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Find video protocols related to scientific articles indexed in Pubmed.
CNV instability associated with DNA replication dynamics: evidence for replicative mechanisms in CNV mutagenesis.
Hum. Mol. Genet.
PUBLISHED: 11-16-2014
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Copy number variation (CNV) in the human genome is of vital importance to genomic disorders, human health and evolution of our species. However, much of the molecular basis of CNV mutagenesis remains to be elucidated. Considering the DNA replication model of 'fork stalling and template switching' for CNV formation, we hypothesized that replication fork progression could be important to CNV mutagenesis. However, molecular assays of replication fork progression at the genome level are technically challenging. Instead, we conducted an estimation of DNA replication dynamics, as the statistic R, using the readily available data of replication timing. Small R values can reflect 'slowed' replication, which could result from less fork initiation, reduced fork speed, or fork barriers. We generated genome-wide profiles of R in the genomes of human, mouse and Drosophila. Intriguingly, the CNV breakpoints in all three genomes show significantly biased distributions toward the genomic regions with small R values, suggesting potential replication stress-induced CNV instability. Notably, among the human CNVs with distinct breakpoint junction characteristics, the homology-mediated and VNTR-mediated CNVs contribute mostly to the correlation between CNV instability and the statistic R, consistent with the recent findings in the C. elegans and yeast genomes of repeat-induced DNA replication error and consequent CNV formation. The statistic R may reflect both replication stress and the effect of local genome architecture on fork progression. Our concordant observations suggest an important role for DNA replicative mechanisms in CNV mutagenesis and genome instability.
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Genome editing using cas9 nickases.
Meth. Enzymol.
PUBLISHED: 11-16-2014
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The RNA-guided, sequence-specific endonuclease Cas9 has been widely adopted as genome engineering tool due to its efficiency and ease of use. Derived from the microbial CRISPR (clustered regularly interspaced short palindromic repeat) type II adaptive immune system, Cas9 has now been successfully engineered for genome editing applications in a variety of animal and plant species. To reduce potential off-target mutagenesis by wild-type Cas9, homology- and structure-guided mutagenesis of Streptococcus pyogenes Cas9 catalytic domains has produced "nicking" enzymes (Cas9n) capable of inducing single-strand nicks rather than double-strand breaks. Since nicks are generally repaired with high fidelity in eukaryotic cells, Cas9n can be leveraged to mediate highly specific genome editing, either via nonhomologous end-joining or homology-directed repair. Here we describe the preparation, testing, and application of Cas9n reagents for precision mammalian genome engineering.
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Toxic Effects of Levofloxacin on Rat Annulus Fibrosus Cells: An In-vitro Study.
Med. Sci. Monit.
PUBLISHED: 11-09-2014
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Background Fluoroquinolones are in wide clinical use as safe and effective antibiotics. Articular cartilage, tendons, and epiphyseal growth plates have been recognized as targets of fluoroquinolone-induced connective tissue toxicity. However, the effects of fluoroquinolones on annulus fibrosus (AF) cells are still unknown. Material and Methods The main objective of this study was to investigate the effects of levofloxacin, a typical fluoroquinolone antibiotic drug, on rat AF cells in vitro. Rat annulus fibrosus (RAF) cells were treated with levofloxacin at different concentrations (0, 10, 20, 30, 40, 60, 80, and 90 ?g/ml) and were assessed to determine the possible cytotoxic effects of levofloxacin. Inverted phase-contrast microscopy was used to accomplish the morphological observation of apoptosis of treated cells. Western blot and real-time quantitative RT-PCR (qPCR) was used to explore the expression of active caspase-3 and MMP-3. Flow cytometry was used to measure the apoptotic incidences. Results Our study showed that levofloxacin, with concentrations at 30, 60, and 90 ?g/ml, induced dose-dependent RAF cell apoptosis and higher expression of caspase-3 and MMP-3. More apoptotic cells were observed by inverted phase-contrast microscopy. Moreover, levofloxacin increased the activity of caspase-3, and it also reduced cell viability with different concentrations ranging from 10 to 80 ?g/ml. Conclusions Our study results suggest that levofloxacin has cytotoxic effects on RAF cells, characterized by enhancing apoptosis and reducing cell viability, and indicate a potential toxic effect of fluoroquinolones on RAF cells.
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Discrete Nanoparticle-BSA Conjugates Manipulated by Hydrophobic Interaction.
ACS Appl Mater Interfaces
PUBLISHED: 11-06-2014
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Nanoparticle-protein conjugates are promising probes for biological diagnostics as well as versatile building blocks for nanotechnology. Here we demonstrate a facile method to prepare nanoparticles bearing discrete numbers of BSA simply by physical adsorption and electrophoretic isolation, in which the specific amphiphilic properties of BSA play important roles and the number of adsorbed BSA molecules can also be manipulated by tuning the coating extent of nanoparticles by amphiphilic polymer.
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Effects of tetrahydroxystilbene glucoside on mouse liver cytochrome P450 enzyme expressions.
Xenobiotica
PUBLISHED: 10-29-2014
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Abstract 1. To investigate the effects of tetrahydroxystilbene glucoside (TSG), the main active component of Polygonum multiflorum, on mouse liver cytochrome P450 (Cyp) enzyme protein expressions. Male mice were randomly divided into the control, TSG low (10?mg/kg) and high dose (40?mg/kg) groups. After TSG intragastrical administration for 3, 5 and 7 d, mice were sacrificed and the mouse body and liver weight were detected. The Cyp enzymes and various transcription factors such as AhR, PXR and PPAR? protein expressions in mouse livers were measured by Western blotting assay. 2. No significant difference of mouse body and liver weight between the control and TSG treatment groups was detected. Additionally, TSG decreased Cyp1a2 and Cyp2e1 protein expressions after TSG treatment for 3, 5 and 7 d, respectively. Moreover, TSG suppressed Cyp3a11 protein expression after TSG treatment for 5 and 7 d. Furthermore, TSG high dose inhibited AhR and PXR protein expressions after TSG treatment for 5 and 7 d, while both TSG low dose and high dose obviously decreased PPAR? protein level from TSG treatment for 3 d. 3. TSG has inhibitory effects on mouse liver Cyp1a2, Cyp2e1 and Cyp3a11 protein expressions through the suppression of AhR, PXR and PPAR? activation.
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Caldicellulosiruptor changbaiensis sp. nov., a cellulolytic and hydrogen-producing bacterium from a hot spring in the Changbai Mountains, China.
Int. J. Syst. Evol. Microbiol.
PUBLISHED: 10-25-2014
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A novel thermophilic bacterial strain, CBS-ZT, was isolated from a terrestrial hot spring in the Changbai Mountains, China. Its cells are short straight rods without flagella and have Gram-positive cell walls. It grows at temperatures of 40-90 °C with an optimum at 75 °C, and at pH values of 4.5-8.6 with an optimum at pH 7.8. The primary end products from the fermentation of filter paper by the CBS-ZT strain were acetate, lactate, H2, and CO2. Its main cellular fatty acids were iso-C17:0, iso-C14:03-OH and C16:0. The G+C content of its genomic DNA was 36.08 mol%. Multiple sequence alignment of its 16S rRNA gene sequence and phylogenetic analyses indicated that the CBS-ZT isolate belongs to the genus Caldicellulosiruptor and the most similar microorganism was C. saccharolyticus DSM 8903 (96.36% similarity). The sequence similarity of the CBS-ZT strain to other species is below 95%. Based on its phylogenetic and phenotypic characteristics, the strain CBS-ZT represents a novel species of the genus Caldicellulosiruptor, for which the name Caldicellulosiruptor changbaiensis sp. nov. is proposed. The type strain is CBS-ZT (=DSM 26941T = CGMCC 1.5180T).
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A Prospective Randomized Double Blind Clinical Trial Using a Combination of Olfactory Ensheathing Cells and Schwann Cells for the Treatment of Chronic Complete Spinal Cord Injuries.
Cell Transplant
PUBLISHED: 10-22-2014
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The aim of this prospective randomized double blind clinical study is to examine the benefits of using olfactory ensheathing cells (OECs) combined with or without Schwann cells (SCs) in treating chronic complete spinal cord injuries (SCI). This would offer patients a better alternative for neurological functional recovery. According to the initial design, 28 eligible participants with cervical chronic complete SCI were recruited and randomly allocated into 4 groups of 7 participants each. The neurological assessments were to be performed according to the American Spinal Injury Association (ASIA) and International Association of Neurorestoratology (IANR) Functional Rating Scales, in combination with the electrophysiological tests e.g., electromyography (EMG) and paraspinal somatosensory evoked potentials (PSSEPs). Here, we have summarized the data from 7 patients; 3 patients received an OEC intraspinal transplantation, one underwent SC implantation, whereas one received a combination of OECs and SCs. The remaining 2 patients were used as controls. The scores were evaluated independently by at least 2 neurologists in a blinded fashion for comparing the neurological functional changes during pre- and post- cell transplantation (six month follow-up). All patients who received OECs, SCs alone, and a combination of them showed functional improvement. Mild fever occurred in one of the patients with OEC transplant that subsided after symptomatic treatments. All treated patients except one showed improvement in the electrophysiological tests. The functional improvement rate comprises 5/5 (100%) in the treated group, but 0/2 (0%) in the control group (p=0.008). These preliminary findings show that transplanting OECs, SCs, or a combination of them is well tolerated and that they have beneficial effects in patients. Thus, further studies in larger patient cohorts are warranted to assess the benefits and risks of these intervention strategies. This manuscript is published as part of the International Association of Neurorestoratology (IANR) special issue of Cell Transplantation.
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Correlation between frequency of non-allelic homologous recombination and homology properties: evidence from homology-mediated CNV mutations in the human genome.
Hum. Mol. Genet.
