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1.  Discovery of Novel Wall Teichoic Acid Inhibitors as Effective anti-MRSA β-lactam Combination Agents 
Chemistry & biology  2013;20(2):272-284.
Innovative strategies are needed to combat drug resistance associated with methicillin-resistant Staphylococcus aureus (MRSA). Here, we investigate the potential of wall teichoic acid (WTA) biosynthesis inhibitors as combination agents to restore β-lactam efficacy against MRSA. Performing a whole cell pathway-based screen we identified a series of WTA inhibitors (WTAIs) targeting the WTA transporter protein, TarG. Whole genome sequencing of WTAI resistant isolates across two methicillin-resistant Staphylococci spp. revealed TarG as their common target, as well as a broad assortment of drug resistant bypass mutants mapping to earlier steps of WTA biosynthesis. Extensive in vitro microbiological analysis and animal infection studies provide strong genetic and pharmacological evidence of the potential effectiveness of WTAIs as anti-MRSA β-lactam combination agents. This work also highlights the emerging role of whole genome sequencing in antibiotic mode-of-action and resistance studies.
PMCID: PMC3762323  PMID: 23438756
Staphylococcus aureus; MRSA; MRSE; imipenem; wall teichoic acid; antibiotic resistance; β-lactam potentiation; combination agent; chemical biology; Next Generation Sequencing
2.  Targeting the Biophysical Properties of the Myeloma Initiating Cell Niches: A Pharmaceutical Synergism Analysis Using Multi-Scale Agent-Based Modeling 
PLoS ONE  2014;9(1):e85059.
Multiple myeloma, the second most common hematological cancer, is currently incurable due to refractory disease relapse and development of multiple drug resistance. We and others recently established the biophysical model that myeloma initiating (stem) cells (MICs) trigger the stiffening of their niches via SDF-1/CXCR4 paracrine; The stiffened niches then promote the colonogenesis of MICs and protect them from drug treatment. In this work we examined in silico the pharmaceutical potential of targeting MIC niche stiffness to facilitate cytotoxic chemotherapies. We first established a multi-scale agent-based model using the Markov Chain Monte Carlo approach to recapitulate the niche stiffness centric, pro-oncogenetic positive feedback loop between MICs and myeloma-associated bone marrow stromal cells (MBMSCs), and investigated the effects of such intercellular chemo-physical communications on myeloma development. Then we used AMD3100 (to interrupt the interactions between MICs and their stroma) and Bortezomib (a recently developed novel therapeutic agent) as representative drugs to examine if the biophysical properties of myeloma niches are drugable. Results showed that our model recaptured the key experimental observation that the MBMSCs were more sensitive to SDF-1 secreted by MICs, and provided stiffer niches for these initiating cells and promoted their proliferation and drug resistance. Drug synergism analysis suggested that AMD3100 treatment undermined the capability of MICs to modulate the bone marrow microenvironment, and thus re-sensitized myeloma to Bortezomib treatments. This work is also the first attempt to virtually visualize in 3D the dynamics of the bone marrow stiffness during myeloma development. In summary, we established a multi-scale model to facilitate the translation of the niche-stiffness centric myeloma model as well as experimental observations to possible clinical applications. We concluded that targeting the biophysical properties of stem cell niches is of high clinical potential since it may re-sensitize tumor initiating cells to chemotherapies and reduce risks of cancer relapse.
PMCID: PMC3903473  PMID: 24475036
3.  CASME II: An Improved Spontaneous Micro-Expression Database and the Baseline Evaluation 
PLoS ONE  2014;9(1):e86041.
A robust automatic micro-expression recognition system would have broad applications in national safety, police interrogation, and clinical diagnosis. Developing such a system requires high quality databases with sufficient training samples which are currently not available. We reviewed the previously developed micro-expression databases and built an improved one (CASME II), with higher temporal resolution (200 fps) and spatial resolution (about 280×340 pixels on facial area). We elicited participants' facial expressions in a well-controlled laboratory environment and proper illumination (such as removing light flickering). Among nearly 3000 facial movements, 247 micro-expressions were selected for the database with action units (AUs) and emotions labeled. For baseline evaluation, LBP-TOP and SVM were employed respectively for feature extraction and classifier with the leave-one-subject-out cross-validation method. The best performance is 63.41% for 5-class classification.
