Lipidomic analysis identifies age-disease-related changes and potential new biomarkers in brain-derived extracellular vesicles from metachromatic leukodystrophy mice.

BACKGROUND: Recent findings show that extracellular vesicle constituents can exert short- and long-range biological effects on neighboring cells in the brain, opening an exciting avenue for investigation in the field of neurodegenerative diseases. Although it is well documented that extracellular vesicles contain many lipids and are enriched in sphingomyelin, cholesterol, phosphatidylserines and phosphatidylinositols, no reports have addressed the lipidomic profile of brain derived EVs in the context of Metachromatic Leukodystrophy, a lysosomal storage disease with established metabolic alterations in sulfatides. METHODS: In this study, we isolated and characterized the lipid content of brain-derived EVs using the arylsulfatase A knockout mouse as a model of the human condition. RESULTS: Our results suggest that biogenesis of brain-derived EVs is a tightly regulated process in terms of size and protein concentration during postnatal life. Our lipidomic analysis demonstrated that sulfatides and their precursors (ceramides) as well as other lipids including fatty acids are altered in an age-dependent manner in EVs isolated from the brain of the knockout mouse. CONCLUSIONS: In addition to the possible involvement of EVs in the pathology of Metachromatic Leukodystrophy, our study underlines that measuring lipid signatures in EVs may be useful as biomarkers of disease, with potential application to other genetic lipidoses.

Cost Awareness of Common Supplies Is Severely Impaired Among All Members of the Surgical Team.

BACKGROUND: Increased health care spending concerns have generated interest in reducing operating room (OR) costs, but the cost awareness of the surgical team selecting intraoperative supplies remains unclear. This work characterizes knowledge of supply cost among surgeons and OR staff in a large academic hospital and seeks to examine the role of experience and training with regards to cost insight. METHODS: This work is a cross-sectional study of surgeons, trainees, nurses, and surgical technicians (n = 372) across all surgical specialties at a large academic hospital. Participants completed a survey reporting frequency of use and estimated cost for 11 common surgical supplies as well as opinions on access to cost information in the OR. Cost estimation error was expressed as the ratio of estimated-to-actual cost, and groups were compared with one-way analysis of variance and chi-squared testing. Spearman correlation (ρ) was used to describe the relationship between monotonic variables. RESULTS: Overestimation error was universal and ranged widely (3.80-49.79). There was no significant difference in estimation accuracy when stratified by role or years of experience. Less expensive items had higher rates of estimation error than more expensive items (P < 0.001), and a moderately strong relationship was found between decreased item cost and increased estimation error (ρ: 0.49). The overwhelming majority (91%) of respondents expressed a desire to learn more about supply pricing. CONCLUSIONS: Price knowledge of common supplies is globally impaired for entire surgical team but coexists with a strong desire to augment cost awareness. Improved access to cost information has a high potential to inform surgical decision-making and decrease OR waste.

Mortality in African-Americans Following Cardiac Resynchronization Therapy: A Single Center Experience.

BACKGROUND: Cardiac resynchronization therapy (CRT) improves clinical outcomes and reduces mortality in heart failure patients who remain symptomatic despite optimal medical therapy. CRT trials have reported significant hemodynamic benefits, improvement in functional status, and reduced mortality and heart failure hospitalizations. However, African-American patient representation in these studies is limited thus the results may not be applicable to them. We described baseline clinical characteristics of African-American patients undergoing CRT and determined their outcomes relative to those reported in clinical trials. METHODS: We evaluated 131 African-American patients with New York Heart Association functional class II-IV heart failure undergoing CRT and determined predictors of all-cause mortality. Kaplan-Meier survival estimates and a Cox proportional hazards model determined mortality and risk of death. RESULTS: The mean age was 65 ± 12 years. Over a 6-year period, total mortality in African-Americans was 23% as compared with 29% in the MADIT-CRT trial. Increased mortality was associated with older age (hazard rate (HR) 1.04, 95% confidence interval (CI) 1.01-1.07, P=.01), ischemic cardiomyopathy (HR 2.86, 95% CI 1.36-6.04, P=.006), and absence of treatment with either an angiotensin-converting enzyme inhibitor or angiotensin II receptor blocker (HR 2.75, 95% CI 1.30-5.80, P=.008), or beta-blocker (HR 2.56, 95% CI 0.98-6.69, P=.05). Hydralazine plus nitrate therapy was used in a small number of patients and did not influence mortality outcomes. CONCLUSION: African-Americans experience the same survival benefits from CRT as Caucasian patients reported in major clinical trials. Publication indices used to find publications listed in references: PubMed.

