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1.  The compatibility of inactivated-Enterovirus 71 vaccination with Coxsackievirus A16 and Poliovirus immunizations in humans and animals 
Human Vaccines & Immunotherapeutics  2015;11(11):2723-2733.
Enterovirus 71 (EV71) is the key pathogen for Hand, Foot, and Mouth Disease (HFMD) and can result in severe neurological complications and death among young children. Three inactivated-EV71 vaccines have gone through phase III clinical trials and have demonstrated good safety and efficacy. These vaccines will benefit young children under the threat of severe HFMD. However, the potential immunization-related compatibility for different enterovirus vaccines remains unclear, making it hard to include the EV71 vaccine in Expanded Program on Immunization (EPI). Here, we measured the neutralizing antibodies (NTAbs) against EV71, Coxsackievirus A16 (CA16) and Poliovirus from infants enrolled in those EV71 vaccine clinical trials. The results indicated that the levels of NTAb GMTs for EV71 increased significantly in all 3 vaccine groups (high, middle and low dosages, respectively) post-vaccination. Seroconversion ratios and Geometric mean fold increase were significantly higher in the vaccine groups (≥7/9 and 8.9~228.1) than in the placebo group (≤1/10 and 0.8~1.7, P < 0.05). But no similar NTAb response trends were found in CA16 and 3 types of Poliovirus. The decrease of 3 types of Poliovirus NTAb GMTs and an increase of CA16 GMTs post-EV71-vaccination were found in vaccine and placebo groups. Further animal study on CA16 and poliovirus vaccine co-immunization or pre-immunization with EV71 vaccine in mice indicated that there was no NTAb cross-activity between EV71 and CA16/Poliovirus. Our research showed that inactivated-EV71 vaccine has good specific-neutralizing capacity and can be included in EPI.
doi:10.1080/21645515.2015.1011975
PMCID: PMC4685687  PMID: 25715318
coxsackievirus A16 (CA16); enterovirus 71 (EV71); foot; and mouth disease (HFMD); hand; neutralizing antibody (NTAb); poliovirus; vaccine
2.  Wireless wearable range-of-motion sensor system for upper and lower extremity joints: a validation study 
Healthcare Technology Letters  2015;2(1):12-17.
Range-of-motion (ROM) assessment is a critical assessment tool during the rehabilitation process. The conventional approach uses the goniometer which remains the most reliable instrument but it is usually time-consuming and subject to both intra- and inter-therapist measurement errors. An automated wireless wearable sensor system for the measurement of ROM has previously been developed by the current authors. Presented is the correlation and accuracy of the automated wireless wearable sensor system against a goniometer in measuring ROM in the major joints of upper (UEs) and lower extremities (LEs) in 19 healthy subjects and 20 newly disabled inpatients through intra (same) subject comparison of ROM assessments between the sensor system against goniometer measurements by physical therapists. In healthy subjects, ROM measurements using the new sensor system were highly correlated with goniometry, with 95% of differences < 20° and 10° for most movements in major joints of UE and LE, respectively. Among inpatients undergoing rehabilitation, ROM measurements using the new sensor system were also highly correlated with goniometry, with 95% of the differences being < 20° and 25° for most movements in the major joints of UE and LE, respectively.
doi:10.1049/htl.2014.0100
PMCID: PMC4613894  PMID: 26609398
wireless sensor networks; biomechanics; motion measurement; patient rehabilitation; goniometers; biomedical equipment; goniometer measurements; rehabilitation process; range-of-motion assessment; lower extremity joints; upper extremity joints; wireless wearable range-of-motion sensor system
3.  Type I interferon related genes are common genes on the early stage after vaccination by meta-analysis of microarray data 
The objective of this study was to find common immune mechanism across different kinds of vaccines. A meta-analysis of microarray datasets was performed using publicly available microarray Gene Expression Omnibus (GEO) and Array Express data sets of vaccination records. Seven studies (out of 35) were selected for this meta-analysis. A total of 447 chips (145 pre-vaccination and 302 post-vaccination) were included. Significance analysis of microarrays (SAM) program was used for screening differentially expressed genes (DEGs). Functional pathway enrichment for the DEGs was conducted in DAVID Gene Ontology (GO) database. Twenty DEGs were identified, of which 10 up-regulated genes involved immune response. Six of which were type I interferon (IFN) related genes, including LY6E, MX1, OAS3, IFI44L, IFI6 and IFITM3. Ten down-regulated genes mainly mediated negative regulation of cell proliferation and cell motion. Results of a subgroup analysis showed that although the kinds of genes varied widely between days 3 and 7 post vaccination, the pathways between them are basically the same, such as immune response and response to viruses, etc. For an independent verification of these 6 type I IFN related genes, peripheral blood mononuclear cells (PBMCs) were collected at baseline and day 3 after the vaccination from 8 Enterovirus 71(EV71) vaccinees and were assayed by RT-PCR. Results showed that the 6 DEGs were also upregulated in EV71 vaccinees. In summary, meta-analysis methods were used to explore the immune mechanism of vaccines and results indicated that the type I IFN related genes and corresponding pathways were common in early immune responses for different kinds of vaccines.
doi:10.1080/21645515.2015.1008884
PMCID: PMC4514163  PMID: 25839220
immune mechanism; meta-analysis; microarray; type I interferon; vaccine
4.  Facile Synthesis of Non-Graphitizable Polypyrrole-Derived Carbon/Carbon Nanotubes for Lithium-ion Batteries 
Scientific Reports  2016;6:19317.
