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1.  Inhibitional Effects of Metal Zn2+ on the Reproduction of Aphis medicaginis and Its Predation by Harmonia axyridis 
PLoS ONE  2014;9(2):e87639.
Background
Contamination, including metals, can disturb the reproductive processes of many organisms, including both prey and predatory insects. However, there is virtually no information on the effects of high level Zinc (Zn) pollution on aphids and ladybirds. The high concentrations of Zn2+ or Zn pollution inhibit reproduction in the phytophagous aphid, Aphis medicaginis, and the predatory ladybird Harmonia axyridis could provide important information.
Results
It was observed in this study that Zn concentrations in Vicia faba (broad bean) seeds and seedlings in all Zn2+ treatments were significantly higher than that in the control group, and increased with increasing Zn2+ concentrations in the solution. The rate of reproduction in A. medicaginis declined significantly (p<0.05) over time in the five groups fed on broad bean seedlings treated with different concentrations of Zn2+ solution compared with the control group. These results showed that higher concentrations of Zn2+ significantly inhibited the reproductive capacity of A. medicaginis. We also cloned and identified a gene encoding vitellogenin (Vg) from A. medicaginis, which has an important role in vitellogenesis, and therefore, reproduction was affected by exposure to Zn2+. Expression of AmVg was reduced with increasing exposure to Zn2+ and also in the F1–F3 generations of aphids exposed to different Zn2+ concentrations. Predation by H. axyridis was also reduced in aphids exposed to high-levels of Zn2+. Similarly, ovipositioning by H. axyridis was also reduced.
Conclusions
Our results suggest that Zn2+ can significantly affect the reproductive capacity of both A. medicaginis and its predator H. axyridis, the former through effects on the expression of AmVg and the latter through avoidance of aphids containing high levels of Zn2+.
doi:10.1371/journal.pone.0087639
PMCID: PMC3922717  PMID: 24533059
2.  Behavioral and transcriptome alterations in male and female mice with postnatal deletion of TrkB in dorsal striatal medium spiny neurons 
Background
The high affinity tyrosine kinase receptor, TrkB, is the primary receptor for brain derived neurotrophic factor (BDNF) and plays an important role in development, maintenance and plasticity of the striatal output medium size spiny neuron. The striatal BDNF/TrkB system is thereby implicated in many physiologic and pathophysiologic processes, the latter including mood disorders, addiction, and Huntington’s disease. We crossed a mouse harboring a transgene directing cre-recombinase expression primarily to postnatal, dorsal striatal medium spiny neurons, to a mouse containing a floxed TrkB allele (fB) mouse designed for deletion of TrkB to determine its role in the adult striatum.
Results
We found that there were sexually dimorphic alterations in behaviors in response to stressful situations and drugs of abuse. Significant sex and/or genotype differences were found in the forced swim test of depression-like behaviors, anxiety-like behaviors on the elevated plus maze, and cocaine conditioned reward. Microarray analysis of dorsal striatum revealed significant dysregulation in individual and groups of genes that may contribute to the observed behavioral responses and in some cases, represent previously unidentified downstream targets of TrkB.
Conclusions
The data point to a set of behaviors and changes in gene expression following postnatal deletion of TrkB in the dorsal striatum distinct from those in other brain regions.
doi:10.1186/1750-1326-8-47
PMCID: PMC3880973  PMID: 24369067
TrkB.FL; Medium spiny neuron; Dorsal striatum; BDNF; DARPP-32
3.  Dynamic QTL Analysis and Candidate Gene Mapping for Waterlogging Tolerance at Maize Seedling Stage 
PLoS ONE  2013;8(11):e79305.
Soil waterlogging is one of the major abiotic stresses adversely affecting maize growth and yield. To identify dynamic expression of genes or quantitative trait loci (QTL), QTL associated with plant height, root length, root dry weight, shoot dry weight and total dry weight were identified via conditional analysis in a mixed linear model and inclusive composite interval mapping method at three respective periods under waterlogging and control conditions. A total of 13, 19 and 23 QTL were detected at stages 3D|0D (the period during 0–3 d of waterlogging), 6D|3D and 9D|6D, respectively. The effects of each QTL were moderate and distributed over nine chromosomes, singly explaining 4.14–18.88% of the phenotypic variation. Six QTL (ph6-1, rl1-2, sdw4-1, sdw7-1, tdw4-1 and tdw7-1) were identified at two consistent stages of seedling development, which could reflect a continuous expression of genes; the remaining QTL were detected at only one stage. Thus, expression of most QTL was influenced by the developmental status. In order to provide additional evidence regarding the role of corresponding genes in waterlogging tolerance, mapping of Expressed Sequence Tags markers and microRNAs were conducted. Seven candidate genes were observed to co-localize with the identified QTL on chromosomes 1, 4, 6, 7 and 9, and may be important candidate genes for waterlogging tolerance. These results are a good starting point for understanding the genetic basis for selectively expressing of QTL in different stress periods and the common genetic control mechanism of the co-localized traits.
doi:10.1371/journal.pone.0079305
PMCID: PMC3828346  PMID: 24244474
4.  Sex differences in spinal processing of transient and inflammatory colorectal stimuli in the rat 
Pain  2012;153(9):1965-1973.