Biventricular paced QRS predictors of left ventricular lead locations in relation to mortality in cardiac resynchronization therapy.

BACKGROUND: Left ventricular (LV) lead location during cardiac resynchronization therapy (CRT) has influenced mortality and heart failure events; however the biventricular paced QRS morphology has not been established as a predictor of LV lead location or mortality. METHODS: We evaluated the biventricular paced QRS morphology in 306 patients undergoing CRT in relation to specific anatomic locations. A logistic regression model and Kaplan-Meier survival estimates were used to determine predictors of LV lead location and survival. RESULTS: The mean age was 68±13years. Predictors of LV lead location from anterior, lateral, and posterior segments were: absence of R in V1, QS in aVL; and R in aVL, respectively. Absence of an R in II, III, or aVF predicted an inferior site. A QS in V4-V6 differentiated apical from basal sites (p=0.01). LV pacing from sites along the middle cardiac vein revealed a higher mortality (34%), than lateral sites (20%, p=0.02). CONCLUSIONS: Biventricular paced QRS criteria were predictive of LV lead locations. The proposed algorithm enhanced the predictive accuracy of these criteria. LV pacing sites along the middle cardiac vein were associated with increased mortality.

Mannitol-coated hydroxypropyl methylcellulose as an alternative directly compressible controlled release excipient.

One of the most common forms of controlled release technology for oral drug delivery comprises an active ingredient dispersed in a hydrophilic matrix forming polymer such as hydroxypropyl methylcellulose (HPMC), which is tableted via direct compression. However, HPMC may pose problems in direct compression due to its poor flowability. Hence, mannitol syrup was spray-coated over fluidized HPMC particles to produce co-processed HPMC-mannitol at ratios of 20:80, 50:50, and 70:30. Particles of pure HPMC, co-processed HPMC-mannitol, and their respective physical mixtures were evaluated for powder flowability, compression profiles, and controlled release performance. It was found that co-processed HPMC-mannitol consisted of particles with improved flow compared to pure HPMC particles. Sufficiently strong tablets of >2 MPa could be produced at moderate to high compression forces of 150-200 MPa. The dissolution profile could be tuned to obtain desired release profiles by altering HPMC-mannitol ratios. Co-processed HPMC-mannitol offers an interesting addition to the formulator's toolbox in the design of controlled release formulations for direct compression.

Control of malodorous hydrogen sulfide compounds using microbial fuel cell.

In this study, a microbial fuel cell (MFC) was used to control malodorous hydrogen sulfide compounds generated from domestic wastewaters. The electricity production demonstrated a distinct pattern of a two-step increase during 170 h of system run: the first maximum current density was 118.6 ± 7.2 mA m⁻² followed by a rebound of current density increase, reaching the second maximum of 176.8 ± 9.4 mA m⁻². The behaviors of the redox potential and the sulfate level in the anode compartment indicated that the microbial production of hydrogen sulfide compounds was suppressed in the first stage, and the hydrogen sulfide compounds generated from the system were removed effectively as a result of their electrochemical oxidation, which contributed to the additional electricity production in the second stage. This was also directly supported by sulfur deposits formed on the anode surface, which was confirmed by analyses on those solids using a scanning electron microscope equipped with energy dispersive X-ray spectroscopy as well as an elemental analyzer. To this end, the overall reduction efficiencies for HS⁻ and H2S(g) were as high as 67.5 and 96.4 %, respectively. The correlations among current density, redox potential, and sulfate level supported the idea that the electricity signal generated in the MFC can be utilized as a potential indicator of malodor control for the domestic wastewater system.

Muscle Tension Dysphagia: Contributing Factors and Treatment Efficacy.

OBJECTIVE: To determine factors contributing to disease etiology and treatment efficacy. STUDY DESIGN: Original Report. SETTING: Tertiary academic center. METHODS: IRB approved prospective study of 20 patients with reported dysphagia who exhibited normal oropharyngeal and esophageal swallowing function as evidenced by videofluoroscopic swallow study, esophagogastroduodenoscopy, high-resolution esophageal manometry with stationary impedance, and Bravo pH probe off proton pump inhibitor. Patients underwent speech-language pathology intervention. RESULTS: Atypical laryngeal muscle tension was present in 100% of patients. Forty percent of patients had diagnosed positive gastroesophageal reflux disease. Sixty-five percent of patients showed signs of non-specific laryngeal inflammation and laryngeal hyperresponsiveness during strobolaryngoscopy. All patients reported a mean of 90% recovery by the completion of voice therapy directed toward unloading muscle tension. CONCLUSION: The study results suggest an association between laryngeal muscle tension and these patients' dysphagia symptoms regardless of associated conditions. Speech-language pathology intervention showed high treatment efficacy. LEVEL OF EVIDENCE: 2c- Outcomes research.