PMCID: PMC3903513  PMID: 24475068
4.  Risk Factors and Prevalence of Helicobacter pylori Infection in Persistent High Incidence Area of Gastric Carcinoma in Yangzhong City 
Aim. The aim of this study was to investigate the prevalence and risk factors of H. pylori infection in areas with high prevalence of gastric cancer in Jiangsu Province, China. Methods. A prospective epidemiologic survey of H. pylori infection was accomplished in a natural population of 5417 individuals in Yangzhong city. Questionnaires and 13C-urea breath test for H. pylori infection were performed. Results. Among 5417 subjects who completed questionnaires and 13C-urea breath test, 3435 (63.41%) were H. pylori positive. The prevalence reached a peak at the age of 30–39 years (90.82%). There was significant difference between sexes and women had a higher infection rate than men. The prevalence of H. pylori infection was also associated with eating kipper food and fried food. No association between H. pylori prevalence and smoking or drinking was found. Compared to healthy individuals, people with dyspeptic diseases (peptic ulcer, gastroenteritis) presented a high prevalence of H. pylori infection. Using multivariate logistic regression analysis, age and history of peptic ulcer and gastroenteritis were the independent predictors for H. pylori infection. Conclusions. Yangzhong city had a high prevalence of H. pylori infection and was related to several risk factors. The underlying mechanisms are needed to be further investigated.
PMCID: PMC3914443  PMID: 24550981
5.  An Attenuated Coxsackievirus B3 Vector: A Potential Tool for Viral Tracking Study and Gene Delivery 
PLoS ONE  2013;8(12):e83753.
Cardiomyocytes are quite resistant to gene transfer using standard techniques. We developed an expression vector carrying an attenuated but infectious and replicative coxsackievirus B3 (CVB3) genome, and unique ClaI-StuI cloning sites for an exogenous gene, whose product can be released from the nascent viral polyprotein by 2Apro cleavage. This vector was tested as an expression vehicle for green fluorescent protein (GFP). The vector transiently expressed GFP in cell cultures for at least ten passages and delivered functional GFP to the infected cardiomyocytes for at least 6 days. Moreover, the recombinant viruses showed virulence attenuation in vitro and in vivo. The findings suggest that the recombinant CVB3 vector could be a useful tool for viral tracking study and delivering exogenous proteins to cardiomyocytes.
PMCID: PMC3875476  PMID: 24386270
6.  Contradictory imaging and EEG results in resection surgery of bitemporal lobe epilepsy: A case report 
The present study describes the case of a 27-year-old, right-handed female with bilateral mesial temporal lobe epilepsy. Electroencephalogram (EEG) monitoring from implanted electrodes displayed two different and independent onsets on the two sides of the mesial temporal structures, which specifically included clinical generalized tonic clonic seizure (GTCS) discharges originating from the left mesial temporal lobe, as well as complex partial seizure (CPS) discharges arising from the right mesial region. However, fluid-attenuated inversion recovery magnetic resonance imaging (FLAIR MRI) showed a unilateral abnormality, as in right mesial temporal lobe sclerosis. A decision was made to resect one side of the mesial temporal lobe, in order to avoid memory function impairment, and, relying on the MRI results, the right side was selected. However, surgery did not leave the patient seizure-free. The CPSs gradually eased, while the GTCSs originating from the left side became severely aggravated. In describing this case, the drawbacks of current epileptic diagnostic methods and surgical strategies for bitemporal lobe epilepsy are discussed, and the requirement for more treatment options is emphasized.
PMCID: PMC3919855  PMID: 24520276
epilepsy; bitemporal lobe epilepsy; epileptic surgery
7.  Systematically Studying Kinase Inhibitor Induced Signaling Network Signatures by Integrating Both Therapeutic and Side Effects 
PLoS ONE  2013;8(12):e80832.
Substantial effort in recent years has been devoted to analyzing data based large-scale biological networks, which provide valuable insight into the topologies of complex biological networks but are rarely context specific and cannot be used to predict the responses of cell signaling proteins to specific ligands or compounds. In this work, we proposed a novel strategy to investigate kinase inhibitor induced pathway signatures by integrating multiplex data in Library of Integrated Network-based Cellular Signatures (LINCS), e.g. KINOMEscan data and cell proliferation/mitosis imaging data. Using this strategy, we first established a PC9 cell line specific pathway model to investigate the pathway signatures in PC9 cell line when perturbed by a small molecule kinase inhibitor GW843682. This specific pathway revealed the role of PI3K/AKT in modulating the cell proliferation process and the absence of two anti-proliferation links, which indicated a potential mechanism of abnormal expansion in PC9 cell number. Incorporating the pathway model for side effects on primary human hepatocytes, it was used to screen 27 kinase inhibitors in LINCS database and PF02341066, known as Crizotinib, was finally suggested with an optimal concentration 4.6 uM to suppress PC9 cancer cell expansion while avoiding severe damage to primary human hepatocytes. Drug combination analysis revealed that the synergistic effect region can be predicted straightforwardly based on a threshold which is an inherent property of each kinase inhibitor. Furthermore, this integration strategy can be easily extended to other specific cell lines to be a powerful tool for drug screen before clinical trials.
PMCID: PMC3855094  PMID: 24339888
8.  Control Efficacy of Annual Community-Wide Treatment against Schistosoma japonicum in China: A Meta-Analysis 
PLoS ONE  2013;8(11):e78509.