Sex differences in the spinal processing of somatic and visceral stimuli contribute to greater female sensitivity in many pain disorders. The present study examined spinal mechanisms that contribute to sex differences in visceral sensitivity. The visceromotor response to colorectal distention (CRD) was more robust in normal female rats and following intracolonic mustard oil compared to males. No sex difference was observed in the CRD-evoked response of lumbosacral (LS) and thoracolumbar (TL) colonic afferents in normal and mustard oil-treated rats, but there was a sex difference in spontaneous activity that was exacerbated by intracolonic mustard oil. The response of visceroceptive dorsal horn neurons to CRD was greater in normal females in the LS and TL spinal segments. The effect of intracolonic mustard oil on the CRD-evoked response of different phenotypes of visceroceptive dorsal horn neurons was dependent on sex and segment. The NMDA receptor antagonist APV dose-dependently attenuated the visceromotor response in normal rats with greater effect in males. Correspondingly, there was greater cell membrane expression of the GluN1 subunit in dorsal horn extracts in females. Following intracolonic mustard oil there was no longer a sex difference in the effect of APV nor GluN1 expression in LS segments but greater female expression in TL segments.
These data document a sex difference in spinal processing of nociceptive visceral stimuli from the normal and inflamed colon. Differences in dorsal horn neuronal activity and NMDA receptor expression contribute to the sex differences in the visceral sensitivity observed in awake rats.
doi:10.1016/j.pain.2012.06.019
PMCID: PMC3413769  PMID: 22819535
visceromotor response; hyperalgesia; primary afferent; spinal cord; dorsal horn neuron; visceral pain; gonadal hormone; NMDA receptor
5.  Cell-Penetrating Peptide-Mediated Therapeutic Molecule Delivery into the Central Nervous System 
Current Neuropharmacology  2013;11(2):197-208.
The blood-brain barrier (BBB), a dynamic and complex barrier formed by endothelial cells, can impede the entry of unwanted substances – pathogens and therapeutic molecules alike – into the central nervous system (CNS) from the blood circulation. Taking into account the fact that CNS-related diseases are the largest and fastest growing unmet medical concern, many potential protein- and nucleic acid-based medicines have been developed for therapeutic purposes. However, due to their poor ability to cross the BBB and the plasma membrane, the above-mentioned bio-macromolecules have limited use in treating neurological diseases. Finding effective, safe, and convenient ways to deliver therapeutic molecules into the CNS is thus urgently required. In recent decades, much effort has been expended in the development of drug delivery technologies, of which cell-penetrating peptides (CPPs) have the most promising potential. The present review covers the latest advances in CPP delivery technology, and provides an update on their use in CNS-targeted drug delivery.
doi:10.2174/1570159X11311020006
PMCID: PMC3637673  PMID: 23997754
Central nervous system; blood-brain barrier; cell-penetrating peptides; drug delivery.
6.  Forkhead box protein p1 is a transcriptional repressor of immune signaling in the CNS: implications for transcriptional dysregulation in Huntington disease 
Human Molecular Genetics  2012;21(14):3097-3111.
Forkhead box protein p1 (Foxp1), a transcription factor showing highly enriched expression in the striatum, has been implicated in central nervous system (CNS) development, but its role in the mature brain is unknown. In order to ascertain functional roles for Foxp1 in the CNS, we have identified gene targets for Foxp1 both in vitro and in vivo using genome-wide expression microarrays and chromatin-immunoprecipitation followed by high-throughput sequencing (ChIP-seq) assays. We found that mouse Foxp1 overexpression in striatal cells elicited expression changes of genes related to immune signaling, transcriptional regulation and a manually curated Huntington's disease (HD)-signaling pathway. Similar results were found when the gene expression data set was integrated with Foxp1-binding data determined from ChIP-seq analysis. In vivo lentiviral-mediated overexpression of human FOXP1 in the context of mutant huntingtin (Htt) protein resulted in a robust downregulation of glial cell-associated, immune genes, including those encoding a variety of cytokines and chemokines. Furthermore, Foxp1-induced expression changes were significantly negatively correlated with those changes elicited by mutant Htt protein in several different HD mouse models, and most significantly in post-mortem caudate from human HD subjects. We finally show that Foxp1 interacts with mutant Htt protein in mouse brain and is present in nuclear Htt aggregates in the striatum of R6/1 transgenic mice. These findings implicate Foxp1 as a key repressor of immune signaling in the CNS and suggest that the loss of Foxp1-mediated gene regulation in HD contributes to the immune dysfunction in this disease. We further suggest that Foxp1-regulated pathways might be important mediators of neuronal-glial cell communication.
doi:10.1093/hmg/dds132
PMCID: PMC3384380  PMID: 22492998
7.  Estrogen receptor β activation is antinociceptive in a model of visceral pain in the rat 
The Journal of Pain  2012;13(7):685-694.
The mechanism underlying estrogen modulation of visceral pain remains unclear. Our previous studies indicate activation of estrogen receptor α (ERα) enhances visceral pain. The purpose of the present study was to investigate the role of estrogen receptor β (ERβ) activation in spinal processing of visceral stimuli. The effects of selective ERβ agonists on the visceromotor response (VMR) and dorsal horn neuronal responses to colorectal distention (CRD) were tested in ovariectomized and intact female rats. The magnitude of the VMR to CRD was significantly attenuated by ERβ agonists diarylpropionitrile (DPN) and WAY200070 four hours after subcutaneous injection. Pretreatment with the estrogen receptor antagonist ICI 182,780 obscured the DPN-evoked attenuation. There was no effect of DPN on the VMR at earlier time points. Subcutaneous and spinal administration of DPN attenuated the response of visceroceptive dorsal horn neurons with a comparable time course. DPN attenuated the VMR in intact rats regardless of estrous cycle stage. The timecourse of effect of ERβ activation on the visceromotor response and neuronal activity is consistent with transcriptional or translational modulation of neuronal activity.
