Size and specimen-dependent strategy for x-ray micro-ct and tem correlative analysis of nervous system samples.

Correlative approaches are a powerful tool in the investigation of biological samples, but require specific preparation procedures to maintain the strength of the employed methods. Here we report the optimization of the embedding protocol of nervous system samples for a correlative synchrotron X-ray computed microtomography (micro-CT) and transmission electron microscopy (TEM) approach. We demonstrate that it is possible to locate, with the micrometric resolution of micro-CT, specific volumes of interest for a further ultrastructural characterization to be performed with TEM. This approach can be applied to samples of different size and morphology up to several cm. Our optimized method represents an invaluable tool for investigating those pathologies in which microscopic alterations are localized in few confined regions, rather than diffused in entire tissues, organs or systems. We present a proof of concept of our method in a mouse model of Globoid Cells Leukodistrophy.

On the thermodynamic path enabling a room-temperature, laser-assisted graphite to nanodiamond transformation.

Nanodiamonds are the subject of active research for their potential applications in nano-magnetometry, quantum optics, bioimaging and water cleaning processes. Here, we present a novel thermodynamic model that describes a graphite-liquid-diamond route for the synthesis of nanodiamonds. Its robustness is proved via the production of nanodiamonds powders at room-temperature and standard atmospheric pressure by pulsed laser ablation of pyrolytic graphite in water. The aqueous environment provides a confinement mechanism that promotes diamond nucleation and growth, and a biologically compatible medium for suspension of nanodiamonds. Moreover, we introduce a facile physico-chemical method that does not require harsh chemical or temperature conditions to remove the graphitic byproducts of the laser ablation process. A full characterization of the nanodiamonds by electron and Raman spectroscopies is reported. Our model is also corroborated by comparison with experimental data from the literature.

EXPLOITING THE VERSATILITY OF CHOLESTEROL IN NANOPARTICLES FORMULATION.

The biocompatibility of polymers, lipids and surfactants used to formulate is crucial for the safe and sustainable development of nanocarriers (nanoparticles, liposomes, micelles, and other nanocarriers). In this study, Cholesterol (Chol), a typical biocompatible component of liposomal systems, was formulated in Chol-based solid nanoparticles (NPs) stabilized by the action of surfactant and without the help of any other formulative component. Parameters as type (Solutol HS 15, cholic acid sodium salt, poly vinyl alcohol and Pluronic-F68), concentration (0.2; 0.5 and 1% w/v) of surfactant and working temperature (r.t. and 45°C) were optimized and all samples characterized in terms of size, zeta potential, composition, thermal behavior and structure. Results demonstrated that only Pluronic-F68 (0.5% w/v) favors the organization of Chol chains in structured NPs with mean diameter less than 400nm. Moreover, we demonstrated the pivotal role of working temperature on surfactant aggregation state/architecture/stability of Chol-based nanoparticles. At room temperature, Pluronic-F68 exists in solution as individual coils. In this condition, nanoprecipitation of Chol formed the less stable NPs with a 14±3% (w/w) of Pluronic-F68 prevalently on surface (NP-Chol/0.5). On the contrary, working near the critical micelle temperature (CMT) of surfactant (45°C), Chol precipitates with Pluronic-F68 (9±5% w/w) in a compact stable matricial structure (NP-Chol/0.5-45). In vitro studies highlight the low toxicity and the affinity of NP-Chol/0.5-45 for neuronal cells suggesting their potential applicability in pathologies with a demonstrated alteration of neuronal plasticity and synaptic communication (i.e. Huntington's disease).

Design and optimization of lipid-modified poly(amidoamine) dendrimer coated iron oxide nanoparticles as probes for biomedical applications.

