JoVE Visualize What is visualize?
Stop Reading. Start Watching.
Advanced Search
Stop Reading. Start Watching.
Regular Search
Find video protocols related to scientific articles indexed in Pubmed.
Stable SET knockdown in breast cell carcinoma inhibits cell migration and invasion.
Biochem. Biophys. Res. Commun.
PUBLISHED: 08-28-2014
Show Abstract
Hide Abstract
Breast cancer is the most malignant tumor for women, however, the mechanisms underlying this devastating disease remain unclear. SET is an endogenous inhibitor of protein phosphatase 2A (PP2A) and involved in many physiological and pathological processes. SET could promote the occurrence of tumor through inhibiting PP2A. In this study, we explore the role of SET in the migration and invasion of breast cancer cells MDA-MB-231 and ZR-75-30. The stable suppression of SET expression through lentivirus-mediated RNA interference (RNAi) was shown to inhibit the growth, migration and invasion of breast cancer cells. Knockdown of SET increases the activity and expression of PP2Ac and decrease the expression of matrix metalloproteinase 9 (MMP-9). These data demonstrate that SET may be involved in the pathogenic processes of breast cancer, indicating that SET can serve as a potential therapeutic target for the treatment of breast cancer.
Related JoVE Video
Convergence of Sample Eigenvalues, Eigenvectors, and Principal Component Scores for Ultra-High Dimensional Data.
Biometrika
PUBLISHED: 08-22-2014
Show Abstract
Hide Abstract
The development of high-throughput biomedical technologies has led to increased interest in the analysis of high-dimensional data where the number of features is much larger than the sample size. In this paper, we investigate principal component analysis under the ultra-high dimensional regime, where both the number of features and the sample size increase as the ratio of the two quantities also increases. We bridge the existing results from the finite and the high-dimension low sample size regimes, embedding the two regimes in a more general framework. We also numerically demonstrate the universal application of the results from the finite regime.
Related JoVE Video
Mitochondrial KATP Channels Control Glioma Radioresistance by Regulating ROS-Induced ERK Activation.
Mol. Neurobiol.
PUBLISHED: 07-08-2014
Show Abstract
Hide Abstract
Malignant glioma is the most prevalent form of malignant brain tumor. Although radiotherapy is widely used in glioma treatment, the radioresistance of glioma cells limits the success of the glioma treatment. The lack of effective targets and signaling pathways to reverse glioma radioresistance is the critical obstacle in successful treatment. In this study, we demonstrate that mitochondrial ATP-sensitive potassium channels (mtKATP channels) are overexpressed in glioma cells and are closely related to the malignancy grade and the overall survival of the patients. Importantly, we showed that mtKATP channels could control glioma radioresistance by regulating reactive oxygen species (ROS)-induced ERK activation. The inhibition of mtKATP channels suppresses glioma radioresistance by inhibiting ERK activation both in vitro and in vivo. These findings reveal the important roles of the mitochondria and mtKATP channels as key regulators in the radioresistance of glioma cells, and suggest that mtKATP channel blockers and MAPK/ERK kinase (MEK) inhibitors are potential targets for drug development of glioma treatments.
Related JoVE Video
[Effect of thymic stromal lymphopoietin on human bronchial epithelial permeability].
Nan Fang Yi Ke Da Xue Xue Bao
PUBLISHED: 06-28-2014
Show Abstract
Hide Abstract
To investigate the effect of thymic stromal lymphopoietin (TSLP) on the permeablily of monolayer bronchial epithelial cells in vitro.
Related JoVE Video
BNIP3 upregulation by ERK and JNK mediates cadmium-induced necrosis in neuronal cells.
Toxicol. Sci.
PUBLISHED: 05-13-2014
Show Abstract
Hide Abstract
Cadmium (Cd) is a toxic heavy metal that may cause neurological disorders. We studied the mechanism underlying Cd-mediated cell death in neuronal cells. In Cd-induced neurotoxicity, caspase-3 was only modestly activated, and accordingly, zVAD-fmk, a pan-caspase inhibitor, partially attenuated cell death. However, pretreatment with Necrox-2 or Necrox-5, two novel necrosis inhibitors, suppressed cell death more markedly compared with pretreatment with zVAD-fmk. Moreover, the necrosis inhibitors did not prevent cleavage of caspase-3. These results indicate that caspase-independent necrosis is more prevalent in Cd-induced neurotoxicity. Bcl-2 and adenovirus E1B-19 kDa-interacting protein 3 (BNIP3) has been reported to be related to caspase-independent cell death. Cd treatment caused a dramatic upregulation of BNIP3 mRNA and protein levels in vitro and in vivo. Furthermore, knockdown of BNIP3 greatly inhibited Cd-induced cell death. Importantly, BNIP3 RNAi decreased lactate dehydrogenase release and the percentage of propidium iodide-positive cells, two markers of necrotic cell death due to rupture of the cell membrane, whereas it had no effect on activation of caspase-3 in Cd-treated cells. These data suggest that BNIP3 mediates caspase-independent necrosis, but not apoptosis. Moreover, our results indicate that induction of BNIP3 by Cd may not be related to HIF-1 which is generally regarded as a mediator responsible for BNIP3 expression. Finally, we show that mitogen-activated protein kinases (MAPKs) are activated by Cd in vitro and in vivo; ERK and JNK promote BNIP3 upregulation and subsequent necrosis. Taken together, our results suggest BNIP3, upregulated by activation of ERK and JNK, mediates Cd-induced necrosis in neuronal cells.
Related JoVE Video
Association of STAT3 common variations with obesity and hypertriglyceridemia: protective and contributive effects.
Int J Mol Sci
PUBLISHED: 05-08-2014
Show Abstract
Hide Abstract
Signal transducer and activator of transcription 3 (STAT3) plays an important role in energy metabolism. Here we explore whether STAT3 common variations influence risks of obesity and other metabolic disorders in a Chinese Han population. Two tagging single nucleotide polymorphisms (tagSNPs), rs1053005 and rs957970, were used to capture the common variations of STAT3. Relationships between genotypes and obesity, body mass index, plasma triglyceride and other metabolic diseases related parameters were analyzed for association study in 1742 subjects. Generalized linear model and logistic regression model were used for quantitative data analysis and case-control study, respectively. rs1053005 was significantly associated with body mass index and waist circumference (p=0.013 and p=0.02, respectively). rs957970 was significantly associated with plasma level of triglyceride (p=0.007). GG genotype at rs1053005 had lower risks of both general obesity and central obesity (OR=0.40, p=0.034; OR=0.42, p=0.007, respectively) compared with AA genotype. CT genotype at rs957970 had a higher risk of hypertriglyceridemia (OR=1.43, p=0.015) compared with TT genotype. Neither of the two SNPs was associated with othermetabolic diseases related parameters. Our observations indicated that common variations of STAT3 could significantly affect the risk of obesity and hypertriglyceridemia in Chinese Han population.
Related JoVE Video
Bystander effects of PC12 cells treated with Pb²? depend on ROS-mitochondria-dependent apoptotic signaling via gap-junctional intercellular communication.
Toxicol. Lett.
PUBLISHED: 04-29-2014
Show Abstract
Hide Abstract
The demonstration of bystander effect, which means injured cells propagate damage to neighboring cells, in whole organisms has clear implication of the potential relevance of the non-targeted response to human health. Here we show that 10 ?M lead acetate, the optimum concentration for inducing apoptosis confirmed by the expression levels of Bax and Bcl-2, can also induce rat pheochromocytoma (PC12) cells to exert bystander effects to neighboring cells. In a novel co-culture system, GFP-PC12 (Pb(2+)) cells, which were stable transfected with EF1A-eGFP and pre-exposed with lead acetate, were co-cultured with unexposed PC12 cells at a 1:5 ratio. Parachute assays demonstrated the functional gap-junctional intercellular communication (GJIC) formed between Pb(2+)-exposed and unexposed cells. The Pb(2+)-exposed cells induced very similar effects on neighboring unexposed cells to apoptosis coincide with intracellular ROS generation and the collapse of mitochondrial membrane potential (??m). Furthermore, carbenoxolone (CBX), a blocker of GJIC, inhibited the bystander effects. The results indicate that the Pb(2+)-induced insults propagate through GJIC between PC12 cells, while inducing the bystander cells to apoptosis via ROS-mitochondria-dependent apoptotic signaling.
Related JoVE Video
[1,25-dihydroxyvitamin D3 pretreatment inhibits house dust mite-induced thymic stromal lymphopoietin release by human airway epithelial cells].
Nan Fang Yi Ke Da Xue Xue Bao
PUBLISHED: 04-23-2014
Show Abstract
Hide Abstract
To investigate the effect of 1,25-dihydroxyvitamin D3 (1,25VD3) on house dust mites (HDM)-induced expression of thymic stromal lymphopoietin (TSLP) in human airway epithelial cells in vitro.
Related JoVE Video
TRPC1 is involved in Ca²? influx and cytotoxicity following Pb²? exposure in human embryonic kidney cells.
Toxicol. Lett.
PUBLISHED: 04-21-2014
Show Abstract
Hide Abstract
Lead (Pb(2+)) is a divalent heavy metal ion which causes severe damage to almost all life forms and is therefore considered a notorious toxicant. Exposure to Pb(2+) is associated with poor cognitive development in children at relatively low levels that previously were thought to be safe. The mechanism through which Pb(2+) enters cells, however, is unclear. Previous studies have showed that Ca(2+) release-activated Ca(2+) protein 1 (Orai1), a component of store-operated Ca(2+) channels (SOCs), contributes to Pb(2+) cellular entry. Canonical transient receptor potential (TRPC1) channel 1 is a transient receptor potential (TRP) channel which is sometimes referred to as a SOC. The present study was designed to investigate the role of TRPC1 in Pb(2+) entry and toxicity in human embryonic kidney cells (HEK293). Additionally, changes in intracellular Ca(2+) concentration were determined through Fluo-4 and Mag-fluo-4 fluorescent Ca(2+) imaging. Following Pb(2+) exposure, there was a dose-dependent decrease in cell viability. Overexpression of TRPC1 increased Pb(2+)-induced cell death, while knockdown of this channel attenuated cell death. There was increased entry of Pb(2+), as measured by inductively coupled plasma mass spectrometry (ICP-MS), following overexpression of TRPC1. Conversely, knockdown of TRPC1 led to a decrease in Pb(2+) influx. Down-regulation of STIM1 by RNA interference attenuated the Pb(2+) influx, and transfection with a mutant STIM1, which could not gate TRPC1, had a similar effect. Co-transfection of mutant STIM1 and mutant TRPC1, which restore the electrostatic interaction between STIM1 and TRPC1, resumed Pb(2+) entry in HEK293 cells. Down-regulation of TRPC1 by RNA interference decreased Ca(2+) influx whilst its overexpression increased Ca(2+) entry in HEK293 cells. These results suggest that TRPC1 is involved in the cytotoxicity and entry of Pb(2+) through molecular interactions with STIM1 and subsequent Ca(2+) influx in HEK293 cells.
