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1.  Cardiac Specific Expression of Threonine 5 to Alanine Mutant Sarcolipin Results in Structural Remodeling and Diastolic Dysfunction 
PLoS ONE  2015;10(2):e0115822.
The functional importance of threonine 5 (T5) in modulating the activity of sarcolipin (SLN), a key regulator of sarco/endoplasmic reticulum (SR) Ca2+ ATPase (SERCA) pump was studied using a transgenic mouse model with cardiac specific expression of threonine 5 to alanine mutant SLN (SLNT5A). In these transgenic mice, the SLNT5A protein replaces the endogenous SLN in atria, while maintaining the total SLN content. The cardiac specific expression of SLNT5A results in severe cardiac structural remodeling accompanied by bi-atrial enlargement. Biochemical analyses reveal a selective downregulation of SR Ca2+ handling proteins and a reduced SR Ca2+ uptake both in atria and in the ventricles. Optical mapping analysis shows slower action potential propagation in the transgenic mice atria. Doppler echocardiography and hemodynamic measurements demonstrate a reduced atrial contractility and an impaired diastolic function. Together, these findings suggest that threonine 5 plays an important role in modulating SLN function in the heart. Furthermore, our studies suggest that alteration in SLN function can cause abnormal Ca2+ handling and subsequent cardiac remodeling and dysfunction.
doi:10.1371/journal.pone.0115822
PMCID: PMC4324845  PMID: 25671318
2.  Deep RNA sequencing reveals a high frequency of alternative splicing events in the fungus Trichoderma longibrachiatum 
BMC Genomics  2015;16(1):54.
Background
Alternative splicing is crucial for proteome diversity and functional complexity in higher organisms. However, the alternative splicing landscape in fungi is still elusive.
Results
The transcriptome of the filamentous fungus Trichoderma longibrachiatum was deep sequenced using Illumina Solexa technology. A total of 14305 splice junctions were discovered. Analyses of alternative splicing events revealed that the number of all alternative splicing events (10034), intron retentions (IR, 9369), alternative 5’ splice sites (A5SS, 167), and alternative 3’ splice sites (A3SS, 302) is 7.3, 7.4, 5.1, and 5.9-fold higher, respectively, than those observed in the fungus Aspergillus oryzae using Illumina Solexa technology. This unexpectedly high ratio of alternative splicing suggests that alternative splicing is important to the transcriptome diversity of T. longibrachiatum. Alternatively spliced introns had longer lengths, higher GC contents, and lower splice site scores than constitutive introns. Further analysis demonstrated that the isoform relative frequencies were correlated with the splice site scores of the isoforms. Moreover, comparative transcriptomics determined that most enzymes related to glycolysis and the citrate cycle and glyoxylate cycle as well as a few carbohydrate-active enzymes are transcriptionally regulated.
Conclusions
This study, consisting of a comprehensive analysis of the alternative splicing landscape in the filamentous fungus T. longibrachiatum, revealed an unexpectedly high ratio of alternative splicing events and provided new insights into transcriptome diversity in fungi.
Electronic supplementary material
The online version of this article (doi:10.1186/s12864-015-1251-8) contains supplementary material, which is available to authorized users.
doi:10.1186/s12864-015-1251-8
PMCID: PMC4324775  PMID: 25652134
Alternative splicing; Fungi; RNA-Seq; Intron retention; Transcriptome; Trichoderma longibrachiatum
3.  Multi-frequency complex network from time series for uncovering oil-water flow structure 
Scientific Reports  2015;5:8222.
Uncovering complex oil-water flow structure represents a challenge in diverse scientific disciplines. This challenge stimulates us to develop a new distributed conductance sensor for measuring local flow signals at different positions and then propose a novel approach based on multi-frequency complex network to uncover the flow structures from experimental multivariate measurements. In particular, based on the Fast Fourier transform, we demonstrate how to derive multi-frequency complex network from multivariate time series. We construct complex networks at different frequencies and then detect community structures. Our results indicate that the community structures faithfully represent the structural features of oil-water flow patterns. Furthermore, we investigate the network statistic at different frequencies for each derived network and find that the frequency clustering coefficient enables to uncover the evolution of flow patterns and yield deep insights into the formation of flow structures. Current results present a first step towards a network visualization of complex flow patterns from a community structure perspective.
doi:10.1038/srep08222
PMCID: PMC4316157  PMID: 25649900
4.  Unintentional injuries among Chinese children with different types and severity of disability 
Annals of epidemiology  2013;24(1):23-28.
Purpose
Little research has been done in China to study injury in individuals with disability. We investigated impact of type and severity of disability on injury among children with disability in Hubei Province of China.
Methods
A sample of 1201 children with disability were matched with 1201 healthy children on gender, age, and neighborhood. Disability type and severity were determined using the Chinese national standards. Caregivers were interviewed face-to-face about nonfatal unintentional injuries suffered by the child in the past 12 months prior to the interview. Univariate Chi-square test and logistic regression models were used to investigate association between disability type/severity and nonfatal unintentional injuries.