Human schistosomiasis is caused by schistosome, with annual loss of over 70 million disability adjusted life years in the world. China is endemic with Schistosoma japonicum and large-scale chemotherapy with praziquantel has become the mainstay of control in China since 1990s. However, the control effects of mass treatment in the field have been uneven. Moreover, mass treatment has come into a wide use in other countries with limited health resources. Therefore, a better understanding of the control effect of mass treatment is in an urgent need.
We performed a systematic search of the literature to investigate the control efficiency of annual community-wide treatment (ACWT, treatment to an entire community without any preliminary screening) with a single dose of PZQ (40 mg kg−1 bodyweight) against schistosome in humans in China. Three Chinese literature databases, including China National Knowledge Infrastructure, WanFang and Chinese Scientific Journal Databases, and the PubMed were searched. Pooled prevalence ratios (prevalence after to before treatment) were used to assess effect. Our protocol is available on PROSPERO (No. CRD42013003628).
22 articles were included. Meta-analyses on data from 18 studies on one round of ACWT, 17 studies on two consecutive rounds and 6 studies on three consecutive rounds were performed. The results showed control effects of ACWT plus other measures were statistically significant, with prevalence ratios being 0.38 (0.31, 0.46) for one round, 0.28 (0.22, 0.35) for two rounds and 0.22 (0.10, 0.46) for three rounds. When ACWT was performed alone or with health education only, the values for one and two rounds were 0.389 (0.307, 0.492) and 0.348 (0.300, 0.403), respectively.
The control effect of ACWT alone or with other measures is significant and increases with the number of rounds. Such program is recommended in high endemic areas and the criteria yet merit further assessment.
PMCID: PMC3817216  PMID: 24223819
9.  Cytokine Combination Therapy Prediction for Bone Remodeling in Tissue Engineering Based on the Intracellular Signaling Pathway 
Biomaterials  2012;33(33):8265-8276.
The long-term performance of tissue-engineered bone grafts is determined by a dynamic balance between bone regeneration and resorption. We proposed using embedded cytokine slow-releasing hydrogels to tune this balance toward a desirable final bone density. In this study we established a systems biology model, and quantitatively explored the combinatorial effects of delivered cytokines from hydrogels on final bone density. We hypothesized that: 1) bone regeneration was driven by transcription factors Runx2 and Osterix, which responded to released cytokines, such as Wnt, BMP2, and TGFβ, drove the development of osteoblast lineage, and contributed to bone mass generation; and 2) the osteoclast lineage, on the other hand, governed the bone resorption, and communications between these two lineages determined the dynamics of bone remodeling. In our model, Intracellular signaling pathways were represented by ordinary differential equations, while the intercellular communications and cellular population dynamics were modeled by stochastic differential equations. Effects of synergistic cytokine combinations were evaluated by Loewe index and Bliss index. Simulation results revealed that the Wnt/BMP2 combinations released from hydrogels showed best control of bone regeneration and synergistic effects, and suggested optimal dose ratios of given cytokine combinations released from hydrogels to most efficiently control the long-term bone remodeling. We revealed the characteristics of cytokine combinations of Wnt/BMP2 which could be used to guide the design of in vivo bone scaffolds and the clinical treatment of some diseases such as osteoporosis.
PMCID: PMC3444627  PMID: 22910219
Bone tissue engineering; Bone remodeling; Cytokine combination therapy; Systems biology; Osteogenic differentiation; Signaling pathway
10.  Metabolic Signatures of Kidney Yang Deficiency Syndrome and Protective Effects of Two Herbal Extracts in Rats Using GC/TOF MS 
Kidney Yang Deficiency Syndrome (KDS-Yang), a typical condition in Chinese medicine, shares similar clinical signs of the glucocorticoid withdrawal syndrome. To date, the underlying mechanism of KDS-Yang has been remained unclear, especially at the metabolic level. In this study, we report a metabolomic profiling study on a classical model of KDS-Yang in rats induced by hydrocortisone injection to characterize the metabolic transformation using gas chromatography/time-of-flight mass spectrometry. WKY1, a polysaccharide extract from Astragalus membranaceus and Lycium barbarum, and WKY2, an aqueous extract from a similar formula containing Astragalus membranaceus, Lycium barbarum, Morinda officinalis, Taraxacum mongolicum, and Cinnamomum cassia presl, were used separately for protective treatments of KDS-Yang. The changes of serum metabolic profiles indicated that significant alterations of key metabolic pathways in response to abrupt hydrocortisone perturbation, including decreased energy metabolism (lactic acid, acetylcarnitine), lipid metabolism (free fatty acids, 1-monolinoleoylglycerol, and cholesterol), gut microbiota metabolism (indole-3-propionic acid), biosynthesis of catecholamine (norepinephrine), and elevated alanine metabolism, were attenuated or normalized with different degrees by the pretreatment of WKY1 or WKY2, which is consistent with the observations in which the two herbal agents could ameliorate biochemical markers of serum cortisone, adrenocorticotropic (ACTH), and urine 17-hydroxycorticosteroids (17-OHCS).
