Reduction of cicatricial stenosis after canalplasty for auditory exostoses.

BACKGROUND: Canalplasty for auditory exostoses is reserved for symptomatic patients. This study reviewed the outcomes of our technique regarding cicatricial stenosis. METHOD: A chart review was conducted on patients undergoing canalplasty for auditory exostoses between 2002 and 2017. The surgical technique is described. RESULTS: The study comprised 43 adults (50 operated ears). Exostoses were bilateral in 40 cases (94 per cent) and occlusive in 33 (66 per cent). After drilling, the external auditory meatus was covered with a graft in 34 cases (68 per cent) and a silicone sheet was used in 32 (64 per cent). Cicatricial stenosis appeared in eight cases (16 per cent). Skin grafts were not used in six of these eight cases (p < 0.04), and silicone sheets were used only in one of these eight (p < 0.01). CONCLUSION: Canalplasty is challenging because of its potential complications. Our data showed that the use of skin grafts and silicone sheets to cover the bared external auditory meatus was associated with a lower rate of cicatricial stenosis.

The case for class II bacteriocins: A biophysical approach using "suicide probes" in receptor-free hosts to study their mechanism of action.

Class II bacteriocins are unmodified membrane-active peptides that act over a narrow spectrum of target bacteria. They bind a specific receptor protein on the membrane to form a pore, leading to membrane permeabilization and cell death. However, little is known about the molecular events triggering the pore formation after the bacteriocin recognizes the receptor. It is not clear yet if the pore is the same receptor forced into an open conformation or if the pore results from the bacteriocin insertion and oligomeric assembly in the lipid bilayer. In order to reveal which model is more suitable to explain the toxicity mechanism, in this work we use chimeric peptides, resulting from the fusion of the bitopic membrane protein EtpM with different class II bacteriocins: enterocin CRL35, pediocin PA-1 and microcin V. E. coli strains lacking the specific receptors for these bacteriocins were chosen as expression hosts. As these constructs display a lethal effect when they are heterologously expressed, they are called "suicide probes". The results suggest that, indeed, the specific receptor would act as a docking molecule more than as a structural piece of the pore, as long as the bacteriocin is somehow anchored to the membrane. These set of chimeric peptides also represent an in vivo system that allows to study the interaction of the bacteriocins with real bacterial membranes, instead of model membranes. Hence, the effects of these suicide probes in membrane fluidity and transmembrane potential were also assessed, using fluorescence spectroscopy. The data show that the different suicide probes are able to increase phospholipid order and depolarize the membranes of receptor-free bacterial cells.

Exogenous fibrolytic enzymes and recombinant bacterial expansins synergistically improve hydrolysis and in vitro digestibility of bermudagrass haylage.

