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1.  Construction of a high-density genetic map and QTLs mapping for sugars and acids in grape berries 
BMC Plant Biology  2015;15:28.
Background
QTLs controlling individual sugars and acids (fructose, glucose, malic acid and tartaric acid) in grape berries have not yet been identified. The present study aimed to construct a high-density, high-quality genetic map of a winemaking grape cross with a complex parentage (V. vinifera × V. amurensis) × ((V. labrusca × V. riparia) × V. vinifera), using next-generation restriction site-associated DNA sequencing, and then to identify loci related to phenotypic variability over three years.
Results
In total, 1 826 SNP-based markers were developed. Of these, 621 markers were assembled into 19 linkage groups (LGs) for the maternal map, 696 for the paternal map, and 1 254 for the integrated map. Markers showed good linear agreement on most chromosomes between our genetic maps and the previously published V. vinifera reference sequence. However marker order was different in some chromosome regions, indicating both conservation and variation within the genome. Despite the identification of a range of QTLs controlling the traits of interest, these QTLs explained a relatively small percentage of the observed phenotypic variance. Although they exhibited a large degree of instability from year to year, QTLs were identified for all traits but tartaric acid and titratable acidity in the three years of the study; however only the QTLs for malic acid and β ratio (tartaric acid-to-malic acid ratio) were stable in two years. QTLs related to sugars were located within ten LGs (01, 02, 03, 04, 07, 09, 11, 14, 17, 18), and those related to acids within three LGs (06, 13, 18). Overlapping QTLs in LG14 were observed for fructose, glucose and total sugar. Malic acid, total acid and β ratio each had several QTLs in LG18, and malic acid also had a QTL in LG06. A set of 10 genes underlying these QTLs may be involved in determining the malic acid content of berries.
Conclusion
The genetic map constructed in this study is potentially a high-density, high-quality map, which could be used for QTL detection, genome comparison, and sequence assembly. It may also serve to broaden our understanding of the grape genome.
Electronic supplementary material
The online version of this article (doi:10.1186/s12870-015-0428-2) contains supplementary material, which is available to authorized users.
doi:10.1186/s12870-015-0428-2
PMCID: PMC4329212  PMID: 25644551
Berry quality; Genetic map; Next-generation sequencing (NGS); QTL analysis; Quantitative trait loci; Restriction-site associated DNA (RAD); Vitis
2.  Discovery of Novel Antitumor Dibenzocyclooctatetraene Derivatives and Related Biphenyls as Potent Inhibitors of NF-κB Signaling Pathway 
Bioorganic & medicinal chemistry  2013;22(1):325-333.
Several dibenzocyclooctatetraene derivatives (5-7) and related biphenyls (8-11) were designed, synthesized, and evaluated for inhibition of cancer cell growth and the NF-κB signaling pathway. Compound 5a, a dibenzocyclooctatetraene succinimide, was discovered as a potent inhibitor of the NF-κB signaling pathway with significant antitumor activity against several human tumor cell lines (GI50 1.38–1.45 μM) and was more potent than paclitaxel against the drug-resistant KBvin cell line. Compound 5a also inhibited LPS-induced NF-κB activation in RAW264.7 cells with an IC50 value of 0.52 μM, prevented IκB-α degradation and p65 nuclear translocation, and suppressed LPS-induced NO production in a dose-dependent manner. The antitumor data in cellular assays indicated that relative positions and types of substituents on the dibenzocyclooctatetraene or acyclic biphenyl as well as torsional angles between the two phenyls are of primary importance to antitumor activity.
doi:10.1016/j.bmc.2013.11.018
PMCID: PMC3899348  PMID: 24315191
dibenzocyclooctatetraene derivatives; unsymmetrical biphenyls; anticancer agents; NF-κB inhibitor
3.  Quantitative Hepatitis B Core Antibody Level Is a New Predictor for Treatment Response In HBeAg-positive Chronic Hepatitis B Patients Receiving Peginterferon 
Theranostics  2015;5(3):218-226.
