Development of Vapor Deposited Silica Sol-Gel Particles for a Bioactive Materials System to Direct Osteoblast Behavior

Tissue engineering and regenerative medicine have emerged in an effort to generate replacement tissues capable of restoring native tissue structure and function, but because of the complexity of biologic system, this has proven to be much harder than originally anticipated. Silica based bioactive glasses are popular as biomaterials because of their ability to enhance osteogenesis and angiogenesis. Sol-gel processing methods are popular in generating these materials because it offers: 1) mild processing conditions; 2) easily controlled structure and composition; 3) the ability to incorporate biological molecules; and 4) inherent biocompatibility. The goal of this work was to develop a bioactive vaporization system for the deposition of silica sol-gel particles as a means to modify the material properties of a substrate at the nano- and micro- level to better mimic the instructive conditions of native bone tissue, promoting appropriate osteoblast attachment, proliferation, and differentiation as a means for supporting bone tissue regeneration. The size distribution, morphology and degradation behavior of the vapor deposited sol-gel particles developed here were found to be dependent upon formulation (H2O:TMOS, pH, Ca/P incorporation) and manufacturing (substrate surface character, deposition time). Additionally, deposition of these particles onto substrates can be used to modify overall substrate properties including hydrophobicity, roughness, and topography. Deposition of Ca/P sol particles induced apatite-like mineral formation on both two- and three-dimensional materials when exposed to body fluids. Gene expression analysis suggests that Ca/P sol particles induce upregulation osteoblast gene expression (Runx2, OPN, OCN) in preosteoblasts during early culture time points. Upon further modification-specifically increasing particle stability-these Ca/P sol particles possess the potential to serve as a simple and unique means to modify biomaterial surface properties as a means to direct osteoblast differentiation.

Crossover comparison of the laryngeal mask supreme and the i-gel in simulated difficult airway scenario in anesthetized patients

BACKGROUND: The single-use supraglottic airway devices LMA-Supreme (LMA-S; Laryngeal Mask Company, Henley-on-Thames, United Kingdom) and i-gel (Intersurgical Ltd, Wokingham, Berkshire, United Kingdom) have a second tube for gastric tube insertion. Only the LMA-S has an inflatable cuff. They have the same clinical indications and might be useful for difficult airway management. This prospective, crossover, randomized controlled trial was performed in a simulated difficult airway scenario using an extrication collar limiting mouth opening and neck movement. METHODS: Sixty patients were included. Both devices were placed in random order in each patient. Primary outcome was overall success rate. Other measurements were time to successful ventilation, airway leak pressure, fiberoptic glottic view, and adverse events. RESULTS: Success rate for the LMA-S was 95% versus 93% for the i-gel (P = 1.000). LMA-S needed shorter insertion time (34 +/- 12 s vs. 42 +/- 23 s, P = 0.024). Tidal volumes and airway leak pressure were similar (LMA-S 26 +/- 8 cm H20; i-gel 27 +/- 9 cm H20; P = 0.441). Fiberoptic view through the i-gel showed less epiglottic downfolding. Overall agreement in insertion outcome was 54 (successes) and 1 (failure) or 55 (92%) of 60 patients. The difference in success rate was 1.7% (95% CI -11.3% to 7.6%). CONCLUSIONS: Both airway devices had similar insertion success and clinical performance in the simulated difficult airway situation. The authors found less epiglottic downfolding and better fiberoptic view but longer insertion time with the i-gel. Our study shows that both devices are feasible for emergency airway management in patients with reduced neck movement and limited mouth opening.

Genotyping of Francisella tularensis strains by pulsed-field gel electrophoresis, amplified fragment length polymorphism fingerprinting, and 16S rRNA gene sequencing