Four experiments were conducted to examine the effects of a recombinant bacterial expansin-like protein (BsEXLX1) from Bacillus subtilis and a commercial exogenous fibrolytic enzyme (EFE) preparation for ruminants on hydrolysis of pure substrates (cellulose and xylan) and in vitro digestibility of bermudagrass haylage (BMH). Recombinant Escherichia coli BL21 strain was used to express BsEXLX1; the protein was purified using an affinity column. In experiment 1, carboxymethylcellulose, Whatman #1 filter paper (General Electric, Boston, MA) and oat-spelt xylan substrates were subjected to 4 treatments (1) sodium citrate buffer (control), (2) BsEXLX1 (162 µg/g of substrate), (3) EFE (2.3 mg/g of substrate), and (4) EFE + BsELX1 in 3 independent runs. Samples were incubated at optimal conditions for both additives (pH 5 and 50°C) or at ruminal (pH 6 and 39°C) or ambient (pH 6 and 25°C) conditions for 24 h and sugar release was measured. In experiment 2, digestibility in vitro of BMH was examined after treatment with the following: (1) control (buffer only), (2) BsEXLX1 (162 µg/g of dry matter), (3) EFE (2.2 mg/g of dry matter), and (4) EFE + BsEXLX1 in 3 independent runs at 39°C for 24 h. Experiment 3 examined effects of EFE and BsEXLX1 on simulated preingestive hydrolysis and profile of released sugars from BMH after samples were suspended in deionized water with sodium azide at 25°C for 24 h in 2 independent runs. In experiment 4, the sequence of the BsEXLX1 purified protein was compared with 447 ruminal bacterial genomes to identify similar proteins from the rumen. In experiment 1, compared with EFE alone, EFE and BsEXLX1 synergistically increased sugar release from carboxymethylcellulose and Whatman #1 filter paper under all simulated conditions; however, hydrolysis of xylan was not improved. In experiment 2, compared with EFE alone, treatment with EFE and BsEXLX1 increased neutral detergent fiber and acid detergent fiber digestibility of bermudagrass haylage (by 5.5 and 15%, respectively) and total volatile fatty acid concentrations, and decreased acetate-propionate ratio. In experiment 3, compared with EFE alone. The EFE and BsEXLX1 synergistically reduced concentrations of neutral detergent fiber and acid detergent fiber and increased release of sugars by 9.3%, particularly cellobiose (72.5%). In experiment 4, a similar sequence to that of BsEXLX1 was identified in Bacillus licheniformis, and similar hypothetical protein sequences were identified in Ruminococcus flavefaciens strains along with different protein structures in E. xylanophilum and Lachnospiraceae. This study showed that an expansin-like protein synergistically increased the hydrolysis of pure cellulose substrates and the hydrolysis and digestibility in vitro of BMH.

Blueberries as an additive to increase the survival of Lactobacillus johnsonii N6.2 to lyophilisation.

Effective cultivation methods, total cost, and biomass preservation are key factors that have a significant impact on the commercialisation and effectiveness of probiotics, such as Lactobacillus. Sugar polymers, milk and whey proteins have been suggested as good additives for industrial preparations. Alternative compounds, such as phytophenols, are a more attractive option, given their potential benefits to human health. The overall goal of this study was to determine if the addition of blueberry phytophenols improves the survival of Lactobacillus johnsonii N6.2 during the freeze-drying process. The addition of blueberry aqueous extract (BAE) stimulated the growth of L. johnsonii under aerobic conditions and improved the stationary phase survival of the bacteria. Furthermore, the addition of BAE to the culture media improved the endurance of L. johnsonii N6.2 to freeze-drying stress, as well as to storage at 4 °C for up to 21 weeks. Moreover, blueberry extract performed more effectively as a lyophilising additive compared to skim milk and microencapsulation with whey protein/sodium alginate. In sum, this study demonstrates that BAE is an effective additive to increase the growth and survival of L. johnsonii N6.2 when added to the culture medium and/or used as a lyophilising preservative. Moreover, BAE or other polyphenols sources might likely enhance growth and increase survival of more probiotic lactic acid bacterial strains.

Lactobacillus johnsonii N6.2 diminishes caspase-1 maturation in the gastrointestinal system of diabetes prone rats.

The cells of the gastrointestinal (GI) epithelium are the first to contact the microbiota and food components. As a direct consequence of this, these cells are the first line of defence and key players in priming the immune response. One of the first responses against GI insults is the formation of the inflammasome, a multiprotein complex assembled in response to environmental threats. The formation of the inflammasome regulates caspase-1 by cleaving it into its active form. Once activated, caspase-1 can cleave interleukin-1ß (IL-1ß), which promotes adaptive and humoral immunity. Some strains, like Lactobacillus johnsonii N6.2, are able to modulate the biosynthesis of important host metabolites mediating inflammation. Of these metabolites are the pro-inflammatory kynurenines. L. johnsonii N6.2 is able to downregulate kynurenines biosynthesis via a redox active mechanism negatively affecting indoleamine 2,3-dioxygenase activity. In this study, we evaluated the effects of L. johnsonii N6.2 combined with the natural antioxidant and anti-inflammatory molecule rosmarinic acid (RA). Inflammasome assembly and the kynurenine pathway were evaluated in GI samples of BioBreeding diabetes-prone (BB-DP) rats. In this work, BB-DP rats were fed daily with RA, L. johnsonii N6.2; or both combined. The transcriptional rate and proteins levels of inflammasome and kynurenine pathway components in ileum tissue were evaluated. Elevated levels of pro-caspase-1 were observed in rats fed with L. johnsonii, while RA had no effect on pro-caspase-1 expression. Western blot assays demonstrated that L. johnsonii fed rats showed lower levels of mature caspase-1, when compared to the other treatments. Furthermore, IL-1ß maturation followed a similar pattern across the treatments. Differences were also observed between treatments in expression levels of key enzymes in the kynurenine pathway. These findings support the role of L. johnsonii in modulating the assembly of the inflammasome as well as some steps of the pro-inflammatory kynurenine pathway.