A recent study revealed that quantitative hepatitis B core antibody (qAnti-HBc) level could serve as a novel marker for predicting treatment response. In the present study, we further investigated the predictive value of qAnti-HBc level in HBeAg-positive patients undergoing PEG-IFN therapy. A total of 140 HBeAg-positive patients who underwent PEG-IFN therapy for 48 weeks and follow-up for 24 weeks were enrolled in this study. Serum samples were taken every 12 weeks post-treatment. The predictive value of the baseline qAnti-HBc level for treatment response was evaluated. Patients were further divided into 2 groups according to the baseline qAnti-HBc level, and the response rate was compared. Additionally, the kinetics of the virological and biochemical parameters were analyzed. Patients who achieved response had a significantly higher baseline qAnti-HBc level (serological response [SR], 4.52±0.36 vs. 4.19±0.58, p=0.001; virological response [VR], 4.53±0.35 vs. 4.22±0.57, p=0.005; combined response [CR], 4.50±0.36 vs. 4.22±0.58, p=0.009)). Baseline qAnti-HBc was the only parameter that was independently correlated with SR (p=0.008), VR (p=0.010) and CR(p=0.019). Patients with baseline qAnti-HBc levels ≥30,000 IU/mL had significantly higher response rates, more HBV DNA suppression, and better hepatitis control in PEG-IFN treatment. In conclusion, qAnti-HBc level may be a novel biomarker for predicting treatment response in HBeAg-positive patients receiving PEG-IFN therapy.
doi:10.7150/thno.10636
PMCID: PMC4279186  PMID: 25553110
quantitative anti-HBc;  chronic hepatitis B; PEG-IFN treatment; treatment response prediction; pretreatment biomarker.
4.  Association Study Identifying a New Susceptibility Gene (AUTS2) for Schizophrenia 
Schizophrenia (SCZ) is a severe and debilitating mental disorder, and the specific genetic factors that underlie the risk for SCZ remain elusive. The autism susceptibility candidate 2 (AUTS2) gene has been reported to be associated with autism, suicide, alcohol consumption, and heroin dependence. We hypothesized that AUTS2 might be associated with SCZ. In the present study, three polymorphisms (rs6943555, rs7459368, and rs9886351) in the AUTS2 gene were genotyped in 410 patients with SCZ and 435 controls using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) and forced PCR-RFLP methods. We detected an association between SCZ and the rs6943555 genotype distribution (odds ratio (OR) = 1.363, 95% confidence interval (CI): 0.848–2.191, p = 0.001). The association remained significant after adjusting for gender, and a significant effect (p = 0.001) was observed among the females. In the present study, rs6943555 was determined to be associated with female SCZ. Our results confirm previous reports which have suggested that rs6943555 might elucidate the pathogenesis of schizophrenia and play an important role in its etiology.
doi:10.3390/ijms151119406
PMCID: PMC4264119  PMID: 25347278
AUTS2 gene; SNP; schizophrenia; association study
5.  AGENCY–UNIVERSITY PARTNERSHIP FOR EVIDENCE-BASED PRACTICE IN SOCIAL WORK 
Little is known about evidence-based practice (EBP) in social service agencies beyond studies of researcher, practitioner, and educator opinions. The Bringing Evidence for Social Work Training (BEST) Project involved 16 participants from 3 social service agencies. The experiential training, delivered by 2 doctoral students, focused on a team-identified practice issue and followed the EBP process of motivation, question formulation, search, evaluation, and application planning. Posttraining focus group data were analyzed. Results suggest that university researchers who based in schools of social work can successfully collaborate with agencies to support the process of identifying, evaluating, and discussing the application of research evidence in practice. University–agency training partnerships should be considered as 1 of many potential strategies for advancing EBP in social work.
doi:10.1080/15433714.2011.581545
PMCID: PMC4213455  PMID: 23581802
6.  Down-regulation of miR-181b promotes apoptosis by targeting CYLD in thyroid papillary cancer 
MicroRNAs (miRNAs) are a small class of non-coding RNAs that are widely deregulated in various cancers. They act as either oncogenes or tumor suppressor genes in human cancer. The purpose of this study was to examine the potential role of miR-181b in human thyroid papillary cancer. The expression levels of different miRNAs were measured by micro array analysis in 10 thyroid papillary cancer specimens and adjacent normal thyroid cancer tissues. MTT assays, colony formation assays, apoptosis assays were used to explore the potential function of miR-181b inhibitor in TPC1 human thyroid papillary cancer cells. Luciferase reporter assays were performed to validate the regulation of a putative target of miR-181b, in corroboration with qPCR and western blot assays. We found that the expression of miR-181b was higher in thyroid papillary cancer specimens compared with adjacent normal tissues (P < 0.05). Downregulation of miR-181b inhibited cellular growth and promoted cellular apoptosis. Luciferase assays indicated that miR-181b can bind with its putative target site in the 3’-untranslated region (3’-UTR) of CYLD, suggesting that CYLD is a direct target of miR-181b. Western blot analysis indicated that downregulation of miR-181b results in the upregulation of CYLD at protein levels. Taken together, downregulation of miR-181b expression causes cellular growth inhibition, promoting cellular apoptosis by targeting CYLD. These findings suggest that downregulation of the expression of miR-181b may be a therapeutic target for the treatment of human thyroid papillary cancer.