Perspective
Activation of ERβ is antinociceptive in the colorectal distention model of visceral pain, which may provide a therapeutic target to manage IBS in the clinic.
doi:10.1016/j.jpain.2012.04.010
PMCID: PMC3389154  PMID: 22698981
visceral pain; gonadal hormone; colorectal distention; ERβ agonist; spinal cord; estrogen receptor; visceromotor response
8.  First Genome Sequence of a Burkholderia pseudomallei Isolate in China, Strain BPC006, Obtained from a Melioidosis Patient in Hainan 
Journal of Bacteriology  2012;194(23):6604-6605.
Melioidosis, caused by Burkholderia pseudomallei, is considered to be endemic to Northern Australia and Southeast Asia, with high mortality and relapse rates, regardless of powerful antibiotic therapy. Here we report the first genome sequence of Burkholderia pseudomallei strain BPC006, obtained from a melioidosis patient in Hainan, China. The genome sizes of the 2 chromosomes were determined to be 4,001,777 bp and 3,153,284 bp.
doi:10.1128/JB.01577-12
PMCID: PMC3497521  PMID: 23144371
9.  Histone deacetylase (HDAC) inhibitors targeting HDAC3 and HDAC1 ameliorate polyglutamine-elicited phenotypes in model systems of Huntington's disease 
Neurobiology of disease  2012;46(2):351-361.
We have previously demonstrated amelioration of Huntington's disease (HD)-related phenotypes in R6/2 transgenic mice in response to treatment with the novel histone deacetylase (HDAC) inhibitor 4b. Here we have measured the selectivity profiles of 4b and related compounds against class I and class II HDACs and have tested their ability to restore altered expression of genes related to HD pathology in mice and to rescue disease effects in cell culture and Drosophila models of HD. R6/2 transgenic and wild-type (wt) mice received daily injections of HDAC inhibitors for 3 days followed by real-time PCR analysis to detect expression differences for 13 HD-related genes. We find that HDACi 4b and 136, two compounds showing high potency for inhibiting HDAC3 were most effective in reversing the expression of genes relevant to HD, including Ppp1r1b, which encodes DARPP-32, a marker for medium spiny striatal neurons. In contrast, compounds targeting HDAC1 were less effective at correcting gene expression abnormalities in R6/2 transgenic mice, but did cause significant increases in the expression of selected genes. An additional panel of 4b-related compounds was tested in a Drosophila model of HD and in STHdhQ111 striatal cells to further distinguish HDAC selectivity. Significant improvement in huntingtin-elicited Drosophila eye neurodegeneration in the fly was observed in response to treatment with compounds targeting human HDAC1 and/or HDAC3. In STHdhQ111 striatal cells, the ability of HDAC inhibitors to improve Htt-elicited metabolic deficits correlated with the potency at inhibiting HDAC1 and HDAC3, although the IC50 values for HDAC1 inhibition were typically 10-fold higher than for inhibition of HDAC3. Assessment of HDAC protein localization in brain tissue by Western blot analysis revealed accumulation of HDAC1 and HDAC3 in the nucleus of HD transgenic mice compared to wt mice, with a concurrent decrease in cytoplasmic localization, suggesting that these HDACs contribute to a repressive chromatin environment in HD. No differences were detected in the localization of HDAC2, HDAC4 or HDAC7. These results suggest that inhibition of HDACs 1 and 3 can relieve HD-like phenotypes in model systems and that HDAC inhibitors targeting these isotypes might show therapeutic benefit in human HD.
PMCID: PMC3528106  PMID: 22590724
neurodegenerative; striatum; disease; epigenetic; therapeutic; chromatin; gene expression
10.  Response Surface Methodology for Ultrasound-Assisted Extraction of Astaxanthin from Haematococcus pluvialis 
Marine Drugs  2013;11(5):1644-1655.
Astaxanthin is a novel carotenoid nutraceutical occurring in many crustaceans and red yeasts. It has exhibited various biological activities including prevention or amelioration of cardiovascular disease, gastric ulcer, hypertension, and diabetic nephropathy. In this study, ultrasound-assisted extraction was developed for the effective extraction of astaxanthin from Haematococcus pluvialis. Some parameters such as extraction solvent, liquid-to-solid ratio, extraction temperature, and extraction time were optimized by single-factor experiment and response surface methodology. The optimal extraction conditions were 48.0% ethanol in ethyl acetate, the liquid-to-solid ratio was 20:1 (mL/g), and extraction for 16.0 min at 41.1 °C under ultrasound irradiation of 200 W. Under optimal conditions, the yield of astaxanthin was 27.58 ± 0.40 mg/g. The results obtained are beneficial for the full utilization of Haematococcus pluvialis, which also indicated that ultrasound-assisted extraction is a very useful method for extracting astaxanthin from marine life.
doi:10.3390/md11051644
PMCID: PMC3707165  PMID: 23697948
ultrasound-assisted extraction; astaxanthin; Haematococcus pluvialis; response surface methodology
11.  Differential age- and disease-related effects on the expression of genes related to the arachidonic acid signaling pathway in schizophrenia 
Psychiatry Research  2012;196(2-3):201-206.
