Lea Calò

Trafficking and Postsecretory Events Responsible for the Formation of Secreted Human Salivary Peptides A Proteomics Approach

To elucidate the localization of post-translational modifications of different classes of human salivary proteins and peptides (acidic and basic proline-rich proteins (PRPs), Histatins, Statherin, P-B peptide, and “S type” Cystatins) a comparative reversed phase HPLC-ESI-MS analysis on intact proteins of enriched granule preparations from parotid and submandibular glands as well as parotid, submandibular/sublingual (Sm/Sl), and whole saliva was performed. The main results of this study indicate the following. (i) Phosphorylation of all salivary peptides, sulfation of Histatin 1, proteolytic cleavages of acidic and precursor basic PRPs occur before granule storage. (ii) In agreement with previous studies, basic PRPs are secreted by the parotid gland only, whereas all isoforms of acidic PRPs (aPRPs) are secreted by both parotid and Sm/Sl glands. (iii) Phosphorylation levels of aPRPs, Histatin 1, and Statherin are higher in the parotid gland, whereas the extent of cleavage of aPRP is higher in Sm/Sl glands. (iv) O-Sulfation of tyrosines of Histatin 1 is a post-translational modification specific for the submandibular gland. (v) The concentration of Histatin 3, Histatin 5, and Histatin 6, but not Histatin 1, is higher in parotid saliva. (vi) Histatin 3 is submitted to the first proteolytic cleavage (generating Histatins 6 and 5) during granule maturation, and it occurs to the same relative extent in both glands. (vii) The proteolytic cleavages of Histatin 5 and 6, generating a cascade of Histatin 3 fragments, take place after granule secretion and are more extensive in parotid secretion. (viii) Basic PRPs are cleaved in the oral cavity by unknown peptidases, generating various small proline-rich peptides. (ix) C-terminal removal from Statherin is more extensive in parotid saliva. (x) P-B peptide is secreted by both glands, and its relative quantity is higher in submandibular/sublingual secretion. (xi) In agreement with previous studies, S type Cystatins are mainly the product of Sm/Sl glands.

Damage to Ciliated Epithelium in Chronic Rhinosinusitis: What is the Role of Bacterial Biofilms?

Objectives: We assess the association between the presence of biofilms and cilial damage in patients with chronic rhinosinusitis (CRS), describe the microorganisms associated with samples that exhibited cilial loss and biofilms, and demonstrate the absence of ciliary injury and biofilms in similarly prepared “normal” controls. Methods: We examined samples of ethmoid mucosa obtained from 24 patients who underwent functional endoscopic sinus surgery for CRS. Samples from a control group (20 healthy subjects) were also examined. The specimens were divided into 2 fragments; the first was processed for bacterial cultures, and the second was subjected to scanning electron microscopy. Statistical analysis was performed. Results: All CRS samples had positive bacterial cultures. The scanning electron microscopy analysis showed bacterial biofilms in 10 of the 24 specimens. A marked destruction of the epithelium was observed in samples positive for biofilms (p < 0.001), and the presence of Haemophilus influenzae was associated with ciliary abnormalities (partial damage in 55.6% and absence of cilia in 50%; p = 0.041). Conclusions: The high percentage of biofilms in our specimens confirms the association between biofilms and CRS. Our data support the hypothesis that biofilm formation represents the latter phase of an inflammatory process that leads to complete epithelial destruction.

