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1.  Efficacy of the Modified Endoscopic Frontal Sinus Surgery for Recurrent Chronic Frontal Sinusitis 
To modify the endoscopic frontal sinus surgery and improve the therapeutic effect of recurrent chronic frontal sinusitis (RCFS). Eighty-five patients with RCFS were divided into two groups. Endoscopic frontal sinus surgery through an approach of Frontomaxillary Process-Agger Nasi, a modified Draf IIb procedure, was carried out in 51 patients (Group A), and conservative medication was applied in 34 patients as control (Group B). The therapeutic effect was prospectively evaluated with statistically validated measures of sinusitis-specific quality of life, sino-nasal outcome test-20 questionnaire (SNOT-20). Compared with pre-treatment, the average total score of SNOT-20 in RCFS patients was significantly decreased at the time of 6, 12 months after modified endoscopic frontal sinus surgery and medical treatments (p < 0.05). However, the total score of SNOT20 was significantly lower in group A than group B at the same period of the follow-up after treatments (p < 0.05). The overall efficacy evaluated by patients’ self showed that the rate of “much improved” and “improved” was respectively 68.6 and 17.6 % in group A, and significantly better than group B (p < 0.001). Furthermore, the frontal sinus patency rate in group A was 85 %, and significantly higher than group B (p < 0.001). Endoscopic frontal sinus surgery through an approach of Frontomaxillary Process-Agger Nasi, a modified Draf IIb procedure, is an effective procedure to treat the RCFS.
PMCID: PMC4071437  PMID: 25032109
Frontal sinusitis;  Endoscopy;  Sinus surgery
2.  Truncating mutation in the autophagy gene UVRAG confers oncogenic properties and chemosensitivity in colorectal cancers 
Nature Communications  2015;6:7839.
Autophagy-related factors are implicated in metabolic adaptation and cancer metastasis. However, the role of autophagy factors in cancer progression and their effect in treatment response remain largely elusive. Recent studies have shown that UVRAG, a key autophagic tumour suppressor, is mutated in common human cancers. Here we demonstrate that the cancer-related UVRAG frameshift (FS), which does not result in a null mutation, is expressed as a truncated UVRAGFS in colorectal cancer (CRC) with microsatellite instability (MSI), and promotes tumorigenesis. UVRAGFS abrogates the normal functions of UVRAG, including autophagy, in a dominant-negative manner. Furthermore, expression of UVRAGFS can trigger CRC metastatic spread through Rac1 activation and epithelial-to-mesenchymal transition, independently of autophagy. Interestingly, UVRAGFS expression renders cells more sensitive to standard chemotherapy regimen due to a DNA repair defect. These results identify UVRAG as a new MSI target gene and provide a mechanism for UVRAG participation in CRC pathogenesis and treatment response.
Some colon carcinomas with microsatellite instability carry a frameshift mutation in a tumour suppressor UVRAG. Here the authors show that mutant UVRAG triggers colorectal cancer by antagonizing the activity of normal UVRAG in autophagy and chromosomal stability, but also sensitizes the cancer to DNA damage-inducing chemotherapeutic drugs.
PMCID: PMC4526116  PMID: 26234763
3.  Expression of apoptosis signal-regulating kinase 1 is associated with tight junction protein claudin-6 in cervical carcinoma 
In our previous work, apoptosis signal-regulating kinase 1 (ASK1), a member of the mitogen-activated protein kinase (MAPK) kinase family has been proved to be associated with the pro-apoptosis effect of tight junction protein claudin-6 in breast cancer. However, its expression in cervical carcinoma has not been reported. The ASK1 and claudin-6 expression in cervical carcinoma tissues and adjacent non-neoplastic tissues was examined by immunohistochemistry. The mRNA and protein expression of ASK1 and claudin-6 in cervical cancer carcinoma cells was detected by reverse transcription (RT)-PCR and western blot. The expression level of ASK1 was down-regulated in cervical carcinoma tissues compared with the adjacent non-neoplastic tissues and positively correlated with the level of claudin-6 in cervical carcinoma cells and tissues. Our present study reveals that ASK1 protein expression altered between human cervical carcinoma and adjacent non-neoplastic tissues. The expression of ASK1 is correlated with the level of claudin-6 in cervical carcinoma cells and tissues.
PMCID: PMC4503132  PMID: 26191261
Apoptosis; apoptosis signal-regulating kinase 1; cervical carcinoma; claudin-6
4.  Diastereoselective Three-Component Synthesis of β-Amino Carbonyl Compounds Using Diazo Compounds, Boranes, and Acyl Imines under Catalyst-Free Conditions 
The Journal of Organic Chemistry  2014;79(10):4694-4698.