In vitro and in vivo evaluation of a new zirconia/niobium biocermet for hard tissue replacement.

Metals and ceramics are commonly used in orthopaedics, dentistry and other load bearing applications. However, the use of ceramic matrix composites reinforced with biocompatible metals for heavy load-bearing hard tissue replacement applications has not previously been reported. In order to improve the reliability and the mechanical properties of biomedical implants, new zirconia-Nb composites have been recently developed. The aim of this study was to investigate the biological tolerance of these new zirconia/Nb biocermets implants with both in vitro and in vivo approaches. At first, human bone marrow derived mesenchymal stem cells were cultured on sintered biocermet discs with polished surfaces and were compared with responses to niobium metal. In vitro, the biocermets showed no deleterious effect on cell proliferation, extra-cellular matrix production or on cell morphology. Furthermore, the biocermet showed a higher percentage of cell proliferation than Nb metal. On the other hand, the bone response to these new zirconia/Nb biocermets was studied. Cylinders of biocermets, as well as commercially Nb rod were implanted in the tibiae of New Zealand white rabbits. All the animals were euthanatized after 6 months. The specimens were processed to obtain thin ground sections. The slides were observed in normal transmitted light microscope. A newly formed bone was observed in close contact with material surfaces. No inflamed or multinucleated cells were present. This study concluded that zirconia/Nb composites are biocompatible and osteoconductive. The ceramic-metal composite has even better osteointegration ability than pure Nb. In conclusion, zirconia-Nb biocermet is suitable for heavy load-bearing hard tissue replacement from the point of view of both mechanical properties and biocompatibility.

Effect of adding cofactors to exogenous fibrolytic enzymes on preingestive hydrolysis, in vitro digestibility, and fermentation of bermudagrass haylage.

Our objectives were to examine if adding metal ion cofactors (COF) to exogenous fibrolytic enzymes (EFE) would increase the beneficial effects of the EFE on the preingestive hydrolysis and in vitro digestibility and fermentation of bermudagrass haylage. In experiment 1, 5 COF (Mn(2+), Co(2+), Fe(2+), Ca(2+), and Mg(2+)) were screened to select the best candidates for synergistically enhancing release of water-soluble carbohydrates (WSC) from bermudagrass haylage by 5 EFE. The 5 EFE (1A, 2A, 11C, 13D, and 15D) were sourced from Trichoderma reesei and Aspergillus oryzae and they were the most effective of 12 EFE at increasing the neutral detergent fiber digestibility of bermudagrass haylage in a previous trial. Adding 1mM of each of the COF to EFE 2A or 11C synergistically increased release of WSC from bermudagrass haylage, as did adding (1mM) Fe(2+) to 1A, Mn(2+), Co(2+), or Fe(2+) to 13D, or Co(2+)or Fe(2+) to 15D. The greatest release of WSC responses were obtained by adding Mn(2+) to 11C (38%) or by adding Fe(2+) to 2A or 13D (10 and 21.9%, respectively). In experiment 2, the effect of increasing the COF dose on in vitro digestibility and fermentation of bermudagrass haylage was examined using the best EFE-COF combinations from experiment 1. Effects of adding increasing doses of these COF on EFE-mediated changes in vitro digestibility depended on the COF-EFE combination. Adding 10mM Mn(2+) alone to bermudagrass haylage increased DMD and NDFD by 2.7 and 6.3% and adding 11C alone increased these measures by 6.6 and 15.5%, respectively. However, adding 10mM Mn(2+) with 11C resulted in 3.5 and 8.1% increases in DMD and NDFD, respectively, beyond the increases caused by adding 11C alone. Adding Fe(2+) to 2A had no effects on EFE-mediated digestibility responses, but 2A prevented adverse effects of adding Fe(2+) alone on DMD and NDFD. In contrast, adding Fe(2+) to 13D reduced the increases in DMD and NDFD caused by adding the EFE alone. This study shows that adding COF to EFE can synergistically increase, decrease, or not affect the hydrolytic effects of EFE on bermudagrass haylage cell walls. The outcome depends on the specific EFE-COF combination and the COF dose. More research is required to understand the mechanisms resulting in these outcomes to exploit beneficial effects of COF on EFE.