Results
Injury rate among children with disability was significantly higher than that among children without disability (10.2% vs. 4.4%; P <.001). Children with multiple disabilities had the highest risk of injury after controlling for confounding variables (OR=4.54; 95% CI=2.82, 7.30; P<.001). The magnitude of the association between disability and injury varied by type and severity of disability.
Conclusions
The magnitude of the association between the presence or absence of disability in children and their risk of injury was large and significant, regardless of the type or severity of the children's disabilities.
doi:10.1016/j.annepidem.2013.10.015
PMCID: PMC3897222  PMID: 24331162
Unintentional injury; Children; Disability; China
5.  A Vps21 endocytic module regulates autophagy 
Molecular Biology of the Cell  2014;25(20):3166-3177.
Vps21 plays a role in autophagy in addition to its role in endocytosis. Individual deletions of members of the endocytic Vps21 module, including a GEF and four effectors, result in autophagy defects and accumulation of autophagosomal clusters. Therefore the endocytic Vps21 module regulates autophagy.
In autophagy, the double-membrane autophagosome delivers cellular components for their degradation in the lysosome. The conserved Ypt/Rab GTPases regulate all cellular trafficking pathways, including autophagy. These GTPases function in modules that include guanine-nucleotide exchange factor (GEF) activators and downstream effectors. Rab7 and its yeast homologue, Ypt7, in the context of such a module, regulate the fusion of both late endosomes and autophagosomes with the lysosome. In yeast, the Rab5-related Vps21 is known for its role in early- to late-endosome transport. Here we show an additional role for Vps21 in autophagy. First, vps21∆ mutant cells are defective in selective and nonselective autophagy. Second, fluorescence and electron microscopy analyses show that vps21∆ mutant cells accumulate clusters of autophagosomal structures outside the vacuole. Third, cells with mutations in other members of the endocytic Vps21 module, including the GEF Vps9 and factors that function downstream of Vps21, Vac1, CORVET, Pep12, and Vps45, are also defective in autophagy and accumulate clusters of autophagosomes. Finally, Vps21 localizes to PAS. We propose that the endocytic Vps21 module also regulates autophagy. These findings support the idea that the two pathways leading to the lysosome—endocytosis and autophagy—converge through the Vps21 and Ypt7 GTPase modules.
doi:10.1091/mbc.E14-04-0917
PMCID: PMC4196867  PMID: 25143401
6.  Effect of three common IL-17 single nucleotide polymorphisms on the risk of developing gastric cancer 
Oncology Letters  2014;9(3):1398-1402.
A 1:1 matched case-control study was conducted to analyze the association between three common interleukin (IL)-17A and IL-17F single nucleotide polymorphisms (SNPs) and the risk of developing gastric cancer. Genotyping of SNPs rs2275913, rs763780 and rs3748067 within the IL-17 gene were detected by performing polymerase chain reaction-restriction fragment length polymorphism analysis. Gastric cancer patients were more likely to be cigarette smokers, alcohol drinkers and have a family history of cancer in their first-degree relatives. Patients carrying the rs763780 polymorphism were correlated with a significant increased risk of gastric cancer in codominant, dominant and recessive models. Additionally, individuals with the rs763780 polymorphism were correlated with a markedly increased risk of gastric cancer among alcohol drinkers in codominant, dominant and recessive models. Furthermore, a significant correlation was identified between the rs763780 polymorphism and the consumption of alcohol. However, no association was identified between rs2275913 and rs3748067 polymorphisms and the risk of developing gastric cancer. Thus, the present study reported that the rs763780 polymorphism may be associated with risk of developing gastric cancer in the population studied, particularly in alcohol drinkers.
doi:10.3892/ol.2014.2827
PMCID: PMC4314969  PMID: 25663919
IL-17; gastric cancer; polymorphism
7.  Monoclonal Antibody against Cell Surface GRP78 as a Novel Agent in Suppressing PI3K/AKT Signaling, Tumor Growth and Metastasis 
Purpose
The ER chaperone GRP78 translocates to the surface of tumor cells and promotes survival, metastasis, and resistance to therapy. An oncogenic function of cell surface GRP78 has been attributed to the activation of phosphoinositide 3-kinase (PI3K) pathway. We intend to use a novel anti-GRP78monoclonal antibody (MAb159) to attenuate PI3K signaling and inhibit tumor growth and metastasis.
Experimental Design
MAb159 was characterized biochemically. Anti-tumor activity was tested in cancer cell culture, tumor xenograft models, tumor metastasis models, and spontaneous tumor models. Cancer cells and tumor tissues were analyzed for PI3K activity. MAb159 was humanized and validated for diagnostic and therapeutic application.
Results
MAb159 specifically recognized surface GRP78, triggered GRP78 endocytosis, and localized to tumors but not normal organs in vivo. MAb159 inhibited tumor cell proliferation and enhanced tumor cell death both in vitro and in vivo. In MAb159 treated tumors, PI3K signaling was inhibited without compensatory MAPK pathway activation. Furthermore, MAb159 halted or reversed tumor progression in the spontaneous PTEN loss driven prostate and leukemia tumor models, and inhibited tumor growth and metastasis in xenograft models. Humanized MAb159, which retains high affinity, tumor specific localization, and the anti-tumor activity, was non-toxic in mice and had desirable pharmacokinetics.