Chrono-Proteomics of Human Saliva: Variations of the Salivary Proteome during Human Development

An important contribution to the variability of any proteome is given by the time dimension that should be carefully considered to define physiological modifications. To this purpose, whole saliva proteome was investigated in a wide age range. Whole saliva was collected from 17 preterm newborns with a postconceptional age at birth of 178-217 days. In these subjects sample collection was performed serially starting immediately after birth and within about 1 year follow-up, gathering a total of 111 specimens. Furthermore, whole saliva was collected from 182 subjects aged between 0 and 17 years and from 23 adults aged between 27 and 57 years. The naturally occurring intact salivary proteome of the 316 samples was analyzed by low- and high-resolution HPLC-ESI-MS platforms. Proteins peculiar of the adults appeared in saliva with different time courses during human development. Acidic proline-rich proteins encoded by PRH2 locus and glycosylated basic proline-rich proteins encoded by PRB3 locus appeared following 180 days of postconceptional age, followed at 7 months (±2 weeks) by histatin 1, statherin, and P-B peptide. The other histatins and acidic proline-rich proteins encoded by PRH1 locus appeared in whole saliva of babies from 1 to 3 weeks after the normal term of delivery, S-type cystatins appeared at 1 year (±3 months), and basic proline-rich proteins appeared at 4 years (±1 year) of age. All of the proteinases involved in the maturation of salivary proteins were more active in preterm than in at-term newborns, on the basis of the truncated forms detected. The activity of the Fam20C kinase, involved in the phosphorylation of various proteins, started around 180 days of postconceptional age, slowly increased reaching values comparable to adults at about 2 years (±6 months) of age. Instead, MAPK14 involved in the phosphorylation of S100A9 was fully active since birth also in preterm newborns.

How relevant is the impairment of smell for the quality of life in allergic rhinitis

Purpose of reviewIn the present review, the authors try to evaluate how relevant smell impairment is in patients suffering from allergic rhinitis and how it affects their quality of life. Smell dysfunction has a significant impact on the quality of life as it can lead to a wrong choice of food and intake, a reduction in appetite and eventually to weight loss, malnutrition, immunity reduction and worsening of medical illness. Patients with smell impairment are reported to use larger quantities of sugar and salt to highlight flavours, thus worsening their general health condition and increasing the risk of developing diabetes and hypertension. Recent findingsRecent studies estimate that a complete loss of the sense of smell can be found in at least 1% of the US population, and that an impairment in the olfactory function can be highlighted in about 24% of individuals aged 53–97 years and 19% of individuals aged 20–92 years. Despite the high prevalence, subjective complaints do not accurately reflect the real disturbance experienced by the patient, and usually go unnoticed. SummaryCurrent information in literature highlights the need for additional studies that concentrate on the impact of olfactory dysfunction on the quality of life of patients affected by allergic rhinitis.

Wegener's granulomatosis: a challenging disease for otorhinolaryngologists.

Conclusions Diagnosis of Wegeners granulomatosis (WG) can be delayed because of its aspecific presenting symptoms. Detection of serum circulating antineutrophil cytoplasm antibodies (c-ANCAs), in combination with histology, permits one to identify WG at an early stage and to implement stage-adapted therapy. c-ANCA levels may also help to evaluate the response to medical therapy. Recently, the quality of life of WG patients has been improved by administering cotrimoxazole in order to prevent infections and recurrent diseases during the remission period. Objective WG is of special significance to the otorhinolaryngologist because it is often initially limited to the upper respiratory tract before becoming systemic. The aim of this paper was to describe a series of WG patients and underline the difficulties involved in diagnosing and treating this challenging disease. Material and methods This was a prospective study in 23 consecutive patients with head and neck manifestations of WG (17 systemic, 6 limited). Diagnosis was performed by means of both c-ANCAs detection using indirect immunofluorescence and histology in biopsy specimens. Treatment consisted of daily cyclophosphamide (CYC; 2 mg/kg/day) and glucocorticoids (prednisone; 1 mg/kg/day). If an improvement or toxic events occurred, CYC was discontinued and methotrexate was started. If, during remission of the disease, low serum c-ANCAs levels were detected, CYC was suspended and cotrimoxazole (1 g/day) was introduced. Results Serum c-ANCAs detection was positive for all patients. Biopsy was diagnostic from the beginning in 19/23 cases. The six patients with limited WG did not show a progression to systemic disease. Only 3 patients with a diagnosis of delayed systemic WG died, whereas 19/23 patients were alive with good control of relapses.

