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Find video protocols related to scientific articles indexed in Pubmed.
ATR suppresses apoptosis after UVB light by controlling both translesion synthesis and alternative tolerance pathways.
J. Cell. Sci.
PUBLISHED: 11-09-2014
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Ultraviolet (UV) light can stall replication forks due to the formation of bulky lesions in the DNA. Replication across these blocking lesions occurs via translesion DNA synthesis, and cells activate the ATR damage responses to UV. However, it remains unclear whether lesion bypass requires the replication checkpoint because ATR is not necessary for PCNA ubiquitination. We observed that ATR knockdown by siRNA increased replication stress and promoted early induction of apoptosis following UVB light in SV40-immortalized human cells, including cells from XP-V and XP-C patients. XP-V cells were further sensitized by silencing, indicating that DNA polymerase ? (Pol ?) remains active despite ATR control. However, following UVB irradiation, ATR-depleted cells were unable to achieve mitosis, as would be expected after the loss of a DNA checkpoint control. Thus, ATR also regulates replication arrest recovery following UVB light, independently of Pol ? in SV40-immortalized cell lines. ATR-mediated DNA damage response regulates replication and different tolerance pathways, and in these cells, ATR depletion induces replication catastrophe, which contributes to explain the potential of ATR inhibition to protect against UVB light-induced carcinogenesis.
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Correlates of having never been HIV tested among entrants to substance abuse treatment clinics: empiric findings from real-world New England settings.
J Psychoactive Drugs
PUBLISHED: 07-24-2014
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Routine testing is the cornerstone to identifying HIV, but not all substance abuse treatment patients have been tested. This study is a real-world evaluation of predictors of having never been HIV tested among patients initiating substance abuse treatment. Participants (N = 614) from six New England clinics were asked whether they had ever been HIV tested. Eighty-five patients (13.8%) reported having never been tested and were compared to those who had undergone testing. Clinic, male gender (adjusted odds ratio (AOR) = 1.91, 95% confidence interval (CI) = 1.07-3.41), and having fewer employment (AOR = 0.31; 95% CI = 0.11-0.88) and medical problems (AOR = 0.40, 95% CI = 0.17-0.99) were independently correlated with having never been HIV tested. Thus, there is still considerable room for improved testing strategies as a clinically significant minority of substance abuse patients have never undergone HIV testing when they initiate treatment.
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White problem gamblers discount delayed rewards less steeply than their African American and Hispanic counterparts.
Psychol Addict Behav
PUBLISHED: 06-24-2014
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Impulsivity is a core process underlying addictive behaviors, including nonpharmacological addictive behaviors such as problem gambling. Although considerable attention has been given to the investigation of delay discounting within the context of addiction-related behaviors, relatively little research has examined the relationship between discounting and individual variables, such as race/ethnicity. The purpose of this study was to compare discounting rates in the three most prevalent racial/ethnic groups in the United States: Whites, African Americans, and Hispanics. The study was conducted with 315 problem gamblers. Participants completed a delay-discounting questionnaire involving choices between a smaller amount of money delivered immediately and a larger amount delivered later. A hyperbolic discounting function estimated delay-discounting rates based on participants' indifference points obtained via the questionnaires. Results showed significant effects of race/ethnicity on delay discounting. White gamblers discounted delayed money at lower rates than African Americans and Hispanics, even after controlling for confounding variables. These data suggest that among individuals who develop problem gambling, Whites are less impulsive than African Americans and Hispanics, at least in terms of choosing between delayed and immediate reinforcers. These results have implications for evaluating the onset and treatment of addictive disorders from a health-disparities perspective.
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IncI1/ST113 and IncI1/ST114 conjugative plasmids carrying blaCTX-M-8 in Escherichia coli isolated from poultry in Brazil.
Diagn. Microbiol. Infect. Dis.
PUBLISHED: 05-25-2014
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Extended-spectrum beta-lactamase-producing Escherichia coli isolated from poultry in Brazil showed blaCTX-M-8 gene. IS10 was found upstream of blaCTX-M-8, harbored on plasmids IncI1, ST113/ST114 subtypes. Genomic relationship revealed a heterogeneous E. coli population. The gene blaCTX-M-8 is established in South America in food-producing animals, which represent risk of dissemination for other countries.
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Expansion and evolution of a virulent, extensively drug-resistant (polymyxin B-resistant), QnrS1-, CTX-M-2-, and KPC-2-producing Klebsiella pneumoniae ST11 international high-risk clone.
J. Clin. Microbiol.
PUBLISHED: 05-07-2014
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In this study, we report the early expansion, evolution, and characterization of a multiresistant Klebsiella pneumoniae clone that was isolated with increasing frequency from inpatients in a tertiary-care university hospital in Brazil. Seven carbapenem- and quinolone-resistant and polymyxin B-susceptible or -resistant K. pneumoniae isolates isolated between December 2012 and February 2013 were investigated. Beta-lactamase- and plasmid-mediated quinolone resistance (PMQR)-encoding genes and the genetic environment were investigated using PCR, sequencing, and restriction fragment length polymorphism (RFLP). Clonal relatedness was established using XbaI-pulsed-field gel electrophoresis (PFGE), multilocus sequence typing (MLST), and phylogenetic group characterization. Plasmid analyses included PCR-based replicon typing (PBRT) and hybridization of the S1-PFGE product, plasmid MLST, and conjugation experiments. Virulence potential was assessed by PCR by searching for 10 virulence factor-encoding genes (ureA, fimH, kfuBC, uge, wabG, magA, mrkD, allS, rmpA, and cf29a) and by phenotypic tests to analyze the hypermucoviscous phenotype. The genetic context of a multidrug-resistant and extensively drug-resistant K. pneumoniae ST11-KpI clone harboring IncFIIk-Tn4401a-blaKPC-2, qnrS1, and blaCTX-M-2 was found. Moreover, three isolates displayed high resistance to polymyxin B (MICs = 32, 32, and 128 mg/liter) as well as mucous and hypermucoviscous phenotypes. These bacteria also harbored ureA, fimH, uge, wabG, and mrkD, which code for virulence factors associated with binding, biofilm formation, and the ability to colonize and escape from phagocytosis. Our study describes the association of important coresistance and virulence factors in the K. pneumoniae ST11 international high-risk clone, which makes this pathogen successful at infections and points to the quick expansion and evolution of this multiresistant and virulent clone, leading to a pandrug-resistant phenotype and persistent bacteria in a Brazilian hospital.
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Maintaining high activity levels in sedentary adults with a reinforcement-thinning schedule.
J Appl Behav Anal
PUBLISHED: 04-03-2014
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Physical inactivity is a leading cause of mortality. Reinforcement interventions appear to be useful for increasing activity and preventing adverse consequences of sedentary lifestyles. This study evaluated a reinforcement-thinning schedule for maintaining high activity levels. Sedentary adults (N?=?77) were given pedometers and encouraged to walk ?10,000 steps per day. Initially, all participants earned rewards for each day they walked ?10,000 steps. Subsequently, 61 participants were randomized to a monitoring-only condition or a monitoring-plus-reinforcement-thinning condition, in which frequencies of monitoring and reinforcing walking decreased over 12 weeks. The mean (±?SD) percentage of participants in the monitoring-plus-reinforcement-thinning condition who met walking goals was 83%?±?24% and was 55%?±?31% for participants in the monitoring-only condition, p?
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Ephrin-B2 governs morphogenesis of endolymphatic sac and duct epithelia in the mouse inner ear.
Dev. Biol.
PUBLISHED: 02-18-2014
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Control over ionic composition and volume of the inner ear luminal fluid endolymph is essential for normal hearing and balance. Mice deficient in either the EphB2 receptor tyrosine kinase or the cognate transmembrane ligand ephrin-B2 (Efnb2) exhibit background strain-specific vestibular-behavioral dysfunction and signs of abnormal endolymph homeostasis. Using various loss-of-function mouse models, we found that Efnb2 is required for growth and morphogenesis of the embryonic endolymphatic epithelium, a precursor of the endolymphatic sac (ES) and duct (ED), which mediate endolymph homeostasis. Conditional inactivation of Efnb2 in early-stage embryonic ear tissues disrupted cell proliferation, cell survival, and epithelial folding at the origin of the endolymphatic epithelium. This correlated with apparent absence of an ED, mis-localization of ES ion transport cells relative to inner ear sensory organs, dysplasia of the endolymph fluid space, and abnormally formed otoconia (extracellular calcite-protein composites) at later stages of embryonic development. A comparison of Efnb2 and Notch signaling-deficient mutant phenotypes indicated that these two signaling systems have distinct and non-overlapping roles in ES/ED development. Homozygous deletion of the Efnb2 C-terminus caused abnormalities similar to those found in the conditional Efnb2 null homozygote. Analyses of fetal Efnb2 C-terminus deletion heterozygotes found mis-localized ES ion transport cells only in the genetic background exhibiting vestibular dysfunction. We propose that developmental dysplasias described here are a gene dose-sensitive cause of the vestibular dysfunction observed in EphB-Efnb2 signaling-deficient mice.