PMCID: PMC3789486  PMID: 24159348
11.  Delivery of Therapeutic AGT shRNA by PEG-Bu for Hypertension Therapy 
PLoS ONE  2013;8(7):e68651.
Gene silencing by RNA interference (RNAi) is a promising approach for gene therapy. However, up to today, it is still a major challenge to find safe and efficient non-viral vectors. Previously, we reported PEI-Bu, a small molecular weight PEI derivative, as an efficient non-viral carrier. However, like many PEI-based polymers, PEI-Bu was too toxic. In order to reduce cytotoxicity while maintain or even enhance transfecion efficiency, we modified PEI-Bu with poly(ethylene glycol) (PEG) to obtain PEG-Bu, and used it to delivery a theraputic short hairpin RNA (shRNA) targeting angiotensinogen (AGT) to normal rat liver cells (BRL-3A), which was a key target for the treatment of hypertension. The structure of PEG-Bu was confirmed by proton nuclear magnetic resonance (1H-NMR). Gel permeation chromatography (GPC) showed that the weight average molecular weight (Mw) of PEG-Bu was 5880 Da, with a polydispersity of 1.58. PEG-Bu could condense gene cargo into spherical and uniform nanoparticles with particle size (65–88 nm) and zeta potential (7.3–9.6 mV). Interestingly and importantly, PEG-Bu displayed lower cytotoxicity and enhanced tranfection efficiency than PEI-Bu after PEGylation in both normal cells BRL-3A and tumor cells HeLa. Moreover, PEG-Bu could efficiently delivery AGT shRNA to knockdown the AGT expression. To sum up, PEG-Bu would be a promising non-viral vector for delivering AGT shRNA to BRL-3A cells for hypertension therapy.
PMCID: PMC3716693  PMID: 23894329
12.  Triple pathology in patients with temporal lobe epilepsy: A case report and review of the literature 
The coexistence of three intracranial lesions related to epileptic pathogenesis is known as ‘triple pathology’ and has rarely been reported. In this study we report a case of temporal lobe epilepsy (TLE) with the coexistence of hippocampal sclerosis (HS), focal cortical dysplasia (FCD) and ganglioglioma in the temporal lobe. A 29-year-old male who had experienced recurrent seizures for four years was admitted to hospital. Cerebral magnetic resonance imaging (MRI) was conducted and T2-weighted and fluid-attenuated inversion recovery sequence (FLAIR) images revealed a reduced hippocampal volume with an increased FLAIR signal on the right side and a slightly enlarged temporal horn, which are typical imaging findings for HS and FCD. The patient underwent resectioning of the right anterior temporal lobe, hippocampus and amygdala, in addition to the lesion located in the medial temporal lobe. Immunohistochemical analysis of the medial temporal lobe lesion confirmed a ganglioglioma (WHO grade I) in the medial temporal lobe. During the first eight months following surgery, the patient's seizures were controlled with zonisamide and phenytoin. Electroencephalogram (EEG) assessment post-surgery confirmed the absence of epileptic discharges. Based on a literature review and a detailed review of this case, we postulate two possible explanations for the pathogenesis of ‘triple pathology’: i) ‘triple pathology’ is a combination of pathological progression and occasionality; and ii) ‘triple pathology’ lesions have similar pathological origins.
PMCID: PMC3797311  PMID: 24137291
epilepsy; triple pathology; hippocampal sclerosis; focal cortical dysplasia; ganglioglioma
13.  The BAR domain of amphiphysin is required for cleavage furrow tip–tubule formation during cellularization in Drosophila embryos 
Molecular Biology of the Cell  2013;24(9):1444-1453.
The BAR domain of amphiphysin is required for endocytic tubule formation at the leading edge of cleavage furrows in Drosophila embryos. In the absence of tubules, furrow ingression is faster, whereas elongated tubules slow furrow ingression, suggesting that the tubules may regulate furrow ingression by regulating membrane recycling.