PUBLISHED: 10-16-2014
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Non-allelic homologous recombination (NAHR) is one of the key mechanisms of DNA rearrangement. NAHR occurring between direct homologous repeats can generate genomic copy number variation (CNV) and make significant contributions to both genome evolution and human diseases such as cancer. Intriguingly, previous observations on the rare CNVs at certain genomic disorder loci suggested that NAHR frequency could be dependent on homology properties. However, such a correlation remains unclear at the other NAHR-mediated CNV loci, especially the common CNVs in human populations. Different from the rare CNVs associated with genomic disorders, it is challenging to identify de novo NAHR events at common CNV loci. Therefore, our previously proposed statistic M was employed in estimating relative mutation rate for the NAHR-mediated CNVs in human populations. By utilizing generalized regression neural network and principal component analysis in studying 4330 CNVs ascertained in 3 HapMap populations, we identified the CNVs mediated by NAHR between paired segmental duplications (SDs) and further revealed the correlations between SD properties and NAHR probability. SD length and inter-SD distance were shown to make major contributions to the occurrence of NAHR, whereas chromosomal position and sequence similarity of paired SDs are also involved in NAHR. An integrated effect of SD properties on NAHR frequency was revealed for the common CNVs in human populations. These observations can be well explained by ectopic synapsis in NAHR together with our proposed model of chromosomal compression/extension/looping (CCEL) for homology mis-pairing. Our findings showed the important roles of SDs in NAHR and human genomic evolution.
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Inhibition of aflatoxin metabolism and growth of Aspergillus flavus in liquid culture by a DNA methylation inhibitor.
Food Addit Contam Part A Chem Anal Control Expo Risk Assess
PUBLISHED: 10-15-2014
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Aflatoxins (AFs) are a group of highly oxygenated polyketidese-derived toxins mainly produced by Aspergillus flavus and A. parasiticus, whose biosynthesis mechanisms are extremely sophisticated. Methylation is known as the major form of epigenetic regulation, which is correlated with gene expression. As the DNA methylation inhibitor 5-azacytidine (5-AC) blocks AF production, we studied AFB1 metabolism and morphological changes of A. flavus by treatment with 5-AC in liquid culture. The results show that 5-AC caused a decrease in AF production and concurrent changes in morphology. In addition, we isolated a non-aflatoxigenic mutant of A. flavus, showing a significant reduction in pigment production, after 5-AC treatment. This mutant showed significant reduction in the expression of genes in the AF biosynthesis pathway, and conidia formation. Furthermore, as AF biosynthesis and oxidative stress are intimately related events, we assessed the viability of A. flavus to oxidative stress after treatment with 5-AC, which showed that the mutant was more sensitive to the strong oxidant hydrogen peroxide. We found that the non-aflatoxigenic mutant showed a decrease in reactive oxygen species (ROS) and metabolites indicative of oxidative stress, which may be caused by the disruption of the defence system against excessive ROS formation after 5-AC treatment. These data indicate that 5-AC, as an inactivator of DNA methyltransferase, plays a very important role in AFB1 metabolism and the development of A. flavus, which might provide an effective strategy to pre- or post-harvest control of AFs.
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[Distinct effect of Wansheng Huafeng Dan containing ardisia crenata on renal transporters, mercury accumulation and Kim-1 expression from mercuric chloride].
Zhongguo Zhong Yao Za Zhi
PUBLISHED: 10-07-2014
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To study the effect of Wansheng Huafeng Dan (WSHFD) and mercuric chloride on renal mercury (Hg) extraction transporters (Oat1, Oct2), renal mercury excretion transporters (Mrp4, Mate2K), renal mercury accumulation and kidney injury molecule-1 (Kim-1). The ancient prescription of WSHFD containing 10-fold Hg caused much lower renal mercury accumulation and renal toxicity than HgCl2 in rats, with less effect on renal transporters than HgCl2. The above indicators had no significant difference in WSHFDO, WSHFD2 and WSHFD3 groups, indicating no effect of WSHFD with reduced or no cinnabar.
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Locus-specific epigenetic remodeling controls addiction- and depression-related behaviors.
Nat. Neurosci.
PUBLISHED: 10-02-2014
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Chronic exposure to drugs of abuse or stress regulates transcription factors, chromatin-modifying enzymes and histone post-translational modifications in discrete brain regions. Given the promiscuity of the enzymes involved, it has not yet been possible to obtain direct causal evidence to implicate the regulation of transcription and consequent behavioral plasticity by chromatin remodeling that occurs at a single gene. We investigated the mechanism linking chromatin dynamics to neurobiological phenomena by applying engineered transcription factors to selectively modify chromatin at a specific mouse gene in vivo. We found that histone methylation or acetylation at the Fosb locus in nucleus accumbens, a brain reward region, was sufficient to control drug- and stress-evoked transcriptional and behavioral responses via interactions with the endogenous transcriptional machinery. This approach allowed us to relate the epigenetic landscape at a given gene directly to regulation of its expression and to its subsequent effects on reward behavior.
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Dual regulating effects of gastrodin on extracellular dopamine concentration in rats models of Tourette's syndrome.
Int. J. Neurosci.
PUBLISHED: 10-02-2014
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Purpose: Evaluate the dual regulating effects of gastrodin on striatal extracellular dopamine (DA) concentration in Tourette's syndrome (TS) rat models, and explore the underlying pharmacological mechanisms. Materials and Methods: Seventy Wistar rats were randomly divided into control group and TS model group. The former was intraperitoneally injected with saline (0.9%), while in the later, the rats were injected with Apomorphine (Apo) and 3,3'-iminodipropionitrile (IDPN) respectively to manipulate two kinds of TS rat models. Both Apo and IDPN induced rats were further assigned to three conditions, and the related rats were treated respectively by oral gavage with saline, gastrodin and Haloperidol (Hal). Data of stereotypy of the rats were collected. After 8 weeks, the extracellular content of DA and HVA in striatum were examined by intracerebral microdialysis and follow-up high-performance liquid chromatography (HPLC), and the expression of dopamine transporter (DAT) was probed by Western blot. Results: Gastrodin improved the stereotyped behaviors in TS rats. Furthermore, it down-regulated the elevated striatal extracellular DA concentration in Apo-induced rats and up-regulated the decreased DA content in the rats exposed to IDPN. Meanwhile, a dramatic down-regulation was detected in DAT protein expression in Apo + GAS group, while an opposite profile was showed in the IDPN + GAS group. Conclusions: The dual regulating effects of gastrodin on extracellular DA level have been established, and the related mechanisms would be the dual regulating effects of gastrodin on the expression of DAT, a glycoprotein in the regulation of the extracellular DA concentration.
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Three new species of the genus Otacilia Thorell (Araneae: Phrurolithidae)
from China.
Zootaxa
PUBLISHED: 10-01-2014
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Three new species of the spider genus Otacilia are diagnosed, described and illustrated from China: Otacilia fujiana sp. nov. (male, female), O. pseudostella sp. nov. (male, female) and O. zhangi sp. nov. (male).
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Protection of Momordica charantia polysaccharide against intracerebral hemorrhage-induced brain injury through JNK3 signaling pathway.
J. Recept. Signal Transduct. Res.
PUBLISHED: 09-30-2014
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Abstract It has been well documented that Momordica charantia polysaccharide (MCP) has multiple biological effects such as immune enhancement, anti-oxidation and anti-cancer. However, the potential protective effects of MCP on stroke damage and its relative mechanisms remain unclear. Our present study demonstrated that MCP could scavenge reactive oxygen species (ROS) in intra-cerebral hemorrhage damage, significantly attenuating the neuronal death induced by thrombin in primary hippocampal neurons. Furthermore, we found that MCP prevented the activation of the c-Jun N-terminal protein kinase (JNK3), c-Jun and caspase-3, which was caused by the intra-cerebral hemorrhage injury. Taken together, our study demonstrated that MCP had a neuroprotective effect in response to intra-cerebral hemorrhage and its mechanisms involved the inhibition of JNK3 signaling pathway.
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[Simultaneous determination of twenty psychoactive drugs in pork by solid phase extraction coupled with ultra performance liquid chromatography-tandem mass spectrometry].
Se Pu
PUBLISHED: 09-27-2014
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A method for the simultaneous determination of 20 psychoactive drugs in pork by solid phase extraction coupled with ultra performance liquid chromatography with electrospray ionization tandem mass spectrometry (UPLC-ESI-MS/MS) was established. The samples were extracted with alkalified acetonitrile, and then cleaned up using solid phase extraction with an Oasis MCX column. The UPLC separation was performed on a C18 column (50 mm x 2.1 mm, 1.7 microm) using a gradient elution with the mobile phases of 0.1% (v/v) formic acid in water and acetonitrile. The ESI-MS/MS detection was achieved in positive mode under multiple reaction monitoring (MRM) mode. The calibration curves showed good linearity in the range of 5-100 microg/L with the correlation coefficients more than 0.99 for the 20 psychoactive drugs. The limits of quantification (LOQ, S/N > or = 10) for the 20 psychoactive drugs were 5 microg/kg. The average recoveries of the 20 psychoactive drugs spiked in blank pork at three levels of 5, 10 and 50 microg/kg were 66.8%-97.2% with the relative standard deviations from 4.2% to 12.4%. The meth od is suitable for the simultaneous determination of the 20 psychoactive drugs in pork with the characteristics of easy operation, high accuracy and precision.
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The outcomes of interventional treatment for Budd-Chiari syndrome: systematic review and meta-analysis.
