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1.  Acupuncture at Houxi (SI 3) acupoint for acute neck pain caused by stiff neck: study protocol for a pilot randomised controlled trial 
BMJ Open  2014;4(12):e006236.
Introduction
The use of acupuncture has been suggested for the treatment of acute neck pain caused by stiff neck in China. However, current evidence is insufficient to draw any conclusions about its efficacy. Therefore this pilot study was designed to evaluate the feasibility and efficacy of acupuncture at the Houxi (SI3) acupoint for treatment of acute neck pain.
Methods/analysis
This pilot study will be a two-parallel-group, assessor-blinded, randomised controlled trial. Thirty-six stiff neck participants with acute neck pain will be recruited and randomly divided into two groups in a 1:1 ratio. Participants in the control group will receive massage on the local neck region (5 min each session, three times a day for 3 days). In addition to massage, patients in the treatment group will receive acupuncture (one session a day for 3 days). Measures will be taken at 0, 3 and 15 days. The primary outcome is the Northwick Park Neck Pain Questionnaire (NPQ). The secondary outcome is the Short Form of the McGill Pain Questionnaire (SF-MPQ).
Ethics/dissemination
The protocol for this pilot randomised clinical trial has undergone ethics scrutiny and been approved by the ethics review boards of the First Affiliated Hospital of Heilongjiang University of Traditional Chinese Medicine (Permission number: HZYLL201303502). The findings of this study will provide important clinical evidence on the feasibility and efficacy of acupuncture treatment for stiff neck patients with acute neck pain. In addition, it will explore the feasibility of further acupuncture research.
Trial registration number
ChiCTR-TRC-13003911.
doi:10.1136/bmjopen-2014-006236
PMCID: PMC4275681  PMID: 25537784
acupuncture; acute neck pain; stiff neck; randomized controlled trial
2.  The Ubiquitin Ligase Stub1 Negatively Modulates Regulatory T cell Suppressive Activity by Promoting Degradation of the Transcription Factor Foxp3 
Immunity  2013;39(2):10.1016/j.immuni.2013.08.006.
SUMMARY
Regulatory T (Treg) cells suppress inflammatory immune responses and autoimmunity caused by self-reactive T cells. The key Treg cell transcription factor Foxp3 is downregulated during inflammation to allow for the acquisition of effector T cell-like functions. Here, we demonstrate that stress signals elicited by proinflammatory cytokines and lipopolysaccharide lead to the degradation of Foxp3 through the action of the E3 ubiquitin ligase Stub1. Stub1 interacted with Foxp3 to promote its K48-linked polyubiquitination in an Hsp70-dependent manner. Knockdown of endogenous Stub1 or Hsp70 prevented Foxp3 degradation. Furthermore, the overexpression of Stub1 in Treg cells abrogated their ability to suppress inflammatory immune responses in vitro and in vivo, and conferred a T helper 1 (Th1) cell-like phenotype. Our results demonstrate the critical role of the stress-activated Stub1-Hsp70 complex in promoting Treg cell inactivation, thus providing a potential therapeutic target for the intervention against autoimmune disease, infection and cancer.
doi:10.1016/j.immuni.2013.08.006
PMCID: PMC3817295  PMID: 23973223
3.  Identification and Analysis of Differential miRNAs in PK-15 Cells after Foot-and-Mouth Disease Virus Infection 
PLoS ONE  2014;9(3):e90865.
The alterations of MicroRNAs(miRNAs) in host cell after foot-and-mouth disease virus (FMDV) infection is still obscure. To increase our understanding of the pathogenesis of FMDV at the post-transcriptional regulation level, Solexa high-throu MicroRNAs (miRNAs) play an important role both in the post-transcriptional regulation of gene expression and host-virus interactions. Despite investigations of miRNA expression ghput sequencing and bioinformatic tools were used to identify differentially expressed miRNAs and analyze their functions during FMDV infection of PK-15cells. Results indicated that 9,165,674 and 9,230,378 clean reads were obtained, with 172 known and 72 novel miRNAs differently expressed in infected and uninfected groups respectively. Some of differently expressed miRNAs were validated using stem-loop real-time quantitative RT-PCR. The GO annotation and KEGG pathway analysis for target genes revealed that differently expressed miRNAs were involved in immune response and cell death pathways.