Screening exogenous fibrolytic enzyme preparations for improved in vitro digestibility of bermudagrass haylage.

Our objectives were to evaluate the effects of 12 exogenous fibrolytic enzyme products (EFE) on ruminal in vitro neutral detergent fiber digestibility (NDFD) and preingestive hydrolysis of a 4-wk regrowth of bermudagrass haylage (BH), to examine the accuracy of predicting NDFD with EFE activity measures, and to examine the protein composition of the most and least effective EFE at increasing NDFD. In experiment 1, effects of 12 EFE on NDFD of BH were tested. Enzymes were applied in quadruplicate to culture tubes containing ground BH. The suspension was incubated for 24 h at 25 °C before addition of rumen fluid media and further incubation for 24 h at 39 °C. The experiment was repeated twice. In addition, regression relationships between EFE activity measures and NDFD were examined. Compared with the values for the control, 9 EFE-treated substrates had greater NDFD (37.8 to 40.4 vs. 35.6%), 6 had greater total VFA concentration (59.1 to 61.2 vs. 55.4 mM), and 4 had lower acetate-to-propionate ratios (3.03 to 3.16 vs. 3.24). In experiment 2, EFE effects on preingestive fiber hydrolysis were evaluated by incubating enzyme-treated and untreated bermudagrass suspensions in quadruplicate for 24 h at 25 °C and examining fiber hydrolysis measures. Compared with values for the control, 3 EFE reduced neutral detergent fiber concentration (62.8 to 63.7 vs. 67.3%), 10 increased release of water-soluble carbohydrates (26.8 to 58.5 vs. 22.8 mg/g), and 8 increased release of ferulic acid (210 to 391 vs. 198 µg/g). Regression analyses revealed that enzyme activities accurately [coefficient of determination (R(2)) = 0.98] predicted preingestive hydrolysis measures (water-soluble carbohydrates, ferulic acid), moderately (R(2) = 0.47) predicted neutral detergent fiber hydrolysis, but poorly (R(2) ≤ 0.1) predicted dry matter and NDFD. In experiment 3, proteomic tools were used to examine the protein composition of the most and least effective EFE at improving NDFD. Relative to the least effective, the most effective EFE at increasing NDFD contained 10 times more endoglucanase III, 17 times more acetylxylan esterase with a cellulose-binding domain 1, 33 times more xylanase III, 25 times more ß-xylosidase, and 7.7 times more polysaccharide monooxygenase with cellulose-binding domain 1 and 3 times more swollenin. The most effective EFE had a much greater quantity of fibrolytic enzymes and key proteins necessary for hemicellulose and lignocellulase deconstruction. This study identified several EFE that increased the NDFD and in vitro fermentation of 4-wk BH and revealed why some EFE are more effective than others.

Interfaces in graded coatings on titanium-based implants.