Abdom Imaging
PUBLISHED: 09-25-2014
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The purpose of this study was to conduct a systematic review with meta-analysis quantitatively assesses the outcomes of interventional treatment for Budd-Chiari syndrome (BCS). We evaluated the published studies on interventional treatment for BCS and reviewed reference lists from retrieved articles. Meta-analysis was applied to calculate the combined rates and their 95% confidence intervals. The risk of bias was assessed by the Egger test. As many as 29 articles on interventional treatment with BCS were selected according to the eligibility criteria and included in the meta-analysis, for a total of 2,255 BCS patients. The pooled results (95 % CI) were 93.7 % (92.6-4.8 %) for successful rate of interventional operation, 6.5 % (5.3-7.7 %) for restenosis rate of interventional treatment, and 92.0 % (89.8-94.3 %) and 76.4 % (72.5-80.4 %) for the survival rate at 1 and 5 years, respectively. The interventional therapy of major BCS patients is safe with successful operation, good patency, and long-term survival. Moreover, a step-wise management of BCS is proposed to manage and cure all BCS patients with personalized treatment.
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Levofloxacin increases the effect of serum deprivation on anoikis of rat nucleus pulposus cells via Bax/Bcl-2/caspase-3 pathway.
Toxicol. Mech. Methods
PUBLISHED: 09-23-2014
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Abstract Levofloxacin, a fluoroquinolone, is a widely-used and effective antibiotic. However, various adverse side effects are associated with levofloxacin. The purpose of this study was to further explore the effects of levofloxacin on rat nucleus pulposus cells (NPCs). Inverted phase-contrast microscopy, flow cytometry and caspase-3 activity assays were used and revealed that serum deprivation induced apoptosis, which was markedly increased by levofloxacin in a dose-dependent manner. Simultaneously, levofloxacin decreased cell binding to type II collagen (COL2). Thus, levofloxacin-induced apoptosis exhibits characteristics of anoikis, the process by which cell death is triggered by separation from the extracellular matrix, which contains COL2. Furthermore, real-time quantitative RT-PCR was used to further confirm that levofloxacin downregulates COL2 expression in a dose-dependent manner. At last, western blot was used to find that levofloxacin increased the ratio of Bax/Bcl-2 and active caspase-3 in a dose-dependent manner. Levofloxacin therefore increases the effects of serum deprivation on anoikis by downregulating COL2 in rat NPCs in vitro via Bax/Bcl-2/caspase-3 pathway. This research provides a novel insight into the mechanisms of levofloxacin-induced toxicity and may potentially lead to a better understanding of the clinical effects of levofloxacin, especially in terms of intervertebral disc degeneration.
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Evaluation of the after-effects of cyanobacterial cell removal and lysis by photocatalysis using Ag/AgBr/TiO2.
Water Sci. Technol.
PUBLISHED: 09-17-2014
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This study aimed to evaluate the after-effects of cyanobacterial cell removal and lysis by photocatalysis in water. A low concentration of 50 mg/L Ag/AgBr/TiO2 was applied to inactivate Microcystis aeruginosa under visible light irradiation. Most of the M. aeruginosa was killed within 5 h while microcystins-LR (MC-LR) was released into water and accumulated to a high concentration of 100 ?g/L. Organic constituents released from cell damage led to 70 mg/L of total organic carbon (TOC) in water. The release of MC-LR and TOC would affect the biostability in the receiving water. Further, mineralization of cell lysis after photocatalysis over a long time resulted in the release of nutrients in water which would be a risk to cause cyanobacterial blooming again. Therefore, these after-effects should not be ignored when photochemical catalysis is applied to mitigate cyanobacterial blooming. Perhaps the best treatment is to remove intact cyanobacterial cells from water and then treat them off-site, for example by anaerobic digestion.
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Individual and joint toxic effects of cadmium sulfate and ?-naphthoflavone on the development of zebrafish embryo.
J Zhejiang Univ Sci B
PUBLISHED: 09-04-2014
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This paper aims to evaluate the individual and joint toxicities of cadmium sulfate (CdSO4) and ?-naphthoflavone (ANF) in zebrafish embryos. As a result, CdSO4 caused both lethal and sub-lethal effects, such as 24 h post-fertilization (hpf) death and 72 hpf delayed hatching. However, ANF only caused sub-lethal effects, including 48 hpf cardiac edema and 72 hpf delayed hatching. Taking 24 hpf death and 48 hpf cardiac edema as endpoints, the toxicities of CdSO4 and ANF were significantly enhanced by each other. Consistently, both CdSO4 and ANF caused significant oxidative stress, including decreases in the reduced glutathione (GSH) level, inhibition of superoxide dismutase (SOD) activity, as well as increases in malondialdehyde (MDA) content in zebrafish embryos, but these mixtures produced much more significant alterations on the biomarkers. Co-treatment of CdSO4 and ANF significantly down-regulated the mRNA level of multidrug resistance-associated protein (mrp) 1 and cytochrome P450 (cyp) 1a, which constituted the protective mechanisms for zebrafish embryos to chemical toxins. In conclusion, co-treatment of CdSO4 and ANF exhibited a much more severe damage in zebrafish embryos than individual treatment. Meanwhile, production of oxidative stress and altered expression of mrp1 and cyp1a could be important components of such joint toxicity.
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Glucocorticoids transcriptionally regulate miR-27b expression promoting body fat accumulation via suppressing the browning of white adipose tissue.
Diabetes
PUBLISHED: 09-03-2014
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Long-term glucocorticoid treatment induces central fat accumulation and metabolic dysfunction. We demonstrate that microRNA-27b plays a central role in the pathogenesis of glucocorticoid-induced central fat accumulation. Overexpression of miR-27b had the same effects as dexamethasone treatment in the inhibition of brown adipose differentiation and the energy expenditure of primary adipocyte. Conversely, antagonizing miR-27b function prevented dexamethasone suppression of the expression of BAT-specific genes. GCs transcriptionally regulate the miR-27b expression via a GR-mediated direct DNA binding mechanism, and miR-27b suppresses browning white adipose tissue by targeting the 3'UTR of Prdm16. In vivo, antagonizing miR-27b function in dexamethasone treated mice resulted in the efficient induction of brown adipocytes within white adipose tissue (WAT), improved GC-induced central fat accumulation. Collectively, our results indicate that miR-27b functions as a central target of glucocorticoid and as an upstream regulator of Prdm16 to control browning of WAT and consequently may represent a potential target in preventing obesity.
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[A twin study in Qingdao and Lishui: heritability of exercise participation and sedentary behavior].
Zhonghua Liu Xing Bing Xue Za Zhi
PUBLISHED: 09-02-2014
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To investigate both genetic and environmental influences on the exercise participation and sedentary behavior in Qingdao, Shandong province and 326 in Lishui, Zhejiang province.
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CD82 Restrains Pathological Angiogenesis by Altering Lipid Raft Clustering and CD44 Trafficking in Endothelial Cells.
Circulation
PUBLISHED: 08-22-2014
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Angiogenesis is crucial for many pathological processes and becomes a therapeutic strategy against diseases ranging from inflammation to cancer. The regulatory mechanism of angiogenesis remains unclear. Although tetraspanin CD82 is widely expressed in various endothelial cells (ECs), its vascular function is unknown.
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The construction of hierarchical structure on Ti substrate with superior osteogenic activity and intrinsic antibacterial capability.
Sci Rep
PUBLISHED: 08-22-2014
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The deficient osseointegration and implant-associated infections are pivotal issues for the long-term clinical success of endosteal Ti implants, while development of functional surfaces that can simultaneously overcome these problems remains highly challenging. This study aimed to fabricate sophisticated Ti implant surface with both osteogenic inducing activity and inherent antibacterial ability simply via tailoring surface topographical features. Micro/submciro/nano-scale structure was constructed on Ti by three cumulative subtractive methods, including sequentially conducted sandblasting as well as primary and secondary acid etching treatment. Topographical features of this hierarchical structure can be well tuned by the time of the secondary acid treatment. Ti substrate with mere micro/submicro-scale structure (MS0-Ti) served as a control to examine the influence of hierarchical structures on surface properties and biological activities. Surface analysis indicated that all hierarchically structured surfaces possessed exactly the same surface chemistry as that of MS0-Ti, and all of them showed super-amphiphilicity, high surface free energy, and high protein adsorption capability. Biological evaluations revealed surprisingly antibacterial ability and excellent osteogenic activity for samples with optimized hierarchical structure (MS30-Ti) when compared with MS0-Ti. Consequently, for the first time, a hierarchically structured Ti surface with topography-induced inherent antibacterial capability and excellent osteogenic activity was constructed.
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Drug-coated balloon angioplasty for drug-eluting stent restenosis: Insight from randomized controlled trials.
Ann. Med.
PUBLISHED: 08-19-2014
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Background. The best treatment option for drug-eluting stent (DES) restenosis has not been established. We performed a meta-analysis to assess the clinical efficacy of drug-coated balloon (DCB) for the treatment of DES restenosis. Methods. Trials were identified through a literature search from January 2005 through April 2014. All randomized controlled trials were eligible for inclusion if they compared DCB with a control treatment (plain old balloon angioplasty [POBA] or DES) in patients with DES restenosis. Results. Five studies and a total of 864 patients were included in this analysis. Most end-points were significantly reduced for DCB compared with the control groups. For major adverse cardiac events, the relative risk (RR) was 0.49 (P = 0.012); for target lesion revascularization, it was 0.50 (P = 0.044); for recurrent restenosis, it was 0.41 (P = 0.002). There was a lower mortality for DCB (RR 0.29; P = 0.017). The incidence of myocardial infarction was numerically lower, but without statistical significance (RR 0.76; P = 0.55). The DCB effect was more pronounced when compared with POBA than when compared with DES. Conclusions. This meta-analysis showed that DCB was superior to POBA and comparable to DES for treatment of DES restenosis. The findings in this meta-analysis cannot be extrapolated to DCB in general, because all DCB used in trials included was a single brand of paclitaxel-coated balloon.
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Kinetics of homoallylic/homobenzylic rearrangement reactions under combustion conditions.