doi:10.1371/journal.pone.0090865
PMCID: PMC3968000  PMID: 24675746
4.  Activation of circulated immune cells and inflammatory immune adherence are involved in the whole process of acute venous thrombosis 
Objective: To investigate localization and distribution of integrin subunit β1, β2 and β3 and morphological changes of ligand-recepter binding in thrombi of acute pulmonary embolism (PE) patients and explore activation of circulated immune cells, inflammatory immune adherence and coagulation response in acute venous thrombosis. Methods: Thrombi were collected from patients with acute PE. Immunohistochemistry was done to detect the expression and distribution of integrin β1, β2 and β3 in cells within thrombi, and ligands of integrin subunit β1, β2 and β3 were also determined by immunohistochemistry within the thrombi. Results: 1) Acute venous thrombi were red thrombi composed of skeletons and filamentous mesh containing large amounts of red blood cells and white blood cells; 2) Integrin subunit β1, β2 and β3 were expressed on lymphocytes, neutrophils and platelets; 3) No expression of integrin β1 ligands: Laminin, Fibronectin, Collagen I or Collagen-II on lymphocytes; integrin β2 ligands including ICAM, factor X and iC3b are distributed on neutrophils, and ligand fibrinogen bound to neutrophils; integrin β3 was expressed on platelets which form the skeleton of thrombi and bound to fibrinogen to construct mesh structure; 4) Factor Xa was expressed on the filamentous mesh; 5) Filamentous mesh was fully filled with red blood cell dominant blood cells. Conclusion: Acute venous thrombosis is an activation process of circulated lymphocytes, neutrophils and platelets mainly, and a whole process including integrin subunit β2 and β3 binding with their ligands. Activation of immune cells, inflammatory immune adherence and coagulation response are involved in the acute venous thrombosis.
PMCID: PMC3992394  PMID: 24753749
Pulmonary embolism; venous thrombosis; integrin
5.  Stem Cell-Based Therapies for Ischemic Stroke 
BioMed Research International  2014;2014:468748.
In recent years, stem cell-based approaches have attracted more attention from scientists and clinicians due to their possible therapeutical effect on stroke. Animal studies have demonstrated that the beneficial effects of stem cells including embryonic stem cells (ESCs), inducible pluripotent stem cells (iPSCs), neural stem cells (NSCs), and mesenchymal stem cell (MSCs) might be due to cell replacement, neuroprotection, endogenous neurogenesis, angiogenesis, and modulation on inflammation and immune response. Although several clinical studies have shown the high efficiency and safety of stem cell in stroke management, mainly MSCs, some issues regarding to cell homing, survival, tracking, safety, and optimal cell transplantation protocol, such as cell dose and time window, should be addressed. Undoubtably, stem cell-based gene therapy represents a novel potential therapeutic strategy for stroke in future.
doi:10.1155/2014/468748
PMCID: PMC3955655  PMID: 24719869
6.  MiR-19a is correlated with prognosis and apoptosis of laryngeal squamous cell carcinoma by regulating TIMP-2 expression 
MiRNAs are small, noncoding RNA molecules that act as posttranscriptional regulators of gene expression and function as important regulators in cancer-related processes. The miR-19a is overexpressed in various cancers and has been causally related to cellular proliferation and growth. To determine whether miR-19a plays a role in laryngeal squamous cell carcinoma (LSCC), we used quantitative real time PCR to detect miR-19a expression in LSCC tissues. We found that miR-19a is overexpressed in LSCC and correlated with neck nodal metastasis, poor differentiation and advanced stage. Statistical analysis suggests that higher level of miR-19a was associated with reduced overall survival. In vitro functional study showed that inhibition of miR-19a by antisense oligonucleotides (ASO) led to apoptosis and reduction of cell proliferation in LSCC cells. Furthermore, growth of LSCC xenograft tumors was significantly suppressed by repeated injection of ASO-miR-19a lentivirus. The TUNEL stain and transmission electron microscopy also detected increased apoptotic cells in ASO-miR-19a treated LSCC xenografts. In addition, both realtime PCR and western blot showed ASO-miR-19a can upregulate TIMP-2 expression and this suggests miR-19a is related with TIMP-2 pathway in LSCC cells. Taken together, these data suggest that miR-19a plays an oncogenic role in the progression of LSCC, and may serve as a biomarker or therapeutic target for patients with LSCC.
PMCID: PMC3885460  PMID: 24427326
miR-19a; LSCC; TIMP-2; apoptosis
7.  Association between Mannose-Binding Lectin Gene Polymorphisms and Hepatitis B Virus Infection: A Meta-Analysis 
PLoS ONE  2013;8(10):e75371.
Objective
The results of studies on the relation between Mannose-binding lectin gene (mbl2) polymorphism and HBV infection were contradictory and inconclusive. In order to shed a light on these inconsistent findings and to clarify the role of mbl2 polymorphisms in susceptibility or progression of chronic hepatitis B (CHB), a meta-analysis was performed.