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Detection of chromosomal blaCTX -M-2 in diverse Escherichia coli isolates from healthy broiler chickens.
Clin. Microbiol. Infect.
PUBLISHED: 01-08-2014
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The rise of extended-spectrum beta-lactamase (ESBL)-producing Escherichia coli in food-producing animals is a growing concern for public health. We investigated ESBL producers isolated from broiler chickens in Brazil and characterized 19 CTX-M-2-producing E. coli. The blaCTX -M-2 gene was detected downstream to ISCR1 element, associated with sul-1 type integron, chromosome-located. CTX-M-2-producing E. coli exhibited different PFGE-types and phylogenetic groups, showing a non-clonal dissemination. The sequence types found (ST93, ST155 and ST2309) have been associated to humans and animals worldwide. Herein, we report the chromosomal location of blaCTX -M-2 on E. coli, alerting for the risks of multi-drug resistant bacteria in food-producing animals. This article is protected by copyright. All rights reserved.
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Phylogenetic analysis of phenotypically characterized Cryptococcus laurentii isolates reveals high frequency of cryptic species.
PLoS ONE
PUBLISHED: 01-01-2014
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Although Cryptococcus laurentii has been considered saprophytic and its taxonomy is still being described, several cases of human infections have already reported. This study aimed to evaluate molecular aspects of C. laurentii isolates from Brazil, Botswana, Canada, and the United States.
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Correlation of Actin Crosslinker and Capper Expression Levels with Stereocilia Growth Phases.
Mol. Cell Proteomics
PUBLISHED: 12-07-2013
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During development of the chick cochlea, actin crosslinkers and barbed-end cappers presumably influence growth and remodeling of the actin paracrystal of hair cell stereocilia. We used mass spectrometry to identify and quantify major actin-associated proteins of the cochlear sensory epithelium from E14 to E21, when stereocilia widen and lengthen. Tight actin crosslinkers (i.e., fascins, plastins, and espin) are expressed dynamically during cochlear epithelium development between E7 and E21, with FSCN2 replacing FSCN1 and plastins remaining low in abundance. Capping protein (CAPZ), a barbed-end actin capper, is located at stereocilia tips; it is abundant during growth phase II, when stereocilia have ceased elongating and are increasing in diameter. CAPZ levels then decline during growth phase III, when stereocilia reinitiate barbed-end elongation. Although actin crosslinkers are readily detected by electron microscopy in developing chick cochlea stereocilia, quantitative mass spectrometry of stereocilia isolated from E21 chick cochlea indicated that tight crosslinkers are present there in stoichiometric ratios relative to actin that are much lower than their ratios for vestibular stereocilia. These results demonstrate the value of quantitation of global protein expression in chick cochlea during stereocilia development.
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The effects of alcohol problems and smoking on delay discounting in individuals with gambling problems.
J Psychoactive Drugs
PUBLISHED: 11-02-2013
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Problem gambling is an addictive behavior with high comorbidity with alcohol problems and smoking. A common feature shared by these conditions is impulsivity. Past research shows that individuals with any of these addictions discount delayed money at higher rates than those without, and that the presence of gambling and substance use lead to additive effects on discounting. To date, however, no study examined the impact of smoking on these associations. The goals of this study were to compare the discounting rates of gamblers with and without histories of alcohol problems and smoking, and assess the associations these addictions might have on discounting. We analyzed the discounting rates of treatment-seeking gamblers categorized into four groups based on their histories of alcohol and smoking. Results revealed effects of history of alcohol problems, and an interaction between smoking and alcohol problems, on discounting. Never smokers with histories of alcohol problems discounted money less steeply than the other groups of gamblers. These results suggest that smoking does not produce additional increases on discounting rates in individuals with other addiction problems and the small subpopulation of gamblers with alcohol problems who never smoked is less impulsive and may have unique risk and/or protective behaviors.
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A randomized study of reinforcing ambulatory exercise in older adults.
Psychol Aging
PUBLISHED: 10-14-2013
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Many older adults do not meet physical activity recommendations and suffer from health-related complications. Reinforcement interventions can have pronounced effects on promoting behavior change; this study evaluated the efficacy of a reinforcement intervention to enhance walking in older adults. Forty-five sedentary adults with mild to moderate hypertension were randomized to 12-week interventions consisting of pedometers and guidelines to walk 10,000 steps/day or that same intervention with chances to win $1-$100 prizes for meeting recommendations. Patients walked an average of about 4,000 steps/day at baseline. Throughout the intervention, participants in the reinforcement intervention met walking goals on 82.5% ± 25.8% of days versus 55.3% ± 37.1% of days in the control condition, p < .01. Even though steps walked increased significantly in both groups relative to baseline, participants in the reinforcement condition walked an average of about 2,000 more steps/day than participants in the control condition, p < .02. Beneficial effects of the reinforcement condition relative to the control condition persisted at a 24-week follow-up evaluation, p < .02, although steps/day were lower than during the intervention period in both groups. Participants in the reinforcement intervention also evidenced greater reductions in blood pressure and weight over time and improvements in fitness indices, ps < .05. This reinforcement-based intervention substantially increased walking and improved clinical parameters, suggesting that larger-scale evaluations of reinforcement-based interventions for enhancing active lifestyles in older adults are warranted. Ultimately, economic analyses may reveal reinforcement interventions to be cost-effective, especially in high-risk populations of older adults. (PsycINFO Database Record (c) 2013 APA, all rights reserved).
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Molecular characterization of Klebsiella pneumoniae carbapenemase-producing isolates in southern Brazil.
J. Med. Microbiol.
PUBLISHED: 09-02-2013
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Carbapenem-resistant Enterobacteriaceae have been frequently reported worldwide. They represent a serious concern because of the limited therapeutic options. The aim of this study was to investigate the molecular epidemiology of 14 Klebsiella pneumoniae carbapenemase (KPC) producers among 345 clinical isolates of Enterobacteriaceae with reduced susceptibility to carbapenems recovered from 11 separate hospitals in southern Brazil. The blaKPC-2 gene was detected in 14 isolates (4?%): six Enterobacter cloacae, five K. pneumoniae and three Serratia marcescens. Most of these isolates exhibited high-level resistance against ?-lactams and ciprofloxacin, while the most active drugs were polymyxin B and amikacin. Genetic environment analysis, based on the classical Tn4401 structure, revealed six distinct platforms. Plasmids carrying blaKPC-2 were not typable and most were approximately 20 kb. Only KPC carbapenemases were found among the isolates studied, highlighting the local relevance of these enzymes in acquired resistance to carbapenems in Enterobacteriaceae. Our results contribute to the understanding of carbapenem resistance in Enterobacteriaceae and to the molecular characterization of KPC-2-producing isolates in Brazil.
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CLIC5 stabilizes membrane-actin filament linkages at the base of hair cell stereocilia in a molecular complex with radixin, taperin, and myosin VI.
Cytoskeleton (Hoboken)
PUBLISHED: 08-13-2013
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Chloride intracellular channel 5 protein (CLIC5) was originally isolated from microvilli in complex with actin binding proteins including ezrin, a member of the Ezrin-Radixin-Moesin (ERM) family of membrane-cytoskeletal linkers. CLIC5 concentrates at the base of hair cell stereocilia and is required for normal hearing and balance in mice, but its functional significance is poorly understood. This study investigated the role of CLIC5 in postnatal development and maintenance of hair bundles. Confocal and scanning electron microscopy of CLIC5-deficient jitterbug (jbg) mice revealed progressive fusion of stereocilia as early as postnatal day 10. Radixin (RDX), protein tyrosine phosphatase receptor Q (PTPRQ), and taperin (TPRN), deafness-associated proteins that also concentrate at the base of stereocilia, were mislocalized in fused stereocilia of jbg mice. TPRQ and RDX were dispersed even prior to stereocilia fusion. Biochemical assays showed interaction of CLIC5 with ERM proteins, TPRN, and possibly myosin VI (MYO6). In addition, CLIC5 and RDX failed to localize normally in fused stereocilia of MYO6 mutant mice. Based on these findings, we propose a model in which these proteins work together as a complex to stabilize linkages between the plasma membrane and subjacent actin cytoskeleton at the base of stereocilia. Published 2013-Wiley Periodicals, Inc.(†) This article is a US government work and, as such, is in the public domain in the United States of America.
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[Validation of nursing interventions for patients with spinal cord injury and impaired physical mobility].
Rev Bras Enferm
PUBLISHED: 07-22-2013
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This is a descriptive study that aimed to validate the nursing interventions proposed in the Nursing Interventions Classification for the diagnosis of Impaired physical mobility in adults with sequels of spinal cord injury, based on opinions of experts in rehabilitation about each intervention proposed. The sample consisted of 54 nurses who were asked about how they consider the interventions for the nursing diagnosis. A questionnaire was administered to nurses who work at the Sarah Network of Rehabilitation Hospitals. Among the 46 interventions studied, 17 (37%) were considered essential and should be implemented; others 15 (32.6%) were considered complementary, and 14 (30.4%) should not be used and could be discarded. Nursing interventions not described in the classification and suggested by 22 nurses (40.7%) were: Care with Orthoses: Maintenance; and Care with Orthoses: Prevention.