J Phys Chem A
PUBLISHED: 08-14-2014
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Homoallylic/homobenzylic radicals refer to typical radicals with the radical site located at the ? position from the vinyl/phenyl group. These radicals are largely involved in combustion systems, such as the pyrolysis or oxidation of alkenes, cycloalkanes, and aromatics. The 1,2-vinyl/phenyl migration via two steps (cyclization/fission) is a peculiar reaction type for the homoallylic/homobenzylic radicals, entitled homoallylic/homobenzylic rearrangement, which has been studied by theoretical calculations including the Hirshfeld atomic charge analysis in the present work. With the help of rate constant calculations, the competition between this reaction channel and other possible pathways under combustion temperatures (500-2000 K) were evaluated. Analogous 1,3- and 1,4-vinyl/phenyl migration reactions for similar radicals with the radical sites located at the ? and ? positions from the vinyl/phenyl group were also computed. The results indicate that the 1,2-vinyl/phenyl migration is particularly important for the kinetics of unimolecular reactions of homoallylic radicals under 1500 K; nevertheless, it still has noticeable contribution at higher temperature. For those radicals with the radical site at the ? or ? positions, the respective 1,3- or 1,4-vinyl/phenyl migration channel plays an insignificant role under combustion conditions.
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CRISPR-Cas9 Knockin Mice for Genome Editing and Cancer Modeling.
Cell
PUBLISHED: 08-11-2014
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CRISPR-Cas9 is a versatile genome editing technology for studying the functions of genetic elements. To broadly enable the application of Cas9 in vivo, we established a Cre-dependent Cas9 knockin mouse. We demonstrated in vivo as well as ex vivo genome editing using adeno-associated virus (AAV)-, lentivirus-, or particle-mediated delivery of guide RNA in neurons, immune cells, and endothelial cells. Using these mice, we simultaneously modeled the dynamics of KRAS, p53, and LKB1, the top three significantly mutated genes in lung adenocarcinoma. Delivery of a single AAV vector in the lung generated loss-of-function mutations in p53 and Lkb1, as well as homology-directed repair-mediated Kras(G12D) mutations, leading to macroscopic tumors of adenocarcinoma pathology. Together, these results suggest that Cas9 mice empower a wide range of biological and disease modeling applications.
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Association between thrombomodulin polymorphisms and coronary artery disease risk: a meta-analysis.
Med. Sci. Monit.
PUBLISHED: 08-10-2014
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The associations between the thrombomodulin (TM) polymorphisms and coronary artery disease (CAD) risk remain controversial. The aim of this study was to evaluate the association of TM polymorphisms with CAD susceptibility using a meta-analysis approach.
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Efficient CRISPR-Cas9-mediated genome editing in Plasmodium falciparum.
Nat. Methods
PUBLISHED: 08-10-2014
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Malaria is a major cause of global morbidity and mortality, and new strategies for treating and preventing this disease are needed. Here we show that the Streptococcus pyogenes Cas9 DNA endonuclease and single guide RNAs (sgRNAs) produced using T7 RNA polymerase (T7 RNAP) efficiently edit the Plasmodium falciparum genome. Targeting the genes encoding native knob-associated histidine-rich protein (kahrp) and erythrocyte binding antigen 175 (eba-175), we achieved high (? 50-100%) gene disruption frequencies within the usual time frame for generating transgenic parasites.
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CRISPR-mediated direct mutation of cancer genes in the mouse liver.
Nature
PUBLISHED: 08-06-2014
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The study of cancer genes in mouse models has traditionally relied on genetically-engineered strains made via transgenesis or gene targeting in embryonic stem cells. Here we describe a new method of cancer model generation using the CRISPR/Cas (clustered regularly interspaced short palindromic repeats/CRISPR-associated proteins) system in vivo in wild-type mice. We used hydrodynamic injection to deliver a CRISPR plasmid DNA expressing Cas9 and single guide RNAs (sgRNAs) to the liver that directly target the tumour suppressor genes Pten (ref. 5) and p53 (also known as TP53 and Trp53) (ref. 6), alone and in combination. CRISPR-mediated Pten mutation led to elevated Akt phosphorylation and lipid accumulation in hepatocytes, phenocopying the effects of deletion of the gene using Cre-LoxP technology. Simultaneous targeting of Pten and p53 induced liver tumours that mimicked those caused by Cre-loxP-mediated deletion of Pten and p53. DNA sequencing of liver and tumour tissue revealed insertion or deletion mutations of the tumour suppressor genes, including bi-allelic mutations of both Pten and p53 in tumours. Furthermore, co-injection of Cas9 plasmids harbouring sgRNAs targeting the ?-catenin gene and a single-stranded DNA oligonucleotide donor carrying activating point mutations led to the generation of hepatocytes with nuclear localization of ?-catenin. This study demonstrates the feasibility of direct mutation of tumour suppressor genes and oncogenes in the liver using the CRISPR/Cas system, which presents a new avenue for rapid development of liver cancer models and functional genomics.
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The genus Anahita from Wuyi Mountains, Fujian, China, with description of one new species (Araneae: Ctenidae).
Zootaxa
PUBLISHED: 08-06-2014
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The family Ctenidae is represented by 40 genera and 490 species in the world, distributed mainly in America, Africa and Asia. Sixty-three species have been recorded from Asia (Jäger 2012), however, only nine species in four genera (Anahita Karsch, 1879; Ctenus Walckenaer, 1805; Leptoctenus L. Koch, 1878 and Sinoctenus Marusik, Zhang & Omelko, 2012) have been reported from China (Platnick 2014). The genus Anahita was established by Karsch in 1879 for the type species A. fauna Karsch, 1879. Silva (2003) found that all species in the genus can be recognised by the absence of the retrolateral tibial apophysis and the presence of a hyaline area in the female epigynum. Jäger (2012) reviewed Asian Anahita, illustrated the type species according to the new materials, transferred four species from the genus Ctenus to Anahita and described one new species. Presently, this genus contains 27 species, among them five species are recorded from China (Platnick 2014): A. fauna Karsch, 1879; A. maolan Zhu, Chen & Song, 1999; A. samplexa Yin, Tang & Gong, 2000; A. jianfengensis Zhang, Hu & Han, 2011 and A. jinsi Jäger, 2012 (Jäger 2012; Yin et al. 2000; Zhang et al. 2011; Zhu et al. 1999). 
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Optogenetics: opsins and optical interfaces in neuroscience.
Cold Spring Harb Protoc
PUBLISHED: 08-03-2014
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Optogenetics is defined as the integration of optics and genetics to control well-defined events within specified cells of living tissue. In this introduction, we focus on the basic techniques necessary for employing microbial opsins as optogenetic tools in mammalian brains. We provide a guide for the fundamentals of optogenetic application-selecting an opsin, implementing expression of opsins based on the neuroscientific experimental requirements, and adapting the corresponding optical hardware for delivery of light into mammalian brains.
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Establishing a fiber-optic-based optical neural interface.
Cold Spring Harb Protoc
PUBLISHED: 08-03-2014
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Selective expression of opsins in genetically defined neurons makes it possible to control a subset of neurons without affecting nearby cells and processes in the intact brain, but light must still be delivered to the target brain structure. Light scattering limits the delivery of light from the surface of the brain. For this reason, we have developed a fiber-optic-based optical neural interface (ONI), which allows optical access to any brain structure in freely moving mammals. The ONI system is constructed by modifying the small animal cannula system from PlasticsOne. The system for bilateral stimulation consists of a bilateral cannula guide that has been stereotactically implanted over the target brain region, a screw cap for securing the optical fiber to the animal's head, a fiber guard modified from the internal cannula adapter, and a bare fiber whose length is customized based on the depth of the target region. For unilateral stimulation, a single-fiber system can be constructed using unilateral cannula parts from PlasticsOne. We describe here the preparation of the bilateral ONI system and its use in optical stimulation of the mouse or rat brain. Delivery of opsin-expressing virus and implantation of the ONI may be conducted in the same surgical session; alternatively, with a transgenic animal no opsin virus is delivered during the surgery. Similar procedures are useful for deep or superficial injections (even for neocortical targets, although in some cases surface light-emitting diodes or cortex-apposed fibers can be used for the most superficial cortical targets).
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Efficacy of the End-to-Side Neurorrhaphies with Epineural Window and Partial Donor Neurectomy in Peripheral Nerve Repair: An Experimental Study in Rats.
J Reconstr Microsurg
PUBLISHED: 08-01-2014
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Background?End-to-side (ETS) neurorrhaphy was a useful tool in peripheral nerve repair and "baby-sitter" procedure. The study was designed to evaluate the long-term efficacy of ETS with epineurial window and 40% partial donor neurectomy in rats. Materials and Methods A total of 60 Lewis rats were divided into three groups (n?=?20, each group): following peroneal nerve transection, rats in Group A underwent end-to-end neurorrhaphy; rats in Group B underwent ETS neurorrhaphy of the distal peroneal nerve stump to an epineurial window on the tibial nerve; and rats in Group C underwent ETS neurorrhaphy of the distal peroneal nerve stump to the tibial nerve with 40% partial neurectomy. At 6, 12, 18, and 24 weeks after surgery, electrophysiology, muscle tension, myelinated fiber regeneration, cross-sectional area of muscular fiber, and muscle weight were analyzed. Results?Histology exhibited apparently increased number and size of myelinated fibers in peroneal nerves in Group C, compared with those in Group B. More superior recovery was demonstrated in the electrophysiology and axon regeneration of the peroneal nerves, as well as the maintenance of muscle force, wet weight, and fiber size of the anterior tibial muscles in Group C than those in Group B. Conclusion?ETS neurorrhaphy with partial donor neurectomy can achieve higher efficacy in functional and structural recovery of the recipient system. This study provides the evidence of long-term follow-up for the further investigation of ETS neurorrhaphies with different modalities in peripheral nerve repair and in "baby-sitter" procedure.
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Pre-ischemic exercise alleviates oxidative damage following ischemic stroke in rats.
