Pre-clinical evaluation of a new coral-based bone scaffold.

Coral is used worldwide for bone reconstruction. The favorable characteristics that make this material desirable for implantation are (i) osteoinduction, (ii) and osteoconduction. These proprieties have been demonstrated by in vivo studies with animal models and clinical trials over a twenty-year period. Also poly(2-hydroxyethylmethacrylate) [poly(HEMA)] is a widely used biomaterial. By using coral and poly(HEMA), a scaffold for bone reconstruction application has been recently synthesized. Cytological, histological and genetic analyses were performed to characterize this new alloplastic material. Four samples were analyzed: (a) white coral (WC), (b) red coral (RC), (c) WC plus polymer (WCP) and (d) RC plus polymer (RCP). Quantification of mitochondrial dehydrogenase activity by MTT assay was performed as indirect detector of cytotoxicity. In vivo effects were revealed by implanting corals and coral-based polymers in rabbit tibia. Samples were collected after 4 weeks and subjected to histological analysis. To evaluate the genetic response of cells to corals and coral-derived polymers an osteoblastlike cell line (i.e. MG63) was cultured in wells containing (a) medium, (b) medium plus corals and (c) medium plus two types of scaffolds (RCP or WCP). RNAs extracted from cells were retro-transcribed and hybridized on DNA 19.2K microarrays. No cytotoxicity was detected in corals and coral-based biopolymers. No inflammation or adverse effect was revealed by histological examination. By microarray analysis 154 clones were differentially expressed between RC and WC (81 up and 73 down regulated) whereas only 15 clones were repressed by the polymer. Histological evaluation not only confirmed that coral is a biocompatible material, but also that the polymer has no adverse effect. Microarray results were in agreement with cytological and histological analyses and provided further data regarding the genetic effects of RC, WC and the new polymer.

The contribution of histopathological examination to the diagnosis of cervico-facial actinomycosis: a retrospective analysis of 68 cases.

Cervico-facial actinomycosis is an infectious, suppurative, and granulomatous disease due to Actinomyces species. Usually, the diagnosis is confirmed by microbiological cultures; however, the need for careful anaerobic handling of specimens often makes it difficult to obtain an effective microbial growth. Therefore, we conducted a retrospective study on biopsy samples from patients with a clinical suspicion of cervico-facial actinomycosis, in order to determine whether accurate histopathological examination could reliably confirm the diagnosis. A retrospective revision of formalin-fixed, paraffin-embedded archival material from 68 cases of cervico-facial lesions, with negative culture for anaerobic/microaerophilic microorganisms, was performed. Twelve serial sections for each case were cut from the paraffin blocks, individually collected on positively charged slides to obtain good section-to-slide adhesion, and stained with hematoxylin and eosin (H&E) and periodic acid-Schiff (PAS). Histopathological examination of the serial sections allowed the identification of bacterial colonies consistent with actinomycetes in 22 cases (32 %). The proposed histopathological examination allowed the retrospective diagnosis of cervical actinomycosis in one-third of clinical specimens that remained misdiagnosed following traditional H&E examination.

Expression of poly(ADP-ribose) polymerase in bone regeneration.

Poly(ADP-ribose) polymerase (PARP) is a 116kDa enzyme catalysing the synthesis of ADP-ribose polymers from NAD+. PARP is activated in response to DNA strand breaks and plays a critical role in the maintenance of genomic integrity. However, considering its role also in transcription, proliferation as well as apoptosis in biological process, in the present study the role of PARP in bone regeneration was evaluated, in particular in bone cell proliferation and differentiation processes. Thus, formalin fixed paraffin embedded specimens of 10 human bone samples after sinus lift were collected and investigated by immunohistochemistry using a mouse monoclonal anti-human PARP antibody. PARP was expressed in cells with morphological features of osteoblasts in the areas of new bone formation at the junction between mineralized and unmineralized tissue, between osteoid tissue and bone. Few osteoclasts were observed and showed only focal nuclear expression of PARP, while osteocytes showed no positivity for PARP. Our data showed an overall involvement of PARP enzyme in human bone tissues, in particular during bone regeneration process.

Novel subgingival bacterial phylotypes detected using multiple universal polymerase chain reaction primer sets.