The origin of aerobic methanotrophy within the Proteobacteria.

Aerobic methanotrophs play critical roles in the global carbon cycle, but despite their environmental ubiquity, they are phylogenetically restricted. Via bioinformatic analyses, it is shown that methanotrophy likely arose from methylotrophy from the lateral gene transfer of either of the two known forms of methane monooxygenase (particulate and soluble methane monooxygenases). Moreover, it appears that both known forms of pyrroloquinoline quinone-dependent methanol dehydrogenase (MeDH) found in methanotrophs-the calcium-containing Mxa-MeDH and the rare earth element-containing Xox-MeDH-were likely encoded in the genomes before the acquisition of the methane monooxygenases (MMOs), but that some methanotrophs subsequently received an additional copy of Xox-MeDH-encoding genes via lateral gene transfer. Further, data are presented that indicate the evolution of methanotrophy from methylotrophy not only required lateral transfer of genes encoding for methane monooxygenases, but also likely the pre-existence of a means of collecting copper. Given the emerging interest in valorizing methane via biological platforms, it is recommended that future strategies for heterologous expression of methane monooxygenase for conversion of methane to methanol also include cloning of genes encoding mechanism(s) of copper uptake, especially for expression of particulate methane monooxygenase.

Influence of spray nozzle aperture during high shear wet granulation on granule properties and its compression attributes.

The distribution of granulating liquid is known to affect the high shear wet granulation process but the impact of the spray nozzle attributes is still unclear. While homogenous liquid distribution can be achieved by using a spray nozzle, the effect of different nozzle aperture sizes on granule properties is not well understood. In this study, nozzles of different aperture sizes were used to introduce the granulating liquid in high shear wet granulation using different process parameters. Design of experiment approach was utilised to assess effect of process parameters on granule properties. Granules produced with different spray nozzles were evaluated for binder distribution inhomogeneity, size, shape, flowability and compression attributes such as tabletability and yield pressure. Coarser granules with better flow properties were produced using the smaller aperture size nozzle. On the other hand, granules had better tabletability and lower yield pressure when larger aperture size nozzle was used. Furthermore, size of granules produced by using larger aperture size nozzle was more affected by changes in the process variables which could be influenced by the differences in granulating liquid feed rate and spray droplet size. Although the granules aspect ratios were comparable across the nozzle aperture sizes, granules produced from smaller aperture size nozzle appeared to be rounder. Regardless of the nozzle aperture sizes, homogenous binder distribution was achieved. The findings from this study could be a useful guide to the selection of the appropriate nozzle aperture size in wet granulation.

Oxidative biodegradation of 4-chlorophenol by using recombinant monooxygenase cloned and overexpressed from Arthrobacter chlorophenolicus A6.

In this study, cphC-I and cphB, encoding a putative two-component flavin-diffusible monooxygenase (TC-FDM) complex, were cloned from Arthrobacter chlorophenolicus A6. The corresponding enzymes were overexpressed to assess the feasibility of their utilization for the oxidative decomposition of 4-chlorophenol (4-CP). Soluble CphC-I was produced at a high level (∼50%), and subsequently purified. Since CphB was expressed in an insoluble form, a flavin reductase, Fre, cloned from Escherichia coli was used as an alternative reductase. CphC-I utilized cofactor FADH2, which was reduced by Fre for the hydroxylation of 4-CP. This recombinant enzyme complex exhibited a higher specific activity for the oxidation of 4-CP (45.34U/mg-protein) than that exhibited by CphC-I contained in cells (0.18U/mg-protein). The Michaelis-Menten kinetic parameters were determined as: vmax=223.3µM·min-1, KM=249.4µM, and kcat/KM=0.052min-1·µM-1. These results could be useful for the development of a new biochemical remediation technique based on enzymatic agents catalyzing the degradation of phenolic contaminants.

Muscle Tension Dysphagia: Symptomology and Theoretical Framework.