Graphite is usually used as an anode material in the commercial lithium ion batteries (LIBs). The relatively low lithium storage capacity of 372 mAh g–1 and the confined rate capability however limit its large-scale applications in electrical vehicles and hybrid electrical vehicles. As results, exploring novel carbon-based anode materials with improved reversible capacity for high-energy-density LIBs is urgent task. Herein we present TNGC/MWCNTs by synthesizing tubular polypyrrole (T-PPy) via a self-assembly process, then carbonizing T-PPy at 900 °C under an argon atmosphere (TNGC for short) and finally mixing TNGC with multi-walled carbon nanotubes (MWCNTs). As for TNGC/MWCNTs, the discharge capacity of 561 mAh g−1 is maintained after 100 cycles at a current density of 100 mA g−1. Electrochemical results demonstrate that TNGC/MWCNTs can be considered as promising anode materials for high-energy-density LIBs.
doi:10.1038/srep19317
PMCID: PMC4725918  PMID: 26763296
5.  Genetic polymorphisms of CYP1A1 and risk of leukemia: a meta-analysis 
OncoTargets and therapy  2015;8:2883-2902.
The associations between CYP1A1 polymorphisms and risk of leukemia have been studied extensively, but the results have been inconsistent. Therefore, in this study, we performed a meta-analysis to clarify associations of three CYP1A1 polymorphisms (T3801C, A2455G, and C4887A) with the risks of acute lymphoblastic leukemia (ALL), acute myeloid leukemia (AML), and chronic myeloid leukemia (CML). Medline, EMBASE, and China National Knowledge Infrastructure databases were searched to collect relevant studies published up to April 20, 2015. The extracted data were analyzed statistically, and pooled odds ratios with 95% confidence intervals were calculated to quantify the associations. Overall, 26 publications were included. Finally, T3801C was associated with an increased risk of AML in Asians under the dominant model. For A2455G, the risk of ALL was increased among Caucasians in the recessive model and the allele-contrast model; A2455G was also associated with an increased risk of CML among Caucasians under the recessive model, dominant model, and allele-contrast model. For C4887A, few of the included studies produced data. In conclusion, the results suggest that Asians carrying the T3801C C allele might have an increased risk of AML and that Caucasians with the A2455G GG genotype might have an increased risk of ALL. Further investigations are needed to confirm these associations.
doi:10.2147/OTT.S92259
PMCID: PMC4608596  PMID: 26491362
CYP1A1; ALL; AML; CML; polymorphism
6.  Analysis of a transgenic Oct4 enhancer reveals high fidelity long-range chromosomal interactions 
Scientific Reports  2015;5:14558.
Genome structure or nuclear organization has fascinated researchers investigating genome function. Recently, much effort has gone into defining relationships between specific genome structures and gene expression in pluripotent cells. We previously analyzed chromosomal interactions of the endogenous Oct4 distal enhancer in pluripotent cells. Here, we derive ES and iPS cells from a transgenic Oct4 distal enhancer reporter mouse. Using sonication-based Circularized Chromosome Conformation Capture (4C) coupled with next generation sequencing, we determined and compared the genome-wide interactome of the endogenous and transgenic Oct4 distal enhancers. Integrative genomic analysis indicated that the transgenic enhancer binds to a similar set of loci and shares similar key enrichment profiles with its endogenous counterpart. Both the endogenous and transgenic Oct4 enhancer interacting loci were enriched in the open nucleus compartment, which is associated with active histone marks (H3K4me1, H3K27ac, H3K4me3 and H3K9ac), active cis-regulatory sequences (DNA hypersensitivity sites (DHS)), 5-hydroxymethylcytosine (5-hmc), and early DNA replication domains. In addition, binding of some pluripotency-related transcription factors was consistently enriched in our 4C sites, and genes in those sites were generally more highly expressed. Overall, our work reveals critical features that may function in gene expression regulation in mouse pluripotent cells.
doi:10.1038/srep14558
PMCID: PMC4592970  PMID: 26435056
7.  A common genetic variation in CEBPE and acute lymphoblastic leukemia: a meta-analysis of the available evidence 
OncoTargets and therapy  2015;8:2443-2451.