Superparamagnetic iron oxide nanoparticles with a wide size range (2.6-14.1 nm) were synthesized and coated with the amphiphilic poly(amidoamine) PAMAM-C12 dendrimer. The resulting well dispersed and stable water suspensions were fully characterized in order to explore their possible use in biomedical applications. The structural and magnetic properties of the nanoparticles were preserved during the coating and were related to their relaxometric behaviour. The Nuclear Magnetic Resonance Dispersion (NMRD) profiles were found to be in accordance with the Roch model. The biocompatibility was assessed by means of cell viability tests and Transmission Electron Microscopy (TEM) analysis. The nanoparticles' capability of being detected via Magnetic Resonance Imaging (MRI) was investigated by means of clinical MRI scanners both in water and agar gel phantoms, and in a mouse model.

Autophagy as a new therapeutic target in Duchenne muscular dystrophy.

A resolutive therapy for Duchene muscular dystrophy, a severe degenerative disease of the skeletal muscle, is still lacking. Because autophagy has been shown to be crucial in clearing dysfunctional organelles and in preventing tissue damage, we investigated its pathogenic role and its suitability as a target for new therapeutic interventions in Duchenne muscular dystrophy (DMD). Here we demonstrate that autophagy is severely impaired in muscles from patients affected by DMD and mdx mice, a model of the disease, with accumulation of damaged organelles. The defect in autophagy was accompanied by persistent activation via phosphorylation of Akt, mammalian target of rapamycin (mTOR) and of the autophagy-inhibiting pathways dependent on them, including the translation-initiation factor 4E-binding protein 1 and the ribosomal protein S6, and downregulation of the autophagy-inducing genes LC3, Atg12, Gabarapl1 and Bnip3. The defective autophagy was rescued in mdx mice by long-term exposure to a low-protein diet. The treatment led to normalisation of Akt and mTOR signalling; it also reduced significantly muscle inflammation, fibrosis and myofibre damage, leading to recovery of muscle function. This study highlights novel pathogenic aspects of DMD and suggests autophagy as a new effective therapeutic target. The treatment we propose can be safely applied and immediately tested for efficacy in humans.

Bone regeneration: in vitro evaluation of the behaviour of osteoblast-like MG63 cells placed in contact with polylactic-co-glycolic acid, deproteinized bovine bone and demineralized freeze-dried bone allograft.

Insufficient bone density of the alveolar crests, caused by loss of the dental elements, sometimes impedes the primary stability of an integrated bone implant. The techniques of bone regeneration allow to obtain a sufficient quantity of alveolar bone to permit the implant rehabilitation of the edentulous crests. Today several grafting materials are available and they have different characteristics, according to their structure, which influence the different behaviour of the grafting materials to the bone and the implant surface. The aim of this study is to evaluate the interaction between a human osteosarcoma MG63 cell line and three different biomaterials: polylactic-co-glycolic acid (PLAGA), deproteinized bovine bone and demineralised freeze-dried bone allograft (DFDBA). From this study a different behaviour emerges of the osteoblast-like MG63 cells in relation to the sublayer on which these cells were placed in culture. The results of the study, in fact, demonstrate that the most osteoconductive material of the three analysed is the DFDBA, followed by DPBB. On the contrary, the PLGA, because of its roughness, does not seem to represent a valid support for cell growth, and does not encourage any morphologic modification in tumor cells. Furthermore, deproteinized bovine bone shows a differentiating effect which could lead to hypothesise an osteoconductive capacity of this biomaterial. Further studies should be carried out with the aim of explaining the results obtained.

Mandibular osteomyelitis due to Pseudomonas aeruginosa. Case report.

Osteomyelitis is a relatively frequent bacterial infection of the jaw bones. This report describes a case of mandibular osteomyelitis in a surgical site after enucleation of a follicular cyst and extraction of the associated tooth. This case is unusual because maxillary osteomyelitis generally results from polymicrobial infection. In our patient, however, laboratory analysis identified Pseudomonas aeruginosa as the etiologic agent, an opportunistic pathogen normally found on moist surfaces and vegetation. Notorious for its antibiotic multiresistance, P. aeruginosa is increasingly recognized as a serious problem in hospitalized patients. Isolation of the responsible microbe permitted specific antibiotic treatment with a 10-day course of ciprofloxacin (250 mg/12 h), which fully cleared the infection.