Methods
PubMed and Embase were searched for available articles. A meta-analysis was performed to examine the association between mbl2 polymorphisms and chronicity or progression of hepatitis B infection. Odds ratio (OR) and its 95% confidence interval (CI) served as indexes.
Results
A total of 17 eligible studies were involved, including 2151 healthy controls (HC), 1293 spontaneous recovered (SR) patients with acute infection, 2337cases with chronic hepatitis B (CHB) and 554 cases with progressive hepatitis B. There was no evidence of significant association between mbl2 exon1 polymorphisms and CHB risk in any genetic model or pairwise comparisons when compared with HC group or SR group. In the stratified analysis of ethnic groups, also no obvious relation between mbl2 polymorphism and CHB risk was identified. There was still no significant association between the complete mbl2 genotypic profile (including both the exon1 and the promoter gene) polymorphisms and CHB risk, as compared with SR group. However, it was found that there was an association between the mbl2 AO/OO genotype and severe hepatitis B (SHB) or liver cirrhosis (LC) (LC vs. HC:OR=3.66, 95%CI, 2.38-5.63; SHB vs. HC, OR=3.88, 95%CI, 2.26–6.64), but there was no relationship between the mbl2 AO/OO genotype and hepatocellular carcinoma (HCC) (OR=1.26, 95%CI, 0.82-1.94).
Conclusion
The present meta-analysis indicated that mbl2 exon1 polymorphisms might not significantly associate with chronicity of HBV infection, but might be significantly related to the progressive HBV such as SHB and LC.
doi:10.1371/journal.pone.0075371
PMCID: PMC3792921  PMID: 24116040
8.  Extending the Serum Half-Life of G-CSF via Fusion with the Domain III of Human Serum Albumin 
BioMed Research International  2013;2013:107238.
Protein fusion technology is one of the most commonly used methods to extend the half-life of therapeutic proteins. In this study, in order to prolong the half-life of Granulocyte colony stimulating factor (G-CSF), the domain III of human serum albumin (3DHSA) was genetically fused to the N-terminal of G-CSF. The 3DHSA-G-CSF fusion gene was cloned into pPICZαA along with the open reading frame of the α-factor signal under the control of the AOX1 promoter. The recombinant expression vector was transformed into Pichia pastoris GS115, and the recombinant strains were screened by SDS-PAGE. As expected, the 3DHSA-G-CSF showed high binding affinity with HSA antibody and G-CSF antibody, and the natural N-terminal of 3DHSA was detected by N-terminal sequencing. The bioactivity and pharmacokinetic studies of 3DHSA-G-CSF were respectively determined using neutropenia model mice and human G-CSF ELISA kit. The results demonstrated that 3DHSA-G-CSF has the ability to increase the peripheral white blood cell (WBC) counts of neutropenia model mice, and the half-life of 3DHSA-G-CSF is longer than that of native G-CSF. In conclusion, 3DHSA can be used to extend the half-life of G-CSF.
doi:10.1155/2013/107238
PMCID: PMC3787585  PMID: 24151579
10.  Angioimmunoblastic T-cell lymphoma-associated pure red cell aplasia with abdominal pain 
Angioimmunoblastic T-cell lymphoma (AITL) is a unique type of peripheral T-cell lymphoma with a constellation of clinical symptoms and signs, including weight loss, fever, chills, anemia, skin rash, hepatosplenomegaly, lymphadenopathy, thrombocytopenia and polyclonal hypergammaglobulinemia. The histological features of AITL are also distinctive. Pure red cell aplasia is a bone marrow failure characterized by progressive normocytic anemia and reticulocytopenia without leucopenia or thrombocytopenia. However, AITL with abdominal pain and pure red cell aplasia has rarely been reported. Here, we report a rare case of AITL-associated pure red cell aplasia with abdominal pain. The diagnosis was verified by a biopsy of the enlarged abdominal lymph nodes with immunohistochemical staining.
doi:10.5306/wjco.v4.i3.75
PMCID: PMC3708066  PMID: 23936760
Angioimmunoblastic T-cell lymphoma; Anemia; Pure red cell aplasia; Abdominal pain
11.  The Acute Effects of Grape Polyphenols Supplementation on Endothelial Function in Adults: Meta-Analyses of Controlled Trials 
PLoS ONE  2013;8(7):e69818.
Background
The acute effects of grape polyphenols on endothelial function in adults are inconsistent. Here, we performed meta-analyses to determine these acute effects as measured by flow-mediated dilation (FMD).