De novo formation of cells in the Drosophila embryo is achieved when each nucleus is surrounded by a furrow of plasma membrane. Remodeling of the plasma membrane during cleavage furrow ingression involves the exocytic and endocytic pathways, including endocytic tubules that form at cleavage furrow tips (CFT-tubules). The tubules are marked by amphiphysin but are otherwise poorly understood. Here we identify the septin family of GTPases as new tubule markers. Septins do not decorate CFT-tubules homogeneously: instead, novel septin complexes decorate different CFT-tubules or different domains of the same CFT-tubule. Using these new tubule markers, we determine that all CFT-tubule formation requires the BAR domain of amphiphysin. In contrast, dynamin activity is preferentially required for the formation of the subset of CFT-tubules containing the septin Peanut. The absence of tubules in amphiphysin-null embryos correlates with faster cleavage furrow ingression rates. In contrast, upon inhibition of dynamin, longer tubules formed, which correlated with slower cleavage furrow ingression rates. These data suggest that regulating the recycling of membrane within the embryo is important in supporting timely furrow ingression.
PMCID: PMC3639055  PMID: 23447705
14.  Association between Exposure to Electromagnetic Fields from High Voltage Transmission Lines and Neurobehavioral Function in Children 
PLoS ONE  2013;8(7):e67284.
Evidence for a possible causal relationship between exposure to electromagnetic fields (EMF) emitted by high voltage transmission (HVT) lines and neurobehavioral dysfunction in children is insufficient. The present study aims to investigate the association between EMF exposure from HVT lines and neurobehavioral function in children.
Two primary schools were chosen based on monitoring data of ambient electromagnetic radiation. A cross-sectional study with 437 children (9 to 13 years old) was conducted. Exposure to EMF from HVT lines was monitored at each school. Information was collected on possible confounders and relevant exposure predictors using standardized questionnaires. Neurobehavioral function in children was evaluated using established computerized neurobehavioral tests. Data was analyzed using multivariable regression models adjusted for relevant confounders.
After controlling for potential confounding factors, multivariable regression revealed that children attending a school near 500 kV HVT lines had poorer performance on the computerized neurobehavioral tests for Visual Retention and Pursuit Aiming compared to children attending a school that was not in close proximity to HVT lines.
The results suggest long-term low-level exposure to EMF from HVT lines might have a negative impact on neurobehavioral function in children. However, because of differences in results only for two of four tests achieved statistical significance and potential limitations, more studies are needed to explore the effects of exposure to extremely low frequency EMF on neurobehavioral function and development in children.
PMCID: PMC3700989  PMID: 23843999
15.  Fusion Tensor Subspace Transformation Framework 
PLoS ONE  2013;8(7):e66647.
Tensor subspace transformation, a commonly used subspace transformation technique, has gained more and more popularity over the past few years because many objects in the real world can be naturally represented as multidimensional arrays, i.e. tensors. For example, a RGB facial image can be represented as a three-dimensional array (or 3rd-order tensor). The first two dimensionalities (or modes) represent the facial spatial information and the third dimensionality (or mode) represents the color space information. Each mode of the tensor may express a different semantic meaning. Thus different transformation strategies should be applied to different modes of the tensor according to their semantic meanings to obtain the best performance. To the best of our knowledge, there are no existing tensor subspace transformation algorithm which implements different transformation strategies on different modes of a tensor accordingly. In this paper, we propose a fusion tensor subspace transformation framework, a novel idea where different transformation strategies are implemented on separate modes of a tensor. Under the framework, we propose the Fusion Tensor Color Space (FTCS) model for face recognition.
PMCID: PMC3698091  PMID: 23840864
16.  Brain 3-Mercaptopyruvate Sulfurtransferase (3MST): Cellular Localization and Downregulation after Acute Stroke 
PLoS ONE  2013;8(6):e67322.
3-Mercaptopyruvate sulfurtransferase (3MST) is an important enzyme for the synthesis of hydrogen sulfide (H2S) in the brain. We present here data that indicate an exclusively localization of 3MST in astrocytes. Regional distribution of 3MST activities is even and unremarkable. Following permanent middle cerebral artery occlusion (pMCAO), 3MST was down-regulated in both the cortex and striatum, but not in the corpus collosum. It appears that the down-regulation of astrocytic 3MST persisted in the presence of astrocytic proliferation due to gliosis. Our observations indicate that 3MST is probably not responsible for the increased production of H2S following pMCAO. Therefore, cystathionine β-synthase (CBS), the alternative H2S producing enzyme in the CNS, remains as a more likely potential therapeutic target than 3MST in the treatment of acute stroke through inhibition of H2S production.
PMCID: PMC3689812  PMID: 23805308
17.  Phylogenetically Diverse ureC Genes and Their Expression Suggest the Urea Utilization by Bacterial Symbionts in Marine Sponge Xestospongia testudinaria 
PLoS ONE  2013;8(5):e64848.
