-6/11/18) and evolution of dysplasia. Cidofovir was injected at concentrations from 2.5 to 15 mg/ml, therapeutic response was from 56.25% to 82.3% and relapse-free time was from 10.05 to 49 months. There were 2 cases of dysplasia during therapy. Ten patients had been infected by HPV-6, 4 patients by HPV-11 and 10 patients by HPV-6 and HPV-11. The purposes of our review include the following: to stress that the juvenile form is more aggressive than other forms, to demonstrate than the drug has good adjuvant action although it does not significantly change the final response to the disease, to show that side effects are modest and, finally, to disprove the hypothesis that cidofovir may promote evolution towards dysplasia. In conclusion, combination of surgical removal and injection of cidofovir is associated with good response in recurrent respiratory papillomatosis.

Is vestibular neuritis an immune related vestibular neuropathy inducing vertigo?

OBJECTIVES: To review the current knowledge of the aetiology of vestibular neuritis including viral infections, vascular occlusion, and immunomediated mechanisms and to discuss the pathogenesis with relevance to pharmacotherapy. SYSTEMATIC REVIEW METHODOLOGY: Relevant publications on the aetiology and treatment of vestibular neuritis from 1909 to 2013 were analysed. RESULTS AND CONCLUSIONS: Vestibular neuritis is the second most common cause of peripheral vestibular vertigo and is due to a sudden unilateral loss of vestibular function. Vestibular neuronitis is a disorder thought to represent the vestibular-nerve equivalent of sudden sensorineural hearing loss. Histopathological studies of patients who died from unrelated clinical problems have demonstrated degeneration of the superior vestibular nerve. The characteristic signs and symptoms include sudden and prolonged vertigo, the absence of auditory symptoms, and the absence of other neurological symptoms. The aetiology and pathogenesis of the condition remain unknown. Proposed theories of causation include viral infections, vascular occlusion, and immunomediated mechanisms. The management of vestibular neuritis involves symptomatic treatment with antivertiginous drugs, causal treatment with corticosteroids, and physical therapy. Antiviral agents did not improve the outcomes.

Laryngotracheal stenosis treated with multiple surgeries: experience, results and prognostic factors in 70 patients.

Laryngotracheal stenosis is a complex condition that usually requires multiple procedures to restore physiological respiration. The aim of this study was to evaluate the percentage of decannulation compared to different or multiple surgical treatments. We retrospectively reviewed the charts of 70 patients treated between 1990 and 2005 for laryngotracheal stenosis of various aetiology: iatrogenic stenosis (n = 55), post-traumatic stenosis (n = 11) or other causes (autoimmune disease, n = 3; diphtheria, n = 1). In order to maintain laryngotracheal patency, a Montgomery Safe-T tube was used in all patients as a single dilation treatment or associated with endoscopic and/or open-neck surgery. Fifty-four of the 70 patients (77.1%) were eventually decannulated; 39 of these (72.2%) underwent 3 or fewer surgical procedures, showing a significant difference compared to patients who underwent more than 3 surgeries (p = 0.00002). A total of 257 surgeries were performed. Only seven of 54 patients (13%) were decannulated after more than 5 surgical procedures. Patients over 60 years of age and with a higher grade of stenosis showed a significantly lower success rate (p = 0.0017 and p = 0.007, respectively). There was no significant correlation between the rate of decannulation and gender, aetiology, site of stenosis or surgery. Patients undergoing dilation for laryngotracheal stenosis usually require multiple procedures. The T tube plays an important role in the treatment of this pathology. However, if the tracheostomy is not removed within 3 surgical interventions, the odds of decannulating the patient decrease significantly, and additional surgeries may be of questionable therapeutic benefit.

Development and validation of a liquid chromatography/tandem mass spectrometry method for quantitative determination of amoxicillin in bovine muscle.

A simple, quick and economical liquid chromatographic/tandem mass spectrometry (LC-MS/MS) method for the quantitative determination of amoxicillin in bovine muscle was developed and validated. The sample preparation procedure involved a liquid extraction with water, followed by a protein precipitation step with acetonitrile. The extract was purified by a liquid-liquid partition with dichloromethane and the upper aqueous layer was directly injected into the LC-MS/MS system. Chromatographic separation was achieved on a reversed phase column, using a mixture of acetonitrile, water and 0.005% formic acid in water as mobile phase. Gradient elution was performed at a flow rate of 0.2 mL min⁻¹. Amoxicillin was detected using positive electrospray ionization in selected reaction monitoring (SRM) mode and was quantified using terbutaline as internal standard. The responses for standards prepared in solvent and in matrix were equivalent and additionally the absence of signal suppression was confirmed by the post column infusion technique. Amoxicillin stability in standard solution and in matrix was investigated at different times and storage conditions. Amoxicillin standards prepared in water were stable on storage up to 20 days at -20°C. Amoxicillin stability in matrix (spiked bovine muscle samples) was assessed up to 15 days at -20°C. The method was validated according to the parameters requested by European Commission Decision 2002/657/EC in terms of specificity, linearity, trueness, precision, decision limit (CCα) and detection capability (CCß). All the trueness values fell within a range between 14.5% and 6.3%. Precision values for all levels of concentration tested were lower than the relative limit calculated by the Horwitz equation. The amoxicillin MRL is set at 50 µg kg⁻¹ and the CCα and CCß of the method were 61.2 µg kg⁻¹ and 72.4 µg kg⁻¹, respectively.