PMCID: PMC4270513  PMID: 25550803
MiR-181b inhibitor; thyroid papillary cancer; CYLD
7.  Simultaneous Detection of Five Enteric Viruses Associated with Gastroenteritis by Use of a PCR Assay: a Single Real-Time Multiplex Reaction and Its Clinical Application 
Journal of Clinical Microbiology  2014;52(4):1266-1268.
We developed a highly sensitive reverse transcription and multiplex real-time PCR (rtPCR) assay that can identify five viruses, including six genogroups, in a single reaction: norovirus genogroups I and II; sapovirus genogroups I, II, IV, and V; human rotavirus A; adenovirus serotypes 40 and 41; and human astrovirus. In comparison to monoplex rtPCR assays, the sensitivities and specificities of the multiplex rtPCR ranged from 75% to 100% and from 99% to 100%, respectively, evaluated on 812 clinical stool specimens.
doi:10.1128/JCM.00245-14
PMCID: PMC3993471  PMID: 24478418
8.  Two-Year Outcomes of a Randomized, Family-Based Substance Use Prevention Trial for Asian American Adolescent Girls 
Asian Americans have been largely ignored in the prevention outcome literature. In this study, we tested a parent-child program with a sample of Asian American adolescent girls and their mothers, and evaluated the program’s efficacy on decreasing girls’ substance use, and modifying risk and protective factors at individual, family, and peer levels. One hundred and eight Asian American mother-daughter dyads recruited through online advertisements and from community service agencies were randomly assigned to an intervention arm (n = 56) or to a test-only control arm (n = 52). The intervention consisted of a nine-session substance abuse prevention program, delivered entirely online. Guided by family interaction theory, the prevention program aimed to strengthen the quality of girls’ relationships with their mothers while increasing girls’ resilience to resist substance use. Intent-to-treat analyses showed that at 2-year follow-up, intervention-arm dyads had significantly higher levels of mother-daughter closeness, mother-daughter communication, maternal monitoring, and family rules against substance use compared to the control-arm dyads. Intervention-arm girls also showed sustained improvement in self-efficacy and refusal skills, and had lower intentions to use substances in the future. Most important, intervention-arm girls reported fewer instances of alcohol and marijuana use, and prescription drug misuse relative to the control-arm girls. The study suggests that a culturally generic, family-based prevention program was efficacious in enhancing parent-child relationships, improving girls’ resiliency, and preventing substance use behaviors among Asian American girls.
doi:10.1037/a0030925
PMCID: PMC4135055  PMID: 23276322
Substance use prevention; Asian Americans; adolescent girls; family; web-based
9.  The radial diffusivity and magnetization transfer pool size ratio are sensitive markers for demyelination in a rat model of type III multiple sclerosis (MS) lesions 
NeuroImage  2013;74:298-305.
Determining biophysical sensitivity and specificity of quantitative magnetic resonance imaging is essential to develop effective imaging metrics of neurodegeneration. Among these metrics apparent pool size ratio (PSR) from quantitative magnetization transfer (qMT) imaging and radial diffusivity (RD) from diffusion tensor imaging (DTI) are both known to relate to histological measure of myelin density and integrity. However their relative sensitivities towards quantitative myelin detection are unknown. In this study, we correlated high-resolution quantitative magnetic resonance imaging measures of subvoxel tissue structures with corresponding quantitative myelin histology in a lipopolysacharide (LPS) mediated animal model of MS. Specifically, we acquired quantitative magnetization transfer (qMT) and diffusion tensor imaging (DTI) metrics (on the same tissue sample) in an animal model system of type III oligodendrogliopathy which lacked prominent lymphocytic infiltration, a system that had not been previously examined with quantitative MRI. We find that the qMT measured apparent pool size ratio (PSR) showed the strongest correlation with a histological measure of myelin content. DTI measured RD showed the next strongest correlation, and other DTI and relaxation parameters (such as the longitudinal relaxation rate (R1f) or fractional anisotropy (FA)) showed considerably weaker correlations with myelin content.