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Marijuana use is associated with risky sexual behaviors in treatment-seeking polysubstance abusers.
Am J Drug Alcohol Abuse
PUBLISHED: 07-12-2013
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Multiple types of substance use are associated with HIV risk behaviors, but relatively little research has examined the association between marijuana use and risky sexual activities in treatment-seeking polysubstance abusing patients.
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Engaging in Job-Related Activities Is Associated With Reductions in Employment Problems and Improvements in Quality of Life in Substance Abusing Patients.
Psychol Addict Behav
PUBLISHED: 04-15-2013
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Reinforcement-based interventions are highly efficacious in the treatment of substance use disorders, and their benefits can extend to other areas of functioning as well. In particular, reinforcing participation in job-related activities may be useful for improving employment outcomes, which in turn may enhance quality of life and decrease substance use. These secondary analyses compared substance abusing patients randomized with reinforcement interventions (N = 185) who selected and completed two or more job-related activities during treatment versus those who did not. Patients who completed two or more job-related activities during treatment had significantly greater reductions in employment-related problems and improvements in quality of life than those who completed only one or no job-related activities, even after controlling for baseline differences that may impact employment outcomes. Further, patients who completed employment activities remained in treatment significantly longer and achieved greater durations of abstinence than those who did not. These data suggest that reinforcing job-attainment activities may have broad beneficial effects. Reinforcement interventions should be considered for enhancing employment skills training acquisition more generally. (PsycINFO Database Record (c) 2013 APA, all rights reserved).
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[Autonomic dysreflexia and nursing interventions for patients with spinal cord injury].
Rev Esc Enferm USP
PUBLISHED: 03-22-2013
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This retrospective study, performed in 2009, aimed to identify nursing diagnoses and interventions for the care of patients with spinal cord injury. Data were collected from the nursing records of 465 patients with SCI undergoing rehabilitation. The nursing diagnosis Risk for autonomic dysreflexia was identified in 271 clinical records (58, 3%). Approximately 80 patients developed autonomic dysreflexia, with a predominance in young men around 35.7 years old, who had experienced a trauma as the main cause of the injury. Their neurological injury level was at the sixth thoracic vertebra or above. Nursing interventions were arranged in two groups, one focused on prevention and the other on treatment. An intervention guide was developed and can be used by nurses in their clinical practice of rehabilitation and can be included into information systems. The removal of the stimulus which causes autonomic dysreflexia was identified as the most effective therapy and the best intervention.
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Functional study of TCP23 in Arabidopsis thaliana during plant development.
Plant Physiol. Biochem.
PUBLISHED: 03-04-2013
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The TCP class of genes is found only in plants and is represented by the first three identified genes: teosinte branched1, cycloidea and pcf. Members belonging to this class are important regulators of plant growth, development and control multiple traits in diverse plant species, including flower and petal asymmetry, plant architecture, leaf morphogenesis and senescence, embryo growth and circadian rhythm. Here we described a member of the TCP-P subfamily called AtTCP23. Using qRT-PCR we present evidence that AtTCP23 is ubiquitously express in all organs examined. To ascertain AtTCP23 localization, we fused GFP at the C-terminal position and analyzed stable expression by confocal microscopy. Transgenic lines harboring the full-length protein (OxTCP23:GFP) seems to accumulate GFP in the nucleus. In order to analyze AtTCP23 function, we obtained a T-DNA insertional line and developed AtTCP23 over-expression (OxTCP23) lines. Phenotypic analysis indicates that tcp23-1 knockout line has an early-flowering phenotype while overexpression lines (OxTCP23 and OxTCP23:eGFP) presents opposite phenotype. Besides that those lines have leaf morphology alteration, pale leaf borders and smaller roots. Thus we propose in this study that AtTCP23 may be involved in flowering time control and plant development.
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Nucleotide excision repair activity on DNA damage induced by photoactivated methylene blue.
Free Radic. Biol. Med.
PUBLISHED: 02-20-2013
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The nucleotide excision repair (NER) mechanism is well known to be involved in the removal of UV-induced lesions. Nevertheless, the involvement of this pathway in the repair of lesions generated after DNA oxidation remains controversial. The effects of visible-light-excited methylene blue (MB), known to generate reactive oxygen species (ROS), were examined directly in xeroderma pigmentosum (XP)-A and XP-C NER-deficient human fibroblasts. Initially, MB was confirmed as being incorporated in similar amounts by the cells and that its photoexcitation induces the generation of (1)O2 within cells. The analysis of cell survival indicated that NER-deficient cells were hypersensitive to photoactivated MB. This sensitivity was confirmed with cells silenced for the XPC gene and by host-cell reactivation (HCR) of plasmid exposed to the photosensitizing effects of photoexcited MB. The sensitivity detected by HCR was restored in complemented cells, confirming the participation of XPA and XPC proteins in the repair of DNA lesions induced by photosensitized MB. Furthermore, DNA damage (single- and double-strand breaks and alkali-sensitive sites) was observed in the nuclei of treated cells by alkaline comet assay, with higher frequency of lesions in NER-deficient than in NER-proficient cells. Likewise, NER-deficient cells also presented more ?-H2AX-stained nuclei and G2/M arrest after photoactivated MB treatment, probably as a consequence of DNA damage response. Notwithstanding, the kinetics of both alkali- and FPG-sensitive sites repair were similar among cells, thereby demonstrating not only that MB photoexcitation generates nuclear DNA damage, but also that the removal of these lesions is NER-independent. Therefore, this work provides further evidence that XPA and XPC proteins have specific roles in cell protection and repair/tolerance of ROS-induced DNA damage. Moreover, as XPC-deficient patients do not present neurodegeneration, premature aging, or developmental clinical symptoms, the results indicate that defects in the repair/tolerance of oxidatively generated DNA lesions are not sufficient to explain these severe clinical features of certain XP patients.
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Income received during treatment does not affect response to contingency management treatments in cocaine-dependent outpatients.
Drug Alcohol Depend
PUBLISHED: 02-12-2013
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Prior studies find no effect of baseline income on response to contingency management (CM) interventions. However, income among substance disordered patients is variable, particularly at treatment entry. This study investigated the impact of during-treatment income, a more proximal estimate of economic resources at the time that CM is in effect, on response to standard treatment or the standard treatment plus CM.
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Susceptibility profile of clinical and environmental isolates of Cryptococcus neoformans and Cryptococcus gattii in Uberaba, Minas Gerais, Brazil.
Med. Mycol.
PUBLISHED: 01-23-2013
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Cryptococcus neoformans and C. gattii are the etiologic agents of cryptococcosis, a life-threatening disease in both immunocompromised and immunocompetent hosts. Antifungal resistance has been evaluated using different methods, breakpoints, and sizes of test populations and it is an emerging as a significant issue worldwide. A total of 176 (95 clinical and 81 environmental) C. neoformans and eight clinical C. gattii isolates were evaluated to determine the minimal inhibitory concentration (MIC) according to the Clinical and Laboratory Standards Institute method. A total of 10.5% of the C. neoformans clinical isolates were resistant to amphotericin B (AMB), and 6.2% of the environmental isolates were resistant to fluconazole (FLZ). Environmental and clinical isolates presented epidemiologic cut-off values (ECVs) of 64 and 16 to FLZ and 1 and 2 to AMB, respectively. All of the C. gattii isolates showed high susceptibility to most drugs evaluated. Clinical isolates had lower susceptibility than environmental isolates to AMB and itraconazole whereas environmental isolates had lower susceptibility than the clinical isolates to FLZ, voriconazole, and ketoconazole. However, no difference was found in the susceptibility of the two species. The MICs and ECVs to antifungals can help to select the best therapeutic option for tracking epidemiological resistance among clinical and environmental isolates of Cryptococcus spp. around the world.
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Genome-wide transcriptional effects of the anti-cancer agent camptothecin.
PLoS ONE
PUBLISHED: 01-01-2013
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The anti-cancer drug camptothecin inhibits replication and transcription by trapping DNA topoisomerase I (Top1) covalently to DNA in a "cleavable complex". To examine the effects of camptothecin on RNA synthesis genome-wide we used Bru-Seq and show that camptothecin treatment primarily affected transcription elongation. We also observed that camptothecin increased RNA reads past transcription termination sites as well as at enhancer elements. Following removal of camptothecin, transcription spread as a wave from the 5-end of genes with no recovery of transcription apparent from RNA polymerases stalled in the body of genes. As a result, camptothecin preferentially inhibited the expression of large genes such as proto-oncogenes, and anti-apoptotic genes while smaller ribosomal protein genes, pro-apoptotic genes and p53 target genes showed relative higher expression. Cockayne syndrome group B fibroblasts (CS-B), which are defective in transcription-coupled repair (TCR), showed an RNA synthesis recovery profile similar to normal fibroblasts suggesting that TCR is not involved in the repair of or RNA synthesis recovery from transcription-blocking Top1 lesions. These findings of the effects of camptothecin on transcription have important implications for its anti-cancer activities and may aid in the design of improved combinatorial treatments involving Top1 poisons.