Conclusions
GRP78 specific antibody MAb159 modulates PI3K pathway and inhibits tumor growth and metastasis. Humanized MAb159 will enter human trials shortly.
doi:10.1158/1078-0432.CCR-13-1106
PMCID: PMC4151476  PMID: 24048331
Surface GRP78; monoclonal antibody; targeted cancer therapy; PI3K
8.  EphB4-targeted imaging with antibody h131, h131-F(ab′)2 and h131-Fab 
Molecular pharmaceutics  2013;10(12):4527-4533.
Accumulating evidence suggests that overexpression of the tyrosine kinase receptor EphB4, a mediator of vascular development, is a novel target for tumor diagnosis, prognosis and therapy. Noninvasive imaging of EphB4 expression could therefore be valuable for evaluating disease course and therapeutic efficacy at the earliest stages of anti-EphB4 treatment. In this study, we systematically investigated the use of anti-EphB4 antibody h131 (150 kD) and its fragments (h131-F(ab′)2, 110 kD; h131-Fab, 50 kD) for near-infrared fluorescence (NIRF) imaging of EphB4 expression in vivo. h131-F(ab′)2 and h131-Fab were produced through pepsin and papain digestion of h131 respectively, whose purity was confirmed by FPLC and SDS-PAGE. After conjugation with Cy5.5, in vivo characteristics of h131, h131-F(ab′)2 and h131-Fab were evaluated in EphB4-positive HT29 tumor model. Although h131-Cy5.5 demonstrated highest tumor uptake among these probes, its optimal tumor uptake level was obtained at 2 d post injection (p.i.). For h131-Fab-Cy5.5, maximum tumor uptake was achieved at 4 h p.i.. However, no significant difference was observed between h131-Fab-Cy5.5 and hIgG-Fab-Cy5.5, indicating the tumor accumulation was mainly caused by passive targeting. In contrast, h131-F(ab′)2-Cy5.5 demonstrated prominent tumor uptake at 6 h p.i. The target specificity was confirmed by hIgG-F(ab′)2-Cy5.5 control and immunofluorescent staining. Collectively, h131-F(ab′)2 exhibited prominent and specific tumor uptake at early time points, which suggests it is a promising agent for EphB4-targeted imaging.
doi:10.1021/mp400354y
PMCID: PMC3888823  PMID: 24147882
EphB4 receptor; near-infrared fluorescence (NIRF) imaging; antibody fragment; F(ab′)2; Fab
9.  Pleiotropy of the de novo-originated gene MDF1 
Scientific Reports  2014;4:7280.
MDF1 is a young de novo-originated gene from a non-coding sequence in baker's yeast, S. cerevisiae, which can suppress mating and promote vegetative growth. Our previous experiments successfully demonstrated how Mdf1p binds to the key mating pathway determinant MATα2 to suppress mating. However, how Mdf1p promotes growth and fulfills the crosstalk between the yeast mating and growth pathways are still open questions. Thus, the adaptive significance of this new de novo gene remains speculative. Here, we show that Mdf1p shortens the lag phase of S. cerevisiae by physically interacting with SNF1, the governing factor for nonfermentable carbon source utilization, and thereby confers a selective advantage on yeasts through the rapid consumption of glucose in the early generational stage in rich medium. Therefore, MDF1 functions in two important molecular pathways, mating and fermentation, and mediates the crosstalk between reproduction and vegetative growth. Together, our results provide a comprehensive example of how a de novo-originated gene organizes new regulatory circuits and thereby confers a selective advantage on S. cerevisiae to allow exquisite adaptation to the changing environment.
doi:10.1038/srep07280
PMCID: PMC4250933  PMID: 25452167
10.  Reaction Kinetics of Substrate Transglycosylation Catalyzed by TreX of Sulfolobus solfataricus and Effects on Glycogen Breakdown 
Journal of Bacteriology  2014;196(11):1941-1949.
We studied the activity of a debranching enzyme (TreX) from Sulfolobus solfataricus on glycogen-mimic substrates, branched maltotetraosyl-β-cyclodextrin (Glc4-β-CD), and natural glycogen to better understand substrate transglycosylation and the effect thereof on glycogen debranching in microorganisms. The validation test of Glc4-β-CD as a glycogen mimic substrate showed that it followed the breakdown process of the well-known yeast and rat liver extract. TreX catalyzed both hydrolysis of α-1,6-glycosidic linkages and transglycosylation at relatively high (>0.5 mM) substrate concentrations. TreX transferred maltotetraosyl moieties from the donor substrate to acceptor molecules, resulting in the formation of two positional isomers of dimaltotetraosyl-α-1,6-β-cyclodextrin [(Glc4)2-β-CD]; these were 61,63- and 61,64-dimaltotetraosyl-α-1,6-β-CD. Use of a modified Michaelis-Menten equation to study substrate transglycosylation revealed that the kcat and Km values for transglycosylation were 1.78 × 103 s−1 and 3.30 mM, respectively, whereas the values for hydrolysis were 2.57 × 103 s−1 and 0.206 mM, respectively. Also, enzyme catalytic efficiency (the kcat/Km ratio) increased as the degree of polymerization of branch chains rose. In the model reaction system of Escherichia coli, glucose-1-phosphate production from glycogen by the glycogen phosphorylase was elevated ∼1.45-fold in the presence of TreX compared to that produced in the absence of TreX. The results suggest that outward shifting of glycogen branch chains via transglycosylation increases the number of exposed chains susceptible to phosphorylase action. We developed a model of the glycogen breakdown process featuring both hydrolysis and transglycosylation catalyzed by the debranching enzyme.