Graded bilayered glass-ceramic composite coatings on Ti6Al4V substrates were fabricated using an enameling technique. The layers consisted of a mixture of glasses in the CaO-MgO-Na(2)O-K(2)O-P(2)O(5) system with different amounts of calcium phosphates (CPs). Optimum firing conditions have been determined for the fabrication of coatings having good adhesion to the metal, while avoiding deleterious reactions between the glass and the ceramic particles. The final coatings do not crack or delaminate. The use of high-silica layers (>60 wt % SiO(2)) in contact with the alloy promotes long-term stability of the coating; glass-metal adhesion is achieved through the formation of a nanostructured Ti(5)Si(3) layer. A surface layer containing a mixture of a low-silica glass ( approximately 53 wt % SiO(2)) and synthetic hydroxyapatite particles promotes the precipitation of new apatite during tests in vitro. The in vitro behavior of the coatings in simulated body fluid depends both on the composition of the glass matrix and the CP particles, and is strongly affected by the coating design and the firing conditions.

Mechanism of chromate reduction by the Escherichia coli protein, NfsA, and the role of different chromate reductases in minimizing oxidative stress during chromate reduction.

Chromate [Cr(VI)] is a serious environmental pollutant, which is amenable to bacterial bioremediation. NfsA, the major oxygen-insensitive nitroreductase of Escherichia coli, is a flavoprotein that is able to reduce chromate to less soluble and less toxic Cr(III). We show that this process involves single-electron transfer, giving rise to a flavin semiquinone form of NfsA and Cr(V) as intermediates, which redox cycle, generating more reactive oxygen species (ROS) than a divalent chromate reducer, YieF. However, NfsA generates less ROS than a known one-electron chromate reducer, lipoyl dehydrogenase (LpDH), suggesting that NfsA employs a mixture of uni- and di-valent electron transfer steps. The presence of YieF, ChrR (another chromate reductase we previously characterized), or NfsA in an LpDH-catalysed chromate reduction reaction decreased ROS generation by c. 65, 40, or 20%, respectively, suggesting that these enzymes can pre-empt ROS generation by LpDH. We previously showed that ChrR protects Pseudomonas putida against chromate toxicity; here we show that NfsA or YieF overproduction can also increase the tolerance of E. coli to this compound.

Chromate-reducing properties of soluble flavoproteins from Pseudomonas putida and Escherichia coli.

Cr(VI) (chromate) is a toxic, soluble environmental contaminant. Bacteria can reduce chromate to the insoluble and less toxic Cr(III), and thus chromate bioremediation is of interest. Genetic and protein engineering of suitable enzymes can improve bacterial bioremediation. Many bacterial enzymes catalyze one-electron reduction of chromate, generating Cr(V), which redox cycles, generating excessive reactive oxygen species (ROS). Such enzymes are not appropriate for bioremediation, as they harm the bacteria and their primary end product is not Cr(III). In this work, the chromate reductase activities of two electrophoretically pure soluble bacterial flavoproteins--ChrR (from Pseudomonas putida) and YieF (from Escherichia coli)-were examined. Both are dimers and reduce chromate efficiently to Cr(III) (kcat/Km = approximately 2 x 10(4) M(-1) x s(-1)). The ChrR dimer generated a flavin semiquinone during chromate reduction and transferred >25% of the NADH electrons to ROS. However, the semiquinone was formed transiently and ROS diminished with time. Thus, ChrR probably generates Cr(V), but only transiently. Studies with mutants showed that ChrR protects against chromate toxicity; this is possibly because it preempts chromate reduction by the cellular one-electron reducers, thereby minimizing ROS generation. ChrR is thus a suitable enzyme for further studies. During chromate reduction by YieF, no flavin semiquinone was generated and only 25% of the NADH electrons were transferred to ROS. The YieF dimer may therefore be an obligatory four-electron chromate reducer which in one step transfers three electrons to chromate and one to molecular oxygen. As a mutant lacking this enzyme could not be obtained, the role of YieF in chromate protection could not be directly explored. The results nevertheless suggest that YieF may be an even more suitable candidate for further studies than ChrR.

Independent and Synergistic Activity of Synthetic Peptides Against Thiabendazole-Resistant Fusarium sambucinum.