Related JoVE Video
Effect of hexavalent chromium on histone biotinylation in human bronchial epithelial cells.
Toxicol. Lett.
PUBLISHED: 04-14-2014
Show Abstract
Hide Abstract
Chromium is a potent human mutagen and carcinogen. The capability of chromium to cause cancers has been known for more than a century, and numerous epidemiological studies have been performed to determine its carcinogenicity. In the post-genome era, cancer has been found to relate to epigenetic mutations. However, very few researches have focused on hexavalent chromium (Cr(VI))-induced epigenetic alterations. The present study was designed to investigate whether Cr(VI) would affect the level of a newfound epigenetic modification: histone biotinylation. Histone acetylation and histone biotinylation were studied in detail using human bronchial epithelial (16HBE) cells as an in vitro model after Cr(VI) treatment. Our study showed that Cr(VI) treatment decreased histone acetylation level in 16HBE cells. In addition, low doses of Cr(VI) (?0.6?M) elevated the level of histone biotinylation. Furthermore, immunoblot analysis of biotinidase (BTD), a major protein which maintains homeostasis of histone biotinylation, showed that the distribution of BTD became less even and more concentrated at the nuclear periphery in cells exposed to Cr(VI). Moreover, Cr(VI)-induced histone deacetylation may take part in the regulation of histone biotinylation. Together, our study provides new insight into the mechanisms of Cr(VI)-induced epigenetic regulation that may contribute to the chemoprevention of Cr(VI)-induced cancers and may have important implications for epigenetic therapy.
Related JoVE Video
Sodium butyrate induces apoptosis of human colon cancer cells by modulating ERK and sphingosine kinase 2.
Biomed. Environ. Sci.
PUBLISHED: 04-09-2014
Show Abstract
Hide Abstract
To investigate the role of extracellular signal-regulated kinase (ERK) in apoptosis of human colon cancer (HCT116) cells.
Related JoVE Video
Heritability and genomics of gene expression in peripheral blood.
Nat. Genet.
PUBLISHED: 03-14-2014
Show Abstract
Hide Abstract
We assessed gene expression profiles in 2,752 twins, using a classic twin design to quantify expression heritability and quantitative trait loci (eQTLs) in peripheral blood. The most highly heritable genes (?777) were grouped into distinct expression clusters, enriched in gene-poor regions, associated with specific gene function or ontology classes, and strongly associated with disease designation. The design enabled a comparison of twin-based heritability to estimates based on dizygotic identity-by-descent sharing and distant genetic relatedness. Consideration of sampling variation suggests that previous heritability estimates have been upwardly biased. Genotyping of 2,494 twins enabled powerful identification of eQTLs, which we further examined in a replication set of 1,895 unrelated subjects. A large number of non-redundant local eQTLs (6,756) met replication criteria, whereas a relatively small number of distant eQTLs (165) met quality control and replication standards. Our results provide a new resource toward understanding the genetic control of transcription.
Related JoVE Video
Histone modifications contribute to cellular replicative and hydrogen peroxide-induced premature senescence in human embryonic lung fibroblasts.
Free Radic. Res.
PUBLISHED: 03-10-2014
Show Abstract
Hide Abstract
Histone modifications are major post-translational mechanisms responsible for regulation of gene transcription involved in cellular senescence. By using immunofluorescence and Western blot, we showed that the global acetylated levels of histone H3 and H4 were significantly reduced in both replicative and premature senescence of human embryonic lung fibroblasts. However the whole trimethylated level of histone H4 lysine 20 was higher in senescent cells. The alterations in the mRNA and protein levels of histone acetyltransferases (HATs), histone methyltransferase (HMT), and histone deacetylases (HDACs) indicate that differential expression exists between replicative and premature senescent cells. Meanwhile, the reduced activity of HDACs was accompanied by cellular senescence. By employing the quantitative chromatin immunoprecipitation assay in detecting specific histone modifications in senescence-related genes including p53 and p16, it was demonstrated that the mRNA expression of p53 was associated with increased H4 acetylation in replicative senescence and increased H4 acetylation and trimethylation of histone H3 at lysine 4 (H3K4me3) in premature senescence. Both acetylation and trimethylation of H3 were involved in replicative senescence, while the acetylation of histone H3 and H4 was predominant in premature senescence, contributing to the mRNA expression of p16. In summary, the global hypoacetylation of histone H3 and H4 and the hypertrimethylation of histone H4 lysine 20 account for epigenetic characteristics in senescence, controlled by HATs, HMT, and HDACs differentially between replicative and premature senescence. Taken together, these findings suggest that the specific histone modifications are involved in regulating the expression of genes related to senescence of human embryonic lung fibroblasts.
Related JoVE Video
A novel statistical approach for jointly analyzing RNA-Seq data from F1 reciprocal crosses and inbred lines.
Genetics
PUBLISHED: 02-21-2014
Show Abstract
Hide Abstract
RNA sequencing (RNA-seq) not only measures total gene expression but may also measure allele-specific gene expression in diploid individuals. RNA-seq data collected from F1 reciprocal crosses in mice can powerfully dissect strain and parent-of-origin effects on allelic imbalance of gene expression. In this article, we develop a novel statistical approach to analyze RNA-seq data from F1 and inbred strains. Method development was motivated by a study of F1 reciprocal crosses derived from highly divergent mouse strains, to which we apply the proposed method. Our method jointly models the total number of reads and the number of allele-specific reads of each gene, which significantly boosts power for detecting strain and particularly parent-of-origin effects. The method deals with the overdispersion problem commonly observed in read counts and can flexibly adjust for the effects of covariates such as sex and read depth. The X chromosome in mouse presents particular challenges. As in other mammals, X chromosome inactivation silences one of the two X chromosomes in each female cell, although the choice of which chromosome to be silenced can be highly skewed by alleles at the X-linked X-controlling element (Xce) and stochastic effects. Our model accounts for these chromosome-wide effects on an individual level, allowing proper analysis of chromosome X expression. Furthermore, we propose a genomic control procedure to properly control type I error for RNA-seq studies. A number of these methodological improvements can also be applied to RNA-seq data from other species as well as other types of next-generation sequencing data sets. Finally, we show through simulations that increasing the number of samples is more beneficial than increasing the library size for mapping both the strain and parent-of-origin effects. Unless sample recruiting is too expensive to conduct, we recommend sequencing more samples with lower coverage.
Related JoVE Video
4-(Methylnitrosamino)-1-(3-pyridyl) -1-butanone induces circulating microRNA deregulation in early lung carcinogenesis.
Biomed. Environ. Sci.
PUBLISHED: 02-21-2014
Show Abstract
Hide Abstract
To study the alteration of circulating microRNAs in 4-(methylnitrosamino)-1-(3-pyridyl) -1-butanone (NNK)-induced early stage lung carcinogenesis.
Related JoVE Video
Role of the mitochondrial Ca²? uniporter in Pb²?-induced oxidative stress in human neuroblastoma cells.
Brain Res.
PUBLISHED: 02-14-2014
Show Abstract
Hide Abstract
Lead (Pb(2+)) has been shown to induce cellular oxidative stress, which is linked to changes in intracellular calcium (Ca(2+)) concentration. The mitochondrial Ca(2+) uniporter (MCU) participates in the maintenance of Ca(2+) homeostasis in neurons, but its role in Pb(2+)-induced oxidative stress is unclear. To address this question, oxidative stress was induced in human neuroblastoma SH-SY5Y cells and in newborn rats by Pb(2+) treatment. The results showed that the production of reactive oxygen species is increased in cells upon treatment with Pb(2+) in a dose-dependent manner, while glutathione and MCU expression were reduced. Moreover, neuronal nitric oxide synthase protein expression was elevated in rats exposed to Pb(2+) during gestation, while MCU expression was decreased. Application of the MCU activator spermine or MCU overexpression reversed Pb(2+)-induced oxidative stress and inhibition of mitochondrial Ca(2+) uptake, while the MCU inhibitor Ru360 and MCU knockdown potentiated the effects of Pb(2+). These results indicate that the MCU mediates the Pb(2+)-induced oxidative stress response in neurons through the regulation of mitochondrial Ca(2+) influx.
Related JoVE Video
Ethyl pyruvate decreases airway neutrophil infiltration partly through a high mobility group box 1-dependent mechanism in a chemical-induced murine asthma model.
Int. Immunopharmacol.
PUBLISHED: 02-11-2014
Show Abstract
Hide Abstract
Diisocyanates are one of the leading causes of occupational asthma, which is dominated by granulocytic inflammation in the airway. In this study, we intended to explore the role of ethyl pyruvate (EP) on neutrophil infiltration in a toluene-2,4-diisocyanate (TDI)-induced murine asthma model.
Related JoVE Video
Store-operated Ca²? entry mediated regulation of polarization in differentiated human neutrophil-like HL-60 cells under hypoxia.
Mol Med Rep
PUBLISHED: 01-13-2014
Show Abstract
Hide Abstract
The regulation of neutrophil polarization by calcium entry is critical for maintaining an effective host response. Hypoxia has a major effect on the apoptosis of neutrophils, however the role of store-operated Ca2+ entry (SOCE) in neutrophil polarization under hypoxia remains to be elucidated. In the present study, we examined the polarization of differentiated human neutrophil-like HL-60 (dHL-60) cells exposed to hypoxia (3% O2) and the results demonstrated that the percentage of polarized cells following exposure to an N-formyl-Met-Leu-Phe (fMLP) gradient in the Zigmond chamber was increased. We examined stromal interaction molecule 1 (STIM1) and Orai1 expression in dHL-60 cells during hypoxia, and it was observed that the expression of STIM1 and Orai1 was significantly reduced at day 2. However, no apparent change was observed on the first day, indicating that this effect is dependent on stimulation time. Fluo-4/acetoxymethyl (AM) ester imaging also demonstrated that SOCE was decreased in dHL-60 cells. The plasmid overexpression assay demonstrated that the response of polarization was returned to the control level. We demonstrated the inhibitory role of SOCE on the polarization of dHL-60 cells under hypoxic conditions, which may be the mechanism for the adaptation of neutrophils to hypoxia. SOCE is also suggested to be a key modulator of immune deficiency under hypoxic conditions and is potentially a therapeutic target.