Urea is one of the dominant organic nitrogenous compounds in the oligotrophic oceans. Compared to the knowledge of nitrogen transformation of nitrogen fixation, ammonia oxidization, nitrate and nitrite reduction mediated by sponge-associated microbes, our knowledge of urea utilization in sponges and the phylogenetic diversity of sponge-associated microbes with urea utilization potential is very limited. In this study, Marinobacter litoralis isolated from the marine sponge Xestospongia testudinaria and the slurry of X. testudinaria were found to have urease activity. Subsequently, phylogenetically diverse bacterial ureC genes were detected in the total genomic DNA and RNA of sponge X. testudinaria, i.e., 19 operative taxonomic units (OTUs) in genomic DNA library and 8 OTUs in cDNA library at 90% stringency. Particularly, 6 OTUs were common to both the genomic DNA library and the cDNA library, which suggested that some ureC genes were expressed in this sponge. BLAST and phylogenetic analysis showed that most of the ureC sequences were similar with the urease alpha subunit of members from Proteobacteria, which were the predominant component in sponge X. testudinaria, and the remaining ureC sequences were related to those from Magnetococcus, Cyanobacteria, and Actinobacteria. This study is the first assessment of the role of sponge bacterial symbionts in the regenerated utilization of urea by the detection of transcriptional activity of ureC gene, as well as the phylogenetic diversity of ureC gene of sponge bacterial symbionts. The results suggested the urea utilization by bacterial symbionts in marine sponge X. testudinaria, extending our understanding of nitrogen cycling mediated by sponge-associated microbiota.
PMCID: PMC3669342  PMID: 23741404
18.  Endoplasmic Reticulum Is at the Crossroads of Autophagy, Inflammation, and Apoptosis Signaling Pathways and Participates in the Pathogenesis of Diabetes Mellitus 
Journal of Diabetes Research  2013;2013:193461.
Diabetes mellitus (DM) is a chronic metabolic disease, and its incidence is growing worldwide. The endoplasmic reticulum (ER) is a central component of cellular functions and is involved in protein folding and trafficking, lipid synthesis, and maintenance of calcium homeostasis. The ER is also a sensor of both intra- and extracellular stress and thus participates in monitoring and maintaining cellular homeostasis. Therefore, the ER is one site of interaction between environmental signals and a cell's biological function. The ER is tightly linked to autophagy, inflammation, and apoptosis, and recent evidence suggests that these processes are related to the pathogenesis of DM and its complications. Thus, the ER has been considered an intersection integrating multiple stress responses and playing an important role in metabolism-related diseases including DM. Here, we review the relationship between the ER and autophagy, inflammation, and apoptosis in DM to better understand the molecular mechanisms of this disease.
PMCID: PMC3673337  PMID: 23762873
19.  mRNA expression and hypermethylation of tumor suppressor genes apoptosis protease activating factor-1 and death-associated protein kinase in oral squamous cell carcinoma 
Oncology Letters  2013;6(1):280-286.
Apoptosis protease activating factor-1 (Apaf-1) and death-associated protein kinase (DAPK) are p53 pathway-related genes that play significant roles in the activation of caspases, which are involved in mitochondrial-mediated apoptosis. The present study aimed to confirm the role of hyper-methylation of the Apaf-1 and DAPK gene promoter regions in oral squamous cell carcinoma (OSCC) and the effect of the demethylation drug, 5-aza-2′-deoxycytidine (DAC). mRNA from 53 OSCC samples, 23 normal oral mucosa samples and Tca8113 human tongue carcinoma cell lines was detected using semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR). The DNA from each sample was analyzed using methylation-specific PCR (MSP). The Tca8113 cells were demethylated using DAC and the demethylation and re-expression of Apaf-1 and DAPK were analyzed. The Apaf-1 and DAPK mRNA expression index was decreased in 51 (96.23%) and 50 (94.34%) cases, respectively, in the tumor tissues. Hypermethylation of the Apaf-1 and DAPK promoter regions was detected in 46 (86.79%) and 38 (71.69%) cases, respectively. Promoter hypermethylation of the two genes correlated with a decreased mRNA expression in the tumor tissues. Subsequent to being treated with DAC, Apaf-1 and DAPK were demethylated and re-expressed in the Tca8113 cells. Apaf-1 and DAPK promoter hypermethylation may be associated with low gene expression in OSCC. Furthermore, a loss of Apaf-1 and DAPK expression may recover following demethylation. The data provide evidence that methylation exists in OSCC and may play a role in the development of this disease.