Acute lymphoblastic leukemia (ALL) has been studied intensively for decades, but the details of its etiology and underlying mechanisms have yet to be fully elucidated. It is now generally acknowledged that genetic factors contribute greatly to the development of this disease. The gene encoding CCAAT/enhancer-binding protein ε (CEBPE) is involved in the development of leukemia, and in particular the rs2239633 single nucleotide polymorphism (SNP) of CEBPE. The association between rs2239633 and risk of ALL has been well studied, but remains unclear. Therefore, a meta-analysis was performed in this study to establish a more precise estimation of that relationship. A comprehensive literature search of the PubMed electronic database was conducted, and relevant studies published up to February 20, 2015 were selected for analysis. The references of the retrieved articles were also screened. The extracted data were analyzed statistically, and pooled odds ratios with 95% confidence intervals were calculated using Review Manager (version 5.2) to estimate the association strength. Finally, eleven studies were included in the meta-analysis. The pooled analyses revealed that rs2239633 was associated with an increased risk of childhood ALL in Caucasians under any contrast models (P<0.01). However, this SNP did not affect the risk of ALL in adulthood among Caucasians, or in childhood among East Asians. In conclusion, these findings confirm that the CEBPE rs2239633 SNP could be considered a good marker of pediatric ALL risk in Caucasians, but not in East Asians; it is not a good marker of adult ALL risk in Caucasians.
doi:10.2147/OTT.S89661
PMCID: PMC4571986  PMID: 26388693
CEBPE; rs2239633; ALL; risk; meta-analysis
8.  Effects of Nanowire Length and Surface Roughness on the Electrochemical Sensor Properties of Nafion-Free, Vertically Aligned Pt Nanowire Array Electrodes 
Sensors (Basel, Switzerland)  2015;15(9):22473-22489.
In this paper, vertically aligned Pt nanowire arrays (PtNWA) with different lengths and surface roughnesses were fabricated and their electrochemical performance toward hydrogen peroxide (H2O2) detection was studied. The nanowire arrays were synthesized by electroplating Pt in nanopores of anodic aluminum oxide (AAO) template. Different parameters, such as current density and deposition time, were precisely controlled to synthesize nanowires with different surface roughnesses and various lengths from 3 μm to 12 μm. The PtNWA electrodes showed better performance than the conventional electrodes modified by Pt nanowires randomly dispersed on the electrode surface. The results indicate that both the length and surface roughness can affect the sensing performance of vertically aligned Pt nanowire array electrodes. Generally, longer nanowires with rougher surfaces showed better electrochemical sensing performance. The 12 μm rough surface PtNWA presented the largest sensitivity (654 μA·mM−1·cm−2) among all the nanowires studied, and showed a limit of detection of 2.4 μM. The 12 μm rough surface PtNWA electrode also showed good anti-interference property from chemicals that are typically present in the biological samples such as ascorbic, uric acid, citric acid, and glucose. The sensing performance in real samples (river water) was tested and good recovery was observed. These Nafion-free, vertically aligned Pt nanowires with surface roughness control show great promise as versatile electrochemical sensors and biosensors.
doi:10.3390/s150922473
PMCID: PMC4610575  PMID: 26404303
Pt nanowire array; electrochemical biosensor; nanowire length; surface roughness; Nafion free; hydrogen peroxide
9.  Correlation of Resting Elbow Angle with Spasticity in Chronic Stroke Survivors 
Objective
To evaluate whether resting joint angle is indicative of severity of spasticity of the elbow flexors in chronic stroke survivors.
Methods
Seventeen hemiparetic stroke subjects (male: n = 13; female: n = 4; age: 37–89 years; 11 right and 6 left hemiplegia; averaged 54.8 months after stroke, ranging 12–107 months) participated in the study. The number of subjects with modified Ashworth scale score (MAS) = 0, 1, 1+, 2, and 3 was 3, 3, 5, 3, and 3, respectively. In a single experimental session, resting elbow joint angle, MAS, and Tardieu scale score (Tardieu R1) were measured. A customized motorized stretching device was used to stretch elbow flexors at 5, 50, and 100°/s, respectively. Biomechanical responses (peak reflex torque and reflex stiffness) of elbow flexors were quantified. Correlation analyses between clinical and biomechanical assessments were performed.
Results
Resting elbow joint angle showed a strong positive correlation with Tardieu R1 (r = 0.77, p < 0.01) and a very strong negative correlation with MAS (r = −0.89, p < 0.01). The resting angle also had strong correlations with biomechanical measures (r = −0.63 to −0.76, p < 0.01).
Conclusion
Our study provides experimental evidence for anecdotal observation that the resting elbow joint angle correlates with severity of spasticity in chronic stroke. Resting angle observation for spasticity assessment can and will be an easy, yet a valid way of spasticity estimation in clinical settings, particularly for small muscles or muscles which are not easily measurable by common clinical methods.
doi:10.3389/fneur.2015.00183
PMCID: PMC4549629  PMID: 26379617
stroke; spasticity; resting angle; MAS; Tardieu
10.  Cancer-associated fibroblasts provide a suitable microenvironment for tumor development and progression in oral tongue squamous cancer 
Background
Oral tongue squamous cell carcinoma (OTSCC) is still associated with a poor prognosis due to local recurrence and metastasis. Cancer-associated fibroblasts (CAFs) play an important role in the complex processes of cancer stroma interaction and tumorigenesis. This study aims to determine the role of CAFs in the development and progression of OTSCC.