Related JoVE Video
Environmental and genetic contributors to salivary testosterone levels in infants.
Front Endocrinol (Lausanne)
PUBLISHED: 01-01-2014
Show Abstract
Hide Abstract
Transient activation of the hypothalamic-pituitary-gonadal axis in early infancy plays an important role in male genital development and sexual differentiation of the brain, but factors contributing to individual variation in testosterone levels during this period are poorly understood. We measured salivary testosterone levels in 222 infants (119 males, 103 females, 108 singletons, 114 twins) between 2.70 and 4.80?months of age. We tested 16 major demographic and medical history variables for effects on inter-individual variation in salivary testosterone. Using the subset of twins, we estimated genetic and environmental contributions to salivary testosterone levels. Finally, we tested single nucleotide polymorphisms (SNPs) within ±5?kb of genes involved in testosterone synthesis, transport, signaling, and metabolism for associations with salivary testosterone using univariate tests and random forest (RF) analysis. We report an association between 5?min APGAR scores and salivary testosterone levels in males. Twin modeling indicated that individual variability in testosterone levels was primarily explained by environmental factors. Regarding genetic variation, univariate tests did not reveal any variants significantly associated with salivary testosterone after adjusting for false discovery rate. The top hit in males was rs10923844, an SNP of unknown function located downstream of HSD3B1 and HSD3B2. The top hits in females were two SNPs located upstream of ESR1 (rs3407085 and rs2295190). RF analysis, which reflects joint and conditional effects of multiple variants, indicated that genes involved in regulation of reproductive function, particularly LHCGR, are related to salivary testosterone levels in male infants, as are genes involved in cholesterol production, transport, and removal, while genes involved in estrogen signaling are related to salivary testosterone levels in female infants.
Related JoVE Video
Expression of Hsp90? and cyclin B1 were related to prognosis of esophageal squamous cell carcinoma and keratin pearl formation.
Int J Clin Exp Pathol
PUBLISHED: 01-01-2014
Show Abstract
Hide Abstract
Hsp90? (heat shock protein 90?), one of the important molecular chaperones in cancer cell signal transduction, has been a new candidate target for cancer therapy. Cyclin B1, the client protein of Hsp90?, plays a key role as a mitotic cyclin in the G2-M phase transition during the cell cycle progression. However, the relationship between the level of HSP90? and cyclin B1, the location of Hsp90? and cyclin B1 in prognosis of esophageal squamous cell carcinoma (ESCC) has not been examined. Here, we demonstrate that the diagnostic significance of Hsp90? and cyclin B1 by immunohistochemistry and the association of Hsp90? and cyclin B1 expression in ESCC. In the specimens from 105 ESCC patients (81 stained with Hsp90? antibody by Immunohistochemistry, 65 with cyclin B1 antibody, and among them, 41 paired specimens were stained with Hsp90? and cyclin B1 respectively, and then checked for the correlation of the level and location of Hsp90? and cylcin B1. The positivity rate of Hsp90? and cyclin B1 expression were 96.3% (78 of 81) and 84.6% (55 of 65) respectively. Both of them, the expression levels are associated with the clinical pathological stage (Hsp90?, p=0.027; cyclin B1, p=0.007). No association was found between Hsp90? or cyclin B1 and gender, age, tumor location. As to TMN stage, there is no association with the level of Hsp90?, However, cyclin B1 expression is significantly related to tumor status (p=0.002). Interestingly, Hsp90? expression was negatively correlated to cyclin B1 expression (Gamma=-0.692, p=0.007) in the keratin pearls though there is a positive correlation in the other areas of tumor (Gamma=0.503, p=0.015), which suggest Hsp90? might play diverse roles in the cyclin B1 expression and cyclin B1 related cell cycle regulation in the different area of tumor. These findings demonstrated that the expression of Hsp90?, cyclin B1 protein is associated with tumor malignancy and prognosis for patients with human esophageal squamous cell carcinoma, and Hsp90? might be involved in cyclin B1 expression regulation and cell cycle regulation in keratin peal formation of ESCC.
Related JoVE Video
CRTC3 polymorphisms were associated with the plasma level of total cholesterol and the risks of overweight and hypertriglyceridemia in a Chinese Han population.
Mol. Biol. Rep.
PUBLISHED: 10-26-2013
Show Abstract
Hide Abstract
CREB-regulated transcription coactivator 3 (CRTC3) was a recently identified protein which played an important role in glucose and lipid metabolism. Previous research showed that the polymorphisms of CRTC3 were associated with obesity in Mexican-Americans. Data on that relationship in Chinese was unavailable so far. So we investigated whether the polymorphisms of CRTC3 could confer risks of obesity or other metabolic disorders in Chinese population. 1,550 subjects were recruited from physical examination participants. Two SNPs of CRTC3, rs3862434 and rs11635252, were genotyped with the method of PCR-RFLP. Logistic regression model was applied to calculate the risks of overweight, obesity and dyslipidemias for genotypes. The rs3862434 was significantly associated with the plasma level of total cholesterol (P = 0.026), with the G allele carriers having a lower level compared with the AA genotype (P = 0.018). The rs11635252 was associated with the risks of overweight and hypertriglyceridemia, specifically, T allele had higher risks of overweight and hypertriglyceridemia compared with C allele (OR 1.23, 95 % CI 1.02-1.48, P = 0.024; OR 1.22, 95 % CI 1.00-1.48, P = 0.048, respectively). In conclusion, the CRTC3 polymorphism rs3862434 was associated with the plasma level of total cholesterol, and rs11635252 was associated with the risks of overweight and hypertriglyceridemia in a Chinese Han population, which might strengthen our understanding of the complex heredity of metabolic disorders.
Related JoVE Video
IFN-? in turtle: Conservation in sequence and signalling and role in inhibiting iridovirus replication in Chinese soft-shelled turtle Pelodiscus sinensis.
Dev. Comp. Immunol.
PUBLISHED: 08-16-2013
Show Abstract
Hide Abstract
The IFN-? gene was identified in a turtle, the Chinese soft-shelled turtle, Pelodiscus sinensis, with its genome consisting of 4 exons and 3 introns. The deduced amino acid sequence of this gene contains a signal peptide, an IFN-? family signature motif (130)IQRKAVNELFPT, an NLS motif (155)KRKR and three potential N-glycosylation sites. As revealed by real-time quantitative PCR, the gene was constitutively expressed in all tested organs/tissues, with higher level observed in blood, intestine and thymus. An induced expression of IFN-? at mRNA level was observed in peripheral blood leucocytes (PBLs) in response to in vitro stimulation of LPS and PolyI:C. The overexpression of IFN-? in the Chinese soft-shelled turtle artery (STA) cell line resulted in the increase in the expression of transcriptional regulators, such as IRF1, IRF7 and STAT1, and antiviral genes, such as Mx, PKR, implying possibly the existence of a conserved signalling network and role for IFN-? in the turtle. Furthermore, the infection of soft-shelled turtle iridovirus (STIV) in the cell line transfected with IFN-? may cause the cell death as demonstrated with the elevated lactate dehydrogenase (LDH) level and cell mortality. However, the mechanism involved in the antiviral activity may require further investigation.
Related JoVE Video
Molecular cloning and functional characterization of peptidoglycan recognition protein 6 in grass carp Ctenopharyngodon idella.
Dev. Comp. Immunol.
PUBLISHED: 07-23-2013
Show Abstract
Hide Abstract
Peptidoglycan recognition proteins (PGRPs) are pattern recognition molecules of innate immunity. In this study, a long-form PGRP, designated as gcPGRP6, was identified from grass carp Ctenopharyngodon idella. The deduced amino acid sequence of gcPGRP6 is composed of 464 residues with a conserved PGRP domain at the C-terminus. The gcPGRP6 gene consists of four exons and three introns, spacing approximately 2.7kb of genomic sequence. Phylogenetic analysis demonstrated that gcPGRP6 is clustered closely with zebrafish PGLYRP6, and formed a long-type PGRP subfamily together with PGLYRP2 members identified in teleosts and mammals. Real-time PCR and Western blotting analyses revealed that gcPGRP6 is constitutively expressed in organs/tissues examined, and its expression was significantly induced in liver and intestine of grass carp in response to PGN stimulation and in CIK cells treated with lipoteichoic acid (LTA), polyinosinic polycytidylic acid (Poly I:C) and peptidoglycan (PGN). Immunofluorescence microscopy and Western blotting analyses revealed that gcPGRP6 is effectively secreted to the exterior of CIK cells. The over-expression of gcPGRP6 in CIK cells leads to the activation of NF-?B and the inhibition of intracellular bacterial growth. Moreover, cell lysates from CIK cells transfected with pTurbo-gcPGRP6-GFP plasmid display the binding activity towards Lys-type PGN from Staphylococcus aureus and DAP-type PGN from Bacillus subtilis. Furthermore, proinflammatory cytokine IL-2 and intracellular PGN receptor NOD2 had a significantly increased expression in CIK cells overexpressed with gcPGRP6. It is demonstrated that the PGRP6 in grass carp has a role in binding PGN, in inhibiting the growth of intracellular bacteria, and in activating NF-?B, as well as in regulating innate immune genes.
Related JoVE Video
Association of TGF-?1 -509C/T polymorphisms with breast cancer risk: evidence from an updated meta-analysis.
Tumour Biol.
PUBLISHED: 05-18-2013
Show Abstract
Hide Abstract
Epidemiological studies have evaluated the association between transforming growth factor-?1 (TGF-?1) -509C/T polymorphisms and breast cancer risk. However, the results remain conflicting rather than conclusive. The aim of this study was to comprehensively clarify the association between TGF-?1 -509C/T polymorphisms and breast cancer risk. All relevant studies were searched in the electronic databases. The potential sources of heterogeneity were detected with the chi-square-based Q test. The strength of associations between TGF-?1 -509C/T polymorphisms and breast cancer risk was measured by odds ratio (OR) and 95 % confidence intervals (CI). Publication bias was tested by Beggs test and Eggers test. A total of 10 studies including 10,913 cases and 14,187 controls were included in the meta-analysis. Overall, there was no evidence of significant association of TGF-?1 -509C/T polymorphisms with breast cancer risk (TT vs. CC [OR?=?0.97, 95 % CI?=?0.83-1.14]; CT vs. CC [OR?=?1.05, 95 % CI?=?0.90-1.22]; TT?+?CT vs. CC [OR?=?0.99, 95 % CI?=?0.91-1.08]; T allele vs. C allele [OR?=?0.99, 95 % CI?=?0.93-1.06]). Similar results were also found in the subgroup analyses by ethnicity and source of control. When stratified by estrogen receptor (ER) status, TT genotype and T allele were associated with a decreased ER-positive breast cancer risk (OR?=?0.66, 95 % CI?=?0.49-0.90 and OR?=?0.85, 95 % CI?=?0.75-0.96, respectively). The present meta-analysis results suggest that TGF-?1 -509C/T variants may not contribute to the risk of breast cancer overall. However, T allele may be a potential protective factor for developing ER-positive breast cancer. Well-designed studies with larger sample size were required to verify our findings further.