We evaluated three molecular methods for identification of Francisella strains: pulsed-field gel electrophoresis (PFGE), amplified fragment length polymorphism (AFLP) analysis, and 16S rRNA gene sequencing. The analysis was performed with 54 Francisella tularensis subsp. holarctica, 5 F. tularensis subsp. tularensis, 2 F. tularensis subsp. novicida, and 1 F. philomiragia strains. On the basis of the combination of results obtained by PFGE with the restriction enzymes XhoI and BamHI, PFGE revealed seven pulsotypes, which allowed us to discriminate the strains to the subspecies level and which even allowed us to discriminate among some isolates of F. tularensis subsp. holarctica. The AFLP analysis technique produced some degree of discrimination among F. tularensis subsp. holarctica strains (one primary cluster with three major subclusters and minor variations within subclusters) when EcoRI-C and MseI-A, EcoRI-T and MseI-T, EcoRI-A and MseI-C, and EcoRI-0 and MseI-CA were used as primers. The degree of similarity among the strains was about 94%. The percent similarities of the AFLP profiles of this subspecies compared to those of F. tularensis subsp. tularensis, F. tularensis subsp. novicida, and F. philomiragia were less than 90%, about 72%, and less than 24%, respectively, thus permitting easy differentiation of this subspecies. 16S rRNA gene sequencing revealed 100% similarity for all F. tularensis subsp. holarctica isolates compared in this study. These results suggest that although limited genetic heterogeneity among F. tularensis subsp. holarctica isolates was observed, PFGE and AFLP analysis appear to be promising tools for the diagnosis of infections caused by different subspecies of F. tularensis and suitable techniques for the differentiation of individual strains.

Synthesis of bone-like micro-porous calcium phosphate/iota-carrageenan composites by gel diffusion

Brushite and octacalcium phosphate (OCP) crystals are well-known precursors of hydroxylapatite (HAp), the main mineral found in bone. In this report, we present a new method for biomimicking brushite and OCP using single and double diffusion techniques. Brushite and OCP crystals were grown in an iota-carrageenan gel. The aggregates were analyzed by scanning electron microscopy (SEM), X-ray diffraction (XRD), infrared spectroscopy (IR) and thermal gravimetric analysis (TGA). SEM revealed different morphologies of brushite crystals from highly porous aggregates to plate-shaped forms. OCP crystals grown in iota-carrageenan showed a porous spherical shape different from brushite growth forms. The XRD method demonstrated that the single-diffusion method favors the formation of monoclinic brushite. In contrast, the double diffusion method was found to promote the formation of the triclinic octacalcium phosphate OCP phase. By combining the different parameters for crystal growth in carrageenan, such as ion concentration, gel pH and gel density, it is possible to modify the morphology of composite crystals, change the phase of calcium phosphate and modulate the amount of carrageenan inclusion in crystals. This study suggests that iota-carrageenan is a high-molecular-weight polysaccharide that is potentially applicable for controlling calcium phosphate crystallization.

Identification of the Causative Bacteria in Musculoskeletal Infections Using 16s rDNA - Denaturing Gradient Gel Electrophoresis Analysis