Exp Ther Med
PUBLISHED: 07-31-2014
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Physical exercise has been proved to be neuroprotective in clinical trials and animal experiments. However, the exact mechanism underlying this neuroprotective effect remains unclear. The aim of the present study was to explore whether pre-ischemic treadmill training could act as a form of ischemic preconditioning in a rat following ischemic stroke by reducing oxidative damage. Fifty-four rats were randomly divided into three groups (n=18 per group): Sham surgery, middle cerebral artery occlusion (MCAO) without exercise and MCAO with exercise. Subsequent to treadmill training, ischemic stroke was induced by occluding the MCA for 1.5 h, followed by reperfusion. Six rats in each group were evaluated for neurological deficits and then sacrificed by decapitation to calculate the infarct volume. The remaining rats in each group were sacrificed to detect the level of superoxide dismutase (SOD) activity (n=6) and malondialdehyde (MDA) concentration (n=6). The results indicated that pre-ischemic exercise training reduced brain infarct volume and neurological deficits, increased SOD activity and decreased the concentration of MDA following ischemic stroke. In conclusion, treadmill exercise training prior to MCAO/reperfusion increased the antioxidant ability and decreased the oxidative damage in the brain subsequent to ischemic stroke.
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Submandibular artery: Bilobed platysma myocutaneous flap for total lower lip reconstruction.
J Craniomaxillofac Surg
PUBLISHED: 07-23-2014
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Lower lip reconstruction following cancer resection includes a variety of clinical and microsurgical options.
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2-Guanidinoquinazolines as new inhibitors of the STAT3 pathway.
Bioorg. Med. Chem. Lett.
PUBLISHED: 07-18-2014
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Synthesis and SAR investigation of 2-guanidinoquinazolines, initially identified in a high content screen for selective STAT3 pathway inhibitors, led to a more potent analog (11c) that demonstrated improved anti-proliferative activity against a panel of HNSCC cell lines.
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Overexpression of a cotton annexin gene, GhAnn1, enhances drought and salt stress tolerance in transgenic cotton.
Plant Mol. Biol.
PUBLISHED: 07-06-2014
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Plant annexins are members of a diverse, multigene protein family that has been associated with a variety of cellular processes and responses to abiotic stresses. GhAnn1, which encodes a putative annexin protein, was isolated from a cotton (Gossypium hirsutum L. acc 7235) cDNA library. Tissue-specific expression showed that GhAnn1 is expressed at differential levels in all tissues examined and strongly induced by various phytohormones and abiotic stress. In vivo and in vitro subcellular localization suggested that GhAnn1 is located in the plasma membrane. In response to drought and salt stress, transgenic cotton plants overexpressing GhAnn1 showed significantly higher germination rates, longer roots, and more vigorous growth than wild-type plants. In addition, plants overexpressing GhAnn1 had higher total chlorophyll content, lower lipid peroxidation levels, increased peroxidase activities, and higher levels of proline and soluble sugars, all of which contributed to increased salt and drought stress tolerance. However, transgenic cotton plants in which the expression of GhAnn1 was suppressed showed the opposite results compared to the overexpressing plants. These findings demonstrated that GhAnn1 plays an important role in the abiotic stress response, and that overexpression of GhAnn1 in transgenic cotton improves salt and drought tolerance.
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Numerical study on determining formation porosity using a boron capture gamma ray technique and MCNP.
Appl Radiat Isot
PUBLISHED: 07-02-2014
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Formation porosity can be determined using the boron capture gamma ray counting ratio with a near to far detector in a pulsed neutron-gamma element logging tool. The thermal neutron distribution, boron capture gamma spectroscopy and porosity response for formations with different water salinity and wellbore diameter characteristics were simulated using the Monte Carlo method. We found that a boron lining improves the signal-to-noise ratio and that the boron capture gamma ray counting ratio has a higher sensitivity for determining porosity than total capture gamma.
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Relationship of Serum Heart-Type Fatty Acid-Binding Protein Levels and Cerebral Infarction: a Meta-analysis.
Mol. Neurobiol.
PUBLISHED: 06-20-2014
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Clinical scores are recommended for predicting cardiovascular and cerebrovascular risk in patients with cerebral infarction to inform secondary prevention. Blood biomarkers may improve prediction beyond clinical scores, and we conducted this analysis for the purpose of interpreting the association between serum H-FABP levels and cerebral infarction. Potential relevant studies were identified covering PubMed, Embase, Web of Science, Cochrane Library, CISCOM, CINAHL, Google Scholar, CBM and CNKI databases. Two reviewers extracted data and assessed studies independently. Statistical analyses were conducted with the version 12.0 Stata statistical software. A total of 9 case-control papers that evaluated the correlation of serum H-FABP levels with cerebral infarction, including 1,176 subjects (patients?=?662, healthy controls?=?514) were reviewed. Positive association was detected between serum H-FABP levels and cerebral infarction (SMD?=?1.70, 95 %CI: 1.13-2.27, P?
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Trans-omics pathway analysis suggests that eQTLs contribute to chondrocyte apoptosis of Kashin-Beck disease through regulating apoptosis pathway expression.
Gene
PUBLISHED: 06-18-2014
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Kashin-Beck disease (KBD) is a serious osteoarthropathia, mainly characterized by excessive chondrocyte necrosis and apoptosis. The molecular signaling pathways underlying KBD excessive chondrocyte apoptosis remain unclear, leading to a lack of effective medical interventions now. To clarify whether expression quantitative trait loci (eQTLs) contribute to excessive chondrocyte apoptosis of Kashin-Beck disease through regulating the expression of apoptosis pathways. We conducted a genome-wide eQTLs based pathway association analysis of KBD using Affymetrix Human SNP Array 6.0 in 1717 Chinese Han subjects. PLINK software was used for genome-wide association study (GWAS) of KBD. A modified gene set enrichment algorithm was applied for pathway association analysis based on GWAS results. The KBD-associated pathways were compared with abnormally expressed pathways in KBD articular cartilage, identified by microarray study of KBD. We identified 4 eQTLs pathways, which were not only significantly associated with KBD, but also abnormally expressed in KBD articular cartilage, including REACTOME_INTRINSIC_PATHWAY_FOR_APOPTOSIS (P=0.008), MAHAJAN _RESPONSE_TO_IL1A_UP (P=0.010), KEGG_PEROXISOME (P=0.005) and MARKS_HDAC_TARGETS_UP (P=0.006). Our results suggest that eQTLs contributed to KBD excessive chondrocyte apoptosis through regulating the expression of apoptosis related pathways. This study provides novel insight into the genetic susceptibility and therapeutic rationale of KBD.
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Genomic analysis of Agrobacterium radiobacter DSM 30147(T) and emended description of A. radiobacter (Beijerinck and van Delden 1902) Conn 1942 (Approved Lists 1980) emend. Sawada et al. 1993.
Stand Genomic Sci
PUBLISHED: 06-15-2014
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Agrobacterium radiobacter is the only known non-phytopathogenic species in Agrobacterium genus. In this study, the whole-genome sequence of A. radiobacter type strain DSM 30147(T) was described and compared to the other available Agrobacterium genomes. This bacterium has a genome size of 7,122,065 bp distributed in 612 contigs, including 6,834 protein-coding genes and 41 RNA genes. It harbors a circular chromosome and a linear chromosome but not a tumor-inducing (Ti) plasmid. To the best of our knowledge, this is the first report of a genome from the A. radiobacter species. In addition, an emended description of A. radiobacter is described. This study reveals information that enhances the current understanding of its non-phytopathogenicity and its phylogenetic position within Agrobacterium genus.
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Update on implications and mechanisms of angiogenesis in liver fibrosis.
Hepatol. Res.
PUBLISHED: 06-09-2014
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Liver fibrosis occurs as a compensatory response to the process of tissue repair in a wide range of chronic liver injures. It is characterized by excessive deposition of extracellular matrix in liver tissues. As the pathogenesis progresses without effective management, it will lead to formation of liver fiber nodules and disruption of normal liver structure and function, finally culminating in cirrhosis and hepatocellular carcinoma. A new discovery shows that liver angiogenesis is strictly associated with, and may even favor fibrogenic progression of chronic liver diseases. Recent basic and clinical investigations also demonstrate that liver fibrogenesis is accompanied by pathological angiogenesis and sinusoidal remodeling, which critically determine the pathogenesis and prognosis of liver fibrosis. Inhibition of pathological angiogenesis is considered to be a new strategy for the treatment of liver fibrosis. This review summarizes current knowledge on the process of angiogenesis, the relationships between angiogenesis and liver fibrosis, and on the molecular mechanisms of liver angiogenesis. On the other hand, it also presents the different strategies that have been used in experimental models to counteract excessive angiogenesis and the role of angiogenesis in the prevention and treatment of liver fibrosis.
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Pathological feature and immunoprofile of cystitis glandularis accompanied with upper urinary tract obstruction.
Biomed Res Int
PUBLISHED: 05-29-2014
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To explore the pathological feature and immunoprofile of immunoprofile accompanied with upper urinary tract obstruction and the immunoprofile in various types of glandular cystitis.
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In vivo interrogation of gene function in the mammalian brain using CRISPR-Cas9.
Nat. Biotechnol.
PUBLISHED: 05-27-2014
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Probing gene function in the mammalian brain can be greatly assisted with methods to manipulate the genome of neurons in vivo. The clustered, regularly interspaced, short palindromic repeats (CRISPR)-associated endonuclease (Cas)9 from Streptococcus pyogenes (SpCas9) can be used to edit single or multiple genes in replicating eukaryotic cells, resulting in frame-shifting insertion/deletion (indel) mutations and subsequent protein depletion. Here, we delivered SpCas9 and guide RNAs using adeno-associated viral (AAV) vectors to target single (Mecp2) as well as multiple genes (Dnmt1, Dnmt3a and Dnmt3b) in the adult mouse brain in vivo. We characterized the effects of genome modifications in postmitotic neurons using biochemical, genetic, electrophysiological and behavioral readouts. Our results demonstrate that AAV-mediated SpCas9 genome editing can enable reverse genetic studies of gene function in the brain.