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Susceptibility to DNA damage as a molecular mechanism for non-syndromic cleft lip and palate.
PLoS ONE
PUBLISHED: 01-01-2013
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Non-syndromic cleft lip/palate (NSCL/P) is a complex, frequent congenital malformation, determined by the interplay between genetic and environmental factors during embryonic development. Previous findings have appointed an aetiological overlap between NSCL/P and cancer, and alterations in similar biological pathways may underpin both conditions. Here, using a combination of transcriptomic profiling and functional approaches, we report that NSCL/P dental pulp stem cells exhibit dysregulation of a co-expressed gene network mainly associated with DNA double-strand break repair and cell cycle control (p?=?2.88×10(-2)-5.02×10(-9)). This network included important genes for these cellular processes, such as BRCA1, RAD51, and MSH2, which are predicted to be regulated by transcription factor E2F1. Functional assays support these findings, revealing that NSCL/P cells accumulate DNA double-strand breaks upon exposure to H2O2. Furthermore, we show that E2f1, Brca1 and Rad51 are co-expressed in the developing embryonic orofacial primordia, and may act as a molecular hub playing a role in lip and palate morphogenesis. In conclusion, we show for the first time that cellular defences against DNA damage may take part in determining the susceptibility to NSCL/P. These results are in accordance with the hypothesis of aetiological overlap between this malformation and cancer, and suggest a new pathogenic mechanism for the disease.
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The impact of contingency management on quality of life among cocaine abusers with and without alcohol dependence.
Am J Addict
PUBLISHED: 11-18-2011
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The use of quality of life (QOL) measures in substance abuse treatment research is important because it may lead to a broader understanding of patients health status and effects of interventions. Despite the high rates of comorbid cocaine and alcohol use disorders, little is known about the QOL of this population, and even less about the impact of an efficacious behavioral treatment, contingency management (CM), on QOL. In this study, data from three clinical trials were retrospectively analyzed to examine QOL in outpatient cocaine abusers with and without alcohol dependence (AD) and the impact of CM on QOL over time as a function of AD status. Patients were randomized to standard care (n?= 115) or standard care plus CM (n?= 278) for 12 weeks. QOL was assessed at baseline and Months 1, 3, 6, and 9. At treatment initiation, AD patients had lower QOL total scores and they scored lower on several subscale scores than those without AD. CM treatment was associated with improvement in QOL regardless of AD status. These data suggest that CM produces benefits that go beyond substance abuse outcomes, and they support the use of QOL indexes to capture information related to treatment outcomes.?
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CTGF/CCN2 has a chemoattractive function but a weak adhesive property to embryonic carcinoma cells.
Biochem. Biophys. Res. Commun.
PUBLISHED: 08-29-2011
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Connective tissue growth factor (CTGF/CCN2) is a protein of the CCN family that modulates cell-ECM interactions in a variety of cell types. In this study, we investigated the chemotactic and adhesive properties of CCN2 protein in embryonic teratocarcinoma P19 cells. Initially, P19 cells were attracted to CCN2-coated agarose beads. In Boyden chamber experiments, CCN2-containing medium induced a threefold greater migration of P19 cells. CCN2 adhesion properties were studied by using optical tweezers. The specific adhesion times of P19 cells to polystyrene beads coated with laminin, fibronectin, CCN2 and bovine serum albumin were 1.8 ± 0.5s, 2.7 ± 0.4s, 10 ± 2s and 13 ± 2s, respectively, revealing an unexpectedly low adhesive capacity of CCN2 protein for P19 cells. In conclusion, our findings support the chemoattractive role of CCN2 for P19 cells, but not its adhesive role when compared to laminin or fibronectin.
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Trypanosoma cruzi trypomastigotes induce cytoskeleton modifications during HeLa cell invasion.
Mem. Inst. Oswaldo Cruz
PUBLISHED: 06-14-2011
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It has been recently shown that Trypanosoma cruzi trypomastigotes subvert a constitutive membrane repair mechanism to invade HeLa cells. Using a membrane extraction protocol and high-resolution microscopy, the HeLa cytoskeleton and T. cruzi parasites were imaged during the invasion process after 15 min and 45 min. Parasites were initially found under cells and were later observed in the cytoplasm. At later stages, parasite-driven protrusions with parallel filaments were observed, with trypomastigotes at their tips. We conclude that T. cruzi trypomastigotes induce deformations of the cortical actin cytoskeleton shortly after invasion, leading to the formation of pseudopod-like structures.
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Clinical, epidemiological and outcome features of patients with cryptococcosis in Uberaba, Minas Gerais, Brazil.
Mycopathologia
PUBLISHED: 06-14-2011
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Nearly one million of cryptococcosis cases occur yearly around the world, involving mainly HIV-infected patients who are not receiving antiretroviral therapy (ART) or present poor adherence. This study aims to evaluate epidemiological, clinical and outcome aspects of patients with cryptococcosis from 1998-2010. Patients were prospectively recruited, and their medical and laboratory records were reviewed. A total of 131 cases were included, and of these, 119 (90.83%) had AIDS, 4 received a renal transplant, 2 presented systemic lupus erythematosus and 6 (4.6%) were apparently immunocompetent. Ninety-one (69.46%) were men, and the median age was 38.7 years. Cryptococcal meningitis (CM) was diagnosed in 103 (78.62%), whereas 28 (21.38%) had cryptococcal infection in other sites. Of patients with CM, 94 (91.26%) had AIDS being cryptococcosis the first defining illness in 61 (64.9%), while 37 (60.65%) of them presented simultaneously both diagnosis. Headache, altered mental status, papilledema and seizures at admission were significatively associated with a poor outcome. Of 163 different isolates, 155 (95.09%) were Cryptococcus neoformans and eight (4.88%) Cryptococcus gattii. Antifungal therapy was warranted in 8 (87.4%) patients with CM, but 46 (51.1%) died during the first days or weeks. Of 28 patients without CM, 21 (75%) received treatment, but 6 (28.6%) died. The poor outcome among this case series was similar to that reported from other developing countries, but it is paradoxal in Brazil where the ART is at free disposal in the public health services. Despite, at least 60-70% of patients present advanced immunosuppression when they receive the AIDS diagnosis.
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Fine structure and molecular content of human chondrocytes encapsulated in alginate beads.
Cell Biol. Int.
PUBLISHED: 06-11-2011
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Preservation of the chondrocytic phenotype in vitro requires a 3D (three-dimensional) culture model. Diverse biomaterials have been tested as scaffolds for culture of animal chondrocytes; however, to date, none is considered a gold standard in regenerative medicine. Here, we studied the fine structure and the GAGs (glycosaminoglycans) content of human chondrocytes encapsulated in alginate beads by using electron microscopy and radioactive sulfate [35S] incorporation, respectively. Cells were obtained from human cartilage, encapsulated in alginate beads and cultured for 28 days. [35S]Na2SO4 was added to the culture media and later isolated for quantification of the sulfated GAGs found in three compartments: IC (intracellular), IB (intra-bead) and EB (extra-bead). Round cells were seen isolated or forming small groups throughout the alginate. Human chondrocytes presented the features of active cells such as euchromatic nuclei, abundant RER (rough endoplasmic reticulum) and many transport vesicles. We observed an extracellular matrix rich in collagen fibres and electrondense material adjacent to the cells. Most of the GAGs produced (74%) were found in the culture medium (EB), indicating that alginate has a limited capacity to retain the GAGs. CS (chondroitin sulfate), the major component of aggrecan, was the most prominent GAG produced by the encapsulated cells. Human chondrocytes cultured in alginate can sustain their phenotype, confirming the potential application of this biomaterial for cartilage engineering.
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Antifungal susceptibility, enzymatic activity, PCR-fingerprinting and ITS sequencing of environmental Cryptococcus laurentii isolates from Uberaba, Minas Gerais, Brazil.