ABSTRACT Two heptapeptides with broad antifungal activity were identified and assessed for their ability to act synergistically with thiabendazole. The hexapeptide 66-10 was the progenitor of the heptapeptides and exhibited minimal inhibitory concentrations (MICs) of 9.3 to 9.8 mug/ml for thiabendazole (TBZ) resistant Fusarium sambucinum strains (MIC of 186 to 312 mug/ml). Heptapeptide derivatives 77-3 and 77-12 exhibited MICs between 3.8 and 7.5 mug/ml against the same strains. Incubation of conidia or mycelia with the peptide 77-3 showed that treated fungal structures were stained by the membrane impermeant dye SYTOX Green indicating disruption of membranes. Conidia incubated with peptide 77-3 at 10 mug/ml showed a 91 +/- 3.6% reduction in viability in 15 min. A checkerboard method was used to test the peptides and TBZ individually and in combination to determine potential synergistic activity. The results indicate that small peptides can act synergistically with TBZ against TBZ-resistant F. sambucinum.

Increased differentiation of intracranial white matter lesions by multispectral 3D-tissue segmentation: preliminary results.

MRI is a very sensitive imaging modality, however with relatively low specificity. The aim of this work was to determine the potential of image post-processing using 3D-tissue segmentation technique for identification and quantitative characterization of intracranial lesions primarily in the white matter. Forty subjects participated in this study: 28 patients with brain multiple sclerosis (MS), 6 patients with subcortical ischemic vascular dementia (SIVD), and 6 patients with lacunar white matter infarcts (LI). In routine MR imaging these pathologies may be almost indistinguishable. The 3D-tissue segmentation technique used in this study was based on three input MR images (T(1), T(2)-weighted, and proton density). A modified k-Nearest-Neighbor (k-NN) algorithm optimized for maximum computation speed and high quality segmentation was utilized. In MS lesions, two very distinct subsets were classified using this procedure. Based on the results of segmentation one subset probably represent gliosis, and the other edema and demyelination. In SIVD, the segmented images demonstrated homogeneity, which differentiates SIVD from the heterogeneity observed in MS. This homogeneity was in agreement with the general histological findings. The LI changes pathophysiologically from subacute to chronic. The segmented images closely correlated with these changes, showing a central area of necrosis with cyst formation surrounded by an area that appears like reactive gliosis. In the chronic state, the cyst intensity was similar to that of CSF, while in the subacute stage, the peripheral rim was more prominent. Regional brain lesion load were also obtained on one MS patient to demonstrate the potential use of this technique for lesion load measurements. The majority of lesions were identified in the parietal and occipital lobes. The follow-up study showed qualitatively and quantitatively that the calculated MS load increase was associated with brain atrophy represented by an increase in CSF volume as well as decrease in "normal" brain tissue volumes. Importantly, these results were consistent with the patient's clinical evolution of the disease after a six-month period. In conclusion, these results show there is a potential application for a 3D tissue segmentation technique to characterize white matter lesions with similar intensities on T(2)-weighted MR images. The proposed methodology warrants further clinical investigation and evaluation in a large patient population.

A clinical comparison of the safety and efficacy of MultiHance (gadobenate dimeglumine) and Omniscan (Gadodiamide) in magnetic resonance imaging in patients with central nervous system pathology.

RATIONALE AND OBJECTIVES: The safety and diagnostic efficacy of MultiHance (gadobenate dimeglumine) in the central nervous system (CNS) were evaluated in a double-blind, multicenter, phase III clinical trial. METHODS: Two hundred five patients highly suspected of having a CNS lesion (by previous imaging exam) were enrolled at 16 sites in the United States. Patients were randomized to one of three incremental dosing regimens. Magnetic resonance imaging with Omniscan (gadodiamide) at doses of 0.1 and 0.3 mmol/kg was compared with MultiHance (gadobenate dimeglumine) at doses of 0.05 and 0.15 mmol/kg and at 0.1 and 0.2 mmol/kg. RESULTS: Compared with predose images alone, efficacy was demonstrated in each of the gadobenate dimeglumine and gadodiamide groups (single and cumulative doses) as indicated by the level of diagnostic information, number of lesions detected, and contrast-to-noise ratio measurements. The level of diagnostic information from gadobenate dimeglumine at 0.1 mmol/kg was equivalent to that with gadodiamide at the same dose. One of the two blinded reviewers found equivalence between the gadobenate dimeglumine 0.05 mmol/kg dose and gadodiamide at 0.1 mmol/kg. Both reviewers found the level of diagnostic information to be equivalent after the second dose of contrast for all three dosing regimens. The cumulative doses of gadobenate dimeglumine were well tolerated and as safe as gadodiamide. CONCLUSIONS: Gadobenate dimeglumine is comparable to gadodiamide in terms of safety and efficacy for imaging of CNS lesions, with a possible advantage in imaging applications owing to enhanced T1 relaxivity.