We have previously identified differential effects of age on global brain gene expression profiles in subjects with schizophrenia compared to normal controls. Here, we have focused on age-related effects of genes associated with the arachidonic acid-related inflammation pathway. Linear correlation analysis of published microarray expression data reveal strong age- and cell-type specific-effects on the expression of genes related to the arachidonic acid signaling pathway, which differed in control subjects compared to those with schizophrenia. Using real-time qPCR analysis, we validated age- and disease-effects of arachidonic acid-related genes in a large cohort of subjects with schizophrenia and matched controls (n=76 subjects in total). We found that levels of prostaglandin-endoperoxide synthase 1 (PTGS1; aka COX-1) and prostaglandin-endoperoxide receptor 3 (PTGER3) mRNA are increased, and levels of prostaglandin-endoperoxide synthase 2 (PTGS2; aka COX-2) mRNA are decreased, in older subjects with schizophrenia (>40 years of age) compared to matched normal controls or younger subjects with schizophrenia (<40 years of age). These findings contribute to the accumulating evidence suggesting that inflammatory processes in the CNS contribute to pathophysiology of schizophrenia and further suggest that age may be an important factor in the potential use of anti-inflammatory therapies.
doi:10.1016/j.psychres.2011.09.026
PMCID: PMC3361581  PMID: 22397921
psychiatric disorder; inflammation; prostaglandin; microarray; COX-2; Celecoxib
12.  Autophagy Protects against Oxaliplatin-Induced Cell Death via ER Stress and ROS in Caco-2 Cells 
PLoS ONE  2012;7(11):e51076.
Oxaliplatin is included in a number of effective combination regimens used as first and subsequent lines of therapy for metastatic colorectal cancer. Accumulating evidence indicates that autophagy plays a significant role in response to cancer therapy. However, the role of autophagy in oxaliplatin-induced cell death remains to be clarified. In this study, we showed that oxaliplatin induced cell death and autophagy in Caco-2 colorectal cancer cells. The suppression of autophagy using either pharmacologic inhibitors (3-methyladenine, bafilomycin A1) or RNA interference in essential autophagy genes (ATG5 or Beclin1) enhanced the cell death and reactive oxygen species (ROS) production induced by oxaliplatin in Caco-2 cells. Blocking oxaliplatin-induced ROS production by using ROS scavengers (NAC or Tiron) decreased autophagy. Furthermore, numerous dilated endoplasmic reticula (ER) were present in oxaliplatin-treated Caco-2 cells, and blocking ER stress by RNA interference against candidate of metastasis-1 (P8) and C/EBP-homologous protein (CHOP) decreased autophagy and ROS production. Taken together, these data indicate that oxaliplatin activates autophagy as a cytoprotective response via ER stress and ROS in human colorectal cancer cells.
doi:10.1371/journal.pone.0051076
PMCID: PMC3511352  PMID: 23226467
13.  Development and characterization of a novel nanoemulsion drug-delivery system for potential application in oral delivery of protein drugs 
Background:
The stability of protein drugs remains one of the key hurdles to their success in the market. The aim of the present study was to design a novel nanoemulsion drug-delivery system (NEDDS) that would encapsulate a standard-model protein drug – bovine serum albumin (BSA) – to improve drug stability.
Methods:
The BSA NEDDS was prepared using a phase-inversion method and pseudoternary phase diagrams. The following characteristics were studied: morphology, size, zeta potential, drug loading, and encapsulation efficiency. We also investigated the stability of the BSA NEDDS, bioactivity of BSA encapsulated within the NEDDS, the integrity of the primary, secondary, and tertiary structures, and specificity.
Results:
The BSA NEDDS consisted of Cremophor EL-35, propylene glycol, isopropyl myristate, and normal saline. The average particle diameter of the BSA NEDDS was about 21.8 nm, and the system showed a high encapsulation efficiency (>90%) and an adequate drug-loading capacity (45 mg/mL). The thermodynamic stability of the system was investigated at different temperatures and pH levels and in room-temperature conditions for 180 days. BSA NEDDS showed good structural integrity and specificity for the primary, secondary, and tertiary structures, and good bioactivity of the loaded BSA.
Conclusions:
BSA NEDDS showed the properties of a good nanoemulsion-delivery system. NEDDS can greatly enhance the stability of the protein drug BSA while maintaining high levels of drug bioactivity, good specificity, and integrity of the primary, secondary, and tertiary protein structures. These findings indicate that the nanoemulsion is a potential formulation for oral administration of protein drugs.
Video abstract
Video
doi:10.2147/IJN.S36071
PMCID: PMC3484902  PMID: 23118537
nanoemulsion; drug-delivery system; protein drug; oral administration; stability
14.  Global Gene Expression Analysis of the Zoonotic Parasite Trichinella spiralis Revealed Novel Genes in Host Parasite Interaction 
Background
Trichinellosis is a typical food-borne zoonotic disease which is epidemic worldwide and the nematode Trichinella spiralis is the main pathogen. The life cycle of T. spiralis contains three developmental stages, i.e. adult worms, new borne larva (new borne L1 larva) and muscular larva (infective L1 larva). Stage-specific gene expression in the parasites has been investigated with various immunological and cDNA cloning approaches, whereas the genome-wide transcriptome and expression features of the parasite have been largely unknown. The availability of the genome sequence information of T. spiralis has made it possible to deeply dissect parasite biology in association with global gene expression and pathogenesis.