Adeno-tonsillar surgery in Italy.

Indications to surgery for adeno-tonsillar inflammatory disorders and analysis of the effectiveness of surgical treatment, compared with watchful waiting strategy, continue to be the subject of scientific debate. The present investigation focuses on the surgical activity of 14 Italian Otorhinolaryngological Units between 1999 and 2004. Surgical interventions (adeno-tonsillectomy, adenoidectomy, tonsillectomy) on 26915 children (age range: 2-11 years) were considered. Data on adeno-tonsillar interventions were analysed in relation to other interventions of ENT interest, performed in the same units and in the same period. Adeno-tonsillar interventions accounted for 35.4% of all operations of ENT interest. Adeno-tonsillectomy accounted for 56.6% of overall adeno-tonsillar operations, adenoidectomy 31.6%, tonsillectomy 11.8%. The percentage for the three interventions was homogeneous in the period of the study and in the recruited units. The percentage of children who underwent adeno-tonsillar surgery in paediatric units was higher as compared to general units, as far as concerns the overall number of operations performed. In southern Italy, the number of adeno-tonsillar interventions, in general, and of adeno-tonsillectomy, in particular, was higher compared to that in northern Italy. Results of the present study suggest that environmental factors, cultural issues and local health demands, may influence indications and, therefore, the different incidence of the operations under consideration in the units taking part in the investigation.

Bone substitutes in oral surgery.

Osseous defects pose a clinical challenge the operator can meet with the aid of techniques that promote bone tissue regeneration. The current gold standard is autologous bone harvested from intra- and extraoral donor sites; however, autologous bone grafting requires two surgical sites (donor and recipient), resulting in greater morbidity and prolonged operating times, particularly for extraoral sites, with greater discomfort for the patient. Such disadvantages can be overcome with the use of bone substitute materials. There is a notable variety of so-called intelligent biomaterials that can modulate bone response in regeneration. Based on origin, bone substitute materials are classified as allogenic, heterologous and alloplastic materials. The first refer to bone from same-species donors, the second are obtained through processing of bone from different species, while alloplastic materials are synthetic composites. Besides different resorption rates, they possess different chemical and structural characteristics that influence the stimulation or support of bone regeneration. In daily clinical practice, before selecting from the wide variety of biomaterials, a wise step is to analyze and compare the clinical and histological results obtained with these materials. This article examines the clinical applications and osteoconductive and/or osteoinductive properties of some currently available biomaterials.

[The treatment of acute recurrent pharyngotonsillitis]. / Il trattamento delle flogosi acute ricorrenti faringo-tonsillari.