doi:10.1128/JB.01442-13
PMCID: PMC4010986  PMID: 24610710
11.  Impact of Age on Clinicopathological Features and Survival of Patients with Noncardia Gastric Adenocarcinoma 
Journal of Gastric Cancer  2014;14(4):238-245.
Purpose
Gastric cancer often occurs in the elderly but is uncommon in young individuals. Whether young patients have different clinical behaviors and outcomes from those of older patients remain unclear.
Materials and Methods
We identified 1,366 cases of newly diagnosed noncardia gastric adenocarcinoma from the Kaiser Permanente Northern California Cancer Registry between 2000 and 2010. We then compared the clinicopathological features and survival among the different age groups.
Results
The male : female ratio differed significantly between the younger and older patient groups (0.84 in age <50 years vs. 1.52>60 years, P<0.01). More younger patients were Hispanic (54% patients <40 years vs. 19% patients ≥70 years, P<0.0001), while more older patients were Caucasian (49% patients ≥70 years vs. 15% patients <40 years; P<0.0001). The diffuse/mixed histological type was more prevalent in younger patients (70% patients <40 years vs. 27% patients ≥70 years; P<0.0001), whereas the intestinal type was more frequent in older patients (71% in patients ≥70 years vs. 30% in patients <40 years; P<0.0001). Poorly differentiated adenocarcinoma was more common in the younger patients (80% in patients <40 years vs. 60% in patients ≥70 years; P=0.016). Survival rates at 1, 2, and 5 years gradually declined with increasing age (overall P=0.0002).
Conclusions
Young patients with gastric cancer had more aggressive disease but higher overall survival rates than older patients. Younger Hispanic patients and older Caucasian patients were more likely to be diagnosed with gastric cancer. These differences may be due to biological predisposition and/or environmental exposure.
doi:10.5230/jgc.2014.14.4.238
PMCID: PMC4286902  PMID: 25580355
Stomach neoplasms; Outcome assessment (health care); Age groups; Ethnic groups
12.  Homocysteine, methylenetetrahydrofolate reductase C677T polymorphism, and risk of retinal vein occlusion: an updated meta-analysis 
BMC Ophthalmology  2014;14(1):147.
Background
To evaluate the role of plasma total homocysteine (tHcy) and homozygosity for the thermolabile variant of the methylenetetrahydrofolate reductase (MTHFR) C677T genotype in the risk of retinal vein occlusion (RVO).
Methods
Relevant studies were selected through an extensive search of PubMed, EMBASE, and the Web of Science databases. Summary weighted mean differences (WMDs) or odds ratios (ORs) and 95% confidence intervals (CI) were calculated with a random-effects model.
Results
Forty-two studies with 6445 participants were included in this updated systematic review and meta-analysis. The mean plasma tHcy level in the RVO patients was significantly higher than in the controls (WMD =2.13 μmol/L; 95% CI: 1.29 to 2.98, P < 0.001), but there was evidence of between-study heterogeneity (P < 0.001). No significant association between MTHFR C677T genotype and RVO was found under all genetic models.
Conclusion
There was some evidence that plasma tHcy is associated with an increased risk of RVO. There was no evidence to suggest an association between homozygosity for the MTHFR C677T genotype and RVO.
doi:10.1186/1471-2415-14-147
PMCID: PMC4255450  PMID: 25428529
Homocysteine; Methylenetetrahydrofolate reductase; Retinal vein occlusion
13.  Chronic Exposure to Perfluorooctane Sulfonate Induces Behavior Defects and Neurotoxicity through Oxidative Damages, In Vivo and In Vitro 
PLoS ONE  2014;9(11):e113453.
Perfluorooctane sulfonate (PFOS) is an emerging persistent pollutant which shows multiple adverse health effects. However, the neurotoxicity of PFOS and its mechanisms have not been fully elucidated. Using a combination of in vivo and in vitro methods, the present study provides a detailed description of PFOS-induced neurotoxicity. Results showed that the median lethal concentration of PFOS was 2.03 mM in Caenorhabditis elegans for 48 h exposure. 20 µM PFOS caused decrease of locomotor behaviors including forward movement, body bend and head thrash. Additionally, PFOS exposure reduced chemotaxis index of C. elegans, which indicates the decline of chemotaxis learning ability. Using green fluorescent protein (GFP) labelled transgenic strains, we found that PFOS caused down-regulated expression of a chemoreceptor gene, gcy-5, in ASE chemosensory neurons, but did not affect cholinergic neurons and dopaminergic neurons. In SH-SY5Y cells, 48 h exposure to 25 µM and 50 µM PFOS induced cell damage, apoptosis and the reactive oxygen species (ROS) generation. PFOS caused significant increases of lipid peroxidation and superoxide dismutase activity, but an actual decrease of glutathione peroxidase activity. Furthermore, antioxidant N-acetylcysteine rescued cells from PFOS-induced apoptosis via blocking ROS. Our results demonstrate that chronic exposure to PFOS can cause obvious neurotoxicity and behavior defects. Oxidative damage and anti-oxidative deficit are crucial mechanisms in neurotoxicity of PFOS.
doi:10.1371/journal.pone.0113453
PMCID: PMC4239059  PMID: 25412474
14.  Investigating the role of introns in the regulation of regenerating gene 1 expression 
Oncology Letters  2014;9(2):875-880.