Diazo compounds, boranes, and acyl imines undergo a three-component Mannich condensation reaction under catalyst-free conditions to give the anti β-amino carbonyl compounds in high diastereoselectivity. The reaction tolerates a variety of functional groups, and an asymmetric variant was achieved using the (−)-phenylmenthol as chiral auxiliary in good yield and selectivity. These β-amino carbonyl compounds are valuable intermediates, which can be transformed to many potential bioactive molecules.
PMCID: PMC4033649  PMID: 24787904
5.  Angiotensin-converting enzyme insertion/deletion polymorphism and gastric cancer: a systematic review and meta-analysis 
Previous case-control studies on the association of the angiotensin-converting enzyme (ACE) gene insertion/deletion (I/D) polymorphism with gastric cancer were controversial. A meta-analysis was conducted to further evaluate the association between polymorphism in the ACE gene I/D and gastric cancer. We searched MEDLINE (PubMed), EMBASE, Web of Science, and CBM without language restrictions to Nov 20, 2014. Odds ratios (ORs) with 95% confidence intervals (CIs) were used to assess the strength of association. Eight studies involving 1480 gastric cancer cases and 3773 cancer-free controls were included. Overall, no significant association between ACE I/D polymorphism and gastric cancer risk was observed (OR = 1.15; 95% CI 0.90-1.46, P = 0.26). The subgroup analysis on the basis of H. Pylori status showed the decreased gastric cancer risk in H. Pylori negative subgroup (OR = 0.40; 95% CI: 0.27-0.59; P < 0.00001) rather than in H. Pylori positive subgroup (OR = 1.82, 95% CI: 0.87-3.82, P = 0.11). Subgroup analysis was performed according to ethnicity (Caucasian and Asian). The results showed no genetic effects between ACE I/D polymorphism and gastric cancer risk. This meta-analysis suggested that the ACE gene I/D polymorphism was associated gastric cancer risk in H. Pylori negative subjects.
PMCID: PMC4483819  PMID: 26131166
Gastric cancer; ACE; gene polymorphism; meta-analysis
6.  A Comprehensive Alanine-Scanning Mutagenesis Study Reveals Roles for Salt Bridges in the Structure and Activity of Pseudomonas aeruginosa Elastase 
PLoS ONE  2015;10(3):e0121108.
The relationship between salt bridges and stability/enzymatic activity is unclear. We studied this relationship by systematic alanine-scanning mutation analysis using the typical M4 family metalloprotease Pseudomonas aeruginosa elastase (PAE, also known as pseudolysin) as a model. Structural analysis revealed seven salt bridges in the PAE structure. We constructed ten mutants for six salt bridges. Among these mutants, six (Asp189Ala, Arg179Ala, Asp201Ala, Arg205Ala, Arg245Ala and Glu249Ala) were active and four (Asp168Ala, Arg198Ala, Arg253Ala, and Arg279Ala) were inactive. Five mutants were purified, and their catalytic efficiencies (kcat/Km), half-lives (t1/2) and thermal unfolding curves were compared with those of PAE. Mutants Asp189Ala and Arg179Ala both showed decreased thermal stabilities and increased activities, suggesting that the salt bridge Asp189-Arg179 stabilizes the protein at the expense of catalytic efficiency. In contrast, mutants Asp201Ala and Arg205Ala both showed slightly increased thermal stability and slightly decreased activity, suggesting that the salt bridge Asp201-Arg205 destabilizes the protein. Mutant Glu249Ala is related to a C-terminal salt bridge network and showed both decreased thermal stability and decreased activity. Furthermore, Glu249Ala showed a thermal unfolding curve with three discernable states [the native state (N), the partially unfolded state (I) and the unfolded state (U)]. In comparison, there were only two discernable states (N and U) in the thermal unfolding curve of PAE. These results suggest that Glu249 is important for catalytic efficiency, stability and unfolding cooperativity. This study represents a systematic mutational analyses of salt bridges in the model metalloprotease PAE and provides important insights into the structure-function relationship of enzymes.
PMCID: PMC4376888  PMID: 25815820
7.  Transdifferentiation of Human Hair Follicle Mesenchymal Stem Cells into Red Blood Cells by OCT4 
Stem Cells International  2015;2015:389628.
