Collagen-chondroitin sulfate-based PLLA-SAIB-coated rhBMP-2 delivery system for bone repair

Bone morphogenetic proteins (BMPs) are osteoinductive proteins used intensively in clinical investigations involving various bone-related treatments. Owing to their high potential in new bone formation they require local application at the treatment site. For this purpose various controlled delivery systems with BMPs as the excipients have been prepared in recent years. Focusing on this clinical need a disc-shaped BMP carrier was designed as a local delivery system using soluble collagen and chondroitin sulfate. In situ release studies carried out with a model protein (FITC-labeled Protein A) presented a very high rate of release; with most of the protein content being released within 24 h. This rate could be decreased by providing a poly(L-lactide) (PLLA) and sucrose acetate isobutyrate-based (SAIB-based) coat around the release system, applied after BMP loading. In this way, it was possible to extend the release period from 24 h to about 12 days. In situ release of BMP from the same carriers, as quantitated using an ELISA kit, was even slower, with 50% of the protein being released in 15 days. In order to be able to secure the BMP delivery system at the bone defect site and to provide support a mesh knitted using Vicryl sutures and bonded with poly(L-lactide-co-glycolide) (PLGA) was tested in in vivo. Two time periods, 1 and 3 weeks, were used to evaluate the healing process. Osteoinduction by the BMP carrier system was assessed by histology-based bone scoring and X-ray examinations. PLLA-SAIB-coated collagen discs containing BMP presented good biocompatibility and optimum osteogenic stimulation. Structural changes in histological micrographs at week 1 indicated dose-dependent periosteal ossification. At the end of week 3 histological findings with both BMP (1 and 2 microg) doses were almost the same.     

Neuronal nuclear antigen (NeuN): a new tool in the diagnosis of central neurocytoma

The use of neuronal nuclear antigen (NeuN) as a reliable neuronal marker in the differential diagnosis of clear cell neoplasms of the central nervous system was determined in a biopsy series of 23 cases. Immunohistochemical analyses were carried out by antisera against neuronal nuclear antigen, synaptophysin, neuron-specific enolase, microtubule-associated protein 2, and glial fibrillary acidic protein. All eight central neurocytomas were characteristically immunolabeled by NeuN. NeuN immunoreactivity was uniformly strong and basically located in the nuclei of neurocytes. Despite this uniform staining pattern of central neurocytomas, 12 cases of oligodendrogliomas and three cases of ependymoma were negative for NeuN. As the diagnostic criteria for central neurocytoma include immunohistochemical and/or ultrastructural evidence for neuronal differentiation, NeuN as a sensitive and specific neuronal marker in formalin-fixed, paraffin-embedded tissues may greatly facilitate the differential diagnosis of central neurocytomas.     

Diagnosis of gastric carcinoma by classification on feature projections

A new classification algorithm, called benefit maximizing classifier on feature projections (BCFP), is developed and applied to the problem of diagnosis of gastric carcinoma. The domain contains records of patients with known diagnosis through gastroscopy results. Given a training set of such records, the BCFP classifier learns how to differentiate a new case in the domain. BCFP represents a concept in the form of feature projections on each feature dimension separately. Classification in the BCFP algorithm is based on a voting among the individual predictions made on each feature. In the gastric carcinoma domain, a lesion can be an indicator of one of nine different levels of gastric carcinoma, from early to late stages. The benefit of correct classification of early levels is much more than that of late cases. Also, the costs of wrong classifications are not symmetric. In the training phase, the BCFP algorithm learns classification rules that maximize the benefit of classification. In the querying phase, using these rules, the BCFP algorithm tries to make a prediction maximizing the benefit. A genetic algorithm is applied to select the relevant features. The performance of the BCFP algorithm is evaluated in terms of accuracy and running time. The rules induced are verified by experts of the domain.     