Gastrin is a hormone that physiologically regulates gastric acid secretion and contributes to the maintenance of gastric epithelial architecture by regulating the expression of genes such as regenerating gene 1 (Reg1). Reg1 is involved in gastric carcinogenesis as an antiapoptotic factor. The current study explores the molecular mechanism of gastrin-regulated Reg1 expression in human gastric cancer cells. In total, five intron fragments of the Reg1 gene were cloned by polymerase chain reaction and inserted into luciferase reporter vector pGL3 to construct intron-luciferase reporter vectors. After confirmation by Xho I/Hind III digestion and DNA sequencing, the five constructs were transfected into the SGC7901 gastric cancer cell line. The luciferase activity of the cells transfected with each of the five constructs was detected following incubation without or with gastrin. The five intron fragments of Reg1 were also randomly labeled with digoxin as a probe, and nuclear proteins of gastric cancer cells were extracted following treatment with or without gastrin. Southwestern blotting was subsequently performed to detect transcription factors that bind to the introns. The results indicated that the luciferase activity was significantly higher in cells transfected with recombinant vectors containing introns 2, 3, 4 or 5 than that in the cells transfected with an empty vector (P<0.05). However, no statistically significant difference in luciferase activity was identified between cells transfected with pGL3-intron 1 and those transfected with pGL3-Basic (P>0.05). Following incubation with gastrin, no significant difference was identified (P>0.05). The five introns of Reg1 can bind a number of transcription factors and gastrin may affect this interaction. Introns 2–5 of Reg1 potentially have transcriptional control over gene expression in gastric cancer cells. In conclusion, gastrin may regulate the expression of the Reg1 gene via the interaction of the introns by binding to the transcription factors.
doi:10.3892/ol.2014.2712
PMCID: PMC4301469  PMID: 25621062
human gastric cancer; regenerating gene 1; gastrin; intron; transcription factors
15.  Consumption of garlic and risk of colorectal cancer: An updated meta-analysis of prospective studies 
World Journal of Gastroenterology : WJG  2014;20(41):15413-15422.
AIM: To conduct an updated meta-analysis of prospective studies addressing the association between garlic consumption and colorectal cancer.
METHODS: Eligible cohort studies were identified by searching MEDLINE (PubMed) and screening the references of related articles published up to October 2013. Meta-analyses were conducted for colorectal cancer in relation to consumption of raw and cooked (RC) garlic and garlic supplements, separately. The summary relative risks (RR) with 95%CI were calculated using fixed-effects or random-effects model depending on the heterogeneity among studies.
RESULTS: A total of 5 prospective cohort studies were identified. In contrast to the previous meta-analysis, no significant associations were found between consumption of RC garlic (RR: 1.06; 95%CI: 0.95-1.19) or garlic supplements (RR: 1.12; 95%CI: 0.96-1.31) and risk of colorectal cancer. A non-significant protective effect of garlic supplement intake against colorectal cancer was observed in females (RR: 0.84; 95%CI: 0.64-1.11), but the opposite was the case in males (RR: 1.24; 95%CI: 0.96-1.59).
CONCLUSION: Consumption of RC garlic or garlic supplements is not significantly associated with reduced colorectal cancer risk.
doi:10.3748/wjg.v20.i41.15413
PMCID: PMC4223276  PMID: 25386091
Colorectal cancer; Garlic; Garlic supplement; Cancer prevention; Meta-analysis
16.  Iron Metabolism Regulates p53 Signaling through Direct Heme-p53 Interaction and Modulation of p53 Localization, Stability, and Function 
Cell reports  2014;7(1):180-193.
SUMMARY
Iron excess is closely associated with tumorigenesis in multiple types of human cancers, with underlying mechanisms yet unclear. Recently, iron deprivation has emerged as a major strategy for chemotherapy, but it exerts tumor suppression only on select human malignancies. Here, we report that the tumor suppressor protein p53 is downregulated during iron excess. Strikingly, the iron polyporphyrin heme binds to p53 protein, interferes with p53-DNA interactions, and triggers both nuclear export and cytosolic degradation of p53. Moreover, in a tumorigenicity assay, iron deprivation suppressed wild-type p53-dependent tumor growth, suggesting that upregulation of wild-type p53 signaling underlies the selective efficacy of iron deprivation. Our findings thus identify a direct link between iron/heme homeostasis and the regulation of p53 signaling, which not only provides mechanistic insights into iron-excess-associated tumorigenesis but may also help predict and improve outcomes in iron-deprivation-based chemotherapy.