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Qualitative and quantitative analysis of the major constituents in Acorus tatarinowii Schott by HPLC/ESI-QTOF-MS/MS.
Biomed. Chromatogr.
PUBLISHED: 05-21-2014
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Acorus tatarinowii Schott (ATS) is a well-known traditional Chinese medicine (TCM) for the treatment of epilepsy, amnesia and insomnia. In this study, a methodology utilizing high-performance liquid chromatography (HPLC) coupled with electrospray ionization quadrupole time-of-flight mass spectrometry (ESI-QTOF-MS/MS) was established for the separation and structural identification of the major chemical constituents in ATS for the first time. Overall, 46 major constituents including flavonoid glycosides, phenylpropane derivatives, amides and lignans were identified or tentatively characterized. Seven major constituents, including four phenylpropane derivatives and three lignans, were further quantified as marker substances, which showed good linearity within the test ranges. These results indicated that the developed quantitative method was linear, sensitive, and precise for quality control of ATS. Copyright © 2014 John Wiley & Sons, Ltd.
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Protective effects of icariin?mediated SIRT1/FOXO3 signaling pathway on intestinal ischemia/reperfusion?induced acute lung injury.
Mol Med Rep
PUBLISHED: 05-15-2014
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Acute lung injury (ALI) is a common complication following intestinal ischemia/reperfusion (I/R) and is a major contributing factor to its high mortality rate. Sirtuin 1 (SIRT1), a NAD+?dependent deacetylase, has been reported to have an important role in apoptosis inhibition, oxidative stress resistance and cell lifespan extension through its deacetylation of forkhead box protein O3 (FOXO3). It has been demonstrated that icariin (ICA), a flavonoid extracted from Epimedium, upregulates SIRT1 expression. The aim of the present study was to examine whether ICA?mediated SIRT1/FOXO3 signaling pathway activation had a protective effect on intestinal I/R?induced ALI. The effects of ICA on intestinal I/R?induced ALI and its regulation of the SIRT1/FOXO3 signaling pathway on intestinal I/R?induced ALI were investigated in rats. The results demonstrated that ICA pretreatment markedly reduced intestinal I/R?induced ALI as indicated by histological alterations, including decreased tumor necrosis factor?? (TNF??), interleukin 6 (IL?6), reduced oxidative stress, acetylated FOXO3 and B?cell lymphoma 2 (Bcl?2)?interacting mediator of cell death levels, and increased glutathione (GSH), GSH peroxidase, SIRT1, manganese superoxide dismutase and Bcl?2 levels in rat lung tissues. Furthermore, ICA pretreatment upregulated SIRT1 expression, which then downregulated FOXO3 acetylation. In conclusion, ICA exhibited significant protective effects in intestinal I/R?induced ALI. The protective effect of ICA may be attributed to the upregulation of SIRT1, which contributed to FOXO3 deacetylation and the modulation of downstream antioxidative and anti?apoptotic factors.
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Regeneration of high-quality silk fibroin fiber by wet spinning from CaCl2-formic acid solvent.
Acta Biomater
PUBLISHED: 04-29-2014
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Silks spun by silkworms and spiders feature outstanding mechanical properties despite being spun under benign conditions. The superior physical properties of silk are closely related to its complicated hierarchical structures constructed from nanoscale building blocks, such as nanocrystals and nanofibrils. Here, we report a novel silk dissolution behavior, which preserved nanofibrils in CaCl2-formic acid solution, that enables spinning of high-quality fibers with a hierarchical structure. This process is characterized by simplicity, high efficiency, low cost, environmental compatibility and large-scale industrialization potential, as well as having utility and potential for the recycling of silk waste and the production of silk-based functional materials.
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Genome-wide association study identifies ITPR2 as a susceptibility gene for Kashin-Beck disease in Han Chinese.
PUBLISHED: 04-09-2014
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Objective. Kashin-Beck Disease (KBD) is a chronic osteochondropathy, the pathogenesis of which remains elusive. To identify susceptibility genes for KBD, we conducted a two-stage genome-wide association study (GWAS) of KBD. Methods. 90 grade II or III KBD patients with extreme KBD phenotypes and 1,627 healthy controls were enrolled in initial GWAS. Affymetrix Genome Wide Human SNP Array 6.0 was applied for genotyping. For replication study, 9 SNPs of significant ITPR2 gene identified by the GWAS were tested in an independent validation sample containing 559 KBD patients and 467 healthy controls. Results. We identified a novel associated locus for KBD (ITRP2, rs10842750, P = 1.58×10(-) (8) ) in the GWAS. Replication study observed significant associations between KBD and the 9 validation SNPs of ITPR2 gene, including rs10842750 (P = 5.97×10(-3) ), rs16931011 (P = 1.29×10(-3) ), rs1531928 (P = 4.95×10(-3) ), rs4414322 (P = 4.40×10(-3) ), rs11048570 (P = 4.53×10(-3) ), rs11048572 (P = 4.43×10(-3) ), rs2017510 (P = 4.58×10(-3) ), rs9669395 (P = 5.77×10(-3) ) and rs1002835 (P = 4.85×10(-3) ). In KBD patients, the genotype score of rs10842750 (P = 0.013) was also correlated with KBD clinical severity grades. Conclusion. Our results strongly suggest that ITPR2 was a novel susceptibility gene for KBD in Han Chinese. This study may provide new insight into the pathogenesis and rationale of therapies for KBD as well as other osteoarthrosis with similar articular cartilage lesions. © 2014 American College of Rheumatology.
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N-acetylcysteine attenuates reactive-oxygen-species-mediated endoplasmic reticulum stress during liver ischemia-reperfusion injury.
World J. Gastroenterol.
PUBLISHED: 04-08-2014
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To investigate the effects of N-acetylcysteine (NAC) on endoplasmic reticulum (ER) stress and tissue injury during liver ischemia reperfusion injury (IRI).
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Enhancer Activation Requires trans-Recruitment of a Mega Transcription Factor Complex.
Cell
PUBLISHED: 04-03-2014
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Enhancers provide critical information directing cell-type-specific transcriptional programs, regulated by binding of signal-dependent transcription factors and their associated cofactors. Here, we report that the most strongly activated estrogen (E2)-responsive enhancers are characterized by trans-recruitment and in situ assembly of a large 1-2 MDa complex of diverse DNA-binding transcription factors by ER? at ERE-containing enhancers. We refer to enhancers recruiting these factors as mega transcription factor-bound in trans (MegaTrans) enhancers. The MegaTrans complex is a signature of the most potent functional enhancers and is required for activation of enhancer RNA transcription and recruitment of coactivators, including p300 and Med1. The MegaTrans complex functions, in part, by recruiting specific enzymatic machinery, exemplified by DNA-dependent protein kinase. Thus, MegaTrans-containing enhancers represent a cohort of functional enhancers that mediate a broad and important transcriptional program and provide a molecular explanation for transcription factor clustering and hotspots noted in the genome.
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C/EBP homologous protein (CHOP) contributes to hepatocyte death via the promotion of ERO1? signaling in acute liver failure.
Biochem. J.
PUBLISHED: 03-31-2014
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C/EBP Homologous Protein (CHOP) has been shown to be a key molecule in endoplasmic reticulum (ER) stress-mediated apoptosis. ERO1?, a target of CHOP, is an important oxidizing enzyme to regulate ROS, which play a prominent role in hepatocellular death during acute liver failure (ALF). However, little is known how CHOP facilitates ROS-induced hepatocellular injury. This study was designed to investigate the roles and molecular mechanisms of CHOP in ALF. In the liver tissues from ALF patients, the expression of CHOP was significantly increased, which accompanied by increased expression of PERK signaling, ATF6 signaling, IRE1 signaling and ERO1a, as compared with healthy controls. In the mouse model of GaIN/LPS-induced ALF, the hepatocellular injury was accompanied by upregulated PERK signaling, ATF6 signaling, IRE1 signaling, CHOP and ERO1a. In contrast, CHOP deficiency decreased hepatocellular apoptosis/necrosis and increased animal survival. Furthermore, disruption of CHOP decreased ERO1a expression leading to reducing ROS-induced cell death in vivo and in vitro. Interestingly, ERO1a overexpression restored GaIN/LPS-induced hepatocellular injury in CHOP-deficient mice. Our studies demonstrate for the first time that CHOP promotes liver damage during ALF through activation of ERO1a, a key mediator to link ER stress and ROS. Therefore, targeting CHOP/ERO1a signaling could be a novel therapeutic approach during ALF.
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Genome-wide pathway-based association study implicates complement system in the development of Kashin-Beck disease in Han Chinese.
Bone
PUBLISHED: 03-14-2014
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Kashin-Beck disease (KBD) is a chronic osteochondropathy. The pathogenesis of KBD remains unknown. To identify relevant biological pathways for KBD, we conducted a genome-wide pathway-based association study (GWPAS) following by replication analysis, totally using 2,743 Chinese Han adults. A modified gene set enrichment algorithm was used to detect association between KBD and 963 biological pathways. Cartilage gene expression analysis and serum complement measurement were performed to evaluate the functional relevance of identified pathway with KBD. We found that the Complement and Coagulation Cascades (CACC) pathway was significantly associated with KBD (P value=3.09×10(-5), false-discovery rate=0.042). Within the CACC pathway, the most significant association was observed at rs1656966 (P value=1.97×10(-4)) of KNG1 gene. Further replication study observed that rs1656966 (P value=0.037) was significantly associated with KBD in an independent validation sample of 1,026 subjects. Gene expression analysis observed that CFD (ratio=3.39±2.68), A2M (ratio=3.67±5.63), C5 (ratio=2.65±2.52) and CD46 (ratio=2.29±137) genes of the CACC pathway were up-regulated in KBD articular cartilage compared to healthy articular cartilage. The serum level of complement C5 in KBD patients were significantly higher than that in healthy controls (P value=0.038). Our study is the first to suggest that complement system-related CACC pathway contributed to the development of KBD.