Related JoVE Video
[Role of calcium dyshomeostasis in 1-methyl-4-phenylpyridinium ion-induced apoptosis of human neuroblastoma SH-SY5Y cells].
Nan Fang Yi Ke Da Xue Xue Bao
PUBLISHED: 05-07-2013
Show Abstract
Hide Abstract
To investigate the role of calcium dyshomeostasis in 1-methyl-4-phenylpyridinium ion (MPP?)-induced apoptosis of human neuroblastoma SH-SY5Y cells.
Related JoVE Video
Development and validation of a hydrophilic interaction liquid chromatography-tandem mass spectrometry method for the simultaneous determination of five first-line antituberculosis drugs in plasma.
Anal Bioanal Chem
PUBLISHED: 05-06-2013
Show Abstract
Hide Abstract
A new, sensitive and fast method for the simultaneous determination of pyrazinamide, isoniazid, streptomycin, ethambutol, and rifampicin in human plasma was developed and validated. The method required only 100 ?L of plasma and one step for sample preparation by protein precipitation. The drugs were separated by using a hydrophilic interaction liquid chromatography (HILIC) column. The mobile phase was methanol and water (0.1% formic acid and 5 mM ammonium acetate, pH 3.0 ± 0.1) in a ratio of 65:35 (v/v), which was eluted at an isocratic flow rate of 0.5 mL/min. Tandem mass spectrometry was performed with a triple-quadrupole tandem mass spectrometer. By use of the HILIC column, the detection was free of ion-pair reagents in the mobile phase, with no significant matrix effects. The total run time was less than 2 min for each sample. The method was validated by evaluating its selectivity, sensitivity, linearity, accuracy, and precision according to US Food and Drug Administration guidelines. The lower limit of quantification was 4.0 ng/mL for pyrazinamide, isoniazid, and rifampicin, 0.5 ng/mL for ethambutol, and 10.0 ng/mL for streptomycin. The intraday precision and interday precision were less than 9%, with the accuracy ranging between -9.3 and 7.3%. The method was successfully applied to therapeutic drug monitoring of 33 patients with tuberculosis after administration of standard antituberculosis drugs. The method has been proved to meet the high-throughput requirements in therapeutic drug monitoring.
Related JoVE Video
Activation of PI3K/Akt pathway by CD133-p85 interaction promotes tumorigenic capacity of glioma stem cells.
Proc. Natl. Acad. Sci. U.S.A.
PUBLISHED: 04-08-2013
Show Abstract
Hide Abstract
The biological significance of a known normal and cancer stem cell marker CD133 remains elusive. We now demonstrate that the phosphorylation of tyrosine-828 residue in CD133 C-terminal cytoplasmic domain mediates direct interaction between CD133 and phosphoinositide 3-kinase (PI3K) 85 kDa regulatory subunit (p85), resulting in preferential activation of PI3K/protein kinase B (Akt) pathway in glioma stem cell (GSC) relative to matched nonstem cell. CD133 knockdown potently inhibits the activity of PI3K/Akt pathway with an accompanying reduction in the self-renewal and tumorigenicity of GSC. The inhibitory effects of CD133 knockdown could be completely rescued by expression of WT CD133, but not its p85-binding deficient Y828F mutant. Analysis of glioma samples reveals that CD133 Y828 phosphorylation level is correlated with histopathological grade and overlaps with Akt activation. Our results identify the CD133/PI3K/Akt signaling axis, exploring the fundamental role of CD133 in glioma stem cell behavior.
Related JoVE Video
Mast cell expression of the serotonin1A receptor in guinea pig and human intestine.
Am. J. Physiol. Gastrointest. Liver Physiol.
PUBLISHED: 03-21-2013
Show Abstract
Hide Abstract
Serotonin [5-hydroxytryptamine (5-HT)] is released from enterochromaffin cells in the mucosa of the small intestine. We tested a hypothesis that elevation of 5-HT in the environment of enteric mast cells might degranulate the mast cells and release mediators that become paracrine signals to the enteric nervous system, spinal afferents, and secretory glands. Western blotting, immunofluorescence, ELISA, and pharmacological analysis were used to study expression of 5-HT receptors by mast cells in the small intestine and action of 5-HT to degranulate the mast cells and release histamine in guinea pig small intestine and segments of human jejunum discarded during Roux-en-Y gastric bypass surgeries. Mast cells in human and guinea pig preparations expressed the 5-HT1A receptor. ELISA detected spontaneous release of histamine in guinea pig and human preparations. The selective 5-HT1A receptor agonist 8-hydroxy-PIPAT evoked release of histamine. A selective 5-HT1A receptor antagonist, WAY-100135, suppressed stimulation of histamine release by 5-HT or 8-hydroxy-PIPAT. Mast cell-stabilizing drugs, doxantrazole and cromolyn sodium, suppressed the release of histamine evoked by 5-HT or 8-hydroxy-PIPAT in guinea pig and human preparations. Our results support the hypothesis that serotonergic degranulation of enteric mast cells and release of preformed mediators, including histamine, are mediated by the 5-HT1A serotonergic receptor. Association of 5-HT with the pathophysiology of functional gastrointestinal disorders (e.g., irritable bowel syndrome) underlies a question of whether selective 5-HT1A receptor antagonists might have therapeutic application in disorders of this nature.
Related JoVE Video
The apoptosis of non-small cell lung cancer induced by cisplatin through modulation of STIM1.
Exp. Toxicol. Pathol.
PUBLISHED: 02-14-2013
Show Abstract
Hide Abstract
Cis-diamminedichloroplatinum (II) (cisplatin) is one of the most active antitumor agents used in human chemotherapy of non-small cell lung cancer. Cisplatin forms crosslinked DNA adducts and its cytotoxicity has been shown to be mediated by propagation of DNA damage recognition signals to downstream pathways prompting apoptosis. The steps involved in the process include changes in Ca(2+) signaling with dysregulated tumor cell turn-over. Stromal interaction molecules 1 (STIM1), as one of the most potent tumor suppressor genes, are identified as the endoplasmic-reticulum (ER) Ca(2+) sensor controlling store-operated Ca(2+) entry (SOCE) in non-excitable cells, which is main pathway to extracellular Ca(2+) influx. Its role in STIM1 cisplatin-induced apoptosis of non-small cell lung cancer was the focus of study with focus on SOCE inhibitors 2-APB- and SKF96365-cisplatin-induced apoptosis in the non-small cell lung cancer (NSCLC) cell lines A549 and H460. In this experimental model, cisplatin-induced apoptosis and decreased concentration of intracellular Ca(2+) was demonstrated. The expression of STIM1 was significantly higher in carcinoma tissue than in the adjacent non-neoplastic lung tissue. These findings support the conclusion that STIM1 may play an important role in the development of NSCLC which makes drugs that repress the expression of STIM1 to be a potential target for lung cancer therapy.
Related JoVE Video
Induction of thymic stromal lymphopoietin expression in 16-HBE human bronchial epithelial cells by 25-hydroxyvitamin D3 and 1,25-dihydroxyvitamin D3.
Int. J. Mol. Med.
PUBLISHED: 02-08-2013
Show Abstract
Hide Abstract
Vitamin D exerts profound effects on airway epithelial cells. Thymic stromal lymphopoietin (TSLP) derived from airway epithelial cells plays a role in the innate and antigen?specific adaptive immune responses. However, the effect of vitamin D on TSLP expression in airway epithelial cells is unclear. In this study, 16-HBE human bronchial epithelial (HBE) cells were cultured with various concentrations of 25-hydroxyvitamin D(3) (25 D(3)) and 1,25-dihydroxyvitamin D(3) (1,25 D(3)). The expression of TSLP in the 16-HBE human bronchial epithelial cell line was analyzed by PCR and enzyme-linked immunosorbent assay (ELISA). We found that the 16-HBE cells converted inactive 25 D(3) to active 1,25 D(3) and that TSLP mRNA and protein expression levels were significantly increased, peaking at 2 or 12 h in the cells exposed to 500 nM 25 D(3) and 50 nM 1,25 D(3) respectively. Since vitamin D(3) upregulated protein 1 (VDUP1) plays a multifunctional role in a variety of cellular responses, we hypothesized that VDUP1 is involved in the induction of TSLP production by 25 D(3). The results showed that the mRNA and protein levels of VDUP1 were significantly upregulated by vitamin D. Furthermore, the silencing of VDUP1 by small interfering RNA (siRNA) significantly inhibited the 25 D(3)- and 1,25 D(3)-mediated induction of TSLP expression. To characterize the metabolic properties of vitamin D in airway epithelial biology, we used the chemical inhibitor of 1?-hydroxylase, itraconazole. The results revealed that itraconazole (10-6 M) reduced the 25 D(3)- but not the 1,25 D(3)-induced TSLP expression in 16-HBE cells. Based on these data, it can be concluded that vitamin D increases TSLP expression in 16-HBE cells through the VDUP1 pathway, which suggests a novel mechanism by which vitamin D alters immune function in the lungs.
Related JoVE Video
Association of tumor-associated fibroblasts with progression of hepatocellular carcinoma.
Med. Oncol.
PUBLISHED: 02-04-2013
Show Abstract
Hide Abstract
Interaction between tumor and stromal cells plays an important role in cancer progression. The aim of this study was to explore the effects of tumor-associated fibroblasts on regulation of hepatocellular carcinoma (HCC) progression. Sixty-five cases of HCC and the corresponding normal liver tissues were recruited for immunohistochemical assessment of ?-smooth muscle actin (?-SMA) expression, a biomarker for activated fibroblasts. Clinicopathological data were also collected from HCC patients for association with ?-SMA expression. Primary cell culture of fibroblasts from HCC tissues was used to generate conditioned medium for testing the effect on regulation of HCC cell migration capacity in the transwell cell migration assay. ?-SMA protein was expressed in 84.0 % (21 out 25 cases) of the fibroblasts from the metastatic HCCs, 45 % (18/40) from HCCs without metastasis, and 19.2 % (5/26) from normal liver tissues, difference of which was statistically significant (P < 0.01). The expression of ?-SMA protein in HCC tissues was associated with tumor thrombosis, poor pathology grade, advanced clinical stages, and lymph node metastasis. The conditioned medium from the primary cultured fibroblasts with ?-SMA expression significantly promoted the migration capacity of HCC Hep3B cells compared to the heat-inactivated conditioned medium. The data from the current study demonstrated that expression of ?-SMA protein in HCC fibroblasts associated with tumor metastasis and advanced clinical stages and that the conditioned medium from ?-SMA-positive fibroblasts enhanced HCC cell migration. This study indicates that ?-SMA protein might serve as a biomarker to predict HCC progression.