AIM OF THE STUDY: In paediatric patients, recurrent febrile pharyngo-tonsillitis is usually due to chronic inflammation of the tonsils and/or adenoids: these episodes are often associated with other clinical manifestations (respiratory obstruction of naso-pharyngeal origin; auricular inflammation, especially effusive otitis media and acute otitis media; streptococcal beta-haemolytic Group A (SBEGA) infection, causing a distant disorder of varying severity and which may, therefore, give rise to serious pathological conditions). A retrospective multicentric investigation has, therefore, been carried out in order not only to further elaborate findings emerging from earlier studies, performed in other countries, in which random methods were used, but also to focus on: 1. the possible association of these clinical manifestations with recurrent febrile pharyngo-tonsillar inflammation in relation to frequency of the condition; 2. results obtained with adenoidectomy or with adeno-tonsillectomy, referring to the overall clinical outcome of the pathological condition, bearing in mind data emerging from the control groups, submitted to strict follow-up, associated with any medical treatment deemed necessary. MATERIAL AND METHODS: The study population comprised 692 patients (394 male, 298 female), age range 2-11 years and with a minimum follow-up of 2 years. Considering the clinical manifestations emerging from the case history, the following two investigations were carried out: 1. the first, on 501 patients, all previously submitted to adeno-tonsillectomy in whom the recurrent febrile pharyngo-tonsillar inflammation had occurred, respectively > 4 or < or = 4 episodes during the last 12 months; 2. the second, on 455 patients in whom recurrent febrile pharyngo-tonsillar inflammation, in the 12 months prior to the beginning of the study, had been < or = 4 episodes: in 264 cases, adeno-tonsillectomy was carried out; in 144, adenoidectomy; 47 children were studied as controls and, therefore, submitted to watchful waiting. The results obtained were statistically analysed by chi2 and Fisher test: a "p" value of < 0.05 was considered statistically significant. RESULTS: The first investigation showed that all symptomatological manifestations considered were resolved following adeno-tonsillectomy, in a significantly high percentage of cases (recurrent febrile pharyngo-tonsillar in flammation: 91.8%; naso-pharyngeal respiratory obstruction: 92.2%; pathological ASO titre: 69.8%; distant disorders probably due to SBEGA: 76%; effusive otitis media: 76%; acute recurrent otitis media: 87.5%). The most interesting clinical findings obtained can be summarized as follows: naso-pharyngeal respiratory obstruction is a frequent finding; the incidence is statistically higher in cases in which recurrent febrile pharyngo-tonsillar inflammation is less frequent (p < 0.05): in these patients, this is an important pathological manifestation indicating the need for surgery; the higher incidence is associated with at significant finding of pathological auricular processes; the percentages of recovery for naso-pharyngeal respiratory obstruction are statistically greater in the group of patients in whom the recurrent febrile pharyngo-tonsillar inflammation is more frequent: an identical result is found in patients with the effusive form of otitis media and in those with recurrent acute otitis media; a pathological ASO titre is more frequently found, the difference being statistically significant (p < 0.05), in the group of patients with more frequent recurrent febrile pharyngo-tonsillar inflammation, albeit this factor does not affect the percentage of recovery; also the favourable evolution and recovery of clinical distant manifestations, related to streptococcal beta-haemolytic Group A infection, are correlated with the return to normal of the ASO titre. The second investigation demonstrated the efficacy of surgical treatment, adeno-tonsillectomy and adenoidectomy, in all the clinical manifestations studied, the possibility of achieving resolution of recurrent febrile pharyngo-tonsillar inflammation even in the control subjects submitted to medical treatment, but also the lack of success, in the latter, as far as concerns naso-pharygeal respiratory obstruction, the pathological ASO titre and the inflammatory auricular processes. CONCLUSIONS: The present investigations confirm previous observations emerging from randomized clinical studies, regarding the possibility to achieve valid prevention of recurrent febrile pharyngo-tonsillar inflammation, also in a large percentage of control patients submitted to watchful waiting; these investigations, however, also demonstrated that, in the control group, the percentage of subjects in whom recovery of other symptoms was achieved was very low compared to that in the groups submitted to surgery. In other words, clearly emerges as far as concerns the overall symptomatological picture of the patients, the efficacy of the surgical treatment, but also the lack of success of the medical treatments in all clinical manifestations associated with recurrent febrile inflammations. Therefore, it is necessary to select and advise the most suitable treatment strategy, not in relation to the Guidelines elaborated on the basis of results emerging from statistical research, that, due to the intrinsic methodology limits cannot be applied to all cases in an acritical fashion, but in relationship to the specific clinical picture of each individual patient.

[Acute recurrent pharyngotonsillitis and otitis media]. / Le flogosi faringo-tonsillari acute ricorrenti e le manifestazioni infiammatorie auricolari.