Methodology and Principal Findings
In this study, we analyzed the global gene expression patterns in the three developmental stages of T. spiralis using digital gene expression (DGE) analysis. Almost 15 million sequence tags were generated with the Illumina RNA-seq technology, producing expression data for more than 9,000 genes, covering 65% of the genome. The transcriptome analysis revealed thousands of differentially expressed genes within the genome, and importantly, a panel of genes encoding functional proteins associated with parasite invasion and immuno-modulation were identified. More than 45% of the genes were found to be transcribed from both strands, indicating the importance of RNA-mediated gene regulation in the development of the parasite. Further, based on gene ontological analysis, over 3000 genes were functionally categorized and biological pathways in the three life cycle stage were elucidated.
Conclusions and Significance
The global transcriptome of T. spiralis in three developmental stages has been profiled, and most gene activity in the genome was found to be developmentally regulated. Many metabolic and biological pathways have been revealed. The findings of the differential expression of several protein families facilitate understanding of the molecular mechanisms of parasite biology and the pathological aspects of trichinellosis.
Author Summary
Trichinellosis of human and other mammals was caused through the ingestion of the parasite Trichinella sparilis in contaminated meat. It is a typical zoonotic disease that affects more than 10 million people world-wide. Parasites of the genus Trichinella are unique intracellular pathogens. Adult Trichinella parasites directly release newborn larvae which invade striated muscle cells and causes diseases. In this study, we profiled the global transcriptome in the three developmental stages of T. spiralis. The transcriptomic analysis revealed the global gene expression patterns from newborn larval stage through muscle larval stage to adults. Thousands of genes with stage-specific transcriptional patterns were described and novel genes involving host-parasite interaction were identified. More than 45% of the protein-coding genes showed evidence of transcription from both sense and antisense strands which suggests the importance of RNA-mediated gene regulation in the parasite. This study presents a first deep analysis of the transcriptome of T. spiralis, providing insight information of the parasite biology.
doi:10.1371/journal.pntd.0001794
PMCID: PMC3429391  PMID: 22953016
15.  Plasma microRNAs, miR-223, miR-21 and miR-218, as Novel Potential Biomarkers for Gastric Cancer Detection 
PLoS ONE  2012;7(7):e41629.
Background
MicroRNAs (miRNAs), endogenous small non-coding RNAs, are stably detected in human plasma. Early diagnosis of gastric cancer (GC) is very important to improve the therapy effect and prolong the survival of patients. We aimed to identify whether four miRNAs (miR-223, miR-21, miR-218 and miR-25) closely associated with the tumorigenesis or metastasis of GC can serve as novel potential biomarkers for GC detection.
Methodology
We initially measured the plasma levels of the four miRNAs in 10 GC patients and 10 healthy control subjects by quantitative reverse transcription polymerase chain reaction (qRT-PCR), and then compared plasma miRNA results with the expressions in cancer tissues from eight GC patients. Finally, the presence of miR-223, miR-21 and miR-218 in the plasma was validated in 60 GC patients and 60 healthy control subjects, and the areas under the receiver operating characteristic (ROC) curves of these miRNAs were analyzed.
Results
We found that the plasma levels of miR-223 (P<0.001) and miR-21 (P<0.001) were significantly higher in GC patients than in healthy controls, while miR-218 (P<0.001) was significantly lower. The ROC analyses yielded the AUC values of 0.9089 for miR-223, 0.7944 for miR-21 and 0.7432 for miR-218, and combined ROC analysis revealed the highest AUC value of 0.9531 in discriminating GC patients from healthy controls. Moreover, the plasma levels of miR-223 (P<0.001) and miR-21 (P = 0.003) were significantly higher in GC patients with stage I than in healthy controls. Furthermore, the plasma levels of miR-223 were significantly higher in GC patients with helicobacter pylori (Hp) infection than those without (P = 0.014), and significantly higher in healthy control subjects with Hp infection than those without (P = 0.016).
Conclusions
Plasma miR-223, miR-21 and miR-218 are novel potential biomarkers for GC detection.
doi:10.1371/journal.pone.0041629
PMCID: PMC3408505  PMID: 22860003
16.  Compromised autophagy by MIR30B benefits the intracellular survival of Helicobacter pylori 
Autophagy  2012;8(7):1045-1057.
Helicobacter pylori evade immune responses and achieve persistent colonization in the stomach. However, the mechanism by which H. pylori infections persist is not clear. In this study, we showed that MIR30B is upregulated during H. pylori infection of an AGS cell line and human gastric tissues. Upregulation of MIR30B benefited bacterial replication by compromising the process of autophagy during the H. pylori infection. As a potential mechanistic explanation for this observation, we demonstrate that MIR30B directly targets ATG12 and BECN1, which are important proteins involved in autophagy. These results suggest that compromise of autophagy by MIR30B allows intracellular H. pylori to evade autophagic clearance, thereby contributing to the persistence of H. pylori infections.
doi:10.4161/auto.20159
PMCID: PMC3429542  PMID: 22647547
Helicobacter pylori; MIR30B; ATG12; BECN1; autophagy
17.  Gene expression profiling of R6/2 transgenic mice with different CAG repeat lengths reveals genes associated with disease onset and progression in Huntington's disease 
Neurobiology of disease  2011;42(3):459-467.