INTRODUCTION: Molecular ecological analysis based on 16S rRNA gene sequence analysis is well established for the characterisation of complex bacterial communities. However, 'universal' PCR primers can introduce biases into the analysis of the species composition of clone libraries because of mismatches between the primer and target organism sequences. In this study, three universal primer sets were compared for the analysis of the microflora in subgingival plaque. METHODS: Three subgingival plaque samples were collected from two subjects with localised severe chronic periodontitis. Half of each sample was cultured while DNA was extracted from the remaining half and 16S rDNA amplified with universal primer pairs 27F, 1525R (A); 27F, 1492R (B) and 530F, 1525R (C). Amplified genes were cloned, sequenced and identified by comparison with 16S rRNA databases. RESULTS: 137 taxa were identified among 177 isolates and 417 clones sequenced. Of these, 86 were detected only by the molecular technique whereas 26 were found only by culture. Sequences from 81 taxa did not correspond to those of named species and of these, 38 were not represented in the nucleotide databases. 16S RNA genes for these 38 taxa were sequenced and deposited with GenBank. CONCLUSION: The use of three sets of universal primers allowed the identification of 38 novel bacterial phylotypes. There were marked differences in the composition of the libraries generated by the different primer sets. A combination of molecular and cultural techniques is recommended to maximise the coverage of detection of bacterial taxa in oral samples.

Cephalometric evaluation of the upper airway in patients with complete dentures.

AIM: Obstructive sleep apnea syndrome (OSAS) is a disorder characterised by repeated obstructions of the upper airway, with consequent episodes of apnea and hypopnea during sleep, snoring, and daytime sleepiness. Recently, a role of edentulism in the occurrence or in the worsening of this disorder was suspected. The aim of the study was to assess, through a cephalometric analysis, if the removal of denture induces, and where, modifications that can favour the pharynx collapse. METHODS: A total of 27 subjects with complete or partial loss of teeth and with heavy loss of the vertical dimension, were examined. The patients were submitted to radiographs of the cranium in supine position, to simulate as much as possible the night condition, with the denture in intercuspal position (ICP), and in relaxed position both with denture (D-RP) and without it (edentule-relaxed position, E-RP). The radiographs were analysed through specific measures for OSA evaluating the parameters that could be modified by the denture. RESULTS: The pharyngeal airway space (PAS) decreases, at the level of uvula, from ICP (6.7 mm) to RP (5.3 mm) (P< 0.05). The distance between the base and the tip of the tongue significatively decreased both from ICP vs E-RP (7.35 mm vs 6.87 mm; P< 0.05), both from D-RP to E-RP (7.22 mm vs 6.87 mm; P< 0.005). CONCLUSIONS: From the data obtained from the cephalometric analysis it seems that wearing denture induces modifications in the position of the tongue, of the jaw and of the pharyngeal airway space that can favour the reduction of apnea episodes.

Squamous cell carcinoma of the lower lip: FAS/FASL expression, lymphocyte subtypes and outcome.

Squamous cell carcinoma (SCC) of the lip is a relatively common malignancy of the head and neck region. Tumour thickness, grading and perineural invasion are significant prognostic indicators. However, there is still the need of new reliable biological markers able to predict the prognosis of the single cases with an unfavourable biological behaviour unpredictable by the classic clinical-pathological parameters. 32 cases of (SCC) of the lower lip were analysed for their clincopathologic features, and immunohistochemical expression of Fas/FasL in neoplastic cells and in inflammatory infiltrate. Moreover the density and phenotype of tumour-infiltrating lymphocytes (TIL) were analysed. The results were related with the follow-up of the patients ranging from 2 to 6 years. The cases with over-expression of Fas/FasL in neoplastic cells and Fas+ in T cells preferentially showed a more aggressive clinical behaviour (P<0.01). Moreover we found an alteration of the normal expression of CD4 and CD8 lymphocyte types in ten cases. This data suggest that the Fas/FasL pathway is involved in the close relation between neoplastic cells and T cells and so in the biological behaviour of these tumours.

Culture-independent identification of periodontitis-associated Porphyromonas and Tannerella populations by targeted molecular analysis.