Methods
Trials were searched in PubMed, Embase and the Cochrane Library database. Summary estimates of weighted mean differences (WMDs) and 95% CIs were obtained by using random-effects models. Meta-regression and subgroup analyses were performed to identify the source of heterogeneity. The protocol details of our meta-analysis have been submitted to the PROSPERO register and our registration number is CRD42013004157.
Results
Nine studies were included in the present meta-analyses. The results showed that the FMD level was significantly increased in the initial 120 min after intake of grape polyphenols as compared with controls. Meta-regression and subgroup analyses were performed and showed that a health status was the main effect modifier of the significant heterogeneity. Subgroups indicated that intake of grape polyphenols could significantly increase FMD in healthy subjects, and the increased FMD appeared to be more obviously in subjects with high cardiovascular risk factors. Moreover, the peak effect of grape polyphenols on FMD in healthy subjects was found 30 min after ingestion, which was different from the effect in subjects with high cardiovascular risk factors, in whom the peak effect was found 60 min after ingestion.
Conclusions
Endothelial function can be significantly improved in healthy adults in the initial 2 h after intake of grape polyphenols. The acute effect of grape polyphenols on endothelial function may be more significant but the peak effect is delayed in subjects with a smoking history or coronary heart disease as compared with the healthy subjects.
doi:10.1371/journal.pone.0069818
PMCID: PMC3722169  PMID: 23894543
12.  Elevational Gradient in Species Richness Pattern of Epigaeic Beetles and Underlying Mechanisms at East Slope of Balang Mountain in Southwestern China 
PLoS ONE  2013;8(7):e69177.
We report on the species richness patterns of epigaeic beetles (Coleoptera: Carabidae and Staphylinidae) along a subtropical elevational gradient of Balang Mountain, southwestern China. We tested the roles of environmental factors (e.g. temperature, area and litter cover) and direct biotic interactions (e.g. foods and antagonists) that shape elevational diversity gradients. Beetles were sampled at 19 sites using pitfall traps along the studied elevational gradient ranging from 1500 m–4000 m during the 2004 growing season. A total of 74416 specimens representing 260 species were recorded. Species richness of epigaeic beetles and two families showed unimodal patterns along the elevational gradient, peaking at mid-elevations (c. 2535 m), and the ranges of most beetle species were narrow along the gradient. The potential correlates of both species richness and environmental variables were examined using linear and second order polynomial regressions. The results showed that temperature, area and litter cover had strong explanatory power of beetle species richness for nearly all richness patterns, of beetles as a whole and of Carabidae and Staphylinidae, but the density of antagonists was associated with species richness of Carabidae only. Multiple regression analyses suggested that the three environmental factors combined contributed most to richness patterns for most taxa. The results suggest that environmental factors associated with temperature, area and habitat heterogeneity could account for most variation in richness pattern of epigaeic beetles. Additionally, the mid-elevation peaks and the small range size of most species indicate that conservation efforts should give attention to the entire gradient rather than just mid-elevations.
doi:10.1371/journal.pone.0069177
PMCID: PMC3715450  PMID: 23874906
13.  Characterization of Asia 1 sdAb from Camels Bactrianus (C. bactrianus) and Conjugation with Quantum Dots for Imaging FMDV in BHK-21 Cells 
PLoS ONE  2013;8(5):e63500.
Foot-and-mouth disease (FMD), caused by FMD virus (FMDV), is a highly contagious viral disease affecting cloven-hoofed animals. Camelids have a unique immunoglobulin profile, with the smallest functional heavy-chain antibodies (sdAb or VHH) naturally devoid of light chains with antigen-binding capacity. We screened and characterized five sdAbs against FMDV by immunized library from C. bactrianus with Asia 1 virus-like particles (VLPs). Three of five recombinant sdAbs were stably expressed in E.coli, remained highly soluble, and were serotype-specific for VP1 protein of FMDV Asia 1 by ELISA. These failed to completely neutralize the Asia 1 virus. According to the KD value of binding affinity to three sdAbs, which ranged from 0.44 to 0.71 nm by SPR, sdAb-C6 was selected and conjugated with Zn/CdSe quantum dots (QDs) to form a QDs-C6 probe, which was used to trace and image the subcellular location of FMDV in BHK-21 cells. The results show that FMD virions were observed from 3 h.p.i., and most of virions were distributed on one side of the nucleus in the cytoplasm. We demonstrate the utility of sdAbs as functionalized QDs are powerful tools for FMDV research.
doi:10.1371/journal.pone.0063500
PMCID: PMC3667858  PMID: 23737944
14.  Comparison of a loop-mediated isothermal amplification for orf virus with quantitative real-time PCR 
Virology Journal  2013;10:138.