Role of Biofilms in Chronic Inflammatory Diseases of the Upper Airways

UNLABELLED The objective of our studies was to document the presence of bacterial biofilms in recurrent and chronic infectious diseases of the upper airways (UA) (adenoiditis, tonsillitis, chronic rhinosinusitis) and to assess the association between the presence of biofilm and the maintenance of a chronic inflammation. METHODS 16 surgical samples of tonsils and adenoids from patients with UA infections and 24 samples of ethmoid mucosa from patients who underwent endoscopic sinus surgery for chronic rhinosinusitis (CRS) were cultured using conventional methods and subjected to scanning electron microscopy (SEM) to detect evidence of biofilm. RESULTS Bacterial biofilms were observed in 57.5% of chronically infected UA mucosa; in 41.7% of ethmoid mucosa of CRS patients they were significantly (p<0.001) associated with a marked destruction of ciliated epithelium. DISCUSSION Our studies confirm that biofilm formation plays a role in UA infections, it not only explains the resistance of these infections to antibiotic therapy, but also represents an important element that contributes to the maintenance of a chronic inflammatory reaction.

Rhino sinusal bilateral hamartoma: a case report.

The hamartoma is a benign rare tumor constituted by a mixture of tissue. It is very unusual in the nasal cavity. The objective of the study is to describe an unusual case of bilateral nasal hamartoma. We report a 52-year-old male patient with a bilateral paranasal hamartoma of the ethmoid and maxillary sinus. Functional endoscopic sinus surgery was performed to completely remove the masses. The reported localization is unusual because the most common site in the nose is the posterior septum. Although hamartoma arising from the rhino sinusal region is very rare, head and neck surgeons must know this entity in order to differentiate it from inverted papilloma and adenocarcinoma. Misinterpretation of this lesion may result in aggressive surgery for a benign lesion.

Is it sufficient to quantify aspiration for predicting aspiration pneumonia

A 63-year-old man previously treated with a subtotal laryngectomy, presented with a 4 month history of severe dysphagia and intractable aspiration. On the last follow-up he underwent a scintigraphic swallowing procedure that showed severe tracheopulmonary aspiration with visualization of the trachea, of the right and left bronchi, and evidence of good airway clearance on the follow-up scan obtained 15 minutes later. A chest radiograph showed no signs of pneumonia. Oropharyngeal-esophageal scintigraphy is still the gold standard technique to objectively assess and quantify tracheopulmonary aspiration, and to check the airway’s clearance mechanism. Pulmonary clearance represents the most important factor to prevent aspiration pneumonia in the case of intractable aspiration. Quantitative parameters are not sufficient to predict aspiration pneumonia. (Clin Nucl Med 2008;33: 236–239) REFERENCES 1. Prgomet D, Bumber Z, Bilic M, et al. Videofluoroscopy of the swallowing act after partial supraglottic laryngectomy by CO2 laser. Eur Arch Otorhinolaryngol. 2002;259:399–403. 2. Kreuzer SH, Schima W, Schober E, et al. Complications after laryngeal surgery: videofluoroscopic evaluation of 120 patients. Clin Radiol. 2000;55:775–781. 3. Galli J, Valenza V, D’Alatri L, et al. Postoperative dysphagia versus neurogenic dysphagia: scintigraphic assessment. Ann Otol Rhinol Laryngol. 2003;112:20–28. 4. Ekberg O. Diagnostic aspects of dysphagia. Acta Otolaryngol Suppl. 2000;543:225–228. 5. Marik PE. Aspiration pneumonitis and aspiration pneumonia. N Engl J Med. 2001;344:665–671. 6. Perlman AL, Booth BM, Grayhack JP. Videofluoroscopy predictors of aspiration in patients with oropharyngeal dysphagia. Dysphagia. 1994; 9:90–95. 7. Langmore SE, Skarupski KA, Park PS, et al. Predictors of aspiration pneumonia in nursing home residents. Dysphagia. 2002;17:298–307. 8. Bastian RW. Contemporary diagnosis of the dysphagic patient. Otolaryngol Clin North Am. 1998;31:489–506. 9. Kaya M, Inan M, Bedel D. Detection of tracheoesophageal fistula caused by ingestion of a caustic substance by esophageal scintigraphy. Clin Nucl Med. 2005;30:365–366. 10. Mc Adoo A, Leonard JC. Paraesophageal hernia and aspiration of oral secretions demonstrated by nuclear salivagram. Clin Nucl Med. 2007; 32:42–44. 11. Gomes GF, Campos AC, Pisani JC, et al. Diagnostic methods for the detection of anterograde aspiration in enterally fed patients. Curr Opin Clin Nutr Metab Care. 2004;7:285–292. Received for publication March 27, 2007; revision accepted August 6, 2007. From the *Institute of Otolaryngology, and †Nuclear Medicine, Catholic University of Sacred Heart, Rome, Italy; and ‡Don Gnocchi Foundation, Rome, Italy. Reprints: Claudio Parrilla, MD, PhD, Department of Otolaryngology, Catholic University of Sacred Heart, Policlinico “A.Gemelli”-Largo A. Gemelli n.8, 00168 Rome, Italy. E-mail: claudioparrilla@yahoo.com. Copyright