BACKGROUND: In the more recent randomized clinical studies, the surgical procedures adopted (adenoidectomy and adeno-tonsillectomy) for the treatment of the effusive and the acute recurrent forms of otitis media have generally shown a relatively modest and not persistent efficacy. It should be pointed out, however, that the cases studied displayed no clinical elements suggesting the existence of any aetio-pathogenic relationship between the auricular inflammation and a possible pathological disorder localized in the tonsils and/or adenoids. AIM OF THE STUDY: The present investigation aimed to establish: a) the frequency with which the two forms of tympanic inflammation, under study, were detected in children with recurrent febrile pharyngo-tonsillar inflammation, also considering the possible presence of naso-pharyngeal respiratory obstruction and the severity of these two pathological manifestations; b) in relationship to these parameters, what might be the outcome with respect to the pathological auricular processes following surgical treatment of the pharyngeal disorder (adeno-tonsillectomy or adenoidectomy) vs. watchful waiting, c) the possible advantages offered by ventilation tube, inserted during surgical treatment, in the prevention of the two forms of otitis media studied. MATERIAL AND METHODS: This retrospective multi-centre investigation involved 678 subjects (388 male, 290 female), age range 2 - 11 years: all subjects were followed-up for a minimum period of 2 years. Patients in this study population were submitted to 3 groups of investigations: a) in the first (487 children submitted to adeno-tonsillectomy), the study population was subdivided into 2 groups according to the frequency of the complaints of febrile pharyngo-tonsillar inflammation reported during the previous 12 months (> 4 or < or = 4); b) in the second (443 children with recurrent febrile pharyngo-tonsillar inflammation during the previous 12 months < or = 4), three groups of cases were compared; in the 1st (252 cases), patients were submitted to adeno-tonsillectomy; in the 2nd (144 subjects), adenoidectomy was carried out; in the 3rd (47 cases), the watchful waiting approach was adopted; c) in the third (193 cases, with both pathological auricular manifestations), patients were submitted to adeno-tonsillectomy (119 cases) or to adenoidectomy 74 cases); during surgery, ventilation tube was applied bilaterally in 61 cases, while 132 patients formed the control group. The results obtained were statistically analysed by chi2 and Fisher test: a "p" value of < 0.05 was considered statistically significant. RESULTS: The first investigation showed: a) the incidence of cases with auricular disorders was greater in the group presenting recurrent febrile pharyngo-tonsillar inflammation < or = 4 (effusive otitis media: p < 0.05); in this group, the presence of cases with naso-pharyngeal respiratory obstruction was statistically greater (p < 0.05); b) the number of cases in which the pathological auricular process was resolved following adeno-tonsillectomy, appeared high (effusive otitis media: 136/179 = 76%; acute recurrent otitis media: 189/216 = 87.5%), not unlike the percentage of patients with positive results on recurrent febrile pharyngo-tonsillar inflammation and on naso-pharyngeal respiratory obstruction (91.6% and 92.4%, respectively); c) the percentages of recovery, for auricular disorders, were significantly higher in the group presenting more frequent recurrent febrile pharyngo-tonsillar inflammation, i.e. > 4 (p < 0.05); in this group, also the incidence of cases, in which the naso-pharyngeal respiratory obstruction was resolved, was greater. The second investigation demonstrated, in the groups in which surgical treatment was carried out, compared to controls: a) a significantly greater percentage of cases achieving recovery, for both parameters (p < 0.05); as far as concerns the effusive form of otitis media, a total of 72 (67.3%) subjects were cured following adeno-tonsillectomy; 67 (81.7%), following adenoidectomy; in acute recurrent otitis media, the subjects achieving recovery were; for adeno-tonsillectomy, 97 (82.2%); for adenoidectomy; 85 (81.7%); b) in the subjects in whom, following surgical treatment, the inflammatory auricular disorder was resolved, it was generally found that recovery had been achieved not only for the recurrent febrile pharyngo-tonsillar inflammation but also for the naso-pharyngeal respiratory obstruction; c) in the controls, the results appeared to be significantly less satisfactory (< 0.001); together with the lack of success in the watchful waiting approach, as far as concerns auricular disorders, we found, likewise, a significant lack of efficacy (p < 0.001) of this treatment in the management of naso-pharyngeal respiratory obstruction (5/35 cases cured: 14.3%). The third investigation showed that the application of the ventilation tube during adeno-tonsillectomy or adenoidectomy did not lead to any improvement in the results; indeed, the percent recovery of the pathological tympanic processes was significantly greater (p < 0.05) in those subjects in whom transtympanic drainage had not been adopted. CONCLUSIONS: The results of these investigations reveal a clear relationship between the naso-pharyngeal respiratory obstruction and the pathological auricular processes in subjects with episodes of recurrent febrile pharyngo-tonsillar inflammation. These results also demonstrate that adeno-tonsillectomy and/or adenoidectomy are able to resolve the auricular inflammatory manifestations in a very large percentage of cases. These data are apparently in contrast with those described in recent reports related to randomised investigations. In effect, the two groups of investigations had different aims: the studies carried out according to a randomised approach were performed on study populations in which the findings of pathological auricular manifestations were not linked by the demonstration of a correlation with possible adenoid or adeno-tonsillar inflammation; the present study, on the other hand, was performed on patients in whom the auricular disorder was part of the clinical picture of recurrent febrile pharyngo-tonsillar inflammation. Therefore we believe that each of these study methods has precise possibilities, but also limitations; thus, one should not exclude the other, but rather be integrated.