Background
Orf virus (ORFV) causes orf (also known as contagious ecthyma or contagious papular dermatitis), a severe infectious skin disease in goats, sheep and other ruminants. Therefore, a rapid, highly specific and accurate method for the diagnosis of ORFV infections is essential to ensure that the appropriate treatments are administered and to reduce economic losses.
Methods
A loop-mediated isothermal amplification (LAMP) assay based on the identification of the F1L gene was developed for the specific detection of ORFV infections. The sensitivity and specificity of the LAMP assay were evaluated, and the effectiveness of this method was compared with that of real-time PCR.
Results
The sensitivity of this assay was determined to be 10 copies of a standard plasmid. Furthermore, no cross-reactivity was found with either capripox virus or FMDV. The LAMP and real-time PCR assays were both able to detect intracutaneous- and cohabitation-infection samples, with a concordance of 97.83%. LAMP demonstrated a sensitivity of 89.13%.
Conclusion
The LAMP assay is a highly efficient and practical method for detecting ORFV infection. This LAMP method shows great potential for monitoring the prevalence of orf, and it could prove to be a powerful supplemental tool for current diagnostic methods.
doi:10.1186/1743-422X-10-138
PMCID: PMC3651318  PMID: 23634981
Orf virus; Loop-mediated isothermal amplification (LAMP); Real-time PCR
15.  Plasma miR-200c and miR-18a as potential biomarkers for the detection of colorectal carcinoma 
Molecular and Clinical Oncology  2013;1(2):379-384.
It has been demonstrated that there are abundant stable microRNAs (miRNAs) in plasma/serum, which can be detected and are potentially disease-specific. The aim of this study was to investigate whether plasma miR-200c and miR-18a can be used as biomarkers for the detection of colorectal carcinoma (CRC). This study was divided into three parts: i) confirmation of higher miR-200c and miR-18a levels in primary CRC tissues compared to normal colorectal tissues; ii) evaluation of plasma miR-200c and miR-18a expression by comparing 78 patients with 86 healthy volunteers and iii) comparison of miR-200c and miR-18a levels in paired pre-and post-operative plasma in cancer patients who underwent curative CRC resection. Results showed that the expression of miR-200c and miR-18a was significantly higher in CRC compared to normal tissues. The plasma levels of miR-200c and miR-18a were significantly higher in CRC patients compared to controls. miR-200c yielded an area under the receiver-operating characteristics (ROC) curve (AUC) of 0.749 and miR-18a yielded an AUC of 0.804 when distinguishing CRC patients from the controls. Combined ROC analyses using the two miRNAs revealed an elevated AUC of 0.839 with 84.6% sensitivity and 75.6% specificity in discriminating CRC. Plasma levels of miR-200c and miR-18a were significantly lower in post-operative compared to pre-operative samples. The results of this study suggest that plasma miR-200c and miR-18a are significantly elevated in the plasma of CRC patients and that they may serve as non-invasive molecular markers for CRC screening.
doi:10.3892/mco.2013.61
PMCID: PMC3915707  PMID: 24649179
miR-200c; miR-18a; colorectal cancer; plasma; biomarker
16.  Pathogenic characteristics of three genotype II porcine reproductive and respiratory syndrome viruses isolated from China 
Virology Journal  2013;10:7.
Background
We examined differences in pathogenicity in pigs from China that had been experimentally infected with porcine reproductive and respiratory syndrome virus (PRRSV).
Methods
We compared pathogenic characteristics of a field isolate (GX-1/2008F), two PRRSV isolates (HN-1/2008, YN-1/2008) propagated in cells, and GX-1/2008F that had been propagated in cells (GX-1/2008). The clinical courses, along with humoral and cell-mediated responses, were monitored for 21 days post-infection (DPI). Animals were sacrificed and tissue samples used for gross pathological, histopathological and ultrastructure examination.
Results
At 2–3 DPI, animals infected with cell-propagated viruses exhibited signs of coughing, anorexia and fever. However their rectal temperature did not exceed 40.5°C. Viremia was detectable as early as 3 DPI in animals infected with HN-1/2008 and YN-1/2008. Animals inoculated with GX-1/2008F displayed clinical signs at 6 DPI; the rectal temperature of two animals in this group exceeded 41.0°C, with viremia first detected at 7 DPI. Seroconversion for all challenged pigs, except those infected with GX-1/2008, was seen as early as 7 DPI. All of these pigs had fully seroconverted by 11 DPI. All animals challenged with GX-1/2008 remained seronegative until the end of the experiment. Innate immunity was inhibited, with levels of IFN-α and IL-1 not significantly different between control and infected animals. The cytokines IFN-γ and IL-6 transiently increased during acute infection. All virus strains caused gross lesions including multifocal interstitial pneumonia and hyperplasia of lymph nodes. Inflammation of the stomach and small intestine was also observed. Lesions in the group infected with GX-1/2008F were more serious than in other groups. Transmission electron microscopy revealed that alveolar macrophages, plasmacytes and lymphocytes had fractured cytomembranes, and hepatocytes had disrupted organelles and swollen mitochondria.