Musculoskeletal infections are infections of the bone and surrounding tissues. They are currently diagnosed based on culture analysis, which is the gold standard for pathogen identification. However, these clinical laboratory methods are frequently inadequate for the identification of the causative agents, because a large percentage (25-50%) of confirmed musculoskeletal infections are false negatives in which no pathogen is identified in culture. My data supports these results. The goal of this project was to use PCR amplification of a portion of the 16S rRNA gene to test an alternative approach for the identification of these pathogens and to assess the diversity of the bacteria involved. The advantages of this alternative method are that it should increase sample sensitivity and the speed of detection. In addition, bacteria that are non-culturable or in low abundance can be detected using this molecular technique. However, a complication of this approach is that the majority of musculoskeletal infections are polymicrobial, which prohibits direct identification from the infected tissue by DNA sequencing of the initial 16S rDNA amplification products. One way to solve this problem is to use denaturing gradient gel electrophoresis (DGGE) to separate the PCR products before DNA sequencing. Denaturing gradient gel electrophoresis (DGGE) separates DNA molecules based on their melting point, which is determined by their DNA sequence. This analytical technique allows a mixture of PCR products of the same length that electrophoreses through agarose gels as one band, to be separated into different bands and then used for DNA sequence analysis. In this way, the DGGE allows for the identification of individual bacterial species in polymicrobial-infected tissue, which is critical for improving clinical outcomes. By combining the 16S rDNA amplification and the DGGE techniques together, an alternative approach for identification has been used. The 16S rRNA gene PCR-DGGE method includes several critical steps: DNA extraction from tissue biopsies, amplification of the bacterial DNA, PCR product separation by DGGE, amplification of the gel-extracted DNA, and DNA sequencing and analysis. Each step of the method was optimized to increase its sensitivity and for rapid detection of the bacteria present in human tissue samples. The limit of detection for the DNA extraction from tissue was at least 20 Staphylococcus aureus cells and the limit of detection for PCR was at least 0.05 pg of template DNA. The conditions for DGGE electrophoreses were optimized by using a double gradient of acrylamide (6 – 10%) and denaturant (30-70%), which increased the separation between distinct PCR products. The use of GelRed (Biotium) improved the DNA visualization in the DGGE gel. To recover the DNA from the DGGE gels the gel slices were excised, shredded in a bead beater, and the DNA was allowed to diffuse into sterile water overnight. The use of primers containing specific linkers allowed the entire amplified PCR product to be sequenced and then analyzed. The optimized 16S rRNA gene PCR-DGGE method was used to analyze 50 tissue biopsy samples chosen randomly from our collection. The results were compared to those of the Memorial Hermann Hospital Clinical Microbiology Laboratory for the same samples. The molecular method was congruent for 10 of the 17 (59%) culture negative tissue samples. In 7 of the 17 (41%) culture negative the molecular method identified a bacterium. The molecular method was congruent with the culture identification for 7 of the 33 (21%) positive cultured tissue samples. However, in 8 of the 33 (24%) the molecular method identified more organisms. In 13 of the 15 (87%) polymicrobial cultured tissue samples the molecular method identified at least one organism that was also identified by culture techniques. Overall, the DGGE analysis of 16S rDNA is an effective method to identify bacteria not identified by culture analysis.

Genetic Relatedness, Antimicrobial and Biocide Susceptibility Comparative Analysis of Methicillin-Resistant and -Susceptible Staphylococcus pseudintermedius from Portugal

Forty methicillin-resistant and -susceptible Staphylococcus pseudintermedius (MRSP and MSSP, respectively) from colonization and infection in dogs and cats were characterized for clonality, antimicrobial, and biocide susceptibility. MSSP were genetically more diverse than MRSP by multi-locus sequence typing and pulsed-field gel electrophoresis. Three different spa types (t06, t02, t05) and two SCCmec types (II-III and V) were detected in the MRSP isolates. All MRSP and two MSSP strains were multidrug-resistant. Several antibiotic resistance genes (mecA, blaZ, tet(M), tet(K), aac(6')-Ie-aph(2')-Ia, aph(3')-III, ant(6)-Ia, sat4, erm(B), lnu(A), dfr(G), and catpC221) were identified by microarray and double mutations in the gyrA and grlA genes and a single mutation in the rpoB gene were detected by sequence analysis. No differences were detected between MSSP and MRSP in the chlorhexidine acetate (CHA) minimum inhibitory concentrations (MICs). However, two MSSP had elevated MIC to triclosan (TCL) and one to benzalkonium chloride and ethidium bromide. One MSSP isolate harboured a qacA gene, while in another a qacB gene was detected. None of the isolates harboured the sh-fabI gene. Three of the biocide products studied had high bactericidal activity (Otodine(®), Clorexyderm Spot Gel(®), Dermocanis Piocure-M(®)), while Skingel(®) failed to achieve a five log reduction in the bacterial counting. S. pseudintermedius have become a serious therapeutic challenge in particular if methicillin- resistance and/or multidrug-resistance are involved. Biocides, like CHA and TCL, seem to be clinically effective and safe topical therapeutic options.

Maxillary sinus grafting with a synthetic, nanocrystalline hydroxyapatite-silica gel in humans: histologic and histomorphometric results.