Methods
Immunohistochemistry was performed to evaluate the frequency and distribution of CAFs in 178 paraffin specimens from patients with OTSCC. Immunofluorescence, a cell proliferation assay, flow cytometry, migration and invasion assays and western blot analysis were used to study the effects of CAFs and the corresponding conditioned medium (CM) on the proliferation and invasion of OTSCC cell lines.
Results
Statistical analysis showed a strong correlation between the frequency and distribution of CAFs and the clinicopathological characteristics of patients with cN0 OTSCC, including pathological stage (P = 0.001), T classification (P = 0.001), and N classification (P = 0.009). Survival analysis demonstrated a negative correlation of the frequency and distribution of CAFs with the overall survival and disease-free survival of patients with cN0 tongue squamous cell cancer (P = 0.009, 0.002, respectively); Cox regression analysis showed that the presence of CAFs (relative risk: 2.113, CI 1.461–3.015, P = 0.023) is an independent prognostic factor. A functional study demonstrated that CAFs and CM from CAFs could promote the growth, proliferation, mobility, invasion and even Epithelial Mesenchymal Transition (EMT) of OTSCC cells compared with NFs and CM from NFs.
Conclusions
CAFs were an independent prognostic factor for patients with OTSCC. Compared with NFs, CAFs and their CM have the ability to promote the growth, proliferation, metastasis and even EMT of OTSCC cells.
Electronic supplementary material
The online version of this article (doi:10.1186/s12967-015-0551-8) contains supplementary material, which is available to authorized users.
doi:10.1186/s12967-015-0551-8
PMCID: PMC4475624  PMID: 26094024
Oral tongue squamous cell cancer; Microenvironment; Cancer-associated fibroblast; Progression
11.  RPdb: a database of experimentally verified cellular reprogramming records 
Bioinformatics  2015;31(19):3237-3239.
Summary: Many cell lines can be reprogrammed to other cell lines by forced expression of a few transcription factors or by specifically designed culture methods, which have attracted a great interest in the field of regenerative medicine and stem cell research. Plenty of cell lines have been used to generate induced pluripotent stem cells (IPSCs) by expressing a group of genes and microRNAs. These IPSCs can differentiate into somatic cells to promote tissue regeneration. Similarly, many somatic cells can be directly reprogrammed to other cells without a stem cell state. All these findings are helpful in searching for new reprogramming methods and understanding the biological mechanism inside. However, to the best of our knowledge, there is still no database dedicated to integrating the reprogramming records. We built RPdb (cellular reprogramming database) to collect cellular reprogramming information and make it easy to access. All entries in RPdb are manually extracted from more than 2000 published articles, which is helpful for researchers in regenerative medicine and cell biology.
Availability and Implementation: RPdb is freely available on the web at http://bioinformatics.ustc.edu.cn/rpdb with all major browsers supported.
Contact: aoli@ustc.edu.cn
Supplementary information: Supplementary data are available at Bioinformatics online.
doi:10.1093/bioinformatics/btv331
PMCID: PMC4576693  PMID: 26026167
12.  Ligation-anchored PCR unveils immune repertoire of TCR-beta from whole blood 
BMC Biotechnology  2015;15:39.
Background
As one of the genetic mechanisms for adaptive immunity, V(D)J recombination generates an enormous repertoire of T-cell receptors (TCRs). With the development of high-throughput sequencing techniques, systematic exploration of V(D)J recombination becomes possible. Multiplex PCR has been previously developed to assay immune repertoire; however, the use of primer pools leads to inherent biases in target amplification. In our study, we developed a “single-primer" ligation-anchored PCR method that may amplify the repertoire with much less biases.
Results
By utilizing a universal primer paired with a single primer targeting the conserved constant region, we amplified TCR-beta (TRB) variable regions from total RNA extracted from blood. Next-generation sequencing libraries were then prepared for Illumina HiSeq 2500 sequencer, which generates 151-bp read length to cover the entire V(D)J recombination region. We evaluated this approach on blood samples from healthy donors and from patients with malignant and benign meningiomas. Mapping of sequencing data showed that 64% to 96% of mapped TCRV-containing reads belong to TRB subtype. An increased usage of specific V segments and V-J pairing were observed in malignant meningiomas samples. The CDR3 sequences of the expanded V-J pairs were distinct in each malignant individual, even for pairing of TRBV7-3 with TRBJ2-2 that showed increased usage in both cases.
Conclusions
We demonstrated the technical feasibility and effectiveness of ligation-anchored PCR approach in capturing the TCR-beta landscapes. Further development of this technology may enable a comprehensive delineation of immune repertoire, including other forms of TCRs as well as immunoglobulins.