Conclusions
The pathogenicity of the PRRSV field isolate became attenuated when propagated in MARC-145 cells. Tissue tropism of highly pathogenic strains prevailing in China was altered compared with classical PRRSV strains. The observed damage to immune cells and modulation of cytokine production could be mechanisms that PRRSV employs to evade host immune responses.
doi:10.1186/1743-422X-10-7
PMCID: PMC3616938  PMID: 23282224
Immune response; Pathogenic characteristics; Porcine reproductive and respiratory syndrome virus
17.  Construction and expression of a recombinant eukaryotic expression plasmid containing the preS1-preS2-S genes of hepatitis B virus and the granulocyte-macrophage colony stimulating factor gene: A study of its immunomodulatory effects 
Biomedical Reports  2012;1(2):251-256.
A total of 10–20% of the population remains unresponsive or weakly responsive to hepatitis B vaccine, which is composed of hepatitis B surface antigen HBsAg (S protein). Therefore, it is necessary to develop a hepatitis B vaccine with a better penetrating and responsive rate. In the present study, a plasmid pVAX1-L-GM was constructed and its immunomodulatory effect of as hepatitis B virus (HBV) DNA vaccine was analyzed through the immunization of BALB/c mice. Immune responses were measured after immunization by anti-HBsAg, proliferation of splenocytes, the number of CD4+ and CD8+ molecules, CTL cytotoxicity, cytokines of IFN-γ and IL-2 secretion assays. Following the immunization, mice in the pVAX1-L-GM group produced antibody 2 weeks earlier compared to the control plasmid pVAX1 and pVAX1HBsAg groups and antibody levels showed significant differences. Enhanced HBsAg-specific splenocyte proliferation as well as specific cytotoxic activities of splenic CTLs were also detected. Furthermore, pVAX1-L-GM plasmid increased the number of CD4+ and CD8+ molecules on the surface of the spleen T cell and the level of IFN-γ, IL-2 secretion. pVAX1-L-GM induced a specific immune response in mice and enhanced the immune effect. Thus, a foundation was laid for developing immunogenicity of a better prevention and treatment of HBV via a hepatitis B vaccine.
doi:10.3892/br.2012.47
PMCID: PMC3956218  PMID: 24648930
hepatitis B virus; preS1-; preS2/S gene; large envelope protein; granulocyte-macrophage colony stimulating factor
18.  Development of a SYBR green real-time PCR method for rapid detection of sheep pox virus 
Virology Journal  2012;9:291.
Background
In this study, we developed a SYBR Green-based real-time PCR assay for the detection of sheep pox virus using a plasmid construct carrying one of the highly conserved genes encoding the virion envelope protein (P32) as a template.
Results
The method was demonstrated to be highly sensitive, allowing a precise SPV DNA quantitation over a range of nine orders of magnitude (from 101 to 109 copies of standard DNA). Then, specimens from SPV suspected sheep were analyzed by conventional gel-based PCR, real-time PCR and sequence analysis.
Conclusion
Comparison between these different techniques revealed that real-time PCR is more sensitive than conventional gel-based PCR, allowing detection low viral titers of SPV in infected sheep.
doi:10.1186/1743-422X-9-291
PMCID: PMC3546957  PMID: 23186407
Sheep pox virus; SYBR Green I based quantitative PCR
19.  High expression of ZEB1 correlates with liver metastasis and poor prognosis in colorectal cancer 
Oncology Letters  2012;5(2):564-568.
Zinc finger E-box binding homeobox 1 (ZEB1) has been shown to promote invasion and metastasis in several types of human cancer and to have a prognostic role in certain cancers. However, the clinical significance of ZEB1 in colorectal cancer (CRC) has not been sufficiently investigated. This study aimed to address this issue. In this study, we compared the expression of ZEB1 between CRC tissues and normal adjacent mucosa using quantitative real-time RT-PCR. The association of ZEB1 expression with clinicopathological characteristics was analyzed by appropriate statistical analyses. Kaplan-Meier analysis and Cox proportional hazards regression models were used to investigate the association of ZEB1 expression with survival of patients. The results showed that the relative expression levels of ZEB1 were significantly higher in CRC tissues compared to the normal adjacent mucosa and higher expression of ZEB1 correlated with liver metastasis. Kaplan-Meier analysis indicated that patients with high ZEB1 had a poor overall survival. Moreover, the multivariate analysis showed that high expression of ZEB1 was an independent predictor of overall survival. Our data indicate the potential of ZEB1 as a novel prognostic biomarker for CRC.
doi:10.3892/ol.2012.1026
PMCID: PMC3573155  PMID: 23420790
zinc finger e-box binding homeobox 1; colorectal cancer; prognostic factor
20.  Association study of HLA-B alleles with idiopathic male infertility in Han population of China 
Purpose
To investigate the distributions of HLA-B alleles and estimate their associations with idiopathic male infertility in Chinese Han population.