R6/2 transgenic mice with expanded CAG repeats (>300) have a surprisingly prolonged disease progression and longer lifespan than prototypical parent R6/2 mice (carrying 150 CAGs), however, the mechanism of this phenotype amelioration is unknown. We compared gene expression profiles in the striatum of R6/2 transgenic mice carrying ~300 CAG repeats (R6/2Q300 transgenic mice), those carrying ~150 CAG repeats (R6/2Q150 transgenic mice) and littermate wt controls in order to identify genes that may play determinant roles in the time course of phenotypic expression in these mice. Of the top genes showing concordant expression changes in the striatum of both R6/2 lines, 85% were decreased in expression, while discordant expression changes were observed mostly for genes upregulated in R6/2Q300 transgenic mice. Upregulated genes in the R6/2Q300 mice were associated with the ubiquitin ligase complex, cell adhesion, protein folding and establishment of protein localization. We qPCR-validated increases in expression of genes related to the latter category, including Lrsam1, Erp29, Nasp, Tap1, Rab9b and Pfdn5 in R6/2Q300 mice, changes that were not observed in R6/2 mice with shorter CAG repeats, even in late stages (i.e. 12 weeks of age). We further tested Lrsam1 and Erp29, the two genes showing the greatest upregulation in R6/2Q300 transgenic mice, for potential neuroprotective effects in primary striatal cultures overexpressing a mutated human huntingtin (htt) fragment. Overexpression of Lrsam1 prevented the loss of NeuN-positive cell bodies in htt171-82Q cultures, concomitant with a reduction of nuclear htt aggregates. Erp29 showed no significant effects in this model. This is consistent with the distinct pattern of htt inclusion localization observed in R6/2Q300 transgenic mice, in which smaller cytoplasmic inclusions represent the major form of insoluble htt in the cell, as opposed to large nuclear inclusions observed in R6/2Q150 transgenic mice. We suggest that the prolonged onset and disease course observed in R6/2 mice with greatly expanded CAG repeats might result from differential upregulation of genes related to protein localization and clearance. Such genes may represent novel therapeutic avenues to decrease htt aggregate toxicity and cell death in HD patients, with Lrsam1 being a promising, novel candidate disease modifier.
doi:10.1016/j.nbd.2011.02.008
PMCID: PMC3079804  PMID: 21334439
Huntington's disease; striatum; gene expression; inclusion; neuroprotection
18.  Spinal estrogen receptor alpha mediates estradiol-induced pronociception in a visceral pain model in the rat 
Pain  2011;152(5):1182-1191.
We previously reported that 17β – estradiol (E2) is pronociceptive in a visceral pain model in the rat. Subcutaneously (s.c.) administered E2 reversed the decrease in the colorectal distention (CRD)-evoked visceromotor response produced by ovariectomy (OVx) and CRD-induced nociceptive responses were greater in proestrous rats compared to met/diestrous rats. The site of action, the type of estrogen receptors activated and the possible intracellular signaling pathway involved are yet to be established. In the present study, intrathecal (i.t.) E2 administered to OVx rats mimicked the effects of s.c. E2, suggesting spinal E2 receptors are involved. This is further supported by the observations that the anti-estrogen ICI 182,780 injected i.t. in intact female rats significantly decreased the visceromotor response to CRD, the response of colonic afferents was not affected by OVx and colonic afferents did not label for estrogen receptor α (ERα). The ERα selective agonist, 4,4',4"-[4-propyl-(1H)-pyrazole-1,3,5-triyl]tris-phenol (PPT; s.c. or i.t.) facilitated the visceromotor response similar to E2, suggesting ERα activation is involved in mediating the pronociceptive effect of E2. PPT (s.c. or i.t.) increased the response of spinal dorsal horn neurons to CRD, indicating a spinal site of action. In addition, s.c. E2 or PPT increased CRD-induced spinal extracellular-signal-regulated kinase (ERK) phosphorylation that was not observed in OVx rats and a mitogen-activated protein kinase (MAPK) kinase (MEK) inhibitor blocked facilitation of the visceromotor response by PPT. Taken together, the present study demonstrates that spinal ERα mediates the pronociceptive effect of E2 on visceral signal processing through activation of the MAPK pathway.
doi:10.1016/j.pain.2011.01.046
PMCID: PMC3079062  PMID: 21392887
colorectal distention; visceromotor response; gonadal hormones; estrogen receptor alpha; spinal cord; visceral pain; pERK; dorsal horn neurons; estradiol; PPT; colonic afferent
19.  In vivo cell-autonomous transcriptional abnormalities revealed in mice expressing mutant huntingtin in striatal but not cortical neurons 
Human Molecular Genetics  2010;20(6):1049-1060.