Related JoVE Video
Mechanism of E-cadherin redistribution in bronchial airway epithelial cells in a TDI-induced asthma model.
Toxicol. Lett.
PUBLISHED: 01-28-2013
Show Abstract
Hide Abstract
E-cadherin (epithelial cadherin), a transmembrane protein, provides essential architecture and immunological function to the airway epithelium, a barrier structure that plays an essential role in asthma pathogenesis. Toluene diisocyanate (TDI) is currently one of the leading causes of occupational asthma. However, relatively few studies have been undertaken to determine the biological effects of TDI on the barrier properties of airway epithelium, but it is known that TDI can damage airway epithelial tight junctions in vitro. Here, we hypothesize that TDI can injure E-cadherin both in normal and allergic-induced airway epithelium. To test this, we developed a murine model of TDI-induced asthma characterized by neutrophil-dominated airway inflammation, epithelial shedding, and obvious aberrant distribution of E-cadherin. Pretreatment with dexamethasone (DEX) significantly rescued the immunoreactivity of E-cadherin, accompanied by increased neutrophils in bronchoalveolar lavage fluid (BALF). In vitro, TDI-human serum albumin (HSA)-induced redistribution of E-cadherin was associated with extracellular signal-regulated kinase (ERK)1/2 activation. The inhibition of phospho-ERK (p-ERK)1/2 by DEX can partly reverse this reaction. These results indicate that E-cadherin redistribution may be an important contributor in the generation of TDI-induced asthma.
Related JoVE Video
The genetic polymorphisms of cathepsin S were associated with metabolic disorders in a Chinese Han population.
Gene
PUBLISHED: 01-25-2013
Show Abstract
Hide Abstract
Cathepsin S (CTSS) played an important role in the etiology of cardiovascular disease and metabolic syndrome. Few studies had been reported on the association between the polymorphisms of CTSS and metabolic disorders in Asian population. Therefore we explored the association between the polymorphisms of CTSS and metabolic disorders in a Chinese Han population. The subjects were a Chinese Han cohort with 1160 participants, and the genotyping was performed with PCR-RFLP. Polymorphism rs16827671 was associated with BMI and serum total cholesterol (P=0.001; P=0.02, respectively). Subjects with CT genotype of rs16827671 had a higher risk of hypercholesterolemia (OR=1.64, 95% CI: 1.15-2.33, P=0.006) compared with TT genotype. Subjects with AG genotype of rs11576175 had lower risks of hypertriglyceridemia and borderline hypercholesterolemia (OR=0.52, 95% CI: 0.36-0.73, P=0.0001; OR=0.52, 95% CI: 0.35-0.77, P=0.001, respectively) compared with GG genotype. Compared with the haplotype TG, haplotype TA had a lower risk of hypertriglyceridemia and a higher risk of borderline hypercholesterolemia (OR=0.62, 95% CI: 0.44-0.88, P=0.002; OR=1.59, 95% CI: 1.10-2.31, P=0.008, respectively), and haplotype CA had a lower risk of hypercholesterolemia (OR=0.35, 95% CI: 0.18-0.68, P=0.002). In conclusion, we found that the genetic polymorphisms of CTSS were associated with metabolic disorders in a Chinese Han population, which would enrich the knowledge on genetic mechanisms of the pathogenesis of metabolic disorders.
Related JoVE Video
Efficacy of intradiscal hepatocyte growth factor injection for the treatment of intervertebral disc degeneration.
Mol Med Rep
PUBLISHED: 01-18-2013
Show Abstract
Hide Abstract
A poor nutritional supply to the cells of the avascular intervertebral discs, caused by dehydration of the extracellular matrix, is a major cause of intervertebral disc degeneration (IDD). Since hepatocyte growth factor (HGF) has been shown to exert antifibrotic effects, we hypothesized that HGF treatment may be capable of retarding IDD. The present study aimed to evaluate the efficacy of HGF treatment in retarding IDD in a rat tail model of disc degeneration. The disc degeneration models were induced by needle puncture of the rat tail discs. Four weeks following needle puncture, a triblock poly(lactide-co-glycolide)-poly(ethyleneglycol)-poly(lactide-co-glycolide; PLGA-PEG-PLGA) polymer gel loaded with HGF or the gel alone was injected into rat tail discs. The efficacy of HGF in retarding IDD was assessed by magnetic resonance imaging (MRI), histological and immunohistochemical evaluation of the type I collagen, type II collagen and bone morphogenetic protein-2 (BMP?2) expression levels. Following injection of the HGF-loaded gel into the nucleus pulposus (NP), a significant trend towards an increase in T2 signal intensity (P=0.028), type II collagen staining in the NP and the number of BMP-2-positive cells in the annulus fibrosus was observed. In addition, the results demonstrated a significant trend towards a decrease in the histological score (P=0.025) and type I collagen staining in the NP compared with segments treated with the gel alone, following the induction of disc degeneration by stab injury. Following treatment with HGF, a tendency for the level of disc height to be maintained was also observed (no statistical significance). By MRI, histological and immunohistochemical evaluation, the present study demonstrated that HGF-loaded PLGA-PEG-PLGA gel was able to retard disc degeneration when injected into the degenerative discs of rat tail models.
Related JoVE Video
TRPM7 mediates breast cancer cell migration and invasion through the MAPK pathway.
Cancer Lett.
PUBLISHED: 01-11-2013
Show Abstract
Hide Abstract
Metastasis is an inherent feature of breast cancer and transient receptor potential (TRP) channels were found to be potentially implicated in this process. Particularly, TRPM7 may regulate cell motility. We therefore examined the expression of TRPM7 mRNA in the Oncomine database and found that TRPM7 is correlated to metastasis and invasive breast cancer. Silencing TRPM7 with RNA interference resulted in a significant decrease in migration and invasion capability of MDA-MB-435 breast cancer cells, and phosphorylation levels of Src and MAPK but not AKT. Our results suggest that TRPM7 regulates migration and invasion of metastatic breast cancer cells via MAPK pathway.
Related JoVE Video
Association of X-ray Repair Cross Complementing Group 1 Arg399Gln Polymorphisms with the Risk of Squamous Cell Carcinoma of the Head and Neck: Evidence from an Updated Meta-Analysis.
PLoS ONE
PUBLISHED: 01-01-2013
Show Abstract
Hide Abstract
Epidemiologic studies have reported the association of X-ray repair cross-complementary group 1 (XRCC1) Arg399Gln polymorphisms with susceptibility to squamous cell carcinoma of the head and neck (HNSCC). However, the results were conflictive rather than conclusive. The purpose of this study was to clarify the association of XRCC1 Arg399Gln variants with HNSCC risk.
Related JoVE Video
Fine mapping of dental fluorosis quantitative trait loci in mice.
Eur. J. Oral Sci.
PUBLISHED: 12-21-2011
Show Abstract
Hide Abstract
Genetic factors underlie the susceptibility and the resistance to dental fluorosis (DF). The A/J (DF susceptible) and 129P3/J (DF resistant) mouse strains have previously been used to detect quantitative trait loci (QTLs) associated with DF on chromosome (Chr) 2 and Chr 11. In the present study, increased marker density genotyping followed by interval mapping was performed to narrow the QTL intervals and improve the logarithm of the odds (to the base 10) (LOD) scores. Narrower intervals were obtained on Chr 2 where LOD ? 6.0 (57-84 cM or ? 51 Mb), LOD ? 7.0 (62-79 cM or ? 32 Mb), and LOD ? 8.0 (65-74 cM or ? 17 Mb); and on Chr 11 where LOD ? 6.0 (18-51 cM or ? 53 Mb), LOD ? 7.0 (28-48 cM or ? 34 Mb), and LOD ? 8.0 (31-45 cM or ? 22 Mb). Haplotype analysis between A/J and 129P3/J mice further reduced the QTL intervals. Accn1 was selected as a candidate gene based upon its location near the peak LOD score on Chr 11 and distant homology with the Caenorhabditis elegans fluoride-resistance gene, flr1. The severity of DF between Accn1(-/-) and wild-type mice was not significantly different. Hence, the loss of ACCN1 function does not modify DF severity in mice. Narrowing the DF QTL intervals will facilitate additional candidate gene selections and interrogation.
Related JoVE Video
[Surveillance of echinococcosis in high risk areas of Yixing City from 2008 to 2009].
Zhongguo Xue Xi Chong Bing Fang Zhi Za Zhi
PUBLISHED: 12-16-2011
Show Abstract
Hide Abstract
An investigation was conducted retrospectively for echinococcosis (hydatid disease) in Yixing City from 2008 to 2009, the serum or fecal samples of suspected patients during the past 5 years and host animals were assessed by ELISA. There were 2 cases reported by network and 4 confirmed patients by investigation, and among the total 6 cases, 5 cases may be infected in the locality. The positive rates were 0.52%, 0.14% and 1.08% in adults aged over 20 years, children aged 7 to 12 years (by serum tests) and dogs (by fecal tests) respectively.
Related JoVE Video
seeQTL: a searchable database for human eQTLs.
Bioinformatics
PUBLISHED: 12-13-2011
Show Abstract
Hide Abstract
seeQTL is a comprehensive and versatile eQTL database, including various eQTL studies and a meta-analysis of HapMap eQTL information. The database presents eQTL association results in a convenient browser, using both segmented local-association plots and genome-wide Manhattan plots.
Related JoVE Video
Effect of total body X-ray irradiation on lymph node in Tibet minipig.