doi:10.1016/j.neuroimage.2013.02.034
PMCID: PMC3995162  PMID: 23481461
Quantitative magnetization transfer (qMT); Diffusion Tensor Imaging (DTI); 9.4T; White matter; Myelin; demyelination; lippopolysaccharide (LPS); Rat brain; Multiple Sclerosis (MS)
10.  Alterations of microRNAs are associated with impaired growth of MCF-7 breast cancer cells induced by inhibition of casein kinase 2 
Background and aim: Protein Kinase (casein kinase 2, CK2) is a pleiotropic serine-threonine kinase that is frequently dysregulated in many human tumors; microRNAs (miRNAs) are a class of small noncoding RNAs which play important roles in human cancers. This study aimed to investigate the role of CK2 and miRNA expression in breast cancer. Methods: Casein kinase 2 in MCF-7 breast cancer cell line was inhibited by the CK2 inhibitor TBB (4,5,6,7-tetrabromobenzotriazole), then cell proliferation was studied using MTT assay, and cell cycle distribution and apoptosis were detected by flow cytometry. The alteration of microRNAs expression profile was determined by microarray technology, followed by RT-PCR confirmation. Results: Here, we for the first time showed that inhibition of CK2 in MCF-7 breast cancer cells causes suppressed cell growth, which was related with dysregulation of the miRNA profile and altered expression. CK2 inhibition induced the up-regulated expression of 17 miRNAs and 10 down-regulated microRNAs which contributed to the impaired growth, inhibited cell cycle progress and increased apoptosis of MCF-7 cells by a CK2 inhibitor. Conclusions: These findings highlight the potential role of dysregulated miRNA expression regulated by CK2 in breast cancer.
PMCID: PMC4129013  PMID: 25120778
TBB; casein kinase 2; microRNA; breast cancer
11.  Clinical Study on Prospective Efficacy of All-Trans Acid, Realgar-Indigo Naturalis Formula Combined with Chemotherapy as Maintenance Treatment of Acute Promyelocytic Leukemia 
Objectives. To test the efficiency and safety of sequential application of retinoic acid (ATRA), Realgar-Indigo naturalis formula (RIF) and chemotherapy (CT) were used as the maintenance treatment in patients with acute promyelocytic leukemia (APL). Methods. This was a retrospective study of 98 patients with newly diagnosed APL who accepted two different maintenance treatments. After remission induction and consolidation chemotherapy according to their Sanz scores, patients received two different kinds of maintenance scheme. The first regimen was using ATRA, RIF, and standard dose of CT sequentially (ATRA/RIF/CT regimen), while the second one was using ATRA and low dose of chemotherapy with methotrexate (MTX) plus 6-mercaptopurine (6-MP) alternately (ATRA/CTlow regimen). The OS, DFS, relapse rate, minimal residual disease, and adverse reactions in two groups were monitored and evaluated. Results. ATRA/RIF/CT regimen could effectively reduce the chance of relapse in different risk stratification of patients, but there was no significant difference in 5-year DFS rate and OS rate between the two groups. Besides, the patients in the experimental group suffered less severe adverse reactions than those in the control group. Conclusions. The repeated sequential therapeutic regimen to APL with ATRA, RIF, and chemotherapy is worth popularizing for its high effectiveness and low toxicity.
doi:10.1155/2014/987560
PMCID: PMC4055014  PMID: 24963332
12.  Broad-Spectrum Transgenic Resistance against Distinct Tospovirus Species at the Genus Level 
PLoS ONE  2014;9(5):e96073.