Huntington's disease (HD), caused by a CAG repeat expansion in the huntingtin (HTT) gene, is characterized by abnormal protein aggregates and motor and cognitive dysfunction. Htt protein is ubiquitously expressed, but the striatal medium spiny neuron (MSN) is most susceptible to dysfunction and death. Abnormal gene expression represents a core pathogenic feature of HD, but the relative roles of cell-autonomous and non-cell-autonomous effects on transcription remain unclear. To determine the extent of cell-autonomous dysregulation in the striatum in vivo, we examined genome-wide RNA expression in symptomatic D9-N171-98Q (a.k.a. DE5) transgenic mice in which the forebrain expression of the first 171 amino acids of human Htt with a 98Q repeat expansion is limited to MSNs. Microarray data generated from these mice were compared with those generated on the identical array platform from a pan-neuronal HD mouse model, R6/2, carrying two different CAG repeat lengths, and a relatively high degree of overlap of changes in gene expression was revealed. We further focused on known canonical pathways associated with excitotoxicity, oxidative stress, mitochondrial dysfunction, dopamine signaling and trophic support. While genes related to excitotoxicity, dopamine signaling and  trophic support were altered in both DE5 and R6/2 mice, which may be either cell autonomous or non-cell autonomous, genes related to mitochondrial dysfunction, oxidative stress and the peroxisome proliferator-activated receptor are primarily affected in DE5 transgenic mice, indicating  cell-autonomous mechanisms. Overall, HD-induced dysregulation of the striatal transcriptome can be largely attributed to intrinsic effects of mutant Htt, in the absence of expression in cortical neurons.
doi:10.1093/hmg/ddq548
PMCID: PMC3043657  PMID: 21177255
20.  Elongation Factor 1β′ Gene from Spodoptera exigua: Characterization and Function Identification through RNA Interference 
Elongation factor (EF) is a key regulation factor for translation in many organisms, including plants, bacteria, fungi, animals and insects. To investigate the nature and function of elongation factor 1β′ from Spodoptera exigua (SeEF-1β′), its cDNA was cloned. This contained an open reading frame of 672 nucleotides encoding a protein of 223 amino acids with a predicted molecular weight of 24.04 kDa and pI of 4.53. Northern blotting revealed that SeEF-1β′ mRNA is expressed in brain, epidermis, fat body, midgut, Malpighian tubules, ovary and tracheae. RT-PCR revealed that SeEF-1β′ mRNA is expressed at different levels in fat body and whole body during different developmental stages. In RNAi experiments, the survival rate of insects injected with SeEF-1β′ dsRNA was 58.7% at 36 h after injection, which was significantly lower than three control groups. Other elongation factors and transcription factors were also influenced when EF-1β′ was suppressed. The results demonstrate that SeEF-1β′ is a key gene in transcription in S. exigua.
doi:10.3390/ijms13078126
PMCID: PMC3430225  PMID: 22942694
elongation factor; cloning; expression pattern; RNAi; Spodoptera exigua
21.  Preparation of a Cu(II)-PVA/PA6 Composite Nanofibrous Membrane for Enzyme Immobilization 
PVA/PA6 composite nanofibers were formed by electrospinning. Cu(II)-PVA/PA6 metal chelated nanofibers, prepared by the reaction between PVA/PA6 composite nanofibers and Cu2+ solution, were used as the support for catalase immobilization. The result of the experiments showed that PVA/PA6 composite nanofibers had an excellent chelation capacity for Cu2+ ions, and the structures of nanofibers were stable during the reaction with Cu2+ solution. The adsorption of Cu(II) onto PVA/PA6 composite nanofibers was studied by the Langmuir isothermal adsorption model. The maximum amount of coordinated Cu(II) (qm) was 3.731 mmol/g (dry fiber), and the binding constant (Kl) was 0.0593 L/mmol. Kinetic parameters were analyzed for both immobilized and free catalases. The value of Vmax (3774 μmol/mg·min) for the immobilized catalases was smaller than that of the free catalases (4878 μmol/mg·min), while the Km for the immobilized catalases was larger. The immobilized catalases showed better resistance to pH and temperature than that of free form, and the storage stabilities, reusability of immobilized catalases were significantly improved. The half-lives of free and immobilized catalases were 8 days and 24 days, respectively.
doi:10.3390/ijms131012734
PMCID: PMC3497296  PMID: 23202922
PVA/PA6; nanofibrous membranes; metal chelation; enzyme; immobilization
22.  Transcriptome of Small Regulatory RNAs in the Development of the Zoonotic Parasite Trichinella spiralis 
PLoS ONE  2011;6(11):e26448.
Background
Trichinella spiralis is a parasite with unique features. It is a multicellular organism but with an intracellular parasitization and development stage. T. spiralis is the helminthic pathogen that causes zoonotic trichinellosis and afflicts more than 10 million people worldwide, whereas the parasite's biology, especially the developmental regulation is largely unknown. In other organisms, small non-coding RNAs, such as microRNAs (miRNA) and small interfering RNAs (siRNA) execute post-transcriptional regulation by translational repression or mRNA degradation, and a large number of miRNAs have been identified in diverse species. In T. spiralis, the profile of small non-coding RNAs and their function remains poorly understood.