Mycopathologia
PUBLISHED: 06-06-2011
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Cryptococcus laurentii has been classically considered a saprophytic species, although several cases of human infection have been already reported. This study aimed to evaluate the phospholipase, proteinase and hemolysins activity, the antifungal susceptibility profile, the genetic variability by M13 and (GACA)(4) fingerprinting and the internal transcribe spacer (ITS) sequencing of 38 C. laurentii isolates recovered from captive bird droppings and surrounding hospital areas. All of them exhibited phospholipase activity, while the hemolytic activity was evidenced in 34 (89.4%) isolates. None of them exhibited proteinase activity. Twenty-seven isolates (71.1%) presented susceptibility dose dependent to fluconazole. Most isolates (94.7%) were susceptible to voriconazole, while one (2.65%) was resistant to this drug. Twenty-one (55.3%) isolates showed reduced susceptibility to itraconazole while nine (23.7%) were resistant. Three (7.9%) and five (13.1%) isolates exhibited resistance to ketoconazole and amphotericin B, respectively. Most C. laurentii fingerprinting obtained with M13 and (GACA)(4) showed high heterogeneity. By using the two primers, seven (18.4%) isolates grouped as A (CL2, CL7, and CL8), B (CL35, CL38) and C (CL29, CL30) with 100% similarity. Different from most variable surrounding hospital isolates, all but one of the pet shops strains clustered with the two primers, although they had been recovered from different neighborhoods. All isolates were identified as C. laurentii phylogenetic group I by ITS sequencing. Thus, the presence of virulence factors, a decreased antifungal susceptibility and a heterogeneous molecular pattern of the C. laurentii isolates here described suggests this species can be a potential pathogen in the context of the immunocompromised population.
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Germ cell transplantation in felids: a potential approach to preserving endangered species.
J. Androl.
PUBLISHED: 05-19-2011
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With the exception of the domestic cat, all members of the family Felidae are considered either endangered or threatened. Although not yet used for this purpose, spermatogonial stem cell (SSC) transplantation has a high potential to preserve the genetic stock of endangered species. However, this technique has not previously been established in felids. Therefore, we developed the necessary procedures to perform syngeneic and xenogeneic SSC transplants (eg, germ cell [GC] depletion in the recipient domestic cats, enrichment and labeling of donor cell suspension, and the transplantation method) in order to investigate the feasibility of the domestic cat as a recipient for the preservation and propagation of male germ plasm from wild felids. In comparison with busulfan treatment, local x-ray fractionated radiation was a more effective approach to depleting endogenous spermatogenesis. The results of both syngeneic and xenogeneic transplants revealed that SSCs were able to successfully colonize and differentiate in the recipient testis, generating elongated spermatids several weeks posttransplantation. Specifically, ocelot spermatozoa were observed in the cat epididymis 13 weeks following transplantation. As donor GCs from domestic cats and ocelots were able to develop and form mature GCs in the recipient environment seminiferous tubules, these findings indicate that the domestic cat is a suitable recipient for SSC transplantation. Moreover, as modern cats descended from a medium-size cat that existed approximately 10 to 11 million years ago, these results strongly suggest that the domestic cat could be potentially used as a recipient for generating and propagating the genome of wild felids.
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Dissemination of blaKPC-2 by the spread of Klebsiella pneumoniae clonal complex 258 clones (ST258, ST11, ST437) and plasmids (IncFII, IncN, IncL/M) among Enterobacteriaceae species in Brazil.
Antimicrob. Agents Chemother.
PUBLISHED: 05-16-2011
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This article reports the spread of bla(KPC-2) in the Sao Paulo and Rio de Janeiro states, facilitated by globally spread K. pneumoniae clonal complex 258 (CC258) clones (ST258, ST11, and ST437) and a diversity of plasmids (IncFII, IncN, and IncL/M, two untypeable plasmids carrying Tn4401a or Tn4401b) successfully disseminated among species of the Enterobacteriaceae (Enterobacter cloacae, Serratia marcescens, and Citrobacter freundii). It also constitutes the first description of sequence type 258 (ST258) in Brazil, which was associated with a nosocomial hospital outbreak in Ribeirao Preto city.
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Delay and probability discounting in pathological gamblers with and without a history of substance use problems.
Psychopharmacology (Berl.)
PUBLISHED: 04-29-2011
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Pathological gambling and substance use disorders are highly comorbid, possibly because they both stem from a similar process-impulsivity. Although much data exist regarding the association between delay discounting and these psychiatric disorders, relatively little research has examined probability discounting and its relationship with either substance use or gambling.
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Regulation of PCDH15 function in mechanosensory hair cells by alternative splicing of the cytoplasmic domain.
Development
PUBLISHED: 03-24-2011
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Protocadherin 15 (PCDH15) is expressed in hair cells of the inner ear and in photoreceptors of the retina. Mutations in PCDH15 cause Usher Syndrome (deaf-blindness) and recessive deafness. In developing hair cells, PCDH15 localizes to extracellular linkages that connect the stereocilia and kinocilium into a bundle and regulate its morphogenesis. In mature hair cells, PCDH15 is a component of tip links, which gate mechanotransduction channels. PCDH15 is expressed in several isoforms differing in their cytoplasmic domains, suggesting that alternative splicing regulates PCDH15 function in hair cells. To test this model, we generated three mouse lines, each of which lacks one out of three prominent PCDH15 isoforms (CD1, CD2 and CD3). Surprisingly, mice lacking PCDH15-CD1 and PCDH15-CD3 form normal hair bundles and tip links and maintain hearing function. Tip links are also present in mice lacking PCDH15-CD2. However, PCDH15-CD2-deficient mice are deaf, lack kinociliary links and have abnormally polarized hair bundles. Planar cell polarity (PCP) proteins are distributed normally in the sensory epithelia of the mutants, suggesting that PCDH15-CD2 acts downstream of PCP components to control polarity. Despite the absence of kinociliary links, vestibular function is surprisingly intact in the PCDH15-CD2 mutants. Our findings reveal an essential role for PCDH15-CD2 in the formation of kinociliary links and hair bundle polarization, and show that several PCDH15 isoforms can function redundantly at tip links.
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The Arabidopsis translocator protein (AtTSPO) is regulated at multiple levels in response to salt stress and perturbations in tetrapyrrole metabolism.
BMC Plant Biol.
PUBLISHED: 01-14-2011
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The translocator protein 18 kDa (TSPO), previously known as the peripheral-type benzodiazepine receptor (PBR), is important for many cellular functions in mammals and bacteria, such as steroid biosynthesis, cellular respiration, cell proliferation, apoptosis, immunomodulation, transport of porphyrins and anions. Arabidopsis thaliana contains a single TSPO/PBR-related gene with a 40 amino acid N-terminal extension compared to its homologs in bacteria or mammals suggesting it might be chloroplast or mitochondrial localized.
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Regulation of stereocilia length by myosin XVa and whirlin depends on the actin-regulatory protein Eps8.
Curr. Biol.
PUBLISHED: 01-13-2011
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Myosin XVa (MyoXVa) and its cargo whirlin are implicated in deafness and vestibular dysfunction and have been shown to localize at stereocilia tips and to be essential for the elongation of these actin protrusions [1-4]. Given that whirlin has no known actin-regulatory activity, it remains unclear how these proteins work together to influence stereocilia length. Here we show that the actin-regulatory protein Eps8 [5] interacts with MyoXVa and that mice lacking Eps8 show short stereocilia compared to MyoXVa- and whirlin-deficient mice. We show that Eps8 fails to accumulate at the tips of stereocilia in the absence of MyoXVa, that overexpression of MyoXVa results in both elongation of stereocilia and increased accumulation of Eps8 at stereocilia tips, and that the exogenous expression of MyoXVa in MyoXVa-deficient hair cells rescues Eps8 tip localization. We find that Eps8 also interacts with whirlin and that the expression of both Eps8 and MyoXVa at stereocilia tips is reduced in whirlin-deficient mice. We conclude that MyoXVa, whirlin, and Eps8 are integral components of the stereocilia tip complex, where Eps8 is a central actin-regulatory element for elongation of the stereocilia actin core.
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Primary cutaneous cryptococcosis caused by Cryptococcus gattii in an immunocompetent host.
Med. Mycol.
PUBLISHED: 10-28-2010
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This paper presents the case of a 75-year-old Brazilian man who developed inflammatory skin lesions with nodules and ulcerations on the right forearm after an injury caused by handling barbed-wire and Eucalyptus spp. logs. Histopathological assessment of the lesions showed granulomatous processes with yeasts similar to Cryptococcus spp. Tissue fragments yielded yeasts when cultured that were identified as Cryptococcus gattii VGII through biochemical reactions and URA5-RFLP genotype. No evidence of systemic involvement or any underlying immunosuppressive diseases were identified, which supported the diagnosis of primary cutaneous cryptococcosis. After 5 months on therapy with high fluconazole doses, the skin lesions had fully healed.
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[Role of nursing in physical rehabilitation].
Rev Bras Enferm
PUBLISHED: 07-10-2010
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This study reviews the literature concerning improvement of rehabilitation nurse role, suggesting paths to reach this goal and evoking a still poorly discussed area. Some questions were raised to provoke reflection and discussion. It is evident, based on the literature, that nurses have a total role in rehabilitation, however the boundaries are not clearly defined. Professional nurses must must join rehabilitation practice with permanent education, that will contribute for a more efficient and influence in processes and rehabilitation services.
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Genotyping of Cryptococcus neoformans isolated from captive birds in Uberaba, Minas Gerais, Brazil.