Electronic supplementary material
The online version of this article (doi:10.1186/s12896-015-0153-9) contains supplementary material, which is available to authorized users.
doi:10.1186/s12896-015-0153-9
PMCID: PMC4446965  PMID: 26018870
Next-generation sequencing; Immune repertoire; Blood
13.  Complete Genome Sequence Analysis of Echovirus 24 Associated with Hand-Foot-and-Mouth Disease in China in 2012 
Genome Announcements  2015;3(1):e01456-14.
Echovirus 24 belongs to human enterovirus B species in the family Picornaviridae. Here, we report the whole-genome sequences of a novel complete genome sequence of a recombinant (echovirus 24) Echo 24 strain, PZ18/JS/2012, which was isolated from a patient with hand-foot-and-mouth disease in China.
doi:10.1128/genomeA.01456-14
PMCID: PMC4319619  PMID: 25657272
14.  Immunogenicity, Safety, and Lot Consistency of a Novel Inactivated Enterovirus 71 Vaccine in Chinese Children Aged 6 to 59 Months 
Clinical and Vaccine Immunology : CVI  2013;20(12):1805-1811.
The determination of lot-to-lot consistency in the manufacturing process is a mandatory step in the clinical development of the novel enterovirus 71 (EV71) vaccine. A phase III, randomized, placebo-controlled, double-blind trial assessed the lot consistency, immunogenicity, and safety of the EV71 vaccine in children aged 6 to 59 months. Healthy children (n = 1,400) received one of three lots of the EV71 vaccine containing 400 U of EV71 antigen or a placebo at days 0 and 28. Blood samples were collected before dose 1 and at 28 days after dose 2 (day 56) for an anti-EV71 neutralizing antibody (NTAb) assay. The geometric mean titer (GMT) and the seropositivity rates (with titers of ≥1:8) were compared at day 56. After each dose, the solicited injection site and general adverse events (AEs) were recorded for 7 days, and unsolicited AEs were recorded for 28 days. At day 56, the seropositivity rates ranged from 99.7% to 100% for the vaccine groups. The NTAb GMTs for the vaccine were 140.3 (95% confidence interval [CI], 117.8 to 167.1), 141.5 (95% CI, 118.0 to 169.6), and 146.6 (95% CI, 122.5 to 175.3). The two-sided 95% CI of the log difference in GMTs between the pairs of lots were between −0.176 and 0.176, therefore meeting the predefined equivalence criteria. The percentages of subjects reporting any injection site AEs, general AEs, or serious AEs were similar across the four vaccination groups. In conclusion, the demonstration of consistency between the manufacturing lots confirms for the purposes of clinical development the reliability of the EV71 vaccine production process. (This study has been registered at ClinicalTrials.gov under registration no. NCT01636245.)
doi:10.1128/CVI.00491-13
PMCID: PMC3889509  PMID: 24108780
15.  The Cross-Neutralizing Activity of Enterovirus 71 Subgenotype C4 Vaccines in Healthy Chinese Infants and Children 
PLoS ONE  2013;8(11):e79599.
Background
EV71 is one of major etiologic causes of hand-foot-mouth disease (HFMD) and leads to severe neurological complications in young children and infants. Recently inactivated EV71 vaccines have been developed by five manufactures and clinically show good safety and immunogenicity. However, the cross-neutralizing activity of these vaccines remains unclear, and is of particular interest because RNA recombination is seen more frequently in EV71 epidemics.
Methodology/Principal Findings
In this post-hoc study, sera from a subset of 119 infants and children in two clinical trials of EV71 subgenotype C4 vaccines (ClinicalTrials.gov Identifier: NCT01313715 and NCT01273246), were detected for neutralizing antibody (NTAb) titres with sera from infected patients as controls. Cytopathogenic effect method was employed to test NTAb against EV71 subgenotype B4, B5, C2, C4 and C5, which were prominent epidemic strains worldwide over the past decade. To validate the accuracy of the results, ELISpot assay was employed in parallel to detect NTAb in all the post-vaccine sera. After two-dose vaccination, 49 out of 53 participants in initially seronegative group and 52 out of 53 participants in initially seropositive group showed less than 4-fold differences in NTAb titers against five EV71 strains, whereas corresponding values among sera from pediatric patients recovering from EV71-induced HFMD and subclinically infected participants were 8/8 and 41/43, respectively. The geometric mean titers of participants against five subgenotypes EV71 all grew significantly after vaccinations, irrespective of the baseline NTAb titer. The relative fold increase in antibody titers (NTAb-FI) against B4, B5, C2, and C5 displayed a positive correlation to the NTAb-FI against C4.
Conclusions/Significance
The results demonstrated broad cross-neutralizing activity induced by two C4 EV71 vaccines in healthy Chinese infants and children. However, the degree of induced cross-protective immunity, and the potential escape evolution for EV71 still need to be monitored and researched in future for these new vaccines.
doi:10.1371/journal.pone.0079599
PMCID: PMC3834186  PMID: 24260259
16.  Characterizing Immunoglobulin Repertoire from Whole Blood by a Personal Genome Sequencer 
PLoS ONE  2013;8(9):e75294.