Biofilm’s Role in Chronic Cholesteatomatous Otitis Media A Pilot Study

Cholesteatoma is a destructive lesion involving the temporal bone, which may induce severe complications due to its expansion and erosion of adjacent structures. Bacterial biofilm plays a crucial role in the pathogenesis of many otolaryngologic inflammatory/infectious chronic diseases. In this pilot study, we investigated, by means of cultural examination and with scanning electron microscope, the presence of bacterial biofilm in a series of samples from the epitympanic and mastoid region in patients affected by cholesteatoma and from the promontory region in patients with healthy mucosa who were undergoing to stapes surgery. The preliminary data support the association between biofilm and cholesteatoma (81.3% of the cases) and allow us to hypothesize that keratinized matrix of cholesteatoma may represent the ideal substrate for biofilm colonization and survival; this finding is consistent with the clinical course of aural cholesteatoma, characterized by recurrent exacerbations and recalcitrant course.

Vestibular evoked myogenic potentials in evaluation of whiplash syndrome disability

Abstract Objective: Whiplash is an acceleration-deceleration mechanism of energy transfer to the neck that may result from a rear-end or side impact, predominantly in motor vehicle accidents but also from other mishaps. Vertigo and dizziness after whiplash injury are the most frequent, persistent and disabling symptoms, even in the absence of anatomical and cervical spine damage. Vestibular disturbances in whiplash patients have been widely investigated; nevertheless, physiopathology of vertigo and dizziness still remains an unsolved problem. The aim of our study was to demonstrate that cVEMPs (cervical Vestibular Evoked Myogenic Potentials) represent a valid and non-invasive technique able to investigate vestibular function, in particular the vestibulo-collic reflex and macular function. Study design: We examined 25 patients affected by disabling post-whiplash injury by means of audiovestibular tests: audiometric and impedance tests, nystagmus evaluation, caloric test, dynamic posturography and cervical VEMPs. Data obtained were compared with those of a normal control group. Results: Audiological tests, impedance tests, nystagmus evaluation, and caloric tests were normal in all patients. Comparison of dynamic posturography values obtained in whiplash patients and in the normal control group demonstrated significant differences in the Composite Equilibrium Score and Visual Component. Finally, cVEMPs were bilaterally absent in 72% of patients, unilaterally absent in 16% and present in 12%. Conclusions: Based upon neurophysiological evidence and our results, we can hypothesize that a concussion of the macular receptors takes place and that the maculae are more sensitive than the ampulla or cochlea receptor to this kind of injury. We conclude that cervical VEMPs represent a valid and non-invasive technique to evaluate disability after whiplash injury.