PMCID: PMC3742820  PMID: 23946818
methylation; apoptosis protease activating factor-1; death-associated protein kinase; oral squamous cell carcinoma; 5-aza-2′-deoxycytidine
20.  Metabonomic Strategy to the Evaluation of Chinese Medicine Compound Danshen Dripping Pills Interfering Myocardial Ischemia in Rats 
Coronary heart disease (CHD) is one of the highest mortality diseases in the world. Traditional Chinese medicine compound Danshen dripping pills (CDDPs) have currently made a great achievement in treating CHD. However, the therapeutic mechanism of CDDP is often poorly interpreted. In this study, a GC-MS-based metabonomic study was conducted to assess the holistic efficacy of CDDP for myocardial infarction in male Sprague-Dawley rats, which were divided into the control group, the sham group, the model group, the control + CDDP group, and the model + CDDP, with CDDP at a dose of 107 mg/kg·d (equal to 1.8 mL/kg·d). The metabonomic findings demonstrated great differences of metabolic pattern among sham, model, and the model + CDDP in the orthogonal partial least squares discriminant analysis (OPLS-DA) models, which coordinated well with the assessment of plasma biochemistry and histopathological assay. Differentially expressed metabolites suggested that energy metabolism, glycolysis, and lipid metabolism might be disrupted by myocardial infarction. Both the potential metabolic biomarkers and the biochemical histopathological indices were regulated effectively by CDDP.
PMCID: PMC3659432  PMID: 23737844
21.  Development of dextran nanoparticles for stabilizing delicate proteins 
Nanoscale Research Letters  2013;8(1):197.
One of the most challenging problems in the development of protein pharmaceuticals is to deal with stabilities of proteins due to its complicated structures. This study aims to develop a novel approach to stabilize and encapsulate proteins into dextran nanoparticles without contacting the interface between the aqueous phase and the organic phase. The bovine serum albumin, granulocyte-macrophage colony-stimulating factor (GM-CSF), granulocyte colony-stimulating factor (G-CSF), β-galactosidase, and myoglobin were selected as model proteins. The proteins were added into an aqueous solution containing the dextran and polyethylene glycol, and then encapsulated into dextran nanoparticles by aqueous-aqueous freezing-induced phase separation. The encapsulation efficiency and recovery of dextran nanoparticles were determined. The dextran nanoparticles loaded with proteins were characterized by scanning electron microscopy and particle size analysis. The protein aggregation was determined by size-exclusion chromatography-high-performance chromatography, and the bioactivity of proteins recovered during formulation steps was determined. The bioactivity of GM-CSF, G-CSF, and β-galactosidase were examined by the proliferation of TF-1 cell, NSF-60 cell, and ortho-nitrophenyl-β-galactoside assay, respectively. The results of bioactivity recovered show that this novel dextran nanoparticle can preserve the protein's bioactivity during the preparation process. LysoSensor™ Yellow/Blue dextran, a pH-sensitive indicator with fluorescence excited at two channels, was encapsulated into dextran nanoparticles to investigate the ability of dextran nanoparticles to resist the acidic microenvironment (pH < 2.5). The result shows that the dextran nanoparticles attenuate the acidic microenvironment in the poly (lactic-co-glycolic acid) microsphere by means of the dilution effect. These novel dextran nanoparticles provided an appealing approach to stabilize the delicate proteins for administration.
PMCID: PMC3655889  PMID: 23622054
Dextran nanoparticles; Protein; Aggregation; Bioactivity; Acidic microenvironment
22.  Epidermal Growth Factor Receptor (EGFR)-RAS Signaling Pathway in Penile Squamous Cell Carcinoma 
PLoS ONE  2013;8(4):e62175.
Penile Squamous Cell Carcinoma (SCC) is a rare cancer with poor prognosis and limited response to conventional chemotherapy. The genetic and epigenetic alterations of Epidermal Growth Factor Receptor (EGFR)-RAS-RAF signaling in penile SCC are unclear. This study aims to investigate four key members of this pathway in penile SCC. We examined the expression of EGFR and RAS-association domain family 1 A (RASSF1A) as well as the mutation status of K-RAS and BRAF in 150 cases of penile SCC. EGFR and RASSF1A expression was evaluated by immunohistochemistry. KRAS mutations at codons 12 and 13, and the BRAF mutation at codon 600 were analyzed on DNA isolated from formalin fixed paraffin embedded tissues by direct genomic sequencing. EGFR expression was positive in all specimens, and its over-expression rate was 92%. RASSF1A expression rate was only 3.42%. Significant correlation was not found between the expression of EGFR or RASSF1A and tumor grade, pT stage or lymph node metastases. The detection of KRAS and BRAF mutations analysis was performed in 94 and 83 tumor tissues, respectively. We found KRAS mutation in only one sample and found no BRAF V600E point mutation. In summary, we found over-expression of EGFR in the majority cases of penile SCC, but only rare expression of RASSF1A, rare KRAS mutation, and no BRAF mutation in penile SCC. These data suggest that anti-EGFR agents may be potentially considered as therapeutic options in penile SCC.
PMCID: PMC3634795  PMID: 23637996
23.  High-Throughput Carrier Screening Using TaqMan Allelic Discrimination 
PLoS ONE  2013;8(3):e59722.