In human immune system, V(D)J recombination produces an enormously large repertoire of immunoglobulins (Ig) so that they can tackle different antigens from bacteria, viruses and tumor cells. Several studies have demonstrated the utility of next-generation sequencers such as Roche 454 and Illumina Genome Analyzer to characterize the repertoire of immunoglobulins. However, these techniques typically require separation of B cell population from whole blood and require a few weeks for running the sequencers, so it may not be practical to implement them in clinical settings. Recently, the Ion Torrent personal genome sequencer has emerged as a tabletop personal genome sequencer that can be operated in a time-efficient and cost-effective manner. In this study, we explored the technical feasibility to use multiplex PCR for amplifying V(D)J recombination for IgH, directly from whole blood, then sequence the amplicons by the Ion Torrent sequencer. The whole process including data generation and analysis can be completed in one day. We tested the method in a pilot study on patients with benign, atypical and malignant meningiomas. Despite the noisy data, we were able to compare the samples by their usage frequencies of the V segment, as well as their somatic hypermutation rates. In summary, our study suggested that it is technically feasible to perform clinical monitoring of V(D)J recombination within a day by personal genome sequencers.
doi:10.1371/journal.pone.0075294
PMCID: PMC3772839  PMID: 24058670
17.  Ionotropic glutamate receptors mediate OFF responses in light-adapted ON bipolar cells 
Vision research  2012;68:48-58.
Previous studies have suggested that photoreceptor synaptic inputs to depolarizing bipolar cells (DBCs or ON bipolar cells) are mediated by mGluR6 receptors and those to hyperpolarizing bipolar cells (HBCs or OFF bipolar cells) are mediated by AMPA/kainate receptors. Here we show that in addition to mGluR6 receptors which mediate the sign-inverting, depolarizing light responses, subpopulations of cone-dominated and rod/cone mixed DBCs use GluR4 AMPA receptors to generate a transient sign-preserving OFF response under light adapted conditions. These AMPA receptors are located at the basal junctions postsynaptic to rods and they are silent under dark-adapted conditions, as tonic glutamate release in darkness desensitizes these receptors. Light adaptation enhances rod-cone coupling and thus allows cone photocurrents with an abrupt OFF depolarization to enter the rods. The abrupt rod depolarization triggers glutamate activation of unoccupied AMPA receptors, resulting in a transient OFF response in DBCs. It has been widely accepted that the DNQX-sensitive, OFF transient responses in retinal amacrine cells and ganglion cells are mediated exclusively by HBCs. Our results suggests that this view needs revision as AMPA receptors in subpopulations of DBCs are likely to significantly contribute to the DNQX-sensitive OFF transient responses in light-adapted third- and higher-order visual neurons.
doi:10.1016/j.visres.2012.07.012
PMCID: PMC3733217  PMID: 22842089
Depolarizing bipolar cells; transient OFF responses; GluR4 receptors; mGluR6 receptors; 6,7-dinitroquinoxaline-2,3-dione (DNQX); cyclothiazide; dark and light adaptation
18.  Progress on the research and development of inactivated EV71 whole-virus vaccines 
Human Vaccines & Immunotherapeutics  2013;9(8):1701-1705.
The prevalence of diseases caused by EV71 infection has become a serious public health problem in the Western Pacific region. Due to a lack of effective treatment options, controlling EV71 epidemics has mainly focused on the research and development (R&D) of EV71 vaccines. Thus far, five organizations have completed pre-clinical studies focused on the development of inactivated EV71 whole-virus vaccines, including vaccine strain screening, process optimization, safety and immunogenicity evaluation, and are in different stages of clinical trials. Among these organizations, three companies in Mainland China [Beijing Vigoo Biological Co., Ltd. (Vigoo), Sinovac Biotech Ltd. (Sinovac) and Institute of Medical Biology, Chinese Academy of Medical Science (CAMS)] have recently completed Phase III trials for the vaccines they developed. In addition, the other two vaccines, developed by National Health Research Institutes (NHRI) of Taiwan and Inviragen Pte., Ltd (Inviragen), of Singapore, have also completed Phase I clinical trials. Published clinical trial results indicate that the inactivated EV71 vaccines have good safety and immunogenicity in the target population (infants) and confer a relatively high rate of protection against EV71 infection-related diseases. The results of clinical trials suggest a promising future for the clinical use of EV71 vaccines. Here, we review and highlight the recent progress on the R&D of inactivated EV71 whole-virus vaccines.
doi:10.4161/hv.24949
PMCID: PMC3906269  PMID: 23744508
enterovirus 71; hand-foot-mouth diseases (HFMD); inactivated whole-virus vaccine; immunogenicity; protective effect; standard
19.  Development and Evaluation of a Pseudovirus-Luciferase Assay for Rapid and Quantitative Detection of Neutralizing Antibodies against Enterovirus 71 
PLoS ONE  2013;8(6):e64116.