Implant surface and osseointegration.

Oral rehabilitation by means of osteointegrated implants is gaining more and more importance, in particular due to the improved characteristics of the fixture: among these, the superficial microscopical morphology plays a role of primary importance, since it can influence the evolution of the osteointegrative process. Therefore, this led to many surface treatments which can improve the quality and the speed of formation of the bony tissue. The aim of this paper is a review of the literature on this topic in order to find out which processes of characterization of the surface implant are now available and which results can be obtained.

Drugs in oral surgery. Brief guidelines for adult patients.

Drugs administrable in oral surgery for adult patients are antiseptics-antibiotics, antiinflammatory-analgesics and sedative-hypnotics. Such drugs can be administered before, during or after oral surgery. Sedative-hypnotics can be administered before or during oral surgery in order to control the patient's anxiety. Anti-inflammatory-analgesics, on the other hand, can be administered before or after oral surgery to lower edema and pain. For this purpose, FANS are the most commonly used drugs but, in more traumatic oral surgery, the administration of a single pre-surgery dose of corticosteroids is suitable. As regards, antibiotics have to be given from 15 min to 1 h before oral surgery and continued or otherwise for 24-48 h depending on the dosage. post-surgery infection onset, in fact, is higher within 3 h after oral surgery.

Synthesis and binding mode of heterocyclic analogues of suramin inhibiting the human basic fibroblast growth factor.

The design, synthesis, and biological evaluation of a series of pyrrole and pyrazole congeners 2 of suramin, directed toward the development and identification of new ligands that complex the human fibroblast growth factor (bFGF), thereby inhibiting tumor-promoted angiogenesis, is reported. Compounds 2 were evaluated for their ability to inhibit binding of bFGF to its receptor, in vivo bFGF-induced angiogenesis, and neovascularization of the chorioallantoic membrane in comparison with suramin. These assays showed that ligands 2 exhibit moderate to good activity, comparable to that of suramin, and are less toxic than suramin itself. In this study, affinity data of ligands in combination with the crystal structure of bFGF were used to explain structure-affinity relationships and to gain an insight into the possible mode of ligand-protein interaction. Due to the lack of experimental structural data on the ligand-bFGF complexes, molecular mechanics techniques were used to obtain putative bioactive conformations and to generate docked complexes with the three-dimensional structure of bFGF. These experiments led to suggest that compounds 2 give rise to 1:1 complexes with bFGF through an unprecedented, bidentate attachment of their naphthylsulfonate groups to two main domains, commonly referred to as the heparin binding site and the receptor binding site, on bFGF, thus preventing the interaction of the growth factor with its receptor.