J. Radiat. Res.
PUBLISHED: 12-01-2011
Show Abstract
Hide Abstract
The purpose of this study was to determine the time-dose-effect of total body X-ray irradiation on lymphocytes in lymph nodes and peripheral blood in Tibet minipigs. Forty-eight Tibet minipigs were assigned into 6 groups including 5 experimental groups with 9 and the control group with 3. The minipigs in experimental groups were subjected to a total body X-ray irradiation of 2, 5, 8, 11, and 14 Gy respectively. Lymph nodes and peripheral blood samples were collected at 6, 24, and 72 hours after X-ray exposure and received histological microscopy examination and apoptosis analysis. Histology observation showed that the number of lymphocytes decreased within the lymph nodes with the increase of radiation doses and exposure time. The observation of transmission electron microscopy (TEM) showed typical apoptotic cells below 11 Gy while at 14 Gy necrotic cells were dominant. The apoptotic rate of lymphocytes in the lymph nodes was positively correlated with radiation dose in the range of 2-11 Gy, and reached the maximal level (39.4 ± 2.8) at 24 hours after 11 Gy irradiation, followed by a decrease in the apoptotic rate, but still higher than that of the control group. The number of lymphocytes in the peripheral blood samples was decreased significantly by increasing of the radiation dose and exposure time. We conclude that early damage of lymphocytes by total body X-ray irradiation is dose and time dependent below 11 Gy and before 24 hours post irradiation, and that the dosage of irradiation less than 11 Gy induced apoptosis, whereas the dose at 14 Gy resulted in necrosis in lymphocytes of the lymph nodes.
Related JoVE Video
A dodecanuclear heterometallic dysprosium-cobalt wheel exhibiting single-molecule magnet behaviour.
Chem. Commun. (Camb.)
PUBLISHED: 09-03-2011
Show Abstract
Hide Abstract
A novel dodecanuclear wheel with ten Dy(III) ions and two Co(II) ions bridged by four Schiff-base ligands and sixteen acetates represents the highest-nuclearity 3d-4f example of its type displaying single-molecule magnet behaviour.
Related JoVE Video
Effects of ZD7288 on firing pattern of thermosensitive neurons isolated from hypothalamus.
Neurosci. Lett.
PUBLISHED: 08-31-2011
Show Abstract
Hide Abstract
The role of the hyperpolarization-activated current (Ih) mediated by HCN channels in temperature sensing by the hypothalamus was addressed. In warm-sensitive neurons (WSNs), exposure to ZD7288, an inhibitor of Ih mediated by hyperpolarization-activated cyclic nucleotide-gated (HCN) channels, decreased their action potential amplitudes and frequencies significantly. By contrast, ZD7288 had little or no effect on temperature-insensitive neurons (TINs). Exposure of WSNs to ZD7288 led to a significant increase in the duration of the inter-spike interval and a reduction of Ih irreversibly. These results suggest that ZD7288 have the contrasting effects on the firing patterns of WSNs versus TINs, which implies HCN channels play a central role in temperature sensing by hypothalamic neurons.
Related JoVE Video
Mucin variable number tandem repeat polymorphisms and severity of cystic fibrosis lung disease: significant association with MUC5AC.
PLoS ONE
PUBLISHED: 07-19-2011
Show Abstract
Hide Abstract
Variability in cystic fibrosis (CF) lung disease is partially due to non-CFTR genetic modifiers. Mucin genes are very polymorphic, and mucins play a key role in the pathogenesis of CF lung disease; therefore, mucin genes are strong candidates as genetic modifiers. DNA from CF patients recruited for extremes of lung phenotype was analyzed by Southern blot or PCR to define variable number tandem repeat (VNTR) length polymorphisms for MUC1, MUC2, MUC5AC, and MUC7. VNTR length polymorphisms were tested for association with lung disease severity and for linkage disequilibrium (LD) with flanking single nucleotide polymorphisms (SNPs). No strong associations were found for MUC1, MUC2, or MUC7. A significant association was found between the overall distribution of MUC5AC VNTR length and CF lung disease severity (p = 0.025; n = 468 patients); plus, there was robust association of the specific 6.4 kb HinfI VNTR fragment with severity of lung disease (p = 6.2×10(-4) after Bonferroni correction). There was strong LD between MUC5AC VNTR length modes and flanking SNPs. The severity-associated 6.4 kb VNTR allele of MUC5AC was confirmed to be genetically distinct from the 6.3 kb allele, as it showed significantly stronger association with nearby SNPs. These data provide detailed respiratory mucin gene VNTR allele distributions in CF patients. Our data also show a novel link between the MUC5AC 6.4 kb VNTR allele and severity of CF lung disease. The LD pattern with surrounding SNPs suggests that the 6.4 kb allele contains, or is linked to, important functional genetic variation.
Related JoVE Video
Sensitization of human colon cancer cells to sodium butyrate-induced apoptosis by modulation of sphingosine kinase 2 and protein kinase D.
Exp. Cell Res.
PUBLISHED: 06-20-2011
Show Abstract
Hide Abstract
Sphingosine kinases (SphKs) have been recognized as important proteins regulating cell proliferation and apoptosis. Of the two isoforms of SphK (SphK1 and SphK2), little is known about the functions of SphK2. Sodium butyrate (NaBT) has been established as a promising chemotherapeutic agent, but the precise mechanism for its effects is unknown. In this study, we investigated the role of SphK2 in NaBT-induced apoptosis of HCT116 colon cancer cells. The results indicated that following NaBT treatment SphK2 was translocated from the nucleus to the cytoplasm, leading to its accumulation in the cytoplasm; in the meantime, only mild apoptosis occurred. However, downregulation of SphK2 resulted in sensitized apoptosis, and overexpression of SphK2 led to even lighter apoptosis; these strongly indicate an inhibitory role of SphK2 in cell apoptosis induced by NaBT. After knocking down protein kinase D (PKD), another protein reported to be critical in cell proliferation/apoptosis process, by using siRNA, blockage of cytoplasmic accumulation of SphK2 and sensitized apoptosis following NaBT treatment were observed. The present study suggests that PKD and SphK2 may form a mechanism for the resistance of cancer cells to tumor chemotherapies, such as HCT116 colon cancer cells to NaBT, and these two proteins may become molecular targets for designation of new tumor-therapeutic drugs.
Related JoVE Video
Increased heat shock protein 70 levels in induced sputum and plasma correlate with severity of asthma patients.
Cell Stress Chaperones
PUBLISHED: 04-26-2011
Show Abstract
Hide Abstract
Damage-associated molecular pattern molecules such as high-mobility group box 1 protein (HMGB1) and heat shock protein 70 (HSP70) have been implicated in the pathogenesis of asthma. The aim of our study was to examine the induced sputum and plasma concentrations of HSP70 in asthmatic patients to determine their relationship with airway obstruction. Thirty-four healthy controls and 56 patients with persistent bronchial asthma matched for gender and age were enrolled in this study. Spirometry measurements were performed before sputum induction. HSP70 levels in induced sputum and plasma were measured using the ELISA Kit. Sputum and plasma concentrations of HSP70 in asthmatics patients were significantly higher than that in control subjects (sputum, (0.88 ng/ml (0.27-1.88 ng/ml) versus 0.42 ng/ml (0.18-0.85 ng/ml), p?
Related JoVE Video
Genome-wide association and linkage identify modifier loci of lung disease severity in cystic fibrosis at 11p13 and 20q13.2.
Nat. Genet.
PUBLISHED: 04-22-2011
Show Abstract
Hide Abstract
A combined genome-wide association and linkage study was used to identify loci causing variation in cystic fibrosis lung disease severity. We identified a significant association (P = 3.34 × 10(-8)) near EHF and APIP (chr11p13) in p.Phe508del homozygotes (n = 1,978). The association replicated in p.Phe508del homozygotes (P = 0.006) from a separate family based study (n = 557), with P = 1.49 × 10(-9) for the three-study joint meta-analysis. Linkage analysis of 486 sibling pairs from the family based study identified a significant quantitative trait locus on chromosome 20q13.2 (log(10) odds = 5.03). Our findings provide insight into the causes of variation in lung disease severity in cystic fibrosis and suggest new therapeutic targets for this life-limiting disorder.
Related JoVE Video
(2-Oxido-1-naphthaldehyde benzoyl-hydrazonato-?N,N,O)pyridine-copper(II).
Acta Crystallogr Sect E Struct Rep Online
PUBLISHED: 03-18-2011
Show Abstract
Hide Abstract
In the mononuclear title compound, [Cu(II)(C(18)H(12)N(2)O(2))(C(5)H(5)N)], the Cu(II) ion is coordinated by two O atoms and one N atom from the dianionic tridentate L(2-) ligand (H(2)L is 2-hy-droxy-1-naphthaldehyde benzoyl-hydrazide) and one N atom from a pyridine mol-ecule in a CuN(2)O(2) distorted square-planar coordination environment.
Related JoVE Video
Bayesian multiple quantitative trait loci mapping for recombinant inbred intercrosses.
Genetics
PUBLISHED: 03-08-2011
Show Abstract
Hide Abstract
The Collaborative Cross (CC) is a renewable mouse resource that mimics the genetic diversity in humans. The recombinant inbred intercrosses (RIX) generated from CC recombinant inbred (RI) lines share similar genetic structures to those of F(2) individuals. In contrast to F(2) mice, genotypes of RIX can be inferred from the genotypes of their RI parents and can be produced repeatedly. Also, RIX mice do not typically share the same degree of relatedness. This unbalanced genetic relatedness requires careful statistical modeling to avoid a large number of false positive findings. For complex traits, mapping multiple genes simultaneously is arguably more powerful than mapping one gene at a time. In this article, we describe how we have developed a Bayesian quantitative trait locus (QTL) mapping method that simultaneously deals with the special genetic architecture of RIX and maps multiple genes. The performance of the proposed method is evaluated by extensive simulations. In addition, for a given set of RI lines, there are numerous ways to generate RIX samples. To provide a general guideline on future RIX studies, we compare several RIX designs through simulations.
Related JoVE Video
Low-level expression of let-7a in gastric cancer and its involvement in tumorigenesis by targeting RAB40C.
Carcinogenesis
PUBLISHED: 02-24-2011
Show Abstract
Hide Abstract
Gastric cancer is the fourth most common cancer and the second leading cause of cancer mortality worldwide but the underlying molecular mechanism is not entirely clear. The objective of this study was to explore the role of let-7a microRNA (miRNA) in gastric tumorigenesis and the possible correlation between RAB40C and let-7a miRNA in gastric cancer. We found that expression of let-7a is reduced in human gastric cancer tissues and cell lines and there was a significant correlation between the level of let-7a expression and the stage of differentiation. Overexpression of let-7a resulted in a decrease in cell proliferation and G(1) arrest, significantly suppressed anchorage-dependent growth in vitro and the tumorigenicity of gastric cancer cells in a nude mouse xenograft model. Furthermore, we demonstrated that RAB40C is regulated directly by let-7a and plays an essential role as a mediator of the biological effects of let-7a in gastric tumorigenesis. This study revealed that let-7a is significant in suppressing gastric cancer growth in vivo and in vitro and provided the first evidence that RAB40C is negatively regulated by let-7a at the posttranscriptional level via binding to the 3-untranslated region of RAB40C messenger RNA in gastric cancer. The results of this study suggest that let-7a and RAB40C are potentially useful targets for gastric cancer diagnosis and therapy.