Members of the Ashkenazi Jewish community are at an increased risk for inheritance of numerous genetic diseases such that carrier screening is medically recommended. This paper describes the development and evaluation of 30 TaqMan allelic discrimination qPCR assays for 29 mutations on 2 different high-throughput platforms. Four of these mutations are in the GBA gene and are successfully examined using short amplicons due to the qualitative nature of TaqMan allelic discrimination. Two systems were tested for their reliability (call rate) and consistency with previous diagnoses (diagnostic accuracy) indicating a call rate of 99.04% and a diagnostic accuracy of 100% (+/−0.00%) from one platform, and a call rate of 94.66% and a diagnostic accuracy of 93.35% (+/−0.29%) from a second for 9,216 genotypes. Results for mutations tested at the expected carrier frequency indicated a call rate of 97.87% and a diagnostic accuracy of 99.96% (+/−0.05%). This study demonstrated the ability of a high throughput qPCR methodology to accurately and reliably genotype 29 mutations in parallel. The universally applicable nature of this technology provides an opportunity to increase the number of mutations that can be screened simultaneously, and reduce the cost and turnaround time for accommodating newly identified and clinically relevant mutations.
PMCID: PMC3608587  PMID: 23555759
24.  Hepatocyte-targeting gene transfer mediated by galactosylated poly(ethylene glycol)-graft-polyethylenimine derivative 
Biscarbamate cross-linked polyethylenimine derivative (PEI-Et) has been reported as a novel nonviral vector for efficient and safe gene transfer in our previous work. However, it had no cell-specificity. To achieve specific delivery of genes to hepatocytes, galactosylated poly(ethylene glycol)-graft-polyethylenimine derivative (GPE) was prepared through modification of PEI-Et with poly(ethylene glycol) and lactobionic acid, bearing a galactose group as a hepatocyte-targeting moiety. The composition of GPE was characterized by proton nuclear magnetic resonance. The weight-average molecular weight of GPE measured with a gel permeation chromatography instrument was 9489 Da, with a polydispersity of 1.44. GPE could effectively condense plasmid DNA (pDNA) into nanoparticles. Gel retardation assay showed that GPE/pDNA complexes were completely formed at weigh ratios (w/w) over 3. The particle size of GPE/pDNA complexes was 79–100 nm and zeta potential was 6–15 mV, values which were appropriate for cellular uptake. The morphology of GPE/pDNA complexes under atomic force microscopy appeared spherical and uniform in size, with diameters of 53–65 nm. GPE displayed much higher transfection efficiency than commercially available PEI 25 kDa in BRL-3A cell lines. Importantly, GPE showed good hepatocyte specificity. Also, the polymer exhibited significantly lower cytotoxicity compared to PEI 25 kDa at the same concentration or weight ratio in BRL-3A cell lines. To sum up, our results indicated that GPE might carry great potential in safe and efficient hepatocyte-targeting gene delivery.
PMCID: PMC3617917  PMID: 23576866
gene delivery; hepatocyte targeting; galactose; cytotoxicity; transfection efficiency
25.  Prevalence of HIV, Syphilis, HCV and Their High Risk Behaviors among Migrant Workers in Eastern China 
PLoS ONE  2013;8(2):e57258.
The goal of this study was to understand the knowledge about AIDS, identify the correlates and determine the prevalence of HIV infection, syphilis, HCV among migrant workers in Zhejiang, China.
A cross-sectional study using face-to-face anonymous questionnaire interviews was conducted and blood samples were collected for HIV, syphilis and Hepatitis C infection screening.
17,377 (92.8%) of 18,730 migrant workers approached were interviewed. Among 17,377 participants, the HIV/AIDS knowledge rate was 66.2%. A total of 12,694 (73%) of the participants reported having ever had sexual intercourse, with 30.1% of single participants reporting having had sexual intercourse. Among those respondents with sexual experiences, 7.5% admitted they had two or more sexual partners and 4.9% reported having had sex with casual (unpaid) partners in the previous 12 months, whilst 3.7% had paid for sex. More than half of those who had paid for sex (59.4%) had not used a condom every time in their sexual acts with the sex workers. Multiple logistic regression analysis indicated that high risk sexual behavior (defined as sex with a casual or commercial sex partner without using a condom consistently) was associated with being divorced or widowed (P<0.05 for single); male gender; shorter duration of stay in Zhejiang; working in factory, market or domestic service (P<0.05 for odd job); having a province of origin inside Zhejiang; and drug use. The prevalence of HIV and HCV infections were 0.02% (95% CI: 0.01%–0.06%) and 0.40% (95%CI: 0.31%–0.51%), respectively. The prevalence of syphilis among those who were sexually active was 0.55% (95% CI: 0.43%–0.70%). Risk factors for syphilis included shorter duration of stay in Zhejiang, ethnic minority status, being divorced or widowed and having had multiple sex partners.
Much greater efforts are needed to promote safer sex, and programs for the control of syphilis need to be tailored for migrant workers in China.
PMCID: PMC3579873  PMID: 23451193

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