Mycoses
PUBLISHED: 05-19-2010
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To evaluate Cryptococcus spp. molecular types isolated from captive birds droppings, an epidemiological survey was carried out in Uberaba, Minas Gerais, Brazil, from December 2006 to September 2008. A total of 253 samples of bird excreta (120 fresh and 133 dry) were collected from pet shop cages and houses in different neighbourhoods. Cryptococcus neoformans was isolated in 19 (14.28%) dry samples and one fresh sample (0.84%). Cryptococcus laurentii was recovered from seven (5.26%) dry samples, but not in the fresh samples. The canavanine-glycine-bromothymol blue test was positive in all but one of the C. laurentii isolates. Cryptococcus neoformans molecular typing was performed using URA5-RFLP and the mating type locus using mating type specific PCR. Nineteen (95.0%) presented genotype VNI and one VNII (5.0%). In addition, all isolates presented mating type ?. Thus, the genotype of the environmental C. neoformans isolates observed in this study is in accordance with others already reported around the world and adds information about its distribution in Brazil. Cryptococcus laurentii strains were typed using URA5-RFLP and M13 fingerprinting, which showed similar profiles among them. Thus, despite the low number of C. laurentii isolates analysed, their molecular profile is different from another already reported.
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[Case for diagnosis. Reticulate acropigmentation of Dohi].
An Bras Dermatol
PUBLISHED: 05-14-2010
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Reticulate acropigmentation of Dohi is a rare dyschromic disorder of autosomal dominant inheritance. Most cases have been originally described in Japan. The case of a girl with lesions of typical distribution and morphology is reported. Skin biopsy was not considered essential for diagnosis. After literature review, it was concluded that this is the third case of the disorder reported in Brazil.
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Microcapsules of alginate/chitosan containing magnetic nanoparticles for controlled release of insulin.
Colloids Surf B Biointerfaces
PUBLISHED: 05-11-2010
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The challenge of this work was to investigate the potential of alginate/chitosan beads containing magnetite nanoparticles as a drug delivery system. The insulin beads were prepared by dripping a solution of sodium alginate containing insulin into a CaCl(2) solution. Magnetite nanoparticles of 5 nm mean size were synthesized inside the alginate egg-box structure by co-precipitation of Fe(III) and Fe(II) in the presence of NH(4)OH. Quantitative analysis revealed that insulin encapsulation depends on the initial protein content and 35% of insulin was entrapped by alginate beads for a protein concentration of 10 wt%. It was verified that approximately 50% of the insulin was released to Milli-Q water in 800 h release experiments. The application of oscillating magnetic field increased three fold the insulin release. The results suggest that the alginate/chitosan system containing magnetite nanoparticles is a promising system for clinical applications of controlled release of insulin in the presence of an oscillating magnetic field in a subcutaneous implant approach.
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Evaluation of the biological behavior of decellularized pulmonary homografts: an experimental sheep model.
Rev Bras Cir Cardiovasc
PUBLISHED: 04-23-2010
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The cryopreserved homograft is a good valve substitute due attributes like excellent hemodynamics, low incidence of thromboembolic events, infection resistance and good mid-term durability. However, progressive homograft degeneration and fibrocalcification may occur, particularly in the childhood and young adults. Their antigenicity triggers an immunological reaction that plays an important role in their degeneration and failure. The decellularization process was proposed to decrease this antigenicity. By the action of detergents and enzymes, this process removes all cellular components from the homograft matrix, diminishing immunogenicity and probably delaying its degeneration.
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Assessment of heavy metals in the particulate matter of two Brazilian metropolitan areas by using Tillandsia usneoides as atmospheric biomonitor.
Environ Sci Pollut Res Int
PUBLISHED: 04-18-2010
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The aims of this paper were to quantify the heavy metals (HM) in the air of different sites in Rio de Janeiro (RJ) and Salvador (SA) using Tillandsia usneoides (Bromeliaceae) as a biomonitor, and to study the morphology and elemental composition of the air particulate matter (PM) retained on the Tillandsia surface.
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Molecular characterization and evaluation of virulence factors of Cryptococcus laurentii and Cryptococcus neoformans strains isolated from external hospital areas.
Fungal Biol
PUBLISHED: 03-01-2010
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Cryptococcosis is a common opportunistic fungal infection that is mainly caused by the species Cryptococcus neoformans and Cryptococcus gattii, but there have recently been several reports of infection by non-neoformans Cryptococcus species. The aims of this study were to genetically characterize Cryptococcus spp. isolated from external hospital areas in Minas Gerais State, Brazil, and to evaluate their pathogenic potential, analyzing their phospholipase and melanin production and the capacity for capsule enlargement. Seventy-three different samples were collected: 62 from bird droppings and 11 from tree detritus. C. neoformans alone was isolated from 43.8% of the samples, Cryptococcus laurentii alone from 23.3% and both fungi were found together in 10.9%. C. laurentii was exclusively isolated from 45% (5/11) of the tree samples (Anacardium occidentale, Guazuma ulmifolia, Mangifera indica and Ficus benjamina). Among the 51 C. neoformans isolates, 47 were classified as type VNI and four as type VNII. All of the C. neoformans isolates were of MAT? type. Among the 21 isolates of C. laurentii genotyped using the URA5-RFLP technique, 16 amplified a 1.6kb amplicon which produced a specific restriction profile in 15 isolates. In C. neoformans, 76.4% of the isolates were capable of capsule enlargement in the induction medium and 92.1% were phospholipase producers. In C. laurentii, 7.4% of the isolates were capable of capsule enlargement and 85.1% were phospholipase producers. Characterization of the genotypes and the pathogenic potential of the Cryptococcus spp. isolates studied may contribute towards better understanding of the epidemiology of cryptococcosis and the ecology of agents causing this disease in our region.
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Brown algae overproduce cell wall polysaccharides as a protection mechanism against the heavy metal toxicity.
Mar. Pollut. Bull.
PUBLISHED: 03-01-2010
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Brown algae are often used as heavy metal biomonitors and biosorbents because they can accumulate high concentrations of metals. Cation-exchange performed by cell wall polysaccharides is pointed out as the main chemical mechanism for the metal sequestration. Here, we biochemically investigated if the brown alga Padina gymnospora living in a heavy metal contaminated area would modify their polysaccharidic content. We exposed non-living biomass to Cd and Pb and studied the metals adsorption and localization. We found that raw dried polysaccharides, sulfate groups, uronic acids, fucose, mannose, and galactose were significantly higher in contaminated algae compared with the control ones. Metal concentrations adsorbed by non-living biomass were rising comparatively to the tested concentrations. Electron microscopy showed numerous granules in the cell walls and X-ray microanalysis revealed Cd as the main element. We concluded that P. gymnospora overproduces cell wall polysaccharides when exposed to high metal concentrations as a defense mechanism.
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Changes in plasma membrane structure and electromotile properties in prestin deficient outer hair cells.
Cytoskeleton (Hoboken)
PUBLISHED: 02-20-2010
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Cochlear outer hair cells (OHCs) rapidly change their length and stiffness when their membrane potential is altered. Prestin, the motor protein for this electromotility, is present along the OHC lateral plasma membrane where there is a high density of intra-membrane protein particles (IMPs). However, it is not known to what extent prestin contributes to this unusual dense population of proteins and overall organization of the membrane to generate the unique electromechanical response of OHCs. We investigated the relationship of prestin with the IMPs, the underlying cortical cytoskeletal lattice, and electromotility in prestin-deficient mice. Using freeze-fracture, we observed a reduction in density and size of the IMPs that correlates with the reduction and absence of prestin in the heterozygous and homozygous mice, respectively. We also observed a reduction or absence of electromotility-related charge density, axial stiffness, and piezoelectric properties of the OHC. A comparison of the charge density with the number of IMPs suggests that prestin forms tetramers in the wild type but is likely to form lower number oligomers in the prestin-deficient OHCs from the heterozygous mice. Interestingly, the characteristic actin-based cortical cytoskeletal lattice that underlies the membrane is absent in the prestin-null OHCs, suggesting that prestin is also required for recruiting or maintaining the cortical cytoskeletal lattice. These results suggest that the majority of the IMPs are indeed prestin and that electrically evoked length and stiffness changes are interrelated and dependent on both prestin and on the cortical actin cytoskeletal lattice of the OHC lateral membrane.
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The airways and gastric contents in obese patients.
Rev Bras Anestesiol
PUBLISHED: 02-20-2010
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The objective of this study was to review the literature and to question the concept that obese patients should be considered as having a potential full stomach and, therefore, question how the airways, considered as of difficult access due to obesity, should be approached during anesthetic induction.
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Sulfated alpha-L-galactans from the sea urchin ovary: selective 6-desulfation as eggs are spawned.