Shortage of red blood cells (RBCs, erythrocytes) can have potentially life-threatening consequences for rare or unusual blood type patients with massive blood loss resulting from various conditions. Erythrocytes have been derived from human pluripotent stem cells (PSCs), but the risk of potential tumorigenicity cannot be ignored, and a majority of these cells produced from PSCs express embryonic ε- and fetal γ-globins with little or no adult β-globin and remain nucleated. Here we report a method to generate erythrocytes from human hair follicle mesenchymal stem cells (hHFMSCs) by enforcing OCT4 gene expression and cytokine stimulation. Cells generated from hHFMSCs expressed mainly the adult β-globin chain with minimum level of the fetal γ-globin chain. Furthermore, these cells also underwent multiple maturation events and formed enucleated erythrocytes with a biconcave disc shape. Gene expression analyses showed that OCT4 regulated the expression of genes associated with both pluripotency and erythroid development during hHFMSC transdifferentiation toward erythroid cells. These findings show that mature erythrocytes can be generated from adult somatic cells, which may serve as an alternative source of RBCs for potential autologous transfusion.
PMCID: PMC4337757  PMID: 25755671
9.  Enhanced photovoltaic property by forming p-i-n structures containing Si quantum dots/SiC multilayers 
Nanoscale Research Letters  2014;9(1):634.
Si quantum dots (Si QDs)/SiC multilayers were fabricated by annealing hydrogenated amorphous Si/SiC multilayers prepared in a plasma-enhanced chemical vapor deposition system. The thickness of amorphous Si layer was designed to be 4 nm, and the thickness of amorphous SiC layer was kept at 2 nm. Transmission electron microscopy observation revealed the formation of Si QDs after 900°C annealing. The optical properties of the Si QDs/SiC multilayers were studied, and the optical band gap deduced from the optical absorption coefficient result is 1.48 eV. Moreover, the p-i-n structure with n-a-Si/i-(Si QDs/SiC multilayers)/p-Si was fabricated, and the carrier transportation mechanism was investigated. The p-i-n structure was used in a solar cell device. The cell had the open circuit voltage of 532 mV and the power conversion efficiency (PCE) of 6.28%.
81.07.Ta; 78.67.Pt; 88.40.jj
PMCID: PMC4256967  PMID: 25489285
Si quantum dots (Si QDs); Silicon carbide; Multilayers; Solar cell
10.  Fruit and vegetable intake and risk of type 2 diabetes mellitus: meta-analysis of prospective cohort studies 
BMJ Open  2014;4(11):e005497.
To clarify and quantify the potential dose–response association between the intake of fruit and vegetables and risk of type 2 diabetes.
Meta-analysis and systematic review of prospective cohort studies.
Data source
Studies published before February 2014 identified through electronic searches using PubMed and Embase.
Eligibility criteria for selecting studies
Prospective cohort studies with relative risks and 95% CIs for type 2 diabetes according to the intake of fruit, vegetables, or fruit and vegetables.
A total of 10 articles including 13 comparisons with 24 013 cases of type 2 diabetes and 434 342 participants were included in the meta-analysis. Evidence of curve linear associations was seen between fruit and green leafy vegetables consumption and risk of type 2 diabetes (p=0.059 and p=0.036 for non-linearity, respectively). The summary relative risk of type 2 diabetes for an increase of 1 serving fruit consumed/day was 0.93 (95% CI 0.88 to 0.99) without heterogeneity among studies (p=0.477, I2=0%). For vegetables, the combined relative risk of type 2 diabetes for an increase of 1 serving consumed/day was 0.90 (95% CI 0.80 to 1.01) with moderate heterogeneity among studies (p=0.002, I2=66.5%). For green leafy vegetables, the summary relative risk of type 2 diabetes for an increase of 0.2 serving consumed/day was 0.87 (95% CI 0.81 to 0.93) without heterogeneity among studies (p=0.496, I2=0%). The combined estimates showed no significant benefits of increasing the consumption of fruit and vegetables combined.
Higher fruit or green leafy vegetables intake is associated with a significantly reduced risk of type 2 diabetes.
PMCID: PMC4225228  PMID: 25377009
11.  MUC1 gene polymorphism rs4072037 and susceptibility to gastric cancer: a meta-analysis 
SpringerPlus  2014;3:599.