doi:10.1016/j.celrep.2014.02.042
PMCID: PMC4219651  PMID: 24685134
17.  Metabolic Changes in Klebsiella oxytoca in Response to Low Oxidoreduction Potential, as Revealed by Comparative Proteomic Profiling Integrated with Flux Balance Analysis 
Oxidoreduction potential (ORP) is an important physiological parameter for biochemical production in anaerobic or microaerobic processes. However, the effect of ORP on cellular physiology remains largely unknown, which hampers the design of engineering strategies targeting proteins associated with ORP response. Here we characterized the effect of altering ORP in a 1,3-propanediol producer, Klebsiella oxytoca, by comparative proteomic profiling combined with flux balance analysis. Decreasing the extracellular ORP from −150 to −240 mV retarded cell growth and enhanced 1,3-propanediol production. Comparative proteomic analysis identified 61 differentially expressed proteins, mainly involved in carbohydrate catabolism, cellular constituent biosynthesis, and reductive stress response. A hypothetical oxidoreductase (HOR) that catalyzes 1,3-propanediol production was markedly upregulated, while proteins involved in biomass precursor synthesis were downregulated. As revealed by subsequent flux balance analysis, low ORP induced a metabolic shift from glycerol oxidation to reduction and rebalancing of redox and energy metabolism. From the integrated protein expression profiles and flux distributions, we can construct a rational analytic framework that elucidates how (facultative) anaerobes respond to extracellular ORP changes.
doi:10.1128/AEM.03327-13
PMCID: PMC3993281  PMID: 24584239
18.  Ginsenoside-Rg5 induces apoptosis and DNA damage in human cervical cancer cells 
Molecular Medicine Reports  2014;11(2):940-946.
Panax ginseng is traditionally used as a remedy for cancer, inflammation, stress and aging, and ginsenoside-Rg5 is a major bioactive constituent of steamed ginseng. The present study aimed to evaluate whether ginsenoside-Rg5 had any marked cytotoxic, apoptotic or DNA-damaging effects in human cervical cancer cells. Five human cervical cancer cell lines (HeLa, MS751, C33A, Me180 and HT-3) were used to investigate the cytotoxicity of ginsenoside-Rg5 using a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. Additionally, the effects of ginsenoside-Rg5 on the apoptosis of HeLa and MS751 cells were detected using DNA ladder assays and flow cytometry. DNA damage was assessed in the HeLa and MS751 cells using alkaline comet assays and by detection of γH2AX focus formation. The HeLa and MS751 cells were significantly more sensitive to ginsenoside-Rg5 treatment compared with the C-33A, HT-3 and Me180 cells. As expected, ginsenoside-Rg5 induced significant concentration- and time-dependent increases in apoptosis. In addition, ginsenoside-Rg5 induced significant concentration-dependent increases in the level of DNA damage compared with the negative control. Consistent with the comet assay data, the percentage of γH2AX-positive HeLa and MS751 cells also revealed that ginsenoside-Rg5 caused DNA double-strands to break in a concentration-dependent manner. In conclusion, ginsenoside-Rg5 had marked genotoxic effects in the HeLa and MS751 cells and, thus, demonstrates potential as a genotoxic or cytotoxic drug for the treatment of cervical cancer.
doi:10.3892/mmr.2014.2821
PMCID: PMC4262516  PMID: 25355274
cervical cancer; ginsenoside-Rg5; DNA damage; apoptosis
19.  Hierarchical Porous Li2Mg(NH)2@C Nanowires with Long Cycle Life Towards Stable Hydrogen Storage 
Scientific Reports  2014;4:6599.
The hierarchical porous Li2Mg(NH)2@C nanowires full of micropores, mesopores, and macropores are successfully fabricated via a single-nozzle electrospinning technique combined with in-situ reaction between the precursors, i.e., MgCl2 and LiN3, under physical restriction upon thermal annealing. The explosive decomposition of LiN3 well dispersed in the electrospun nanowires during carbothermal treatment induces a highly porous structure, which provides a favourable way for H2 delivering in and out of Li2Mg(NH)2 nanoparticles simultaneously realized by the space-confinement of the porous carbon coating. As a result, the thus-fabricated Li2Mg(NH)2@C nanowires present significantly enhanced thermodynamics and kinetics towards hydrogen storage performance, e.g., a complete cycle of H2 uptake and release with a capacity close to the theoretical value at a temperature as low as 105°C. This is, to the best of our knowledge, the lowest cycling temperature reported to date. More interestingly, induced by the nanosize effects and space-confinement function of porous carbon coating, a excellently stable regeneration without apparent degradation after 20 de-/re-hydrogenation cycles at a temperature as low as 130°C was achieved for the as-prepared Li2Mg(NH)2@C nanowires.
doi:10.1038/srep06599
PMCID: PMC4194431  PMID: 25307874
20.  Human white adipose tissue vasculature contains endothelial colony-forming cells with robust in vivo vasculogenic potential 
Angiogenesis  2013;16(4):735-744.