The aim of this study was to evaluate in humans the amount of new bone after sinus floor elevation with a synthetic bone substitute material consisting of nanocrystalline hydroxyapatite embedded in a highly porous silica gel matrix. The lateral approach was applied in eight patients requiring sinus floor elevation to place dental implants. After elevation of the sinus membrane, the cavities were filled with 0.6-mm granules of nanocrystalline hydroxyapatite mixed with the patient's blood. A collagen membrane (group 1) or a platelet-rich fibrin (PRF) membrane (group 2) was placed over the bony window. After healing periods between 7 and 11 months (in one case after 24 months), 16 biopsy specimens were harvested with a trephine bur during implant bed preparation. The percentage of new bone, residual filler material, and soft tissue was determined histomorphometrically. Four specimens were excluded from the analysis because of incomplete biopsy removal. In all other specimens, new bone was observed in the augmented region. For group 1, the amount of new bone, residual graft material, and soft tissue was 28.7% ± 5.4%, 25.5% ± 7.6%, and 45.8% ± 3.2%, respectively. For group 2, the values were 28.6% ± 6.90%, 25.7% ± 8.8%, and 45.7% ± 9.3%, respectively. All differences between groups 1 and 2 were not statistically significant. The lowest and highest values of new bone were 21.2% and 34.1% for group 1 and 17.4% and 37.8% for group 2, respectively. The amount of new bone after the use of nanocrystalline hydroxyapatite for sinus floor elevation in humans is comparable to values found in the literature for other synthetic or xenogeneic bone substitute materials. There was no additional beneficial effect of the PRF membrane over the non-cross-linked collagen membrane.

Mucosal Erosion of the Cricoid Cartilage After the Use of an i-Gel Supraglottic Airway Device in a Patient with Diffuse Idiopathic Skeletal Hyperostosis

After standard hip arthroplasty, an 82-year-old patient with previously undiagnosed diffuse idiopathic skeletal hyperostosis of the cervical spine experienced life-threatening side effects after use of a supraglottic airway device (i-gel). Extensive mucosal erosion and denudation of the cricoid cartilage caused postoperative supraglottic swelling and prolonged respiratory failure requiring tracheostomy. In this case report, we highlight the importance of evaluating risk factors for failure of supraglottic airway devices.

Antiseptic solutions modulate the paracrine-like activity of bone chips: differential impact of chlorhexidine and sodium hypochlorite

AIM: Chemical decontamination increases the availability of bone grafts; however, it is unclear whether antiseptic processing changes the biological activity of bone. MATERIALS AND METHODS: Bone chips were incubated with 4 different antiseptic solutions including (1) povidone-iodine (0.5%), (2) chlorhexidine diguluconate (0.2%), (3) hydrogen peroxide (1%) and (4) sodium hypochlorite (0.25%). After 10 minutes of incubation, changes in the capacity of the bone-conditioned medium to modulate gene expression of gingival fibroblasts was investigated. RESULTS: Conditioned medium obtained from freshly prepared bone chips increased the expression of TGF-β target genes interleukin 11 (IL11), proteoglycan4 (PRG4), NADPH oxidase 4 (NOX4), and decreased the expression of adrenomedullin (ADM), and pentraxin 3 (PTX3) in gingival fibroblasts. Incubation of bone chips with 0.2% chlorhexidine, followed by vigorously washing resulted in a bone-conditioned medium with even higher expression of IL11, PRG4, and NOX4. These findings were also found with a decrease in cell viability and an activation of apoptosis signaling. Chlorhexidine alone, at low concentrations, increased IL11, PRG4 and NOX4 expression, independent of the TGF-β receptor I kinase activity. In contrast, 0.25% sodium hypochlorite almost entirely abolished the activity of bone-conditioned medium, while the other two antiseptic solutions, 1% hydrogen peroxide and 0.5% povidone-iodine, had relatively no impact, respectively. CONCLUSION: These in vitro findings demonstrate that incubation of bone chips with chlorhexidine differentially affects the activity of the respective bone-conditioned medium compared to the other antiseptic solutions. The data further suggest that the main effects are caused by chlorhexidine remaining in the bone-conditioned medium after repeated washing of the bone chips. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved. KEYWORDS: Autografts; TGF-β; antiseptic solution; bone; bone conditioned medium; bone supernatant; chlorhexidine; hydrogen peroxide; povidone-iodine; sodium hypochlorite

Comparison of the capacity of enamel matrix derivative gel and enamel matrix derivative in liquid formulation to adsorb to bone grafting materials.