Related JoVE Video
High mobility group protein B1 (HMGB1) in Asthma: comparison of patients with chronic obstructive pulmonary disease and healthy controls.
Mol. Med.
PUBLISHED: 02-03-2011
Show Abstract
Hide Abstract
High mobility group protein B1 (HMGB1) has been implicated as an important mediator in the pathogenesis of asthma and chronic obstructive pulmonary disease (COPD). However, the expression of HMGB1 in plasma and sputum of patients with asthma and COPD across disease severity needs to be defined. The objective of the study was to examine the induced sputum and plasma concentrations of HMGB1 in COPD and asthmatic patients to determine differences in HMGB1 levels between these diseases and their relationship with airway obstruction and inflammatory patterns. A total of 147 participants were enrolled in this study. The participants included 34 control subjects, 61 patients with persistent asthma (according to the Global Initiative for Asthma [GINA] guidelines) and 47 patients with stable COPD (stratified by Global Initiative for Chronic Obstructive Lung Disease [GOLD] status). Spirometry was performed before sputum induction. HMGB1 levels in induced sputum and plasma were determined by enzyme-linked immunosorbent assay. Sputum and plasma concentrations of HMGB1 in patients with asthma and COPD were significantly higher than concentrations in control subjects and were significantly negatively correlated with forced expiratory volume in 1 s (FEV(1)), FEV(1) (% predicted) in all 147 participants. The levels of HMGB1 in induced sputum of COPD patients were significantly higher than those of asthma patients and healthy controls (P < 0.001). This difference was present even after adjusting for sex, age, smoking status, daily dose of inhaled corticosteroids and disease severity. There were no significant differences in HMGB1 levels between patients with eosinophilic and noneosinophilic asthma. HMGB1 levels in asthmatic and COPD patients were positively correlated with neutrophil counts and percentage of neutrophils. In multivariate analysis, the two diseases (asthma and COPD) and disease severity were independent predictors of sputum HMGB1, but not smoking, age or use of inhaled corticosteroids. In conclusion, these data support a potential role for HMGB1 as a biomarker and diagnostic tool for the differential diagnosis of asthma and COPD. The importance of this observation on asthma and COPD mechanisms and outcomes should be further investigated in large prospective studies.
Related JoVE Video
Trifluoroethanol-induced activity and structural changes in bos taurus copper- and zinc-containing superoxide dismutase.
Protein Pept. Lett.
PUBLISHED: 01-30-2011
Show Abstract
Hide Abstract
Superoxide dismutase (SOD, EC 1.15.1.1) plays an important antioxidant defense role in organisms exposed to oxygen. Copper- and zinc-containing SOD (Cu/Zn-SOD) catalysis and the change in folding behavior of this enzyme in response to inactivators are therefore of interest. We studied the inhibitory effects of trifluoroethanol (TFE) on the activity and conformation of a Cu/Zn-SOD from Bos taurus. We found that TFE inactivated the enzyme and disrupted the tertiary and secondary structures of Cu/Zn-SOD. Kinetic studies showed that TFE-induced inactivation of Cu/Zn-SOD follows first-order reaction kinetics and that TFE binding sites are distinct from the copper- and zinc-containing active site. These structural changes occurred prior to enzyme activity loss. A computational docking simulation of Cu/Zn-SOD and TFE (binding energy of Dock 6.3: -11.52 kcal/mol) suggested that THR37, ASP40, and GLU119, which are located near the active site, interact with TFE. Evaluation of the ligand binding kinetics of Cu/Zn-SOD during unfolding in the presence of TFE combined with computational prediction allowed us to gain insight into the inactivation of Cu/Zn-SOD.
Related JoVE Video
Confident predictability: identifying reliable gene expression patterns for individualized tumor classification using a local minimax kernel algorithm.
BMC Med Genomics
PUBLISHED: 01-24-2011
Show Abstract
Hide Abstract
Molecular classification of tumors can be achieved by global gene expression profiling. Most machine learning classification algorithms furnish global error rates for the entire population. A few algorithms provide an estimate of probability of malignancy for each queried patient but the degree of accuracy of these estimates is unknown. On the other hand local minimax learning provides such probability estimates with best finite sample bounds on expected mean squared error on an individual basis for each queried patient. This allows a significant percentage of the patients to be identified as confidently predictable, a condition that ensures that the machine learning algorithm possesses an error rate below the tolerable level when applied to the confidently predictable patients.
Related JoVE Video
Analysis of the essential oils of Coriandrum sativum Using GC-MS coupled with chemometric resolution methods.
Chem. Pharm. Bull.
PUBLISHED: 01-08-2011
Show Abstract
Hide Abstract
The essential oils extracted from Coriandrum sativum L. were analyzed by GC-MS coupled with chemometric resolution methods. Through the chemometric resolution methods, peak clusters were uniquely resolved into the pure chromatographic profiles and mass spectra of each component. Qualitative analysis was performed by comparing the pure mass spectra with those in the NIST 05 mass spectral library. Quantitative analysis was performed using the total volume integration method. A total of 118 constituents were detected, of which 104 were identified, accounting for 97.27% of the total content. The results indicate that GC-MS combined with chemometric resolution methods can greatly enhance the capability of separation and the reliability of qualitative and quantitative results. The combined method is an economical and accurate approach for the rapid analysis of the complex essential oil samples in Coriandrum sativum L.
Related JoVE Video
Tools for efficient epistasis detection in genome-wide association study.
Source Code Biol Med
PUBLISHED: 01-04-2011
Show Abstract
Hide Abstract
Genome-wide association study (GWAS) aims to find genetic factors underlying complex phenotypic traits, for which epistasis or gene-gene interaction detection is often preferred over single-locus approach. However, the computational burden has been a major hurdle to apply epistasis test in the genome-wide scale due to a large number of single nucleotide polymorphism (SNP) pairs to be tested.
Related JoVE Video
[Effects of different anesthesia and analgesia on erythrocyte immune function of patients with ovarian benign tumor treated by laparoscopic therapeutic].
Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi
PUBLISHED: 12-09-2010
Show Abstract
Hide Abstract
To investigate the effects of different anesthesia and analgesia on erythrocyte immune function of patients with ovarian benign tumor treated by laparoscopic therapeutic.
Related JoVE Video
Control of population stratification by correlation-selected principal components.
Biometrics
PUBLISHED: 12-06-2010
Show Abstract
Hide Abstract
In genome-wide association studies, population stratification is recognized as producing inflated type I error due to the inflation of test statistics. Principal component-based methods applied to genotypes provide information about population structure, and have been widely used to control for stratification. Here we explore the precise relationship between genotype principal components and inflation of association test statistics, thereby drawing a connection between principal component-based stratification control and the alternative approach of genomic control. Our results provide an inherent justification for the use of principal components, but call into question the popular practice of selecting principal components based on significance of eigenvalues alone. We propose a new approach, called EigenCorr, which selects principal components based on both their eigenvalues and their correlation with the (disease) phenotype. Our approach tends to select fewer principal components for stratification control than does testing of eigenvalues alone, providing substantial computational savings and improvements in power. Analyses of simulated and real data demonstrate the usefulness of the proposed approach.
Related JoVE Video
Simultaneous determination of isoniazid, pyrazinamide, rifampicin and acetylisoniazid in human plasma by high-performance liquid chromatography.
Anal Sci
PUBLISHED: 11-17-2010
Show Abstract
Hide Abstract
A sensitive and accurate high-performance liquid-chromatography method was developed and validated for the simultaneous determination of pyrazinamide (PZA), isoniazid (INH), rifampicin (RFP) and acetylisoniazid (AcINH) in human plasma. Separation was performed on a Max-RP C(12) column using gradient elution and a flow-rate program. The mobile phase was methanol-acetonitrile-buffer (20 mM of heptanesulfonic acid sodium, pH 2.5) with a ratio of 10:8:82 (v/v/v) at the initial phase. All calibration curves had good linearity (r(2) > 0.99) between the test ranges. The intra- and inter-day precision was less than 8.8% in good accuracy (<15%). The limit of detection with a signal-to-noise (S/N) of 3 was 0.014, 0.009, 0.023 and 0.054 µg mL(-1) for PZA, AcINH, INH and RFP, respectively. The method was selective, sensitive and reliable, and is a good alternative for routine therapeutic drug monitoring of the four compounds during the treatment of tuberculosis patients.
Related JoVE Video
BIPES, a cost-effective high-throughput method for assessing microbial diversity.
ISME J
PUBLISHED: 10-21-2010
Show Abstract
Hide Abstract
Pyrosequencing of 16S rRNA (16S) variable tags has become the most popular method for assessing microbial diversity, but the method remains costly for the evaluation of large numbers of environmental samples with high sequencing depths. We developed a barcoded Illumina paired-end (PE) sequencing (BIPES) method that sequences each 16S V6 tag from both ends on the Illumina HiSeq 2000, and the PE reads are then overlapped to obtain the V6 tag. The average accuracy of Illumina single-end (SE) reads was only 97.9%, which decreased from ?99.9% at the start of the read to less than 85% at the end of the read; nevertheless, overlapping of the PE reads significantly increased the sequencing accuracy to 99.65% by verifying the 3 end of each SE in which the sequencing quality was degraded. After the removal of tags with two or more mismatches within the medial 40-70 bases of the reads and of tags with any primer errors, the overall base sequencing accuracy of the BIPES reads was further increased to 99.93%. The BIPES reads reflected the amounts of the various tags in the initial template, but long tags and high GC tags were underestimated. The BIPES method yields 20-50 times more 16S V6 tags than does pyrosequencing in a single-flow cell run, and each of the BIPES reads costs less than 1/40 of a pyrosequencing read. As a laborsaving and cost-effective method, BIPES can be routinely used to analyze the microbial ecology of both environmental and human microbiomes.
Related JoVE Video
Up-regulation of the transient A-type K+ current (IA) in the differentiation of neural stem cells of the early postnatal rat hippocampus.
Chin. Med. J.