Periodontitis is the commonest bacterial disease of humans and is the major cause of adult tooth loss. About half of the oral microflora is unculturable; and 16S rRNA PCR, cloning, and sequencing techniques have demonstrated the high level of species richness of the oral microflora. In the present study, a PCR primer set specific for the genera Porphyromonas and Tannerella was designed and used to analyze the bacterial populations in subgingival plaque samples from inflamed shallow and deep sites in subjects with periodontitis and shallow sites in age- and sex-matched controls. A total of 308 clones were sequenced and found to belong to one of six Porphyromonas or Tannerella species or phylotypes, one of which, Porphyromonas P3, was novel. Tannerella forsythensis was found in significantly higher proportions in patients than in controls. Porphyromonas catoniae and Tannerella phylotype BU063 appeared to be associated with shallow sites. Targeted culture-independent molecular ecology studies have a valuable role to play in the identification of bacterial targets for further investigations of the pathogenesis of bacterial infections.

[In vitro evaluation of chromatic stability in new-generation composites]. / Valutazione in vitro della stabilità cromatica di materiali compositi di nuova generazione.

BACKGROUND: Ultraviolet light exposure produces chromatic changes in composites used for esthetic restoration. The aim of the study was to compare the chromatic changes induced by accelerated aging in two composites of new generation: Tetric Ceram and Diamond Crown. METHODS: Five samples were prepared for each composite, each divided into two pieces, one exposed to accelerated aging obtained by ultraviolet-light exposure for 24 hours, and one maintained in dark. Thereafter, the samples were lighted by a light beam and the percentage of reflected light was assessed. The distribution of the levels of red, green and blue was assessed using a camera connected to a PC with a software for image processing. RESULTS: For both the composites, the values of the three colors were significantly lower in the samples artificially aged (p<0.0001). The values concerning to the Tetric Ceram were significantly lower than those of Diamond Crown (p<0.0001). The latter showed aging-induced variation significantly smaller than the former. CONCLUSIONS: Both the composites showed significant chromatic variations after 24 hours artificial aging. Diamond Crown showed significantly better stability.

Influence of mastication on gastric emptying.

The role of mastication on digestion efficiency remains to be demonstrated. This study investigates whether masticatory function influences gastric emptying rate. Twelve normal volunteers were studied on two occasions after ingestion of the same test meal containing ham cubes, crackers, and egg (mixed with 13C-octanoic acid), chewed, in random order, either with 50 masticatory cycles or with 25 cycles, swallowing ham cubes whole. Lag phase (Tlag) and gastric half-emptying time (T1/2) were measured by means of the 13C-octanoic acid breath test. Trituration performance was assessed by the sieve test, and was expressed as the percentage of ham particles < or = 1 mm after 50 masticatory cycles. Tlag and T1/2 were significantly shorter when the meal was chewed with 50 cycles than with 25 cycles (Tlag 25.9+/-3.8 vs. 36.4+/-4.1 min, p=0.017; T1/2 49.1+/-5.7 vs. 62.5+/-6 min, p=0.009). Trituration performance was inversely related to both Tlag (r=0.621, p=0.031) and T1/2 (r=0.699, p=0.012). Comminution of food influences significantly gastric emptying rates.

Functional bowel disorders in the geriatric patient: constipation, fecal impaction, and fecal incontinence.

Functional bowel disorders, including constipation, fecal impaction, and fecal incontinence, are common gastrointestinal diseases in the elderly and a major source of morbidity. Often the etiology is multifactorial. Evaluation should investigate presenting symptoms and important historical clues. An appropriate workup, including radiological studies, endoscopy, and physiological testing should be individualized. Therapy should be directed against relieving the major complaint and treating the underlying pathophysiological mechanism.

[Correlation between edentulism, sleep disorders and arterial hypertension. Preliminary research]. / Correlazione fra edentulismo, disturbi del sonno e ipertensione arteriosa. Indagine preliminare.

BACKGROUND: Respiratory disturbances during sleep are considered risk factors for arterial hypertension and cardiovascular diseases. Edentulism, by decreasing retro-pharyngeal space, may favor upper airway occlusion during sleep. Aim of the study is to evaluate whether edentulism is associated with greater prevalence of sleep disturbances, arterial hypertension and other cardiovascular diseases. METHODS: Eighty edentulous subjects with dentures (removed during sleep) and 57 subjects with natural teeth, matched for age, underwent assessment of oral conditions and recording of questionnaires on diseases and respiratory sleep disturbances. RESULTS: Edentulous subjects had a significantly higher prevalence of arterial hypertension and cardiovascular diseases, than subjects with natural teeth. CONCLUSIONS: In edentulous subjects, removing dentures during sleep may favor respiratory disorders, and increase the risk for hypertension and cardiovascular disease.