Fast tissue segmentation based on a 4D feature map in characterization of intracranial lesions.

The aim of this work was to develop a fast and accurate method for tissue segmentation in magnetic resonance imaging (MRI) based on a four-dimensional (4D) feature map and compare it with that derived from a 3D feature map. High-resolution MRI was performed in 5 normal individuals, in 12 patients with brain multiple sclerosis (MS), and 9 patients with malignant brain tumors. Three inputs (proton-density, T2-weighted fast spin-echo, and T1-weighted spin-echo MR images) were routinely utilized. As a fourth input, either magnetization transfer MRT was used or T1-weighted post-contrast MRI (in patients only). A modified k-nearest neighbor segmentation algorithm was optimized for maximum computation speed and high-quality segmentation. In that regard, we a) discarded the redundant seed points; b) discarded the points within 0.5 standard deviation from the cluster center that were non-overlapping with other tissue; and c) removed outlying seed points outside 5 times the standard deviation from the cluster center of each tissue class. After segmentation, a stack of color-coded segmented images was created. Our new technique utilizing all four MRI inputs provided better segmentation than that based on three inputs (P < 0.001 for MS and P < 0.001 for tumors). The tissues were smoother due to the reduction of statistical noise, and the delineation of the tissues became sharper. Details that were previously blurred or invisible now became apparent. In normal persons a detailed depiction of deep gray matter nuclei was obtained. In malignant tumors, up to five abnormal tissue types were identified: 1) solid tumor core, 2) cyst, 3) edema in white matter 4) edema in gray matter, and 5) necrosis. Delineation of MS plaque in different stages of demyelination became much sharper. In conclusion, the proposed methodology warrants further development and clinical evaluation.

Optimization of tissue segmentation of brain MR images based on multispectral 3D feature maps.

The purpose of this work was to optimize and increase the accuracy of tissue segmentation of the brain magnetic resonance (MR) images based on multispectral 3D feature maps. We used three sets of MR images as input to the in-house developed semi-automated 3D tissue segmentation algorithm: proton density (PD) and T2-weighted fast spin echo and, T1-weighted spin echo. First, to eliminate the random noise, non-linear anisotropic diffusion type filtering was applied to all the images. Second, to reduce the nonuniformity of the images, we devised and applied a correction algorithm based on uniform phantoms. Following these steps, the qualified observer "seeded" (identified training points) the tissue of interest. To reduce the operator dependent errors, cluster optimization was also used; this clustering algorithm identifies the densest clusters pertaining to the tissues. Finally, the images were segmented using k-NN (k-Nearest Neighborhood) algorithm and a stack of color-coded segmented images were created along with the connectivity algorithm to generate the entire surface of the brain. The application of pre-processing optimization steps substantially improved the 3D tissue segmentation methodology.

Moyamoya disease in the United States.

The epidemiology and radiological features of Moyamoya disease (MMD) in the US were investigated. This study encompassed 98 cases; 26 were newly collected from eight US institutions and 72 were previously reported in the US literature. The patients ranged in age from 6 months to 67 years with age peaks in the first, third and fourth decades. MMD was seen in various ethnic groups and females were more commonly involved (71%) than males. A specific etiology could not be determined in most cases but arteriosclerosis and use of oral contraceptives were occasional associations. On angiography and/or magnetic resonance angiography (MRA), carotid arterial stenosis or occlusion was seen bilaterally in 95 cases (97%) and unilaterally in three. On MR or MRA, internal carotid steno-occlusive lesions were well demonstrated in all cases but Moyamoya collateral vessels (MMVs) were visualized in only 65% of the patients. MMVs in the basal ganglia and thalami were best demonstrated on T1 weighted images. Parenchymal lesions were seen in all patients and were often bilateral. With advances in MR techniques and increasing awareness of diagnostic guidelines, MMD will be diagnosed more frequently than before in the US.