The association between MUC1 polymorphism rs4072037 and the risk of gastric cancer has been described in several studies. However, these studies yielded inconsistent results, especially in different pathological type of gastric cancer. Therefore, we performed this meta-analysis to evaluate the relationship between MUC1 gene polymorphism and gastric cancer susceptibility. A comprehensive database search was performed to identify eligible studies. Odds ratios with 95% confidence intervals were calculated to assess the strength of the association between MUC1 rs4072037 and risk of gastric cancer. Subgroup analyses, publication bias, and sensitivity analyses were also conducted. PubMed, EMBASE, Web of Science and CNKI databases were systematically searched to identify relevant studies. A total of 9 studies (12 datasets) were included in the meta-analysis including 10,410 cases and 11,437 controls. Overall, the G allele at rs4072037 of MUC1 gene was associated with a significant decreased gastric cancer risk (OR=0.70, 95% CI: 0.64–0.76). The association was significant in both anatomic location and pathological subtype subgroup analyses. However, the association was detected in Asian rather than Caucasian. Our findings demonstrate that the presence of the G allele at rs4072037 of the MUC1 gene may contribute to protection against gastric cancer in Asian. Further large studies of multiethnic groups are needed to validate these findings.
Electronic supplementary material
The online version of this article (doi:10.1186/2193-1801-3-599) contains supplementary material, which is available to authorized users.
PMCID: PMC4198476  PMID: 25332893
MUC1; Polymorphism; Genetic; Stomach neoplasms; Meta-analysis
12.  Prognostic Value of FGFR Gene Amplification in Patients with Different Types of Cancer: A Systematic Review and Meta-Analysis 
PLoS ONE  2014;9(8):e105524.
Fibroblast growth factor receptor (FGFR) gene amplification has been reported in different types of cancer. We performed an up-to-date meta-analysis to further characterize the prognostic value of FGFR gene amplification in patients with cancer.
A search of several databases, including MEDLINE (PubMed), EMBASE, Web of Science, and China National Knowledge Infrastructure, was conducted to identify studies examining the association between FGFR gene amplification and cancer. A total of 24 studies met the inclusion criteria, and overall incidence rates, hazard risk (HR), overall survival, disease-free survival, and 95% confidence intervals (CIs) were calculated employing fixed- or random-effects models depending on the heterogeneity of the included studies.
In the meta-analysis of 24 studies, the prevalence of FGFR gene amplification was FGFR1: 0.11 (95% CI: 0.08–0.13) and FGFR2: 0.04 (95% CI: 0.02–0.06). Overall survival was significantly worse among patients with FGFR gene amplification: FGFR1 [HR 1.57 (95% CI: 1.23–1.99); p = 0.0002] and FGFR2 [HR 2.27 (95% CI: 1.73–3.00); p<0.00001].
Current evidence supports the conclusion that the outcomes of patients with FGFR gene amplified cancers is worse than for those with non-FGFR gene amplified cancers.
PMCID: PMC4149366  PMID: 25171497
13.  In vivo imaging of protein–protein and RNA–protein interactions using novel far-red fluorescence complementation systems 
Nucleic Acids Research  2014;42(13):e103.
Imaging of protein–protein and RNA–protein interactions in vivo, especially in live animals, is still challenging. Here we developed far-red mNeptune-based bimolecular fluorescence complementation (BiFC) and trimolecular fluorescence complementation (TriFC) systems with excitation and emission above 600 nm in the ‘tissue optical window’ for imaging of protein–protein and RNA–protein interactions in live cells and mice. The far-red mNeptune BiFC was first built by selecting appropriate split mNeptune fragments, and then the mNeptune-TriFC system was built based on the mNeptune-BiFC system. The newly constructed mNeptune BiFC and TriFC systems were verified as useful tools for imaging protein–protein and mRNA–protein interactions, respectively, in live cells and mice. We then used the new mNeptune-TriFC system to investigate the interactions between human polypyrimidine-tract-binding protein (PTB) and HIV-1 mRNA elements as PTB may participate in HIV mRNA processing in HIV activation from latency. An interaction between PTB and the 3′long terminal repeat region of HIV-1 mRNAs was found and imaged in live cells and mice, implying a role for PTB in regulating HIV-1 mRNA processing. The study provides new tools for in vivo imaging of RNA–protein and protein–protein interactions, and adds new insight into the mechanism of HIV-1 mRNA processing.
PMCID: PMC4117741  PMID: 24813442
14.  Human Astrocytic Cells Support Persistent Coxsackievirus B3 Infection 
Journal of Virology  2013;87(22):12407-12421.