Methodology and Principal Findings
Here, the transcriptional profiles of miRNA and siRNA in three developmental stages of T. spiralis in the rat host were investigated, and compared by high-throughput cDNA sequencing technique (“RNA-seq”). 5,443,641 unique sequence tags were obtained. Of these, 21 represented conserved miRNAs related to 13 previously identified metazoan miRNA families and 213 were novel miRNAs so far unique to T. spiralis. Some of these miRNAs exhibited stage-specific expression. Expression of miRNAs was confirmed in three stages of the life cycle by qRT-PCR and northern blot analysis. In addition, endogenous siRNAs (endo-siRNAs) were found mainly derived from natural antisense transcripts (NAT) and transposable elements (TE) in the parasite.
Conclusions and Significance
We provide evidence for the presence of miRNAs and endo-siRNAs in T. spiralis. The miRNAs accounted for the major proportion of the small regulatory RNA population of T. spiralis, while fewer endogenous siRNAs were found. The finding of stage-specific expression patterns of the miRNAs in different developmental stages of T. spiralis suggests that miRNAs may play important roles in parasite development. Our data provide a basis for further understanding of the molecular regulation and functional evolution of miRNAs in parasitic nematodes.
doi:10.1371/journal.pone.0026448
PMCID: PMC3212509  PMID: 22096484
23.  Genome-Wide Identification of Bcl11b Gene Targets Reveals Role in Brain-Derived Neurotrophic Factor Signaling 
PLoS ONE  2011;6(9):e23691.
B-cell leukemia/lymphoma 11B (Bcl11b) is a transcription factor showing predominant expression in the striatum. To date, there are no known gene targets of Bcl11b in the nervous system. Here, we define targets for Bcl11b in striatal cells by performing chromatin immunoprecipitation followed by high-throughput sequencing (ChIP-seq) in combination with genome-wide expression profiling. Transcriptome-wide analysis revealed that 694 genes were significantly altered in striatal cells over-expressing Bcl11b, including genes showing striatal-enriched expression similar to Bcl11b. ChIP-seq analysis demonstrated that Bcl11b bound a mixture of coding and non-coding sequences that were within 10 kb of the transcription start site of an annotated gene. Integrating all ChIP-seq hits with the microarray expression data, 248 direct targets of Bcl11b were identified. Functional analysis on the integrated gene target list identified several zinc-finger encoding genes as Bcl11b targets, and further revealed a significant association of Bcl11b to brain-derived neurotrophic factor/neurotrophin signaling. Analysis of ChIP-seq binding regions revealed significant consensus DNA binding motifs for Bcl11b. These data implicate Bcl11b as a novel regulator of the BDNF signaling pathway, which is disrupted in many neurological disorders. Specific targeting of the Bcl11b-DNA interaction could represent a novel therapeutic approach to lowering BDNF signaling specifically in striatal cells.
doi:10.1371/journal.pone.0023691
PMCID: PMC3164671  PMID: 21912641
24.  Microarchitecture and Nanomechanical Properties of Trabecular Bone After Strontium Administration in Osteoporotic Goats 
Strontium (Sr) ralenate is a new agent used for the prevention and treatment of osteoporosis. As a bone-seeking element, 98% of Sr is deposited in the bone and teeth after oral ingestion. However, the effect of Sr treatment on bone microarchitecture and bone nanomechanical properties remains unclear. In this study, 18 osteoporotic goats were divided into four groups according to the treatment regimen: control, calcium alone (Ca), calcium and Sr at 24 mg/kg (Ca + 24Sr), and calcium and Sr at 40 mg/kg (Ca + 40Sr). The effects of Sr administration on bone microarchitecture and nanomechanical properties of trabecular bones were analyzed with micro-CT and nanoindentation test, respectively. Serum Sr levels increased six- and tenfold in the Ca + 24Sr and Ca + 40Sr groups, respectively. Similarly, Sr in the bone increased four- and sixfold in these two groups. Sr administration significantly increased trabecular bone volume fraction, trabecular thickness, and double-labeled new bone area. Sr administration, however, did not significantly change the nanomechanical properties of trabecular bone (elastic modulus and hardness). The data suggested that Sr administration increased trabecular bone volume and improved the microarchitecture while maintaining the intrinsic tissue properties in the osteoporotic goat model.
doi:10.1007/s12011-011-9158-y
PMCID: PMC3256317  PMID: 21814830
Bone strength; Nanoindentation; Osteoporosis; Ovariectomized goats; Strontium
25.  Optimization of Ultrasound-Assisted Extraction of Anthocyanins from Mulberry, Using Response Surface Methodology 
Mulberry is one of the most widely used traditional Chinese medicines. Anthocyanins are the main bioactive components of mulberry, and possess important biological activities, such as antimicrobial, anti-inflammatory and antioxidant activities. This study investigated the ultrasound-assisted extraction (UAE) of anthocyanins from mulberry by using response surface methodology (RSM). The extraction conditions associated with anthocyanin yield, including extraction solvent, liquid-to-solid rate, temperature and extraction time, are discussed. The optimal conditions obtained by RSM for UAE from mulberry include 63.8% methanol contains 1% (v/v) trifluoroacetic acid (TFA), 43.2 °C temperature, 23.8 (v/w) liquid-to-solid ratio, and 40 min time for the maximum yield (64.70 ± 0.45 mg/g). The results indicated that the UAE can be an effective method for the extraction of some active components from plant materials.
doi:10.3390/ijms12053006
PMCID: PMC3116171  PMID: 21686165
ultrasound-assisted extraction; anthocyanins; mulberry; response surface methodology

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