OBJECTIVE: To identify symptoms, common diagnostic findings, pattern of treatments and referrals offered, and their efficacy in a group of patients with idiopathic functional dysphagia in an otolaryngology setting with multiple providers. STUDY DESIGN: Case series with chart review. SETTING: Tertiary academic center. SUBJECTS AND METHODS: Following Mayo Clinic Institutional Review Board approval, a retrospective chart review was conducted of patients with dysphagia who had a videofluoroscopic swallow study between January 1, 2013, and April 30, 2015. Each patient's dysphagia symptomology, videofluoroscopic swallow study, flexible laryngoscopy, and medical chart were reviewed to identify the treatment paradigms that were utilized. RESULTS: Sixty-seven adult patients met the inclusion criteria. Abnormal laryngeal muscle tension was present in 97% of patients. Eighty-two percent of patients also demonstrated signs of laryngeal hyperresponsiveness. Nonspecific laryngeal inflammation was evident in 52% of patients. Twenty-seven patients were referred to speech-language pathology for evaluation. Thirteen patients completed a course of voice therapy directed toward unloading muscle tension. All 13 patients self-reported resolution of dysphagia symptoms. CONCLUSION: The study results suggest that laryngeal muscle tension may be a factor in the underlying etiology in patients with idiopathic functional dysphagia. We propose the diagnostic term muscle tension dysphagia to describe a subset of patients with functional dysphagia. Further prospective studies are needed to better evaluate potential gastroesophageal confounders in this group of patients and to identify an effective paradigm for treatment. In our limited series, speech-language pathology intervention directed toward unloading muscle tension appears effective.

Noncovalent and covalent immobilization of oxygenase on single-walled carbon nanotube for enzymatic decomposition of aromatic hydrocarbon intermediates.

The decomposition of various aromatic hydrocarbon intermediates was examined using a recombinant oxidative enzyme immobilized on single-walled carbon nanotubes (SWCNTs). Hydroxyquinol 1,2-dioxygenase (CphA-I), which catalyzes ring cleavage of catechol and its analogues, was obtained from Arthrobacter chlorophenolicus A6 via cloning, overexpression, and subsequent purification. This recombinant enzyme was immobilized on SWCNTs by physical adsorption and covalent coupling in the absence and presence of N-hydroxysuccinimide. The immobilization yield was as high as 52.1%, and a high level of enzyme activity of up to 64.7% was preserved after immobilization. Kinetic analysis showed that the substrate utilization rates (vmax) and catalytic efficiencies (kcat/KM) of the immobilized enzyme for all substrates evaluated were similar to those of the free enzyme, indicating minimal loss of enzyme activity during immobilization. The immobilized enzyme was more stable toward extreme pH, temperature, and ionic strength conditions than the free enzyme. Thus, the oxidative enzyme immobilized on SWCNTs can be used as an effective and stable biocatalyst for the biochemical remediation process if further investigations would be carried out under field conditions.

Production of electrically-conductive nanoscale filaments by sulfate-reducing bacteria in the microbial fuel cell.

This study reports that the obligate anaerobic microorganism, Desulfovibrio desulfuricans, a predominant sulfate-reducing bacterium (SRB) in soils and sediments, can produce nanoscale bacterial appendages for extracellular electron transfer. These nanofilaments were electrically-conductive (5.81S·m(-1)) and allowed SRBs to directly colonize the surface of insoluble or solid electron acceptors. Thus, the direct extracellular electron transfer to the insoluble electrode in the microbial fuel cell (MFC) was possible without inorganic electron-shuttling mediators. The production of nanofilaments was stimulated when only insoluble electron acceptors were available for cellular respiration. These results suggest that when availability of a soluble electron acceptor for SRBs (SO4(2-)) is limited, D. desulfuricans initiates the production of conductive nanofilaments as an alternative strategy to transfer electrons to insoluble electron acceptors. The findings of this study contribute to understanding of the role of SRBs in the biotransformation of various substances in soils and sediments and in the MFC.

Enzymatic degradation of aromatic hydrocarbon intermediates using a recombinant dioxygenase immobilized onto surfactant-activated carbon nanotube.

This study examined the enzymatic decomposition of aromatic hydrocarbon intermediates (catechol, 4-chlorocatechol, and 3-methylcatechol) using a dioxygenase immobilized onto single-walled carbon nanotube (SWCNT). The surfaces of SWCNTs were activated with surfactants. The dioxygenase was obtained by recombinant technique: the corresponding gene was cloned from Arthrobacter chlorophenolicus A6, and the enzyme was overexpressed and purified subsequently. The enzyme immobilization yield was 62%, and the high level of enzyme activity was preserved (60-79%) after enzyme immobilization. Kinetic analyses showed that the substrate utilization rates and the catalytic efficiencies of the immobilized enzyme for all substrates (target aromatic hydrocarbon intermediates) tested were similar to those of the free enzyme, indicating that the loss of enzyme activity was minimal during enzyme immobilization. The immobilized enzyme was more stable than the free enzyme against abrupt changes in pH, temperature, and ionic strength. Moreover, it retained high enzyme activity even after repetitive use.

Evaluation of soil flushing of complex contaminated soil: an experimental and modeling simulation study.