Enteroviruses can frequently target the human central nervous system to induce a variety of neurological diseases. Although enteroviruses are highly cytolytic, emerging evidence has shown that these viruses can establish persistent infections both in vivo and in vitro. Here, we investigated the susceptibility of three human brain cell lines, CCF-STTG1, T98G, and SK-N-SH, to infection with three enterovirus serotypes: coxsackievirus B3 (CVB3), enterovirus 71, and coxsackievirus A9. Persistent infection was observed in CVB3-infected CCF-STTG1 cells, as evidenced by prolonged detection of infectious virions, viral RNA, and viral antigens. Of note, infected CCF-STTG1 cells expressed the nonfunctional canonical viral receptors coxsackievirus-adenovirus receptor and decay-accelerating factor, while removal of cell surface chondroitin sulfate from CCF-STTG1 cells inhibited the replication of CVB3, suggesting that receptor usage was one of the major limiting factors in CVB3 persistence. In addition, CVB3 curtailed the induction of beta interferon in infected CCF-STTG1 cells, which likely contributed to the initiation of persistence. Furthermore, proinflammatory chemokines and cytokines, such as vascular cell adhesion molecule 1, interleukin-8 (IL-8), and IL-6, were upregulated in CVB3-infected CCF-STTG1 cells and human progenitor-derived astrocytes. Our data together demonstrate the potential of CCF-STTG1 cells to be a novel cell model for studying CVB3-central nervous system interactions, providing the basis toward a better understanding of CVB3-induced chronic neuropathogenesis.
PMCID: PMC3807905  PMID: 24027313
15.  let-7-repressesed Shc translation delays replicative senescence 
Aging Cell  2013;13(1):185-192.
The p66Shc adaptor protein is an important regulator of lifespan in mammals, but the mechanisms responsible are still unclear. Here, we show that expression of p66Shc, p52Shc, and p46Shc is regulated at the post-transcriptional level by the microRNA let-7a. The levels of let-7a correlated inversely with the levels of Shc proteins without affecting Shc mRNA levels. We identified ‘seedless’ let-7a interaction elements in the coding region of Shc mRNA; mutation of the ‘seedless’ interaction sites abolished the regulation of Shc by let-7a. Our results further revealed that repression of Shc expression by let-7a delays senescence of human diploid fibroblasts (HDFs). In sum, our findings link let-7a abundance to the expression of p66Shc, which in turn controls the replicative lifespan of HDFs.
PMCID: PMC3947057  PMID: 24165399
cellular lifespan; let-7a; p66Shc; replicative senescence; translational regulation
16.  Tunable nonlinear optical properties in nanocrystalline Si/SiO2 multilayers under femtosecond excitation 
The nonlinear optical properties of nanocrystalline-Si/SiO2 (nc-Si/SiO2) multilayers have been investigated through Z-scan technique by using a Ti-sapphire laser with 50-fs pulse duration at 800 nm as a pump laser. It is interesting to note that with increasing the annealing temperature to make the sample change from amorphous phase to nanocrystalline state, the nonlinear absorption turns the reverse saturation absorption into saturation absorption while the nonlinear optical refraction is also changed simultaneously from self-defocusing to self-focusing. We propose that the localized states at the nc-Si/SiO2 interfaces play the key role in the observed switching behaviors. Our results demonstrate that the tunable optical nonlinearities can be achieved by controlling the microstructures of nc-Si, which can be used as engineering different nonlinear optical devices.
PMCID: PMC3896739  PMID: 24423198
Optical nonlinearities; Nanocrystalline Si; Multilayers; Interface state; 42.65.-k; 42.65.Jx; 42.65.Pc
17.  The Combination of RAD001 and MK-2206 Exerts Synergistic Cytotoxic Effects against PTEN Mutant Gastric Cancer Cells: Involvement of MAPK-Dependent Autophagic, but Not Apoptotic Cell Death Pathway 
PLoS ONE  2014;9(1):e85116.
In the current study, we showed that the combination of mammalian target of rapamycin (mTOR) inhibitor RAD001 (everolimus) and Akt inhibitor MK-2206 exerted synergistic cytotoxic effects against low-phosphatase and tensin homolog (PTEN) gastric cancer cells (HGC-27 and SNU-601 lines). In HGC-27 cells, RAD001 and MK-2206 synergistically induced G1/S cell cycle arrest, growth inhibition, cell death but not apoptosis. RAD001 and MK-2206 synergistically induced light chain 3B (LC3B) and beclin-1 expression, two important autophagy indicators. Meanwhile, the autophagy inhibitor 3-methyladenine (3-MA) and chloroquine inhibited the cytotoxic effects by RAD001 and MK-2206, suggesting that autophagic, but not apoptotic cell death was important for the cytotoxic effects by the co-administration. We observed that the combination of RAD001 and MK-2206 exerted enhanced effects on Akt/mTOR inhibition, cyclin D1 down-regulation and ERK/MAPK(extracellular signal-regulated kinase/mitogen-activated protein kinases) activation. Intriguingly, MEK/ERK inhibitors PD98059 and U0126 suppressed RAD001 plus MK-2206-induced beclin-1 expression, autophagy induction and cytotoxicity in HGC-27 cells. In conclusion, these results suggested that the synergistic anti-gastric cancer cells ability by RAD001 and MK-2206 involves ERK-dependent autophagic cell death pathway.