Blood-derived endothelial colony-forming cells (ECFCs) have robust vasculogenic potential that can be exploited to bioengineer long-lasting human vascular networks in vivo. However, circulating ECFCs are exceedingly rare in adult peripheral blood. Because the mechanism by which ECFCs are mobilized into circulation is currently unknown, the reliability of peripheral blood as a clinical source of ECFCs remains a concern. Thus, there is a need to find alternative sources of autologous ECFCs. Here we aimed to determine whether ECFCs reside in the vasculature of human white adipose tissue (WAT) and to evaluate if WAT-derived ECFCs (watECFCs) have equal clinical potential to blood-derived ECFCs. We isolated the complete endothelial cell (EC) population from intact biopsies of normal human subcutaneous WAT by enzymatic digestion and selection of CD31+ cells. Subsequently, we extensively compared WAT-derived EC phenotype and functionality to bonafide ECFCs derived from both umbilical cord blood and adult peripheral blood. We demonstrated that human WAT is indeed a dependable source of ECFCs with indistinguishable properties to adult peripheral blood ECFCs, including hierarchical clonogenic ability, large expansion potential, stable endothelial phenotype, and robust in vivo blood vessel-forming capacity. Considering the unreliability and low rate of occurrence of ECFCs in adult blood and that biopsies of WAT can be obtained with minimal intervention in an ambulatory setting, our results indicate WAT as a more practical alternative to obtain large amounts of readily available autologous ECFCs for future vascular cell therapies.
doi:10.1007/s10456-013-9350-0
PMCID: PMC3762916  PMID: 23636611
endothelial colony-forming cells; endothelial progenitor cells; adipose tissue; peripheral blood; vasculogenesis
21.  Expression and clinical significance of the long non-coding RNA PVT1 in human gastric cancer 
OncoTargets and therapy  2014;7:1625-1630.
Background
Highly sensitive markers are urgently needed for the diagnosis and grading of gastric cancer and for managing drug resistance. The recent identification of long-non-coding RNAs (lncRNAs) has provided new approaches for resolving this challenge. The aim of this study was to screen and identify new biomarkers for human gastric cancer from lncRNAs.
Methods
First, we used lncRNA microarrays to conduct a preliminary screening for candidate lncRNAs of gastric cancer biomarkers in both human gastric cancer tissues and in two gastric cancer cell lines, SGC7901 cells and paclitaxel-resistant SGC7901 cells. The lncRNA plasma-cytoma variant translocation 1 (PVT1) was found to exhibit higher expression in both gastric cancer tissues and the SGC7901 paclitaxel-resistant cell line. Quantitative polymerase chain reaction was used for large-scale analysis in a large number of human gastric cancer tissues to verify the involvement of PVT1 in development of gastric cancer. The relationships between PVT1 expression and clinical features were also analyzed.
Results
PVT1 showed higher expression in human gastric cancer tissues than in adjacent non-cancerous tissues and in SGC7901 paclitaxel-resistant cells compared with SGC7901 cells. PVT1 expression was correlated with lymph node invasion of gastric cancer.
Conclusion
PVT1 is a new biomarker for human gastric cancer and may indicate lymph node invasion. Therefore, PVT1 shows potential as a novel therapeutic target for the treatment of gastric cancer and enhancement of paclitaxel sensitivity.
doi:10.2147/OTT.S68854
PMCID: PMC4172193  PMID: 25258543
microarray analysis; quantitative polymerase chain reaction; lymph node invasion; tumor biomarkers; paclitaxel resistance
22.  Crystal structure of (E)-2-(2-{5-[(2-acet­oxy­eth­yl)(meth­yl)amino]­thio­phen-2-yl}vin­yl)-3-methyl­benzo­thia­zolium iodide monohydrate 
In the cation of the title hydrated salt, C19H21N2O2S2 +·I−·H2O, the benzo­thia­zolium ring system is approximately planar [maximum deviation = 0.0251 (15) Å], and it makes a small dihedral angle of 1.16 (18)° with the plane of the thio­phene ring. In the crystal, the cations, anions and crystalline water mol­ecules are linked by classical O—H⋯O, O—H⋯I and weak C—H⋯O hydrogn bonds, forming a three-dimensional supra­molecular network. π–π stacking is observed between parallel thia­zole rings of adjacent cations [centroid–centroid distance = 3.5945 (16) Å].
doi:10.1107/S1600536814020121
PMCID: PMC4257210  PMID: 25484696
crystal structure; benzo­thia­zolium iodide salt; hydrogen bonding
23.  Age and Sex-Specific Relationships between Phthalate Exposures and Obesity in Chinese Children at Puberty 
PLoS ONE  2014;9(8):e104852.
Objective
To examine the age and sex-specific associations of urine levels of six mono-phthalates with body size and fat distribution in Chinese children at puberty.
Materials and Methods
Four hundred and ninety-three school-aged children (247 boys, 246 girls) were recruited. Obesity related anthropometric indices were measured and body fat proportion (BF%) was calculated. Spot urine samples were collected and phthalate monoesters were detected by an API 2000 electrospray triple quadrupole mass spectrometer (ESI-MS/MS). Associations between phthalate exposure and overweight/obesity measures and their trends were examined by multiple linear regression and Logistic regression analyses, respectively.