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East china sea storm surge modeling and visualization system: the typhoon soulik case.
ScientificWorldJournal
PUBLISHED: 03-11-2014
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East China Sea (ECS) Storm Surge Modeling System (ESSMS) is developed based on Regional Ocean Modeling System (ROMS). Case simulation is performed on the Typhoon Soulik, which landed on the coastal region of Fujian Province, China, at 6?pm of July 13, 2013. Modeling results show that the maximum tide level happened at 6?pm, which was also the landing time of Soulik. This accordance may lead to significant storm surge and water level rise in the coastal region. The water level variation induced by high winds of Soulik ranges from -0.1 to 0.15?m. Water level generally increases near the landing place, in particular on the left hand side of the typhoon track. It is calculated that 0.15?m water level rise in this region can cause a submerge increase of ~0.2?km(2), which could be catastrophic to the coastal environment and the living. Additionally, a Globe Visualization System (GVS) is realized on the basis of World Wind to better provide users with the typhoon/storm surge information. The main functions of GVS include data indexing, browsing, analyzing, and visualization. GVS is capable of facilitating the precaution and mitigation of typhoon/storm surge in ESC in combination with ESSMS.
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Comparison of the clinical effects of open and closed chain exercises after medial patellofemoral ligament reconstruction.
J Phys Ther Sci
PUBLISHED: 03-11-2014
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[Purpose] To compare the effects of open-chain exercise (OCE) and closed-chain exercise (CCE) for patients after medial patellofemoral ligament (MPFL) reconstruction. [Subjects and Methods] Forty patients after MPFL reconstruction were randomly divided into an OCE group and a CCE group. All the patients were evaluated at four different time points. [Results] The mean change of thigh circumference decrease in the CCE group was lower than that in the OCE group at both the 3rd and 6th month after surgery. The Lysholm score of the CCE group was higher than that of the OCE group at both the 3rd and 6th month. At the 3rd month after surgery, the visual analog scale score of the CCE group was lower than that of the OCE group. [Conclusion] CCE is better than OCE for both short and long term outcomes of patients after MPFL reconstruction.
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Exercise pre?conditioning alleviates brain damage via excitatory amino acid transporter 2 and extracellular signal?regulated kinase 1/2 following ischemic stroke in rats.
Mol Med Rep
PUBLISHED: 02-01-2014
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Previous studies have reported that physical exercise may exert a neuroprotective effect in humans as well as animals. However, the detailed mechanisms underlying the neuroprotective effect of exercise has remained to be elucidated. The aim of the present study was to explore the possible signaling pathways involved in the protective effect of pre?ischemic treadmill training for ischemic stroke in rats. A total of 36 male Sprague?Dawley rats were divided at random into three groups as follows (n=12 for each): Sham surgery group; middle cerebral artery occlusion (MCAO) group; and exercise with MCAO group. Following treadmill training for three weeks, the middle cerebral artery was occluded for 90 min in order to induce ischemic stroke, followed by reperfusion. Following 24 h post?reperfusion, six rats from each group were assessed for neurological deficits and then sacrificed to calculate the infarct volume. The remaining rats (n=6 for each group) were sacrificed and the expression levels of excitatory amino acid transporter 2 (EAAT?2) and extracellular signal?regulated kinase 1/2 (ERK1/2) were detected using western blot analysis. The results of the present study demonstrated that rats that underwent pre?ischemic exercise intervention had a significantly decreased brain infarct volume and neurological deficits; in addition, the pre?ischemic exercise group showed decreased overexpression of phosphorylated ERK1/2 and increased expression of EAAT?2 following ischemic stroke. In conclusion, treadmill training exercise prior to ischemic stroke alleviated brain damage in rats via regulation of EAAT?2 and ERK1/2.
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Intrabiliary RF heat-enhanced local chemotherapy of a cholangiocarcinoma cell line: monitoring with dual-modality imaging--preclinical study.
Radiology
PUBLISHED: 01-30-2014
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To determine whether magnetic resonance (MR) imaging heating guidewire-mediated radiofrequency (RF) hyperthermia could enhance the therapeutic effect of gemcitabine and 5-fluorouracil (5-FU) in a cholangiocarcinoma cell line and local deposit doses of chemotherapeutic drugs in swine common bile duct (CBD) walls.
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Should GLP-1 receptor agonists be used with caution in high risk population for colorectal cancer?
Med. Hypotheses
PUBLISHED: 01-16-2014
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Glucagon-like peptide-1 (GLP-1) receptor agonists, with few reported side effects, are a kind of very promising anti-diabetes drugs. They are thus being paid high hopes on by both doctors and patients. GLP-1 receptor agonists, however, have not been used in clinic for a long time. Some unknown potential adverse effects may exist. Because GLP-1 receptor agonists enhance ? cells proliferation in pancreas via influencing Wnt/?-catenin pathway, a pathway which is associated with tumorigenesis in colon, we then assume that GLP-1 receptor agonists may increase the risk for colorectal cancer.
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Paris saponin ? inhibits metastasis by modulating matrix metalloproteinases in colorectal cancer cells.
Mol Med Rep
PUBLISHED: 01-08-2014
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Metastasis is the main cause of mortality of patients with cancer?related disease. Targeting the process of metastasis has been proposed as a potential strategy in cancer treatment. Trillium tschonoskii Maxim., a traditional Chinese medicine, is used for the treatment of numerous diseases, including cancer. The current study aimed to determine the anti?metastatic effect of Paris saponin VII (PS ?), which was extracted from T. tschonoskii Maxim., using SW620 and LoVo cells, two human metastatic colorectal cancer (CRC) cell lines. The present study conducted cell attachment, wound healing and migration assays to detect the anti?metastatic effects of PS VII on colorectal cells. In addition, gelatin zymography assay and western blot analysis were used to detect the possible mechanisms involved. The results of this study demonstrated that PS ? significantly suppresses the viability, attachment, migration and invasive abilities of CRC cells in a concentration?dependent manner. In addition, PS ? reduced the expression levels and activity of matrix metalloproteinase (MMP)?2 and MMP?9. These data indicate that PS ? reduces the metastatic capability of CRC cells, possibly via the downregulation of the expression and activity of MMP?2 and MMP?9. These results demonstrate a novel therapeutic potential for PS ? in anti?metastatic therapy.
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Expression and clinical significance of angiotensin II type 1 receptor in human hepatocellular carcinoma.
Exp Ther Med
PUBLISHED: 01-08-2014
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This study aimed to investigate the expression of angiotensin II type 1 receptor (AT-1R) mRNA and the AT-1R protein in human primary hepatocellular carcinoma (PHC), and to attempt to elucidate their association with pathological and clinical characteristics. Fresh tumor and normal liver tissues were obtained from 44 patients with PHC following hepatectomies. AT-1R mRNA levels were quantitatively analyzed by quantitative polymerase chain reaction (qPCR) while the protein levels were assessed by immunohistochemistry. The expression levels of AT-1R were observed in hepatocellular carcinoma tissues and normal liver tissues. The level of AT-1R protein expression in normal liver tissues was higher compared with that in PHC tissues (P=0.0033). The AT-1R mRNA levels were higher in patients with negative hepatitis B virus surface antigen (HBsAg), normal ?-fetoprotein (AFP) levels and high tumor differentiation, compared with those in patients with positive HBsAg (P=0.0005), upregulated AFP levels (P=0.0008) and poor tumor differentiation (P=0.0290). No significant correlation was identified between the expression levels of AT-1R mRNA and general characteristics such as gender, age, cirrhotic nodules, tumor size, tumor encapsulation, tumor number, carcinoma embolus, tumor metastasis or tumor recurrence. Downregulated levels of AT-1R mRNA and AT-1R protein may indicate a poor prognosis for patients with PHC.
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?-terpineol inhibits cell growth and induces apoptosis in human liver cancer BEL-7402 cells in vitro.
Int J Clin Exp Pathol
PUBLISHED: 01-01-2014
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To investigate the effect of ?-terpineol on cell proliferation and apoptosis of human hepatoma BEL-7402 cells to elucidate its molecular mechanism. Here, BEL-7402 cells were treated with various concentrations (40, 80, 160, 320 and 640 ?g/ml) of ?-terpineol for 48 h, cell proliferation was determined by 3-(4,5-dimethyl-thiazolyl-2)-2,5-diphenyl tetrazolium bromides (MTT) assay. Cell colony inhibition was determined by soft agar assay. Apoptosis and possible molecular mechanisms were evaluated by morphological observation, flow cytometry analysis, and DNA fragmentation assay. The ?-terpineol significantly suppressed BEL-7402 cell proliferation in a dose-dependent manner. Characteristic morphological and biochemical changes associated with apoptosis such as cells shrinkage, deformation and vacuolization of mitochondria, nuclear chromatin condensation and fragmentation, formation of apoptotic bodies were observed after BEL-7402 cells treated with ?-terpineol for 24 h and 48 h. Cell cycle were displayed by flow cytometry analysis, the ?-terpineol treatment resulted in accumulation of cells at G1 or S phase and a blockade of cell proliferation compared to control group. Treating BEL-7402 cells with 320 ?g/ml of ?-terpineol for 36 h and 48 h, a typical apoptotic "DNA ladder" was observed using DNA fragmentation assay. The present study demonstrated that possible anti-cancer mechanism of ?-terpineol on human hematomas cells is through inducing cell apoptosis to suppress tumor cell growth.
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The Neuroprotective Effect of Electro-Acupuncture Against Ischemic Stroke in Animal Model: A Review.