PMCID: PMC3887024  PMID: 24416349
18.  Clinicopathologic significance of claudin-6, occludin, and matrix metalloproteinases −2 expression in ovarian carcinoma 
Diagnostic Pathology  2013;8:190.
Tight junctions (TJs) are mainly composed of claudins, occludin, and tight junction adhesion molecules (JAM). The invasive and metastatic phenotype of highly invasive cancer cells has been related to abnormal structure and function of TJs, and with expression of activated matrix metalloproteinases (MMPs). The relevance of these mechanisms responsible for the invasion and metastasis of ovarian carcinoma is unclear. Similarly, it is not known if the expression of claudin-6, occludin and MMP2 is related with the clinical properties of these tumors.
Expression of claudin-6, occludin, and MMP2 was detected in samples of human ovarian cancer tissues by immunohistochemistry and correlated with the clinical properties of the tumors.
The positive expression rates of claudin-6 and MMP-2 were higher in ovarian papillary serous carcinomas than n ovarian serous adenomas (P < 0.05). There were no differences in the expression of occludin (P > 0.05). The expression of claudin-6 and occludin in ovarian cancer was not correlated with patient age, pathological grade, clinical stage, and metastasis (P > 0.05). MMP-2 expression was enhanced with increased clinical stage and metastasis (P < 0.05), but was unrelated to patient age or tumor grade (P > 0.05). There were no apparent correlations between expression of claudin-6, occludin and MMP-2 in ovarian cancer tissue (P > 0.05).
Our data suggest, for the first time, that the claudin-6 and MMP-2 are up-regulated in ovarian papillary serous carcinomas, MMP-2 expression was enhanced with increased clinical stage and metastasis. Claudin-6 and MMP-2 may play a positive role in the invasion and metastasis of ovarian cancer.
Virtual slides
The virtual slide(s) for this article can be found here:
PMCID: PMC3866569  PMID: 24245968
Tight junctions; Ovarian cancer; Claudin-6; Occludin; MMP-2
19.  Differential reconstructed gene interaction networks for deriving toxicity threshold in chemical risk assessment 
BMC Bioinformatics  2013;14(Suppl 14):S3.
Pathway alterations reflected as changes in gene expression regulation and gene interaction can result from cellular exposure to toxicants. Such information is often used to elucidate toxicological modes of action. From a risk assessment perspective, alterations in biological pathways are a rich resource for setting toxicant thresholds, which may be more sensitive and mechanism-informed than traditional toxicity endpoints. Here we developed a novel differential networks (DNs) approach to connect pathway perturbation with toxicity threshold setting.
Our DNs approach consists of 6 steps: time-series gene expression data collection, identification of altered genes, gene interaction network reconstruction, differential edge inference, mapping of genes with differential edges to pathways, and establishment of causal relationships between chemical concentration and perturbed pathways. A one-sample Gaussian process model and a linear regression model were used to identify genes that exhibited significant profile changes across an entire time course and between treatments, respectively. Interaction networks of differentially expressed (DE) genes were reconstructed for different treatments using a state space model and then compared to infer differential edges/interactions. DE genes possessing differential edges were mapped to biological pathways in databases such as KEGG pathways.
Using the DNs approach, we analyzed a time-series Escherichia coli live cell gene expression dataset consisting of 4 treatments (control, 10, 100, 1000 mg/L naphthenic acids, NAs) and 18 time points. Through comparison of reconstructed networks and construction of differential networks, 80 genes were identified as DE genes with a significant number of differential edges, and 22 KEGG pathways were altered in a concentration-dependent manner. Some of these pathways were perturbed to a degree as high as 70% even at the lowest exposure concentration, implying a high sensitivity of our DNs approach.
Findings from this proof-of-concept study suggest that our approach has a great potential in providing a novel and sensitive tool for threshold setting in chemical risk assessment. In future work, we plan to analyze more time-series datasets with a full spectrum of concentrations and sufficient replications per treatment. The pathway alteration-derived thresholds will also be compared with those derived from apical endpoints such as cell growth rate.
PMCID: PMC3851258  PMID: 24268022
20.  HBP1-Mediated Transcriptional Regulation of DNA Methyltransferase 1 and Its Impact on Cell Senescence 
Molecular and Cellular Biology  2013;33(5):887-903.