Results
Di-2-ethylhexyl phthalate (DEHP) metabolites and monobutyl phthalate (MBP) were found to be the most detectable chemicals. In 8–10 years (yrs) group, concentrations of MEHP and MBP were significantly higher in girls than those in boys. However, concentrations of all phthalate monoesters, except for MEP and MEHP, in 11–13 yrs boys were significantly higher than those in girls. After adjusting for confounders including puberty onset, urinary concentrations of MBP and sum of low molecular-weight phthalate metabolites (∑LMP) were positively associated with boys' obesity in a concentration-effect manner, while concentrations of MEHP, MEHHP and sum of DEHP metabolites (∑MEHP) were negatively associated with girls' obesity. Associations between phthalate exposure levels and BMI z-score changes were age- and sex-specific in school-age children.
Conclusion
There are age and sex-specific concentration-effect associations between phthalate exposure and fat distribution in Chinese children. Urinary phthalate levels in 11–13 yrs boys were about 30 percent higher than those in girls, and ∑MEHP levels in younger boys (<10 yrs) were significantly higher than those in elder boys (>10 yrs). Associations were positive for MBP and ∑LMP with both BMI z-score and fat distribution in boys >10 years of age, and negative for ∑MEHP with fat distribution in girls <10 years of age.
doi:10.1371/journal.pone.0104852
PMCID: PMC4133266  PMID: 25121758
24.  Theranostic nanoparticles based on bioreducible polyethylenimine-coated iron oxide for reduction-responsive gene delivery and magnetic resonance imaging 
Theranostic nanoparticles based on superparamagnetic iron oxide (SPIO) have a great promise for tumor diagnosis and gene therapy. However, the availability of theranostic nanoparticles with efficient gene transfection and minimal toxicity remains a big challenge. In this study, we construct an intelligent SPIO-based nanoparticle comprising a SPIO inner core and a disulfide-containing polyethylenimine (SSPEI) outer layer, which is referred to as a SSPEI-SPIO nanoparticle, for redox-triggered gene release in response to an intracellular reducing environment. We reveal that SSPEI-SPIO nanoparticles are capable of binding genes to form nano-complexes and mediating a facilitated gene release in the presence of dithiothreitol (5–20 mM), thereby leading to high transfection efficiency against different cancer cells. The SSPEI-SPIO nanoparticles are also able to deliver small interfering RNA (siRNA) for the silencing of human telomerase reverse transcriptase genes in HepG2 cells, causing their apoptosis and growth inhibition. Further, the nanoparticles are applicable as T2-negative contrast agents for magnetic resonance (MR) imaging of a tumor xenografted in a nude mouse. Importantly, SSPEI-SPIO nanoparticles have relatively low cytotoxicity in vitro at a high concentration of 100 μg/mL. The results of this study demonstrate the utility of a disulfide-containing cationic polymer-decorated SPIO nanoparticle as highly potent and low-toxic theranostic nano-system for specific nucleic acid delivery inside cancer cells.
doi:10.2147/IJN.S61463
PMCID: PMC4099417  PMID: 25045265
nanoparticles; SSPEI; hTERT; disulfide; RNA interference; tumor; MR imaging
25.  Knockdown of OCT4 suppresses the growth and invasion of pancreatic cancer cells through inhibition of the AKT pathway 
Molecular Medicine Reports  2014;10(3):1335-1342.
Octamer-binding transcription factor 4 (OCT4) is one of the factors associated with self-renewal and differentiation in cancer stem cells, and is crucial for the progression of various types of human malignancy. However, the expression and function of OCT4 in human pancreatic cancer has not been fully elucidated. The purpose of the present study was to investigate the function and molecular mechanisms of OCT4 in pancreatic cancer cells. The clinical significance of OCT4 expression was assessed by an immunohistochemical assay using a tissue microarray procedure in pancreatic cancer tissues and cells with different degrees of differentiation. A loss-of-function approach was used to examine the effects of a lentivirus-mediated OCT4 small hairpin RNA vector on biological behaviors, including cell proliferative activity and invasive potential. The results demonstrated that the expression levels of OCT4 protein in cancer tissues were significantly elevated compared with those in adjacent non-cancerous tissues (65.0 vs. 42.5%; P=0.005), which was correlated with tumor differentiation (P=0.008). The knockdown of OCT4 inhibited the proliferation and invasion of pancreatic cancer cells (Panc-1) expressing high levels of OCT4, accompanied with decreased expression of AKT, proliferating cell nuclear antigen (PCNA) and matrix metalloproteinase-2 (MMP-2). In conclusion, the present study reveals that the increased expression of OCT4 is correlated with the differentiation of pancreatic cancer, while knockdown of OCT4 suppresses the growth and invasion of pancreatic cancer cells through inhibition of AKT pathway-mediated PCNA and MMP-2 expression, suggesting that OCT4 might serve as a potential therapeutic target for the treatment of pancreatic cancer.
doi:10.3892/mmr.2014.2367
PMCID: PMC4121418  PMID: 25017645
octamer-binding transcription factor 4; pancreatic cancer; AKT; growth; invasion

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