The level of neutralizing antibodies (NtAb) induced by vaccine inoculation is an important endpoint to evaluate the efficacy of EV71 vaccine. In order to evaluate the efficacy of EV71 vaccine, here, we reported the development of a novel pseudovirus system expression firefly luciferase (PVLA) for the quantitative measurement of NtAb. We first evaluated and validated the sensitivity and specificity of the PVLA method. A total of 326 serum samples from an epidemiological survey and 144 serum specimens from 3 clinical trials of EV71 vaccines were used, and the level of each specimen's neutralizing antibodies (NtAb) was measured in parallel using both the conventional CPE-based and PVLA-based assay. Against the standard neutralization assay based on the inhibition of the cytopathic effect (CPE), the sensitivity and specificity of the PVLA method are 98% and 96%, respectively. Then, we tested the potential interference of NtAb against hepatitis A virus, Polio-I, Polio-II, and Polio-III standard antisera (WHO) and goat anti-G10/CA16 serum, the PVLA based assay showed no cross-reactivity with NtAb against other specific sera. Importantly, unlike CPE based method, no live replication-competent EV71 is used during the measurement. Taken together, PVLA is a rapid and specific assay with higher sensitivity and accuracy. It could serve as a valuable tool in assessing the efficacy of EV71 vaccines in clinical trials and disease surveillance in epidemiology studies.
doi:10.1371/journal.pone.0064116
PMCID: PMC3673970  PMID: 23755115
20.  Investigation of sit-to-stand and stand-to-sit in an above knee amputee 
the objective of this pilot study is twofold: 1) to extract key factors/features in sit-to-stand and stand-to-sit (STS) performed by an above knee (AK) amputee; 2) to propose a convenient way to quantify symmetry. One male unilateral transfemoral amputee participated in the pilot study. The subject was instructed to rise in a comfortable and natural manner and conduct a series of sit-to-stand, stand-to-sit. We simultaneously measured kinematics, kinetics and muscle activities. Principal component analysis (PCA) was used to reduce the dimension and identify modes and a convenient index of STS symmetry (slope of the major axis of the error ellipse) is proposed using the insole pressure sensors. Based on the preliminary results it is recommended that kinematics and kinetics in both the sagittal and frontal planes be considered for an AK amputee performing STS. The information might be useful for further research on amputee STS.
doi:10.1109/IEMBS.2011.6091712
PMCID: PMC3667594  PMID: 22256034
21.  Comparative analysis of 4C-Seq data generated from enzyme-based and sonication-based methods 
BMC Genomics  2013;14:345.
Background
Circular chromosome conformation capture, when coupled with next-generation sequencing (4C-Seq), can be used to identify genome-wide interaction of a given locus (a “bait” sequence) with all of its interacting partners. Conventional 4C approaches used restriction enzyme digestion to fragment chromatin, and recently sonication approach was also applied for this purpose. However, bioinformatics pipelines for analyzing sonication-based 4C-Seq data are not well developed. In addition, data consistency as well as similarity between the two methods has not been explored previously. Here we present a comparative analysis of 4C-Seq data generated by both methods, using an enhancer element of Pou5f1 gene in mouse embryonic stem (ES) cells.
Results
From biological replicates, we found good correlation (r>0.6) for inter-chromosomal interactions identified in either enzyme or sonication method. Compared to enzyme approach, sonication method generated less distal intra-chromosomal interactions, possibly due to the difference in chromatin fragmentation. From all mapped interactions, we further applied statistical models to identify enriched interacting regions. Interestingly, data generated from the two methods showed 30% overlap of the reproducible interacting regions. The interacting sites in the reproducible regions from both methods are similarly enriched with active histone marks. In addition, the interacting sites identified from sonication-based data are enriched with ChIP-Seq signals of transcription factors Oct4, Klf4, Esrrb, Tcfcp2i1, and Zfx that are critical for reprogramming and pluripotency.
Conclusions
Both enzyme-based and sonication-based 4C-Seq methods are valuable tools to explore long-range chromosomal interactions. Due to the nature of sonication-based method, correlation analysis of the 4C interactions with transcription factor binding should be more straightforward.
doi:10.1186/1471-2164-14-345
PMCID: PMC3679908  PMID: 23705979
Circular chromosome conformation capture; Sonication; 4C-Seq; Bioinformatics; Next-generation sequencing
22.  Use of Multidimensional Fluorescence Resonance Energy Transfer To Establish the Orientation of Cholecystokinin Docked at the Type A Cholecystokinin Receptor† 
Biochemistry  2008;47(36):9574-9581.