The activity of DNA methyltransferase 1 (DNMT1) is associated with diverse biological activities, including cell proliferation, senescence, and cancer development. In this study, we demonstrated that the HMG box-containing protein 1 (HBP1) transcription factor is a new repressor of DNMT1 in a complex mechanism during senescence. The DNMT1 gene contains an HBP1-binding site at bp −115 to −134 from the transcriptional start site. HBP1 repressed the endogenous DNMT1 gene through sequence-specific binding, resulting in both gene-specific (e.g., p16INK4) and global DNA hypomethylation changes. The HBP1-mediated repression by DNMT1 contributed to replicative and premature senescence, the latter of which could be induced by Ras and HBP1 itself. A detailed investigation unexpectedly revealed that HBP1 has dual and complex transcriptional functions, both of which contribute to premature senescence. HBP1 both repressed the DNMT1 gene and activated the p16 gene in premature senescence. The opposite transcriptional functions proceeded through different DNA sequences and differential protein acetylation. While intricate, the reciprocal partnership between HBP1 and DNMT1 has exceptional importance, since its abrogation compromises senescence and promotes tumorigenesis. Together, our results suggest that the HBP1 transcription factor orchestrates a complex regulation of key genes during cellular senescence, with an impact on overall DNA methylation state.
PMCID: PMC3623086  PMID: 23249948
21.  Correction: Altered Reward Circuitry in the Norepinephrine Transporter Knockout Mouse 
PLoS ONE  2013;8(8):10.1371/annotation/76516adc-5541-4941-b4ff-e80d1c4d769f.
PMCID: PMC3747294
22.  The distinct expression patterns of claudin-2, -6, and −11 between human gastric neoplasms and adjacent non-neoplastic tissues 
Diagnostic Pathology  2013;8:133.
Cancers have a multifactorial etiology a part of which is genetic. Recent data indicate that expression of the tight junction claudin proteins is involved in the etiology and progression of cancer.
To explore the correlations of the tight junction proteins claudin-2,-6, and −11 in the pathogenesis and clinical behavior of gastric cancer, 40 gastric cancer tissues and 28 samples of non-neoplastic tissues adjacent to the tumors were examined for expression of claudin-2,-6, and −11 by streptavidin-perosidase immunohistochemical staining method.
The positive expression rates of claudin-2 in gastric cancer tissues and adjacent tissues were 25% and 68% respectively (P < 0.001). The positive expression rates of claudin-6 in gastric cancer tissues and adjacent tissues were 55% and 79% respectively (P = 0.045 < 0.05). In contrast, the positive expression rates of claudin-11 in gastric cancer tissues and gastric cancer adjacent tissues were 80% and 46% (P = 0.004 < 0.01). Thus in our study, the expression of claudin-2, and claudin-6 was down regulated in gastric cancer tissue while the expression of claudin-11 was up regulated. Correlations between claudin expression and clinical behavior were not observed.
Our study provides the first evidence that claudin-2,-6, and −11 protein expression varies between human gastric cancers and adjacent non-neoplastic tissues.
Virtual slides
The virtual slide(s) for this article can be found here:
PMCID: PMC3751254  PMID: 23919729
Gastric cancer; Tight junctions; Claudin-2; Claudin-6; Claudin-11; Immunohistochemistry
23.  Loss of CARM1 is linked to reduced HuR function in replicative senescence 
BMC Molecular Biology  2013;14:15.
The co-activator-associated arginine methyltransferase 1 (CARM1) catalyzes the methylation of HuR. However, the functional impact of this modification is not fully understood. Here, we investigated the influence of HuR methylation by CARM1 upon the turnover of HuR target mRNAs encoding senescence-regulatory proteins.
Changing the methylation status of HuR in HeLa cells by either silencing CARM1 or mutating the major methylation site (R217K) greatly diminished the effect of HuR in regulating the turnover of mRNAs encoding cyclin A, cyclin B1, c-fos, SIRT1, and p16. Although knockdown of CARM1 or HuR individually influenced the expression of cyclin A, cyclin B1, c-fos, SIRT1, and p16, joint knockdown of both CARM1 and HuR did not show further effect. Methylation by CARM1 enhanced the association of HuR with the 3′UTR of p16 mRNA, but not with the 3′UTR of cyclin A, cyclin B1, c-fos, or SIRT1 mRNAs. In senescent human diploid fibroblasts (HDFs), reduced CARM1 was accompanied by reduced HuR methylation. In addition, knockdown of CARM1 or mutation of the major methylation site of HuR in HDF markedly impaired the ability of HuR to regulate the expression of cyclin A, cyclin B1, c-fos, SIRT1, and p16 as well to maintain a proliferative phenotype.