PUBLISHED: 09-08-2010
Show Abstract
Hide Abstract
Neural stem cells (NSCs) not only are essential to cell replacement therapy and transplantation in clinical settings, but also provide a unique model for the research into neurogenesis and epigenesis. However, little attention has been paid to the electrophysiological characterization of NSC development. This work aimed to identify whether the morphological neuronal differentiation process in NSCs included changes in the electrophysiological properties of transient A-type K(+) currents (I(A)).
Related JoVE Video
Simultaneous determination of ropivacaine, bupivacaine and dexamethasone in biodegradable PLGA microspheres by high performance liquid chromatography.
Yakugaku Zasshi
PUBLISHED: 08-06-2010
Show Abstract
Hide Abstract
A simple and rapid high-performance liquid chromatography method coupled with UV detector was developed and validated for the simultaneous determination of ropivacaine, bupivacaine and dexamethasone in biodegradable poly(lactic-co-glycolic acid) (PLGA) microspheres within 11 min. Chromatographic separation was performed on a XDB-C(18) column using a mobile phase comprised of acetonitrile-NaH(2)PO(4) buffer (pH 3.5, 30 mM) (30:70, v/v) with a flow rate gradient program. The method was in good linearity (r>0.999) over the range of 0.025-40.0 microg/ml for ropivacaine and bupivacaine, and 0.05-40 microg/ml for dexamethasone. The method was proved to be precise with intra- and inter-day precision less than 3.0% and 6.0% for all drugs and accurate with intra- and inter-day accuracy between -8.0% to 4.5% and between -5.0% to 5.5% for all drugs. The assay was rapid, simple and easy to apply. Therefore, it was very suitable for routine determination and quality control of ropivacaine, bupivacaine and dexamethasone in PLGA microspheres.
Related JoVE Video
High-dimensional variable selection in meta-analysis for censored data.
Biometrics
PUBLISHED: 08-05-2010
Show Abstract
Hide Abstract
This article considers the problem of selecting predictors of time to an event from a high-dimensional set of candidate predictors using data from multiple studies. As an alternative to the current multistage testing approaches, we propose to model the study-to-study heterogeneity explicitly using a hierarchical model to borrow strength. Our method incorporates censored data through an accelerated failure time model. Using a carefully formulated prior specification, we develop a fast approach to predictor selection and shrinkage estimation for high-dimensional predictors. For model fitting, we develop a Monte Carlo expectation maximization (MC-EM) algorithm to accommodate censored data. The proposed approach, which is related to the relevance vector machine (RVM), relies on maximum a posteriori estimation to rapidly obtain a sparse estimate. As for the typical RVM, there is an intrinsic thresholding property in which unimportant predictors tend to have their coefficients shrunk to zero. We compare our method with some commonly used procedures through simulation studies. We also illustrate the method using the gene expression barcode data from three breast cancer studies.
Related JoVE Video
Integrated lysis procedures reduce extraction biases of microbial DNA from mangrove sediment.
J. Biosci. Bioeng.
PUBLISHED: 07-26-2010
Show Abstract
Hide Abstract
Sufficient lysis of soil or sediment microbes is a critical step for analyzing microbial community structures and for preparing metagenomic DNA libraries. The present study compared lysis methods for recovering archaeal, bacterial, actinomycete, and fungal DNAs from a mangrove sediment sample. PCR results showed that individual procedures using SDS, lysozyme, sonication, freeze-thaw, microwave, and vigorous shaking could extract archaeal or bacterial DNA but failed for actinomycetes or fungi cells. In comparison, an integrated lysis procedure using SDS, lysozyme, and vigorous shaking successfully obtained fungal DNA, and a combination of SDS, lysozyme, vigorous shaking, and microwave treatments recovered DNA from actinomycetes. Denaturing gradient gel electrophoresis (DGGE) results showed that although single lysis procedures can lyse bacterial DNA, all of them assessed the indigenous bacterial community structure with significant biases. The integrated lysis protocols described in the present study could be useful for extracting DNA from various types of sediments.
Related JoVE Video
[Dextran sedimentation for study of neutrophil polarization].
Nan Fang Yi Ke Da Xue Xue Bao
PUBLISHED: 07-24-2010
Show Abstract
Hide Abstract
To determine the optimal method for separating neutrophils for studying neutrophil polarization.
Related JoVE Video
High diversity of extended-spectrum beta-lactamase-producing bacteria in an urban river sediment habitat.
Appl. Environ. Microbiol.
PUBLISHED: 07-16-2010
Show Abstract
Hide Abstract
Antibiotic-resistant bacteria (ARB) have been surveyed widely in water bodies, but few studies have determined the diversity of ARB in sediment, which is the most taxon-abundant habitat in aquatic environments. We isolated 56 extended-spectrum beta-lactamase (ESBL)-producing bacteria from a single sediment sample taken from an urban river in China. All strains were confirmed for ESBL-producing capability by both the clavulanic acid combination disc method and MIC determination. Of the isolated strains, 39 were classified as Enterobacteriaceae (consisting of the genera Escherichia, Klebsiella, Serratia, and Aeromonas) by 16S rRNA gene sequencing and biochemical analysis. The present study identifies, for the first time, ESBL-producing strains from the families Brucellaceae and Moraxellaceae. The bla(CTX-M) gene was the most dominant of the ESBL genes (45 strains), while the bla(TEM) gene was the second-most dominant (22 strains). A total of five types of bla(CTX-M) fragments were identified, with both known and novel sequences. A library of bla(CTX-M) cloned from the sediment DNA showed an even higher diversity of bla(CTX-M) sequences. The discovery of highly diverse ESBL-producing bacteria and ESBL genes, particularly bla(CTX), in urban river sediment raises alarms for potential dissemination of ARB in communities through river environments.
Related JoVE Video
Effects of polymerase, template dilution and cycle number on PCR based 16 S rRNA diversity analysis using the deep sequencing method.
BMC Microbiol.
PUBLISHED: 06-26-2010
Show Abstract
Hide Abstract
The primer and amplicon length have been found to affect PCR based estimates of microbial diversity by pyrosequencing, while other PCR conditions have not been addressed using any deep sequencing method. The present study determined the effects of polymerase, template dilution and PCR cycle number using the Solexa platform.
Related JoVE Video
Overexpression of aspartyl-(asparaginyl)-beta-hydroxylase in hepatocellular carcinoma is associated with worse surgical outcome.
Hepatology
PUBLISHED: 06-26-2010
Show Abstract
Hide Abstract
The association between the overexpression of aspartyl-(asparaginyl)-beta-hydroxylase (AAH) and the invasiveness of hepatocellular carcinoma (HCC) in vitro has been reported. However, the prognostic value of AAH expression in HCC remains unclear. The purpose of this study was to investigate the relationship between AAH expression, tumor recurrence, and patient survival. We identified AAH as the most overexpressed gene in HCC by way of complementary DNA microarray hybridization. A prospective study of 233 patients undergoing curative resection indicated that AAH expression was an independent factor affecting recurrence (hazard ratio [HR] 3.161, 95% confidence interval [CI] 2.115-4.724, P < 0.001) and survival (HR 2.712, 95% CI 1.734-4.241, P < 0.001). Patients with AAH overexpression had a poorer prognosis than those with AAH underexpression (P < 0.001 for both recurrence and survival). In Barcelona Clinic Liver Cancer stage A patients with AAH overexpression or underexpression, the tumor recurrence and survival rates were also statistically different (45% and 85% versus16% and 33% in 1- and 3-year cumulative recurrence rates, respectively; 73% and 37% versus 90% and 80% in 1- and 3-year survival rates, respectively; P < 0.001 for both). Furthermore, in stage A patients with tumors measuring < or =5 cm in diameter, the time to recurrence was 26.7 +/- 1.6 versus 51.9 +/- 2.8 months, and the 1- and 3- year survival rates were 97% and 52% versus 100% and 90% in AAH overexpression and underexpression patients, respectively (P < 0.001 for both).
Related JoVE Video
Nanodiamond as the pH-responsive vehicle for an anticancer drug.
Small
PUBLISHED: 06-23-2010
Show Abstract
Hide Abstract
Cis-dichlorodiammineplatinum(II) (CDDP, cisplatin), a widely used anticancer drug, is successfully loaded onto nanodiamond (ND) by adsorption and complexation. The CDDP-ND composite is characterized by IR spectroscopy, atomic absorption spectroscopy, thermogravimetric analysis, energy-dispersive X-ray spectroscopy, and X-ray photoelectron spectroscopy. CDDP is released from the composite in phosphate-buffered saline (PBS) of pH 6.0 at a rate higher than in PBS of pH 7.4. Therefore, it is predicted that the ND vehicle would deliver low concentrations of CDDP in the blood, but release much more drug after integration into the acidic cytoplasm, thereby reducing toxic side effects. The complexation between CDDP and the carboxyl groups on the ND surface is responsible for the pH-responsive release property. The drug released from the composite retains the same cytotoxicity as free CDDP against human cervical cancer cells.
Related JoVE Video
Nonparametric Bayesian variable selection with applications to multiple quantitative trait loci mapping with epistasis and gene-environment interaction.
Genetics
PUBLISHED: 06-15-2010
Show Abstract
Hide Abstract
The joint action of multiple genes is an important source of variation for complex traits and human diseases. However, mapping genes with epistatic effects and gene-environment interactions is a difficult problem because of relatively small sample sizes and very large parameter spaces for quantitative trait locus models that include such interactions. Here we present a nonparametric Bayesian method to map multiple quantitative trait loci (QTL) by considering epistatic and gene-environment interactions. The proposed method is not restricted to pairwise interactions among genes, as is typically done in parametric QTL analysis. Rather than modeling each main and interaction term explicitly, our nonparametric Bayesian method measures the importance of each QTL, irrespective of whether it is mostly due to a main effect or due to some interaction effect(s), via an unspecified function of the genotypes at all candidate QTL. A Gaussian process prior is assigned to this unknown function. In addition to the candidate QTL, nongenetic factors and covariates, such as age, gender, and environmental conditions, can also be included in the unspecified function. The importance of each genetic factor (QTL) and each nongenetic factor/covariate included in the function is estimated by a single hyperparameter, which enters the covariance function and captures any main or interaction effect associated with a given factor/covariate. An initial evaluation of the performance of the proposed method is obtained via analysis of simulated and real data.
Related JoVE Video

What is Visualize?

JoVE Visualize is a tool created to match the last 5 years of PubMed publications to methods in JoVE's video library.

How does it work?

We use abstracts found on PubMed and match them to JoVE videos to create a list of 10 to 30 related methods videos.

Video X seems to be unrelated to Abstract Y...

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.