Afr J Tradit Complement Altern Med
PUBLISHED: 01-01-2014
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It is well established that electro-acupuncture can exert neuroprotection in animal experiments. However, the exact mechanism of electro-acupuncture against ischemic stroke is not very clear.
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Randomized controlled trial to compare growth parameters and nutrient adequacy in children with picky eating behaviors who received nutritional counseling with or without an oral nutritional supplement.
Nutr Metab Insights
PUBLISHED: 01-01-2014
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In this study, changes in growth parameters and nutrient intake were compared in Chinese children (ages 30-60 months) with picky eating (PE) behaviors and weight-for-height ?25th percentile, who were randomized to receive nutrition counseling alone (NC; n = 76) or with a nutritional milk supplement (NC + NS; n = 77) for 120 days. Increases in weight-for-height z-scores were significantly greater in the NC + NS group at days 30 and 90 and over the entire study period (all P < 0.05), but not at day 120. Increases in weight-for-age z-scores were significantly greater in the NC + NS group at day 90 (P = 0.025) and over the entire study period (P = 0.046). Mean intakes of energy, protein, carbohydrate, docosahexaenoic acid, arachidonic acid, calcium, phosphorous, iron, zinc, and vitamins A, C, D, E, and B6 were significantly higher in the NC + NS group at days 60 and 120 (all P < 0.01). Thus, in young children with PE behaviors, nutritional supplementation given as an adjunct to NC resulted in greater improvements in nutrient intake compared with NC alone. Growth parameters differed between groups at several timepoints during the study, but not at day 120.
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Screening of Duchenne muscular dystrophy (DMD) mutations and investigating its mutational mechanism in Chinese patients.
PLoS ONE
PUBLISHED: 01-01-2014
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Duchenne muscular dystrophy (DMD) is a common X-linked recessive disease of muscle degeneration and death. In order to provide accurate and reliable genetic counseling and prenatal diagnosis, we screened DMD mutations in a cohort of 119 Chinese patients using multiplex ligation-dependent probe amplification (MLPA) and denaturing high performance liquid chromatography (DHPLC) followed by Sanger sequencing. In these unrelated DMD patients, we identified 11 patients with DMD small mutations (9.2%) and 81 patients with DMD deletions/duplications (del/dup) (68.1%), of which 64 (79.0%) were deletions, 16 (19.8%) were duplications, and one (1.2%) was both deletion and duplication. Furthermore, we analyzed the frequency of DMD breakpoint in the 64 deletion cases by calculating exon-deletion events of certain exon interval that revealed a novel mutation hotspot boundary. To explore why DMD rearrangement breakpoints were predisposed to specific regions (hotspot), we precisely characterized junction sequences of breakpoints at the nucleotide level in 21 patients with exon deleted/duplicated in DMD with a high-resolution SNP microarray assay. There were no exactly recurrent breakpoints and there was also no significant difference between single-exon del/dup and multiple-exon del/dup cases. The data from the current study provided a comprehensive strategy to detect DMD mutations for clinical practice, and identified two deletion hotspots at exon 43-55 and exon 10-23 by calculating exon-deletion events of certain exon interval. Furthermore, this is the first study to characterize DMD breakpoint at the nucleotide level in a Chinese population. Our observations provide better understanding of the mechanism for DMD gene rearrangements.
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Carvacrol alleviates ischemia reperfusion injury by regulating the PI3K-Akt pathway in rats.
PLoS ONE
PUBLISHED: 01-01-2014
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Liver ischemia reperfusion (I/R) injury is a common pathophysiological process in many clinical settings. Carvacrol, a food additive commonly used in essential oils, has displayed antimicrobials, antitumor and antidepressant-like activities. In the present study, we investigated the protective effects of carvacrol on I/R injury in the Wistar rat livers and an in vitro hypoxia/restoration (H/R) model.
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[Analysis for Clinicopathological Features, Therapy and Prognosis of 30 Elderly Patients with Non-Hodgkins Lymphoma].
Zhongguo Shi Yan Xue Ye Xue Za Zhi
PUBLISHED: 12-28-2013
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The purpose of this study was to explore the clinicopathological features, therapy and prognostic factors of elderly patients with non-Hodgkins lymphoma (NHL). The clinical data including general clinical characteristics, pathological features, chemotherapy selection and treatment response of 30 patients with NHL in our hospital from January 2003 to December 2012 were analyzed retrospectively. The survival was analyzed by using Kaplan-Meier methods, and the prognosis was evaluated by COX regression multivariate analysis model. The clinical parameters selected include age, Ann Arbor stage, international prognostic index (IPI), B symptom and lactate dehydrogenase (LDH) levels. The results showed that all the patients suffered from underlying disease, and the cardiovascular disease (hypertension, coronary heart disease, arrhythmia) is the most common, and minority (8/30) combined with secondary tumor, the 63% (19/30) cases had B symptoms at diagnosis. only 2 cases were diagnosed as T-cell lymphoma; the 93% (28/30) cases combined with B-cell lymphoma, 57% (17/28) of them combined with diffuse large B-cell lymphoma. Ann-Arbor stage ? IIwas 37% (11/30);10(37%)patients IPI score was ? 2, and 67% (20/30) was scored 3-5; 13(43%) patients serum LDH level was abnormal. Modified R-CHOP chemotherapy was given individually on the basis of clinical features. The patients achieved complete remission, partial remission, stable disease, or progressive disease accounted for 14 (46.7%), 13 (43.3%), 1 (3.3%), and 2 (6.7%), respectively; the total reaction rate was 90% after 4 cycles of chemotherapy; the overall survival (OS) rate at 1 and 2 years was 73.3% and 43.3%, and progression-free survival (PFS)rate at 0.5 and 1 years was 62.2% and 54.9%; multivariate analysis by COX regression showed that B symptoms and Ann-Arbor stage were independent factors (P = 0.014, 0.039; RR = 6.678, 4.939, respectively) affecting the OS of elderly NHL, and IPI score affected PFS independently. It is concluded that elderly patients with NHL usually are of late stage at newly diagnosis and have suffered from underlaying diseases. Besides strengthening supportive treatment, modified R-CHOP chemotherapy should be given individually according to different prognosis. B symptoms and Ann-Arbor stage>IIare indicators for poor prognosis of elderly NHL.
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A plate-based electrochromic approach for the high-throughput detection of electrochemically active bacteria.
Nat Protoc
PUBLISHED: 12-19-2013
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Electrochemically active bacteria (EAB) have the ability to transfer electrons to electron acceptors located outside the cell, and they are widely present in diverse environments. In spite of their important roles in geochemical cycles, environmental remediation and electricity generation, so far, only a limited number and types of EAB have been isolated and characterized. Thus, effective and rapid EAB identification methods are highly desirable. In this protocol, we describe a photometric protocol for the visualization and high-throughput identification and isolation of EAB. The protocol relies on the fast electron acquisition and color change ability of an electrochromic material, namely a tungsten trioxide (WO3) nanorod assembly. The extracellular electron transfer (EET) from EAB to the WO3 nanorod assembly probe is accompanied by a bioelectrochromic reaction made evident by the color change of the probe. This protocol enables researchers to rapidly identify EAB and evaluate their EET ability either qualitatively with the naked eye or quantitatively by image analysis. We have also successfully used this protocol to isolate EAB from environmental samples. The time needed to complete this protocol is ?2 d, with the actual EAB identification process taking about 5 min.
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Associated Analysis of DNA Methylation for Cancer Detection Using CCP-Based FRET Technique.
Anal. Chem.
PUBLISHED: 12-16-2013
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This paper describes an associated analysis method of DNA methylation for the detection of cancer using an optically amplifying cationic conjugated polymer (CCP, poly{(1,4-phenylene)-2,7-[9,9-bis(6-N,N,N-trimethyl ammonium)-hexyl fluorene] dibromide)}. Genomic DNA is digested by methylation-sensitive restriction endonuclease, followed by PCR amplification to incorporate fluorescein-labeled dNTP. Only methylated DNA can be amplified by PCR, and the methylation level is detected through fluorescence resonance energy transfer (FRET) between CCP and fluorescein that is incorporated into the PCR product. The methylation levels of RASSF1A, OPCML, and HOXA9 promoters of 35 ovarian cancer samples and 11 normal samples were assayed. In accordance with the degree of methylation levels, they are clustered to three sections and assigned a value. Through an associated analysis, we acquired a threshold for cancer detection with a sensitivity of 85.7%. The assay takes about 20 h to obtain the detection results and shows great potential as a useful tool for diagnostic and screening of cancer.
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Genome-scale CRISPR-Cas9 knockout screening in human cells.
Science
PUBLISHED: 12-12-2013
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The simplicity of programming the CRISPR (clustered regularly interspaced short palindromic repeats)-associated nuclease Cas9 to modify specific genomic loci suggests a new way to interrogate gene function on a genome-wide scale. We show that lentiviral delivery of a genome-scale CRISPR-Cas9 knockout (GeCKO) library targeting 18,080 genes with 64,751 unique guide sequences enables both negative and positive selection screening in human cells. First, we used the GeCKO library to identify genes essential for cell viability in cancer and pluripotent stem cells. Next, in a melanoma model, we screened for genes whose loss is involved in resistance to vemurafenib, a therapeutic RAF inhibitor. Our highest-ranking candidates include previously validated genes NF1 and MED12, as well as novel hits NF2, CUL3, TADA2B, and TADA1. We observe a high level of consistency between independent guide RNAs targeting the same gene and a high rate of hit confirmation, demonstrating the promise of genome-scale screening with Cas9.
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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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In developing our video relationships, we compare around 5 million PubMed articles to our library of over 4,500 methods videos. In some cases the language used in the PubMed abstracts makes matching that content to a JoVE video difficult. In other cases, there happens not to be any content in our video library that is relevant to the topic of a given abstract. In these cases, our algorithms are trying their best to display videos with relevant content, which can sometimes result in matched videos with only a slight relation.