CARM1 represses replicative senescence by methylating HuR and thereby enhancing HuR’s ability to regulate the turnover of cyclin A, cyclin B1, c-fos, SIRT1, and p16 mRNAs.
PMCID: PMC3718661  PMID: 23837869
CARM1; HuR methylation; mRNA turnover; Replicative senescence
24.  Loss of the miR-144/451 cluster impairs ischaemic preconditioning-mediated cardioprotection by targeting Rac-1 
Cardiovascular Research  2012;94(2):379-390.
While a wealth of data has uncovered distinct microRNA (miR) expression alterations in hypertrophic and ischaemic/reperfused (I/R) hearts, little is known about miR regulation and response to ischaemic preconditioning (IPC).
Methods and results
We analysed miRs in murine hearts preconditioned with six cycles of 4 min ischaemia via coronary artery occlusion, followed by 4 min reperfusion in vivo. Both miRs within the miR-144/451 cluster were the most elevated among a cohort of 21 dysregulated miRs in preconditioned hearts, compared with shams. To investigate the significance of this finding, we examined IPC-mediated cardioprotection within a miR-144/451-knockout (KO) mouse model. Wild-type (WT) hearts exposed to IPC followed by I/R (30 min/24 h) showed a smaller infarction size compared with mice treated with I/R alone. In contrast, IPC failed to protect miR-144/451-KO hearts against infarct caused by I/R treatment. Thus, the miR-144/451 cluster is required for IPC-elicited cardioprotection. Rac-1, a key component of NADPH oxidase, was mostly up-regulated in KO hearts among three bona fide targets (Rac-1, 14-3-3ζ, and CUGBP2) for both miR-144 and miR-451. Accordingly, reactive oxygen species (ROS) levels were markedly increased in KO hearts upon IPC, compared with IPC-WT hearts. Pre-treatment of KO hearts with a Rac-1 inhibitor NSC23766 (20 mg/kg, ip) reduced IPC-triggered ROS levels and restored IPC-elicited cardioprotection. Using antagomiRs, we showed that miR-451 was largely responsible for IPC-mediated cardioprotection.
Loss of the miR-144/451 cluster limits IPC cardioprotection by up-regulating Rac-1-mediated oxidative stress signalling.
PMCID: PMC3331614  PMID: 22354898
MicroRNA-144; MicroRNA-451; Ischaemic preconditioning; Myocardial ischaemia/reperfusion; Rac-1; Oxidative stress
25.  Deficits in axonal transport in hippocampal-based circuitry and the visual pathway in APP knock-out animals witnessed by manganese enhanced MRI 
Neuroimage  2012;60(3):1856-1866.
Mounting evidence implicates axonal transport defects, typified by the presence of axonal varicosities with aberrant accumulations of cargo, as an early event in Alzheimer’s disease (AD) pathogenesis. Work identifying amyloid precursor protein (APP) as a vesicular motor receptor for anterograde axonal transport further implicates axonal transport in AD. Manganese-enhanced MRI (MEMRI) detects axonal transport dynamics in preclinical studies. Here we pursue an understanding of the role of APP in axonal transport in the central nervous system by applying MEMRI to hippocampal circuitry and to the visual pathway in living mice homozygous for either wild type or a deletion in the APP gene (n = 12 for each genotype). Following intra-ocular or stereotaxic hippocampal injection, we performed time-lapse MRI to detect Mn2+ transport. Three dimensional whole brain datasets were compared on a voxel-wise basis using within-group pair-wise analysis. Quantification of transport to structures connected to injection sites via axonal fiber tracts was also performed. Histology confirmed consistent placement of hippocampal injections and no observable difference in glial-response to the injections. APP −/− mice had significantly reduced transport from the hippocampus to the septal nuclei and amygdala after 7 hours and reduced transport to the contralateral hippocampus after 25 hours; axonal transport deficits in the APP −/− animals were also identified in the visual pathway. These data support a system-wide role for APP in axonal transport within the central nervous system and demonstrate the power of MEMRI for assessing neuronal circuitry involved in memory and learning.
PMCID: PMC3328142  PMID: 22500926
Amyloid precursor protein; Manganese enhanced MRI (MEMRI); Alzheimer’s disease; axonal transport; statistical parametric mapping; optic tract; synaptic transmission; hippocampus; visual pathway

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