Somatic mosaicism for a MECP2 mutation associated with classic Rett syndrome in a boy

Rett syndrome is a severe neurodevelopmental disorder that arises from mutations in the X-linked MECP2 gene. It is almost exclusively seen in girls due to the predominant occurrence of the mutations on the paternal X-chromosome, and also the early postnatal lethal effect of the disease causing mutations in hemizygous boys. We identified a boy with features of classic Rett syndrome who is mosaic for the truncating MECP2 mutation R270X. Chromosome analysis showed normal karyotype. These results indicate that a MECP2 mutation associated with Rett syndrome in females could lead to a similar phenotype in males as a result of somatic mosaicism.     

Effect of arachidonic acid on specific binding of [3H]naloxone to opioid receptors

The effects of arachidonic acid (AA) on binding of [3H]naloxone to the agonist and antagonist configurations of opioid receptors were investigated in rat brain. Equilibrium binding parameters of the agonist and antagonist configurations of the receptors were evaluated from homologue displacement data in the presence or absence of AA. Addition of AA at a concentration of 0.6 mM (1.5 mumole/mg of protein) reduced by 22% and 53% the maximal number of binding sites (Bmax) respectively in the absence or presence of 100 mM NaCl. Binding affinity (KD) was not altered significantly (P < 0.05) either in the presence or absence of 100 mM NaCl and AA. We conclude that AA mediated reduction in [3H]naloxone specific binding was chiefly due to a decrease in the number of binding sites.     

Poly(D,L-lactide/epsilon-caprolactone)/hydroxyapatite composites as bone filler: an in vivo study in rats

In this study, a novel composite bone substitute was implanted in animal models (rats) and their in vivo characteristics were examined. A D,L-lactide and E-caprolactone copolymer (Mw: 80,000; Mn:40,000, and PI:2.00) was synthesized by ring-opening polymerization of the respective dimers using stannous octoate as the catalyst. The final ratio of D,L-lactide to epsilon-caprolactone obtained by 1NMR was 60/40. Hydroxyapatite (HA) powder was loaded in the copolymer. The HA/copolymer ratio was 60/40 (w/w). These composites were easily shaped by hand. Animal tests were performed on mature wistar rats (n=30). Defects were created on the proximal, the thickest part of the femur. The bone defects of the first group were filled with polymer/HA composite, the second group filled with only HA and the third group was left empty. Histologic examination of bone tissues showed new bone formation around the yellow-green polymer/HA composite material in the first group of animals whereas no evidence of new bone growth was observed in other groups.     

Pneumatised superior turbinate: a common anatomic variation?

Summary Anatomic variations are important in preendoscopic CT evaluation of the paranasal sinuses. In this study, we investigated whether the superior turbinate could become pneumatised like the middle turbinate, pneumatisation of which is well-known. Images of 52 patients who underwent CT examination prior to endoscopic sinus surgery and who had normally aerated posterior ethmoidal cells and an unobscured nasal cavity were retrospectively evaluated. The patients were 12–68 years old (median age, 35 years); 28 were women and 24 were men. Pneumatisation of the superior turbinates was graded in two groups as minimal or marked. Pneumatisation was evident in 25 patients (48%). 13 unilateral (25%), 8 bilateral (15%) pneumatisations were detected in the group graded as minimal, whereas 1 unilateral and 3 bilateral pneumatisations were present in the markedly (8%) pneumatised group of patients. Superior turbinates were seemingly aerated through the posterior ethmoid cells. The superior turbinates can be pneumatised as the middle turbinate is a not frequent anatomic variation that should be taken into account in preendoscopic CT evaluation of the paranasal sinuses.

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La pneumatisation du cornet nasal supérieur : une variation anatomique courante ?