Methods
Polymerase chain reaction-sequence-based typing (PCR-SBT) method was used for DNA typing at HLA-B locus in 109 patients with idiopathic male infertility and 152 healthy controls in male Han population of Shaanxi Province, situated in northwestern China.
Results
In total, we detected 45 HLA-B alleles in idiopathic infertile patients, 48 HLA-B alleles in control subjects. However, no significant differences of these allelic frequencies were found between the infertile patients and the controls.
Conclusion
HLA-B gene was unlikely a major risk factor of idiopathic male infertility in this sample population. As different populations have different HLA polymorphisms, investigation of the relationship of other HLA genes and idiopathic male infertility with larger sample size, is warranted in the future.
doi:10.1007/s10815-011-9622-9
PMCID: PMC3220441  PMID: 21870185
Human leukocyte antigen (HLA)-B; Polymerase chain reaction-sequence-based typing (PCR-SBT); Idiopathic male infertility
21.  Emergence and Continuous Evolution of Genotype 1E Rubella Viruses in China 
Journal of Clinical Microbiology  2012;50(2):353-363.
In China, rubella vaccination was introduced into the national immunization program in 2008, and a rubella epidemic occurred in the same year. In order to know whether changes in the genotypic distribution of rubella viruses have occurred in the postvaccination era, we investigate in detail the epidemiological profile of rubella in China and estimate the evolutionary rate, molecular clock phylogeny, and demographic history of the predominant rubella virus genotypes circulating in China using Bayesian Markov chain Monte Carlo phylodynamic analyses. 1E was found to be the predominant rubella virus genotype since its initial isolation in China in 2001, and no genotypic shift has occurred since then. The results suggest that the global 1E genotype may have diverged in 1995 and that it has evolved at a mutation rate of 1.65 × 10−3 per site per year. The Chinese 1E rubella virus isolates were grouped into either cluster 1 or cluster 2, which likely originated in 1997 and 2006, respectively. Cluster 1 viruses were found in all provinces examined in this study and had a mutation rate of 1.90 × 10−3 per site per year. The effective number of infections remained constant until 2007, and along with the introduction of rubella vaccine into the national immunization program, although the circulation of cluster 1 viruses has not been interrupted, some viral lineages have disappeared, and the epidemic started a decline that led to a decrease in the effective population size. Cluster 2 viruses were found only in Hainan Province, likely because of importation.
doi:10.1128/JCM.01264-11
PMCID: PMC3264136  PMID: 22162559
22.  Paroxysmal drastic abdominal pain with tardive cutaneous lesions presenting in Henoch-Schönlein purpura 
Henoch-Schönlein purpura (HSP) is a small-vessel vasculitis mediated by IgA-immune complex deposition. It is characterized by the clinical tetrad of non-thrombocytopenic palpable purpura, abdominal pain, arthritis and renal involvement. The diagnosis of HSP is difficult, especially when abdominal symptoms precede cutaneous lesions. We report a rare case of paroxysmal drastic abdominal pain with gastrointestinal bleeding presented in HSP. The diagnosis was verified by renal damage and the occurrence of purpura.
doi:10.3748/wjg.v18.i16.1991
PMCID: PMC3337578  PMID: 22563183
Henoch-Schönlein purpura; Abdominal pain; Gastrointestinal bleeding
23.  Single Endemic Genotype of Measles Virus Continuously Circulating in China for at Least 16 Years 
PLoS ONE  2012;7(4):e34401.