Mobilization, cloning, and sequence determination of a plasmid-encoded polygalacturonase from a phytopathogenic Burkholderia (Pseudomonas) cepacia.

Phytopathogenic strains of Burkholderia cepacia (synonym Pseudomonas cepacia) produce endopolygalacturonase, whereas strains of clinical and soil origin do not. Growth of a phytopathogenic strain (ATCC25416) at elevated temperatures resulted in nonpectolytic derivatives that were either cured of a resident plasmid or contained a plasmid of reduced mass. The resident 200-kb plasmid (pPEC320) in strain ATCC25416 was tagged with Tn5-Mob. The pPEC320::Tn5-Mob (pPEC321) plasmid was mobilized in B. cepacia strains of soil and clinical origin. Transconjugants containing pPEC321 expressed the endopolygalacturonase and showed differential activity on plant tissue. No evidence for self-transfer of pPEC320 or the tagged derivative was observed. A 285-kb cloned fragment from pPEC320 containing the plasmid-borne pehA gene was sequenced and compared to the pehA gene from Erwinia carotovora subsp. carotovora and Ralstonia solanacearum and the polygalacturonase sequence from Lycopersicon esculentum.

Synthetic peptide combinatorial libraries: a method for the identification of bioactive peptides against phytopathogenic fungi.

Synthetic combinatorial libraries were evaluated with an iterative process to identify a hexapeptide with broadspectrum activity against selected phytopathogenic fungi. A D-amino acid hexapeptide (FRLKFH) and pentapeptide (FRLHF) exhibited activity against Fusarium oxysporum f. sp. lycopersici, Rhizoctonia solani (anastomosis group 1), Ceratocystis fagacearum, and Pythium ultimum. The peptides showed no hemolytic or mutagenic activity. Fluorescent microscopy studies with a membrane impermeant dye indicated that fungal cytoplasmic membranes were compromised rapidly and that the nuclear membrane was also affected.

Hemodynamic changes in recurrent intracranial terminal aneurysm after endovascular treatment.

RATIONALE AND OBJECTIVES: Multiple cases of recurrence of aneurysms after endovascular treatment have been reported. The purpose of the current hemodynamic study was to identify changes in shear stress and pressure associated with the recurrence of terminal intracranial aneurysms after endovascular occlusion. METHODS: Using a finite element method, a pulsed flow with a non-Newtonian viscosity of blood was simulated within the aneurysm cavity. A recurrent terminal intracranial aneurysm of a patient originally treated with balloon occlusion was then studied. This was based on a physiologic pulsatile flow, which was observed in the middle cerebral artery. Before and after the balloon occlusion, local maximum wall shear stress and pressure drop at the neck of the aneurysm were calculated and compared with the normal shear stress. RESULTS: Although the maximum shear stress at the right neck of the aneurysm was significantly reduced after balloon insertion, it was still 2.5 times greater than the normal maximum shear stress. This was attributable to the presence of a portion of the aneurysmal neck (residual neck), which was not obliterated by the balloon. The balloon also helped to reduce the maximum pressure inside the aneurysm by approximately 15%. Hemodynamic changes in the residual aneurysm neck, where the shear stress and the pressure are high, and other factors may be responsible for the recurrence of aneurysms after balloon or coil occlusion. CONCLUSION: Residual necks after balloon occlusion, coil occlusion, or both are attributable to the geometric orientation of the aneurysm with respect to parent and daughter vessels and the variety of configurations of the balloon and coils used to occlude the aneurysms. Inadequate reduction in local shear stress found in these residual necks is an important factor in the recurrence and rupture of the aneurysm after endovascular occlusion.