Résumé Les variations anatomiques sont importantes à connaitre dans le cadre du bilan tomodensitométrique précédant une chirurgie endoscopique des sinus paranasaux. Dans cette étude, nous avons recherché si le cornet nasal supérieur pouvait être pneumatisé comme l'est le cornet nasal moyen, entité anatomique bien connue. Les images de 52 patients ayant eu une tomodensitométrie avant chirurgie endoscopique et montrant des cellules ethmoïdales postérieures normalement pneumatisées et une cavité nasale nonopaque, ont été évaluées rétrospectivement. Les patients, 28 femmes et 24 hommes, étaient âgés de 12 à 68 ans (moyenne : 35 ans). La pneumatisation du cornet nasal supérieur a été classée en deux groupes; minime et prononçée. Cette pneumatisation était patente chez 25 patients (48 %). Treize pneumatisations unilatérales (25 %) et 8 pneumatisations bilatérales (15 %) furent retrouvées dans le groupe présentant une pneumatisation minime alors que 1 pneumatisation unilatérale et 3 bilatérales (8 %) étaient présentes dans le groupe dont la pneumatisation était plus marquée. Les cornets nasaux supérieurs semblaient être aérés par les cellules ethmoïdales postérieures. Les cornets nasaux supérieurs peuvent être pneumatisés, comme le sont les cornets nasaux moyens, avec une fréquence non-exceptionnelle. Cette variation anatomique mérite d'être prise en considération dans le bilan tomodensitométrique précédant toute chirurgie endoscopique des sinus paranasaux.

     

Enhancing effect of chitosan on peptide drug delivery across buccal mucosa

The buccal mucosa represents a potentially important topical route for delivery of peptide or protein drugs with some unique advantages such as the avoidance of hepatic first-pass metabolism and the acidity and protease activity encountered in the gastrointestinal tract. However, the bioavailabilities or relative potencies of intraorally administered peptides are usually quite low, unless permeabilizers are employed. Chitosan, a mucopolysaccharide of marine origin, has been claimed to act both as a bioadhesive and permeabilizer, making it a candidate system for mucosal drug delivery. In this study, the enhancement effect of chitosan in gel form for oral mucosa was investigated with a large bioactive peptide, transforming growth factor-beta (TGF-beta). Chitosan gel was prepared at 2% concentration in dilute lactic acid and TGF-beta was incorporated into the gel. The effect of chitosan as a permeabilizer was determined by measuring the flux of TGF-beta across porcine oral mucosa in an in vitro system. The localization of TGF-beta within the oral mucosa was determined by horizontal sectioning and counting. Chitosan was found to exert a marked permeabilizing effect on buccal mucosa for peptide drug.     

Immobilization of acetylcholinesterase and choline oxidase in/on pHEMA membrane for biosensor construction

In this study, acetylcholinesterase (AChE) and choline oxidase (ChO) were co-immobilized on poly(2-hydroxyethyl methacrylate) (pHEMA) membranes with the aim of using them in biosensor construction. pHEMA membranes were prepared with the addition of different salts in different HEMA: aqueous solution ratios and characterized in terms of porosity, thickness. permeability, and mechanical properties. Membranes prepared in the presence of SnCl4 were found to be superior in terms of porosity and permeability and were chosen as the immobilization matrix. Immobilization of the enzymes was achieved both by entrapment and surface attachment via epichlorohydrin (Epi) and Cibacron Blue F36A (CB) activation. The effect of immobilization on enzyme activity was evaluated by the comparison of Km and Vmax values for the free and immobilized bi-enzyme systems. The increase in Km was negligible (1.08-fold) for the bi-enzyme system upon immobilization on surface but was 2.12-fold upon entrapment. Specific activity of the free enzyme system was found to be 0.306 mV s(-1) microg(-1) ChO while it was 0.069 (4.43-fold decrease) for entrapped and 0.198 (1.54 fold decrease) for CB-Epi immobilized enzymes. The performance of immobilized enzymes in different buffer types, pH, and temperature conditions were evaluated. The best enzyme activity was obtained at pH 9.0. Activity of the enzymes was found to increase with increasing temperature (in the range 25-40 degrees C).