[The influence of edentulism on spirometric values]. / Influenza dell'edentulismo sui valori spirometrici.

BACKGROUND: Aim of the paper is to study whether edentulism, by causing a decrease in size and tone of pharyngeal musculature, may affect spirometric measurements. METHODS: Spirometry was recorded with and without dentures in 58 edentulous subjects, 36 asymptomatic normal subjects (N) and 22 patients with chronic obstructive pulmonary disease (COPD). In 10 subjects retropharyngeal space with and without dentures was assessed by cephalometry. RESULTS: In the N group, removing dentures produced a significant decrease in lung volumes and airflow rates, while in COPD patients it was ineffective. In both groups, retropharyngeal space was significantly decreased by removing dentures. CONCLUSIONS: Edentulism, by decreasing extrathoracic airway caliber, influences significantly spirometric measurements in normal subjects but not in those with COPD.

The role of social determinants on men's and women's mobility in Italy. A comparison of discriminant analysis and artificial neural networks.

The paper focuses on the role of the spouse's occupation as a resource for mobile individuals, from the perspective that social positions are held by families, rather than by individuals. Three groups are confronted in terms of the role of the key variables and other relevant factors: men whose spouse does not have a paid job (group 1), men and women whose spouse has a paid job (group 2 and 3). The data set is provided by the national survey on social mobility in Italy, carried out in 1985; social achievements of members of the three groups are considered, including social origins and destinations, social position corresponding to respondent's first job, cultural background (educational achievement of respondent's father and mother found), respondent's education and spouse's social position. The techniques used are discriminant analysis and back propagation Neural Networks. Both techniques traced a clear boundary between group 1 and groups 2 and 3, which were discriminated mainly on the basis of the spouse's occupation; Artificial Neural Networks reached better classification results and allowed a deeper insight into the nonlinear effects of the discriminating variables for the three groups.

Identification of a lactoferrin-binding protein in Prevotella nigrescens.

A 40-kDa lactoferrin-binding protein was identified in a strain of Prevotella nigrescens isolated from a patient with periodontitis. The protein was purified by affinity column chromatography using a Sepharose-lactoferrin column and detergent-solubilized membranes. The N-terminal sequence revealed no apparent similarities with any other sequenced bacterial protein. The native conformation of the 40-kDa protein was a condition to bind either iron-free or iron-saturated lactoferrin. A possible function of this Lf-binding protein could be related with an iron acquisition mechanism in P. nigrescens.

Relationship between antibacterial activity and cell surface binding of lactoferrin in species of genus Micrococcus.

Human lactoferrin was bactericidal in vitro for Micrococcus luteus but not for other Micrococcus species (M. radiophilus, M. roseus and M. varians). A correlation between the binding of lactoferrin to the bacterial surface and the antimicrobial action was observed. Viability assays showed that ferric, but not ferrous, salts prevented binding and consequently M. luteus was not killed. The unsaturated form of lactoferrin showed a greater affinity than that of the iron-saturated molecule for lipomannan, a lipoglycan present on the cell wall of M. luteus, supporting the role for lipomannan as one of the possible binding sites of lactoferrin on M. luteus.

Binding and degradation of lactoferrin by Porphyromonas gingivalis, Prevotella intermedia and Prevotella nigrescens.

The ability of laboratory and clinical strains of Porphyromonas gingivalis, Prevotella intermedia and Prevotella nigrescens to bind and to degrade lactoferrin (Lf) has been assessed. Lf bound readily to whole cells of each species apparently via high-affinity site and one or more low-affinity sites. P. gingivalis showed a lower affinity for Lf than the other two species (P < 0.001). Virtually all strains of P. gingivalis completely degraded Lf under the conditions employed, whereas P. intermedia and P. nigrescens showed only partial degradation. These data suggest that Lf binds to a high-affinity receptor on all these bacteria and, particularly in the case of P. gingivalis, is then degraded by cell-associated proteases. This property may provide protection to the cell against the effects of Lf in periodontal sites and so is a possible virulence factor in disease. There was no association between the ability to degrade Lf and whether the strains had originated from healthy or diseased oral sites.