Fluorescence resonance energy transfer (FRET) represents a powerful tool to establish relative distances between donor and acceptor fluorophores. By utilizing several donors situated in distinct positions within a docked full agonist ligand and several acceptors distributed at distinct sites within its receptor, multiple interdependent dimensions can be determined. These can provide a unique method to establish or confirm three-dimensional structure of the molecular complex. In this work, we have utilized full agonist analogues of cholecystokinin (CCK) with Aladan distributed throughout the pharmacophore in positions 24, 29, and 33, along with receptor constructs derivatized with Alexa546 at positions 94, 102, 204, and 341 in the helical bundle and first, second, and third extracellular loops, respectively. These provided 12 FRET distances to overlay on working models of the CCK-occupied receptor. These established that the carboxyl terminus of CCK resides at the external surface of the lipid bilayer, adjacent to the receptor amino-terminal tail, rather than being inserted into the helical bundle. They also provide important experimentally derived constraints for understanding spatial relationships between the docked ligand and the flexible extracellular loop regions. Multidimensional FRET provides a new independent method to establish and refine structural insights into ligand–receptor complexes.
doi:10.1021/bi800734w
PMCID: PMC3648886  PMID: 18700727
23.  The interactomes of POU5F1 and SOX2 enhancers in human embryonic stem cells 
Scientific Reports  2013;3:1588.
The genes POU5F1 and SOX2 are critical for pluripotency and reprogramming, yet the chromosomal organization around these genes remains poorly understood. We assayed long-range chromosomal interactions on putative enhancers of POU5F1 and SOX2 genes in human embryonic stem cells (hESCs) using 4C-Seq technique. We discovered that their frequent interacting regions mainly overlap with early DNA replication domains. The interactomes are associated with active histone marks and enriched with 5-hydroxymethylcytosine sites. In hESCs, genes within the interactomes have elevated expression. Additionally, some genes associated with the POU5F1 enhancer contribute to pluripotency. Binding sites for multiple DNA binding proteins, including ATF3, CTCF, GABPA, JUND, NANOG, RAD21 and YY1, are enriched in both interactomes. The RARG locus, frequently interacting with the POU5F1 locus, has abundant RAD21 binding sites co-localized with other protein binding sites. Thus the interactomes of these two pluripotency genes could be an important part of the regulatory network in hESCs.
doi:10.1038/srep01588
PMCID: PMC4070749  PMID: 23549118
24.  DNA Methylation in the Malignant Transformation of Meningiomas 
PLoS ONE  2013;8(1):e54114.
Meningiomas are central nervous system tumors that originate from the meningeal coverings of the brain and spinal cord. Most meningiomas are pathologically benign or atypical, but 3–5% display malignant features. Despite previous studies on benign and atypical meningiomas, the key molecular pathways involved in malignant transformation remain to be determined, as does the extent of epigenetic alteration in malignant meningiomas. In this study, we explored the landscape of DNA methylation in ten benign, five atypical and four malignant meningiomas. Compared to the benign tumors, the atypical and malignant meningiomas demonstrate increased global DNA hypomethylation. Clustering analysis readily separates malignant from atypical and benign tumors, implicating that DNA methylation patterns may serve as diagnostic biomarkers for malignancy. Genes with hypermethylated CpG islands in malignant meningiomas (such as HOXA6 and HOXA9) tend to coincide with the binding sites of polycomb repressive complexes (PRC) in early developmental stages. Most genes with hypermethylated CpG islands at promoters are suppressed in malignant and benign meningiomas, suggesting the switching of gene silencing machinery from PRC binding to DNA methylation in malignant meningiomas. One exception is the MAL2 gene that is highly expressed in benign group and silenced in malignant group, representing de novo gene silencing induced by DNA methylation. In summary, our results suggest that malignant meningiomas have distinct DNA methylation patterns compared to their benign and atypical counterparts, and that the differentially methylated genes may serve as diagnostic biomarkers or candidate causal genes for malignant transformation.
doi:10.1371/journal.pone.0054114
PMCID: PMC3551961  PMID: 23349797
25.  Recent Structural and Mechanistic Insights into Endplate Acetylcholine Receptors 
Voluntary movement mediated by skeletal muscle relies on endplate acetylcholine receptors (AChR) to detect nerve-released ACh and depolarize themuscle fiber. Recent structural and mechanistic studies of the endplate AChR have catalyzed a leap in our understanding of the molecular steps in this chemical-to-electrical transduction process. Studies of acetylcholine binding protein (AChBP) give insight into ACh recognition, the first step in activation of the AChR. An atomic structural model of the Torpedo AChR at a resolution of 0.4 nm, together with single-ion channel recording methods, allow tracing of the link between the agonist binding event and gating of the ion channel, as well as determination of how the channel moves when it opens to allow flow of cations. Structural models of the human AChR enable precise mapping of disease-causing mutations, while studies of the speed with which single AChR channels open and close cast light on pathogenic mechanisms.
doi:10.1196/annals.1405.041
PMCID: PMC3478106  PMID: 18567853
acetylcholine receptor; acetylcholine binding protein; agonist recognition; binding-gating coupling mechanism; congenital myasthenic syndrome

Results 1-25 (45)