The incidence of measles in China from 1991 to 2008 was reviewed, and the nucleotide sequences from 1507 measles viruses (MeV) isolated during 1993 to 2008 were phylogenetically analyzed. The results showed that measles epidemics peaked approximately every 3 to 5 years with the range of measles cases detected between 56,850 and 140,048 per year. The Chinese MeV strains represented three genotypes; 1501 H1, 1 H2 and 5 A. Genotype H1 was the predominant genotype throughout China continuously circulating for at least 16 years. Genotype H1 sequences could be divided into two distinct clusters, H1a and H1b. A 4.2% average nucleotide divergence was found between the H1a and H1b clusters, and the nucleotide sequence and predicted amino acid homologies of H1a viruses were 92.3%–100% and 84.7%–100%, H1b were 97.1%–100% and 95.3%–100%, respectively. Viruses from both clusters were distributed throughout China with no apparent geographic restriction and multiple co-circulating lineages were present in many provinces. Cluster H1a and H1b viruses were co-circulating during 1993 to 2005, while no H1b viruses were detected after 2005 and the transmission of that cluster has presumably been interrupted. Analysis of the nucleotide and predicted amino acid changes in the N proteins of H1a and H1b viruses showed no evidence of selective pressure. This study investigated the genotype and cluster distribution of MeV in China over a 16-year period to establish a genetic baseline before MeV elimination in Western Pacific Region (WPR). Continuous and extensive MeV surveillance and the ability to quickly identify imported cases of measles will become more critical as measles elimination goals are achieved in China in the near future. This is the first report that a single endemic genotype of measles virus has been found to be continuously circulating in one country for at least 16 years.
doi:10.1371/journal.pone.0034401
PMCID: PMC3332093  PMID: 22532829
24.  Vena cava thrombosis after vena cava filter placement: Incidence and risk factors 
Background
The objective of this study was to assess the clinical safety and efficacy of vena cava filter (VCF) placement, with particular emphasis on the incidence and risk factors of inferior vena cava thrombosis (VCT) after VCF placement.
Methods
Clinical data of patients with venous thromboembolism (VTE), with or without placement of VCF, were analyzed in a retrospective single-center audit of medical records from January 2005 to June 2009. The collected data included demographics, procedural details, filter type, indications, and complications.
Results
A total of 168 cases of VTE (82 with VCF; 86 without VCF) were examined. Over a median follow-up of 24.2 months, VCT occurred in 18 of 82 patients with VCFs (11 males, 7 females, mean age 55.4 years). In 86 patients without VCFs, VCT occurred in only 6 individuals (4 males, 2 females) during the study period. VCT was observed more frequently in patients fitted with VCFs than in those without VCFs (22% vs. 7.0%).
Conclusions
The incidence of VCT in patients with VTE after VCF implantation was 22% approximately. Anticoagulation therapy should be continued for all patients with VCF placement, unless there is a specific contraindication. Almost all instances of VCT in patients with VCF implants in our study occurred after stopping anticoagulation treatment. The use of VCFs is increasing, and more trials are needed to confirm their benefit and accurately assess their safety.
doi:10.3724/SP.J.1263.2011.00099
PMCID: PMC3390081  PMID: 22783293
vena cava filters; venous thromboembolism; complication
25.  Differentiation of Human Bone Marrow Mesenchymal Stem Cells into Bladder Cells: Potential for Urological Tissue Engineering 
Tissue Engineering. Part A  2010;16(5):1769-1779.
Bone marrow mesenchymal stem cells (BMSCs) are capable of differentiating into multiple cell types, providing an alternative cell source for cell-based therapy and tissue engineering. Simultaneous differentiation of human BMSCs into smooth muscle cells (SMCs) and urothelium would be beneficial for clinical applications in bladder regeneration for patients with bladder exstrophy or cancer who need cystoplasty. We investigated the ability of human BMSCs to differentiate toward both SMCs and urothelium with cocultured or conditioned media and analyzed growth factors from a coculture system. After being cocultured with urothelium or cultured using urothelium-derived conditioned medium, human BMSCs expressed urothelium-specific genes and proteins: uroplakin-Ia, cytokeratin-7, and cytokeratin-13. When cocultured with SMCs or cultured in SMC-conditioned medium, human BMSCs expressed SMC-specific genes and proteins: desmin and myosin. Several growth factors (hepatocyte growth factor, platelet-derived growth factor-homodimer polypeptide of B chain (BB), transforming growth factor-β1, and vascular endothelial growth factor) were detected in the SMC cocultured media and in the urothelium cocultured media (epidermal growth factor, platelet-derived growth factor-BB, transforming growth factor-β1, and vascular endothelial growth factor). BMSC–scaffold constructs significantly improved cell contractility after myogenic differentiation. In vivo-grafted cells displayed significant matrix infiltration and expressed SMC-specific markers in the nanofibrous poly-l-lactic acid scaffolds. In conclusion, smooth muscle- and urothelium-like cells derived from human BMSCs provide an alternative cell source for potential use in bladder tissue engineering.
doi:10.1089/ten.tea.2009.0625
PMCID: PMC2952115  PMID: 20020816

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