BACKGROUND The use of an enamel matrix derivative (EMD) has been shown to enhance periodontal regeneration (e.g., formation of root cementum, periodontal ligament, and alveolar bone). However, in certain clinical situations, the use of EMD alone may not be sufficient to prevent flap collapse or provide sufficient stability of the blood clot. Data from clinical and preclinical studies have demonstrated controversial results after application of EMD combined with different types of bone grafting materials in periodontal regenerative procedures. The aim of the present study is to investigate the adsorption properties of enamel matrix proteins to bone grafts after surface coating with either EMD (as a liquid formulation) or EMD (as a gel formulation). METHODS Three different types of grafting materials, including a natural bone mineral (NBM), demineralized freeze-dried bone allograft (DFDBA), or a calcium phosphate (CaP), were coated with either EMD liquid or EMD gel. Samples were analyzed by scanning electron microscopy or transmission electron microscopy (TEM) using an immunostaining assay with gold-conjugated anti-EMD antibody. Total protein adsorption to bone grafting material was quantified using an enzyme-linked immunosorbent assay (ELISA) kit for amelogenin. RESULTS The adsorption of amelogenin to the surface of grafting material varied substantially based on the carrier system used. EMD gel adsorbed less protein to the surface of grafting particles, which easily dissociated from the graft surface after phosphate-buffered saline rinsing. Analyses by TEM revealed that adsorption of amelogenin proteins were significantly farther from the grafting material surface, likely a result of the thick polyglycolic acid gel carrier. ELISA protein quantification assay demonstrated that the combination of EMD liquid + NBM and EMD liquid + DFDBA adsorbed higher amounts of amelogenin than all other treatment modalities. Furthermore, amelogenin proteins delivered by EMD liquid were able to penetrate the porous surface structure of NBM and DFDBA and adsorb to the interior of bone grafting particles. Grafting materials coated with EMD gel adsorbed more frequently to the exterior of grafting particles with little interior penetration. CONCLUSIONS The present study demonstrates a large variability of adsorbed amelogenin to the surface of bone grafting materials when enamel matrix proteins were delivered in either a liquid formulation or gel carrier. Furthermore, differences in amelogenin adsorption were observed among NBM, DFDBA, and biphasic CaP particles. Thus, the potential for a liquid carrier system for EMD, used to coat EMD, may be advantageous for better surface coating.

Estradiol-induced alopecia in five dogs after contact with a transdermal gel used for the treatment of postmenopausal symptoms in women

BACKGROUND Noninflammatory alopecia is a frequent problem in dogs. Estrogen-induced alopecia is well described in dogs, with estrogen producing testicular tumors and canine female hyperestrogenism. OBJECTIVES To increase awareness that extensive alopecia in dogs can be caused by exposure to estradiol gel used by owners to treat their postmenopausal symptoms. ANIMALS Skin biopsies from five dogs with extensive alopecia were examined. METHODS Owners were asked for a thorough case history, including possible exposure to an estradiol gel. Complete blood work and serum chemistry panel analysis were performed to investigate possible underlying causes. Formalin-fixed skin biopsy samples were obtained from lesional skin and histopathology was performed. RESULTS All owners confirmed the use of a transdermal estradiol gel and close contact with the affected dogs before development of alopecia. Histopathologic examination showed a similar picture in all five dogs. Most hair follicles were predominantly either in kenogen or telogen and hair follicle infundibula showed mild to moderate dilation. Hair regrowth was present in all five dogs after the exposure to the estradiol gel was stopped or minimized. Blood work and serum chemistry panel were within normal limits in all cases. One dog had elevated estradiol concentrations, whereas in another dog estradiol concentrations were within normal limits. CONCLUSION AND CLINICAL IMPORTANCE Alopecia can occur after contact with a transdermal gel used as treatment for postmenopausal symptoms in women. Estradiol gel used by female owners therefore represents a possible cause for noninflammatory alopecia in dogs. Estradiol concentrations are not necessarily elevated in affected dogs.

Flöh Haz Weiber Traz : Der wunder unrichtige, und spotwichtige Rechtshandel der Flöh mit den Weibern Ain Neu geläß, auf das vber kurtzweiligst zu belachen ...

Durch Hultrich Elloposcleron, auf ain neues abgestosen vnd behabelt. ...