Glycobiology
PUBLISHED: 02-10-2010
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The sea urchin eggs are surrounded by a jelly coat, which contains sulfated polysaccharides with unique structures. These molecules are responsible for inducing the species-specific acrosome reaction, an obligatory event for the binding of sperm and fusion with the egg. The mechanism of biosynthesis of these sulfated polysaccharides is virtually unknown. The egg jelly of the sea urchin Echinometra lucunter contains a simple 2-sulfated, 3-linked alpha-L-galactan. Here, we pulse labeled the sea urchin ovary in vitro with (35)S-sulfate to follow the biosynthesis of the sulfated alpha-L-galactan. We found that the ovary contains a 2,6-disulfated, 3-linked alpha-L-galactan, which incorporates (35)S-sulfate more avidly than the 2-sulfated isoform. The 2,6-disulfated alpha-L-galactan was purified by anion exchange chromatography, analyzed by electrophoresis and characterized by 1D and 2D nuclear magnetic resonance spectra. We also investigated the location of the sulfated polysaccharides on the oocytes using histochemical procedures. The stain revealed high amounts of sulfated polysaccharide in mature oocytes and accessory cells. The amount of intracellular sulfated polysaccharides decreased as oocytes are spawned. We speculate that 2,6-disulfated galactan is initially synthesized in the ovary and that 6-sulfate ester is removed when the polysaccharide is secreted into the egg jelly. Similar events related to remodeling of sulfated polysaccharides have been reported in other biological systems.
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Case report: analytical electron microscopy of lung granulomas associated with exposure to coating materials carried by glass wool fibers.
Environ. Health Perspect.
PUBLISHED: 02-04-2010
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Man-made vitreous fibers (MMVFs) are noncrystalline inorganic fibrous material used for thermal and acoustical insulation (e.g., rock wool, glass wool, glass microfibers, and refractory ceramic fibers). Neither epidemiologic studies of human exposure nor animal studies have shown a noticeable hazardous effect of glass wools on health. However, MMVFs have been anecdotally associated with granulomatous lung disease in several case reports.
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Sequential bone healing of immediately loaded mini-implants: histomorphometric and fluorescence analysis.
Am J Orthod Dentofacial Orthop
PUBLISHED: 02-04-2010
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Mini-implants are often immediately loaded for orthodontic treatment; however, changes in interfacial tissues caused by early loading and its effects might compromise the mini-implants function. The purpose of this study was to compare the healing of interfacial tissues 1, 4, and 12 weeks after the placement of titanium-alloy mini-implants in New Zealand rabbits; some of the implants were loaded immediately and others were left unloaded.
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Antimycobacterial activity of natural and semi-synthetic lignans.
Z. Naturforsch., C, J. Biosci.
PUBLISHED: 11-25-2009
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The antimycobacterial activity of (-)-cubebin (1), hinokinin (2), and some of their semisynthetic derivatives, namely (-)-O-acetyl-cubebin (3), (-)-O-methyl-cubebin (4), (-)-O-(N,N-dimethylamine-ethyl)-cubebin (5) and (-)-6,6-dinitrohinokinin (6), was evaluated against Mycobacterium tuberculosis (ATCC 27294), M. kansasii (ATCC 12478), and M. avium (ATCC 15769). The MIC values ranged from 31.25 to 2000 microg/mL. Among the evaluated compounds, 2 displayed a MIC value of 62.5 microg/mL against M. tuberculosis, while 3 and 4 displayed MIC values of 62.5 and 31.25 microg/mL, respectively, against M. avium. All compounds were inactive against M. kansasii. These are promising results concerning the search for biologically active natural products, highlighting that new approaches to the prevention, treatment, and cure of tuberculosis are extremely important.
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A multidimensional high performance liquid chromatography method coupled with amperometric detection using a boron-doped diamond electrode for the simultaneous determination of sulfamethoxazole and trimethoprim in bovine milk.
Anal. Chim. Acta
PUBLISHED: 07-15-2009
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The development and validation of a multidimensional HPLC method using an on-line clean-up column coupled with amperometric detection employing a boron-doped diamond (BDD) electrode for the simultaneous determination of sulfamethoxazole (SMX) and trimethoprim (TMP) in bovine milk are presented. Aliquots of pre-prepared skim-milk samples were directly injected into a RAM octyl-BSA column in order to remove proteins that otherwise would interfere with milk analysis. After exclusion of the milk proteins, SMX and TMP were transferred to the analytical column (an octyl column) and the separation of the compounds from one another and from other endogenous milk components was achieved. SMX and TMP were detected amperometrically at 1.25V vs. Ag/AgCl (3.0molL(-1) KCl). Results with good linearity in the concentration ranges 50-800 and 25-400microgL(-1) for SMX and TMP, respectively, were obtained and no fouling of the BDD electrode was observed within the experimental period of several hours. The intra- and inter-assay coefficients of variation were less than 10% for both drugs and the obtained LOD values for SMX and TMP were 25.0 and 15.0microgL(-1), respectively.
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Systemic levels of metallic ions released from orthodontic mini-implants.
Am J Orthod Dentofacial Orthop
PUBLISHED: 04-14-2009
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Orthodontic mini-implants are a potential source of metallic ions to the human body because of the corrosion of titanium (Ti) alloy in body fluids. The purpose of this study was to gauge the concentration of Ti, aluminum (Al), and vanadium (V), as a function of time, in the kidneys, livers, and lungs of rabbits that had Ti-6Al-4V alloy orthodontic mini-implants placed in their tibia.
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Nitric oxide production by hemocytes of the ascidian Styela plicata.
Cell Tissue Res.
PUBLISHED: 04-09-2009
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Ascidian hemolymph contains various types of blood cells (hemocytes), which are believed to be involved in defense mechanisms. We have studied nitric-oxide (NO) synthase activity in hemocytes of the ascidian Styela plicata after exposure to lipopolysaccharide (LPS). To investigate which cell types are involved in NO production, we first identified, by electron microscopy, the types of hemocytes previously described, mainly by light microscopy, by others. Five types of blood cells could be recognized in the hemolymph: granulocytes, hemoblasts, lymphocyte-like cells, morula cells, and pigment cells. The lymphocyte-like cells produced the most NO. In agreement with studies of other invertebrates, nitrite generation did not change after LPS stimulation in assays in vitro, under either different concentrations of LPS or different time periods. Therefore, we performed an in vivo assay by injecting a known quantity of Escherichia coli into the tunic of the ascidians in order to investigate possible differences in NO levels. No increase of NO occurred accompanying the inflammatory reaction suggesting that another molecule in the pathway was involved. We found that nuclear factor kappaB (NFkappaB) was activated. Since NFkappaB is involved in the production of many substances related to immune responses, additional molecules might also be generated in response to E. coli infection. These observations may improve our understanding of the reaction of animals to eutrophic conditions.
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The septate junction protein caspr is required for structural support and retention of KCNQ4 at calyceal synapses of vestibular hair cells.
J. Neurosci.
PUBLISHED: 03-13-2009
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The afferent innervation contacting the type I hair cells of the vestibular sensory epithelia form distinct calyceal synapses. The apposed presynaptic and postsynaptic membranes at this large area of synaptic contact are kept at a remarkably regular distance. Here, we show by freeze-fracture electron microscopy that a patterned alignment of proteins at the calyceal membrane resembles a type of intercellular junction that is rare in vertebrates, the septate junction (SJ). We found that a core molecular component of SJs, Caspr, colocalizes with the K(+) channel KCNQ4 at the postsynaptic membranes of these calyceal synapses. Immunolabeling and ultrastructural analyses of Caspr knock-out mice reveal that, in the absence of Caspr, the separation between the membranes of the hair cells and the afferent neurons is conspicuously irregular and often increased by an order of magnitude. In these mutants, KCNQ4 fails to cluster at the postsynaptic membrane and appears diffused along the entire calyceal membrane. Our results indicate that a septate-like junction provides structural support to calyceal synaptic contact with the vestibular hair cell and that Caspr is required for the recruitment or retention of KCNQ4 at these synapses.
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[Bougie].
Rev Bras Anestesiol
PUBLISHED: 01-31-2009
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Difficult airways require fast action by the anesthesiologist often requiring complementary devices to ensure patent airways. However, several of those devices are expensive and require training in order to be used. The bougie, a simple and disposable device can also be manufactured by the anesthesiologist, making it a low cost tool.
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Cryo-electron tomography of the magnetotactic vibrio Magnetovibrio blakemorei: insights into the biomineralization of prismatic magnetosomes.
J. Struct. Biol.
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We examined the structure and biomineralization of prismatic magnetosomes in the magnetotactic marine vibrio Magnetovibrio blakemorei strain MV-1 and a non-magnetotactic mutant derived from it, using a combination of cryo-electron tomography and freeze-fracture. The vesicles enveloping the Magnetovibrio magnetosomes were elongated and detached from the cell membrane. Magnetosome crystal formation appeared to be initiated at a nucleation site on the membrane inner surface. Interestingly, while scattered filaments were observed in the surrounding cytoplasm, their association with the magnetosome chains could not be unequivocally established. Our data suggest fundamental differences between prismatic and octahedral magnetosomes in their mechanisms of nucleation and crystal growth as well as in their structural relationships with the cytoplasm and plasma membrane.