The removal of heavy metals (Zn and Pb) and heavy petroleum oils (HPOs) from a soil with complex contamination was examined by soil flushing. Desorption and transport behaviors of the complex contaminants were assessed by batch and continuous flow reactor experiments and through modeling simulations. Flushing a one-dimensional flow column packed with complex contaminated soil sequentially with citric acid then a surfactant resulted in the removal of 85.6% of Zn, 62% of Pb, and 31.6% of HPO. The desorption distribution coefficients, KUbatch and KLbatch, converged to constant values as Ce increased. An equilibrium model (ADR) and nonequilibrium models (TSNE and TRNE) were used to predict the desorption and transport of complex contaminants. The nonequilibrium models demonstrated better fits with the experimental values obtained from the column test than the equilibrium model. The ranges of KUbatch and KLbatch were very close to those of KUfit and KLfit determined from model simulations. The parameters (R, ß, ω, α, and f) determined from model simulations were useful for characterizing the transport of contaminants within the soil matrix. The results of this study provide useful information for the operational parameters of the flushing process for soils with complex contamination.

T-cell responses induced in normal volunteers immunized with a DNA-based vaccine containing HIV-1 env and rev.

OBJECTIVE: An effective HIV-1 vaccine will likely need to induce strong cell-mediated immunity in humans. Therefore, we examined the ability of a DNA HIV-1 vaccine to induce a T-cell response in HIV-1 seronegative humans. DESIGN: Individuals were enrolled in a phase I clinical trial of safety and immune responses to an env/rev-containing plasmid at doses of 100, 300 or 1000 microg. Peripheral blood mononuclear cells (PBMC) samples were analyzed by standard lymphocyte proliferation, cytotoxic T lymphocyte (CTL) and ELISPOT techniques. RESULTS: PBMCs from subjects immunized with doses as low as 300 microg proliferated in vitro to env (four of six) or (three of six) proteins. Importantly, when the dose of vaccine was increased to 1000 microg of DNA, lymphocytes secreted IFN-gamma in an ELISPOT assay following in vitro stimulation with env (three of six) or rev (four of six) proteins. CONCLUSION: We observed HIV-1 DNA plasmid vaccines induce CD4 T-helper cell responses in humans. We observed a discrepancy in the CD4 versus CD8 response suggesting the importance of analyzing both compartments in clinical evaluation. Furthermore, this report demonstrates the high level of immunogenicity of and its importance as a component of a prophylactic vaccine for HIV-1.

Enhanced current production by Desulfovibrio desulfuricans biofilm in a mediator-less microbial fuel cell.

In this study, a mediator-less microbial fuel cell (MFC) inoculated with a sulfate-reducing bacterium (SBR), Desulfovibrio desulfuricans, was equipped with bare and surface-treated graphite felt electrodes. Electrochemical treatment of the anode surface facilitated biofilm formation on the electrode, resulting in rapid and enhanced current production. The maximum current density of the treated anode was 233±24.2mA/m(2), which was 41% higher than that of the untreated anode. The electron transfer rate also increased from 2.45±0.04 to 3.0±0.02µmol of electrons/mg of protein·min. Biofilm formation on the treated anode was mainly due to the strong hydrogen or peptide bonds between the amide groups of bacterial materials (including cytochrome c) and carboxyl groups formed on the electrodes. These results provide useful information on direct electron transfer by SRB in a mediator-less MFC through cytochrome c and the effects of the electrochemical treatment of electrodes on MFC performance.

Effective biochemical decomposition of chlorinated aromatic hydrocarbons with a biocatalyst immobilized on a natural enzyme support.

The enzymatic decomposition of 4-chlorophenol metabolites using an immobilized biocatalyst was investigated in this study. Catechol 1,2-dioxygenase for ortho ring cleavage obtained via cloning of the corresponding gene cphA-I from Arthrobacter chlorophenolicus A6 was overexpressed and purified. It was found that the cphA-I enzyme could catalyze the degradation of catechol, 4-chlorocatechol, and 3-methylcatechol. The expressed enzyme was immobilized onto a natural enzyme support, fulvic acid-activated montmorillonite. The immobilization yield was as high as 63%, and the immobilized enzyme maintained high substrate utilization activity, with only a 15-24% reduction in the specific activity. Kinetic analysis demonstrated marginal differences in νmax and KM values for the free and immobilized enzymes, indicating that inactivation of the immobilized enzyme was minimal. The immobilized enzyme exhibited notably increased stability against changes in the surrounding environment (temperature, pH, and ionic strength). Our results provide useful information for the effective enzymatic biochemical treatment of hazardous organic compounds.