[Isolation of Haemophilus influenzae and Haemophilus parainfluenzae in genitourinary infections: a 4-year review]. / Aislamiento de Haemophilus influenzae y Haemophilus parainfluenzae en infecciones genitourinarias: una revisión de 4 años.

BACKGROUND: Haemophilus spp. had been previously suggested as a potential pathogen in genitourinary infections that could be sexually transmitted. In order to check that suggestions we have determined the incidence, pathogenic role, possible sexual transmission and susceptibility to antibiotics in isolates of Haemophilus parainfluenzae and Haemophilus influenzae from genital tract infections. The microbiological samples were taken during a period of four years from patients attended in a Service of Sexual Transmission Diseases and the data were further reviewed. METHODS: The study included 5,572 genital specimens from 2,182 women prostitutes with different genitourinary infections and from 825 men with urethritis. Microbiological samples were cultured in a non-specific media for genital pathogens and species of Haemophilus spp. and clinical circumstances of isolation were evaluated. Susceptibility tests were performed by using a standard microdilution test. RESULTS: Haemophilus spp. was isolated in 155 samples (2.8%) using a non-selective culture method. H. parainfluenzae was isolated in 100 cases (64.5%), Haemophilus influenzae in 45 cases (29%) and Haemophilus spp. in 10 strains (6.4%). Haemophilus spp. was isolated as a sole pathogen in men with urethritis (8 cases), epididymo-orchitis (2 cases), cervicitis and/or vaginitis (9 cases) and Bartholin's Abscess (2 cases). The most frequent biotypes were H. parainfluenzae biotype II (43%) and III (19%), and H. influenzae biotype IV (35.5%). Beta lactamase activity and ampicillin resistance were present in 29% of the H. parainfluenzae strains and in the 26.7% of clinical isolates of H. influenzae. CONCLUSIONS: 1) Haemophilus spp. was isolated from genitourinary infections at a low frequency in the studied group. 2) The pathogenic role of Haemophilus spp. was suggested when was isolated as a sole pathogen present from some infections of the genitourinary tract such as urethritis in men and Bartholin's abscess in women. 3) The susceptibility to antibiotics in the clinical isolates of Haemophilus spp. from genitourinary infections was similar previously reported studies performed in Spain.

Recurrence of unilateral retinoblastoma following radiation therapy.

A retrospective analysis of 65 unilateral retinoblastoma patients treated initially with external beam radiation, revealed that 25 eyes (38.5%) developed local recurrence of retinoblastoma. The mean age at diagnosis was 1.8 years for patients who developed recurrences vs. 0.9 years for those who did not. Ninety-six percent of the recurrences occurred less than two years from the age at diagnosis; the amount of time from the end of external beam radiation treatment until a tumor recurred was independent of the age at diagnosis. The initial largest basal diameter was 10.7 DD for tumors which later recurred and 5.9 DD for tumors that were cured. Sixty-nine percent of eyes in groups III-V had tumor recurrence, and 10% of eyes in groups I-II had recurrence. All but one eye (24 eyes) that developed recurrence were enucleated. Family history of retinoblastoma, location of the tumor, gender, and laterality did not significantly correlate with the mean age of initial diagnosis for tumors that recurred or the mean time of onset for recurrence.

Interaction of calmodulin with lactoferrin.

Calmodulin, as a major intracellular calcium-binding protein, regulates many Ca(2+)-dependent enzymes and plays an important role in a wide spectrum of cellular functions of the eukaryotes. Interaction between calmodulin and human lactoferrin, a 78 kDa protein with antibacterial properties, was found in the presence of Ca2+ using (i) a method for the detection of calmodulin binding proteins with biotinylated calmodulin, (ii) affinity chromatography on an agarose-calmodulin column with subsequent detection by an enzyme-linked immunosorbent assay (ELISA). The binding of calmodulin to lactoferrin blocked the ability of lactoferrin to agglutinate Micrococcus lysodeikticus.