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Mercury-selenium relationships in liver of Guiana dolphin: the possible role of Kupffer cells in the detoxification process by tiemannite formation.
PLoS ONE
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Top marine predators present high mercury concentrations in their tissues as consequence of biomagnification of the most toxic form of this metal, methylmercury (MeHg). The present study concerns mercury accumulation by Guiana dolphins (Sotalia guianensis), highlighting the selenium-mediated methylmercury detoxification process. Liver samples from 19 dolphins incidentally captured within Guanabara Bay (Rio de Janeiro State, Brazil) from 1994 to 2006 were analyzed for total mercury (THg), methylmercury (MeHg), total organic mercury (TOrgHg) and selenium (Se). X-ray microanalyses were also performed. The specimens, including from fetuses to 30-year-old dolphins, comprising 8 females and 11 males, presented high THg (0.53-132 µg/g wet wt.) and Se concentrations (0.17-74.8 µg/g wet wt.). Correlations between THg, MeHg, TOrgHg and Se were verified with age (p<0.05), as well as a high and positive correlation was observed between molar concentrations of Hg and Se (p<0.05). Negative correlations were observed between THg and the percentage of MeHg contribution to THg (p<0.05), which represents a consequence of the selenium-mediated methylmercury detoxification process. Accumulation of Se-Hg amorphous crystals in Kupffer Cells was demonstrated through ultra-structural analysis, which shows that Guiana dolphin is capable of carrying out the demethylation process via mercury selenide formation.
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Saving the best for last? A cross-species analysis of choices between reinforcer sequences.
J Exp Anal Behav
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Two experiments were conducted to compare choices between sequences of reinforcers in pigeon (Experiment 1) and human (Experiment 2) subjects, using functionally analogous procedures. The subjects made pairwise choices among 3 sequence types, all of which provided the same overall reinforcerment rate, but differed in their temporal patterning. Token reinforcement schedules were used in both experiments and the type of exchange schedule varied across blocks of sessions. Some conditions permitted immediate exchange of tokens for consumable reinforcers (food for pigeons, video access for humans); in other conditions, tokens accumulated and were exchanged for consumable reinforcers only at the end of the sequence. Choice patterns in the immediate-exchange conditions were generally similar across species, with both pigeons and humans preferring sequences with the shortest delay to the initial reinforcer in the series. The results are broadly consistent with models of temporal discounting expanded to include the impact of sequences of delayed reinforcers acting in parallel from the time of the choice. Preferences were less consistent with discounting models in the delayed exchange conditions. Questionnaire data gathered at the end of the experiment were consistent with prior results of questionnaire studies, but showed no straightforward relation to the observed choice patterns, urging caution in the extrapolation of results from one decision-making domain to the other.
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Immunogold TEM of otoconin 90 and otolin - relevance to mineralization of otoconia, and pathogenesis of benign positional vertigo.
Hear. Res.
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Implementation of the deep-etch technique enabled unprecedented definition of substructural elements of otoconia, including the fibrillar meshwork of the inner core with its globular attachments. Subsequently the effects of the principal soluble otoconial protein, otoconin 90, upon calcite crystal growth in vitro were determined, including an increased rate of nucleation, inhibition of growth kinetics and significant morphologic changes. The logical next step, ultrastructural localization of otoconin 90, by means of immunogold TEM in young mature mice, demonstrated a high density of gold particles in the inner core in spite of a relatively low level of mineralization. Here gold particles are typically arranged in oval patterns implying that otoconin 90 is attached to a scaffold consisting of the hexagonal fibrillar meshwork, characteristic of otolin. The level of mineralization is much higher in the outer cortex where mineralized fiber bundles are arranged parallel to the surface. Following decalcification, gold particles, as well as matrix fibrils, presumed to consist of a linear structural phenotype of otolin, are aligned in identical direction, suggesting that they serve as scaffold to guide mineralization mediated by otoconin 90. In the faceted tips, the level of mineralization is highest, even though the density of gold particles is relatively low, conceivably due to the displacement by the dense mineral phase. TEM shows that individual crystallites assemble into iso-oriented columns. Columns are arranged in parallel lamellae which convert into mineralized blocks for hierarchical assembly into the complex otoconial mosaic. Another set of experiments based on immunogold TEM in young mice demonstrates that the fibrils interconnecting otoconia consist of the short chain collagen otolin. By two years of age the superficial layer of mouse otoconia (corresponding to mid-life human) has become demineralized resulting in weakening or loss of anchoring of the fibrils interconnecting otoconia. Consequently, otoconia detached from each other may be released into the endolymphatic space by minor mechanical disturbances. In humans, benign positional vertigo (BPV) is believed to result from translocation of otoconia from the endolymphatic space into the semi-circular canals rendering their receptors susceptible to stimulation by gravity causing severe attacks of vertigo. The combinations of these observations in humans, together with the presented animal experiments, provide a tentative pathogenetic basis of the early stage of BPV.
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Plasma membrane cholesterol depletion disrupts prechordal plate and affects early forebrain patterning.
Dev. Biol.
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Cholesterol-rich membrane microdomains (CRMMs) are specialized structures that have recently gained much attention in cell biology because of their involvement in cell signaling and trafficking. However, few investigations, particularly those addressing embryonic development, have succeeded in manipulating and observing CRMMs in living cells. In this study, we performed a detailed characterization of the CRMMs lipid composition during early frog development. Our data showed that disruption of CRMMs through methyl-?-cyclodextrin (M?CD) cholesterol depletion at the blastula stage did not affect Spemanns organizer gene expression and inductive properties, but impaired correct head development in frog and chick embryos by affecting the prechordal plate gene expression and cellular morphology. The M?CD anterior defect phenotype was recapitulated in head anlagen (HA) explant cultures. Culture of animal cap expressing Dkk1 combined with M?CD-HA generated a head containing eyes and cement gland. Together, these data show that during Xenopus blastula and gastrula stages, CRMMs have a very dynamic lipid composition and provide evidence that the secreted Wnt antagonist Dkk1 can partially rescue anterior structures in cholesterol-depleted head anlagen.
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RAPD analysis with the primer L15996 of Brazilian clinical and environmental Cryptococcus neoformans isolates.
Mycopathologia
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Different methods have been used to perform the molecular characterization of Cryptococcus neoformans. Among them, RAPD analysis is able to separate isolates of the same species and genotypes. This study aimed to evaluate clinical and environmental C. neoformans isolates from Minas Gerais, Brazil by RAPD and correlate the genetic profiles with the ones obtained by URA5-RFLP, virulence factors and antifungal susceptibility patterns. Forty-five environmental (31 from areas surrounding hospital and 14 from captive bird droppings from pet-shops) and 29 clinical C. neoformans isolates were evaluated. Antifungal susceptibility tests (Clinical and Laboratory Standards Institute), URA5-RFLP analysis and the assessment of virulence factors were performed according to their original descriptions. RAPD profiles were obtained using the L15996 primer (5-CTCCACCATTAGCACCCAAAGC-3). RAPD analysis generated two to 20 bands for all studied isolates. The isolates presented similarities ranging from 10.8 to 100.0%. Considering a minimum identity score of 50%, four clusters were formed. Cluster I contained 10 pet-shops bird dropping isolates, cluster II contained 22 clinical isolates most of them recovered from cerebrospinal fluid, cluster III contained 14 isolates from hospital surroundings and cluster IV contained 12 environmental isolates most from hospital surroundings. Fourteen isolates were not grouped. The RAPD profiles were clustered according to their source and URA5-RFLP pattern. No correlation between virulence factors or antifungal susceptibility profile with the obtained RAPD profiles was observed.
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?-Amyloid peptide is internalized into chick retinal neurons and alters the distribution of myosin Vb.
Cytoskeleton (Hoboken)
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The most common neurodegenerative disorder afflicting the aging human population is Alzheimers disease (AD). A major hallmark of AD is dementia from a loss of neuronal function, attributed to the presence and accumulation of ?-amyloid (A?) peptide into senile plaques. Preceding senile plaque formation, abnormalities in axons can be observed as changes in morphologies and intracellular trafficking. Recently, it has been recognized that A? also accumulates within neurons and this intraneuronal A? accumulation has been reported to be critical in the disruption of synapses and cognitive function. Here, we report on the internalization of a fluorescently labeled A? peptide into cultured chick retinal neurons. The pattern of A? distribution during the time course of incubation is reminiscent of the endocytic pathway. Furthermore, the distribution of the internalized A? peptide converges with that of myosin Vb and both relocalize from the axon to cell body. These observations are consistent with the hypothesis that AD proceeds as a result of an imbalance between A? production and A? clearance, suggesting a role for myosin Vb in this process.
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JoVE Visualize is a tool created to match the last 5 years of PubMed publications to methods in JoVE's video library.

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