Thrips-borne tospoviruses cause severe damage to crops worldwide. In this investigation, tobacco lines transgenic for individual WLm constructs containing the conserved motifs of the L RNA-encoded RNA-dependent RNA polymerase (L) gene of Watermelon silver mottle virus (WSMoV) were generated by Agrobacterium-mediated transformation. The WLm constructs included: (i) translatable WLm in a sense orientation; (ii) untranslatable WLmt with two stop codons; (iii) untranslatable WLmts with stop codons and a frame-shift; (iv) untranslatable antisense WLmA; and (v) WLmhp with an untranslatable inverted repeat of WLm containing the tospoviral S RNA 3′-terminal consensus sequence (5′-ATTGCTCT-3′) and an NcoI site as a linker to generate a double-stranded hairpin transcript. A total of 46.7–70.0% transgenic tobacco lines derived from individual constructs showed resistance to the homologous WSMoV; 35.7–100% plants of these different WSMoV-resistant lines exhibited broad-spectrum resistance against four other serologically unrelated tospoviruses Tomato spotted wilt virus, Groundnut yellow spot virus, Impatiens necrotic spot virus and Groundnut chlorotic fan-spot virus. The selected transgenic tobacco lines also exhibited broad-spectrum resistance against five additional tospoviruses from WSMoV and Iris yellow spot virus clades, but not against RNA viruses from other genera. Northern analyses indicated that the broad-spectrum resistance is mediated by RNA silencing. To validate the L conserved region resistance in vegetable crops, the constructs were also used to generate transgenic tomato lines, which also showed effective resistance against WSMoV and other tospoviruses. Thus, our approach of using the conserved motifs of tospoviral L gene as a transgene generates broad-spectrum resistance against tospoviruses at the genus level.
doi:10.1371/journal.pone.0096073
PMCID: PMC4014477  PMID: 24811071
13.  Screening of surface markers on rat intestinal mucosa microfold cells by using laser capture microdissection combined with protein chip technology 
Objective: The objective of this research was to investigate the possibility of screening surface markers on rat intestinal mucosa microfold cells (M cells) by using laser capture microdissection (LCM) combined with protein chip technology. Methods: We labeled rat intestinal mucosa microfold cells with Ulex europaeus agglutinin (UEA)-1 antibody and visualized these by immunofluorescence staining. Using the Proteome Profiler rat protein chip, we analyzed the protein expression profiles of LCM M-cells compared to lymph follicle-associated epithelial (FAE) cells, and we identified potential differences to screen for marker proteins. Results: M cells can be clearly distinguished from lymphoid FAE cells under the fluorescence microscope. We successfully cut, isolated, and obtained microfold and lymph FAE cells with more than 95% homogeneity. Six differentially expressed proteins were identified through comparison of the protein chip profiles of these 2 cell types. Among these, VEGF, LIX, CNTF, and IL-1α/IL-1F1 were found to be at significantly lower levels in M cells, IL-1ra/IL-1F3 and MIG/CXCL9 appeared in significantly higher levels in M cells (P < 0.05). Conclusion: The results presented here clearly demonstrate that the combined use of LCM and protein chip technology is effective in the screening of M cell surface markers with high sensitivity and specificity. This will facilitate isolation, identification, and establishment of M cell lines, allowing further characterization of their functional properties.
PMCID: PMC4057843  PMID: 24955164
Laser capture microdissection (LCM); proteomics; protein chip; M cells; marker
14.  MiRNA-26b inhibits cellular proliferation by targeting CDK8 in breast cancer 
Objectives: MicroRNA-26b (miR-26b) has been reported to be down-regulated in a wide range of malignant tumors, However, the mechanism by which miR-26b is implicated in breast cancer tumorigenesis is incompletely understood. This study was undertaken to evaluate the expression pattern of miR-26b and characterize its biological role in human breast cancer. Methods: Reverse transcription-polymerase chain reaction (RT-PCR) was used to quantify the expression levels of miR-26b in breast cancer and adjacent non-cancerous breast tissues. MTT, colony formation assay and cell cycle assay were carried out to characterize the miR-26b function. Finally, to validate the target gene of miR-26b, luciferase reporter assay was employed, followed by RT-PCR and Western blot confirmation. Results: Here, we found that miR-26b expression was relatively downregulated in breast cancer specimens (P<0.01). Overexpression of miR-26b dramatically suppressed cell proliferation, colony formation and induced G0/G1 cell cycle arrest of MDA-MB-231 and Mcf-7 cells. Luciferase assays revealed that miR-26b directly targeted the 3’UTR of CDK8. Overexpression of miR-26b led to the downregulation of CDK8 and β-catenin expression. Similarly, CDK8 knockdown by siRNA suppressed cell growth and subsequent β-catenin expression. Conclusions: These findings suggest that miR-26b exerts a tumor suppressive role in breast cancer and the miR-26b-mediated growth inhibition is achieved through suppression of a new target gene CDK8
PMCID: PMC3992393  PMID: 24753748
MiR-26b; proliferation; cell cycle; CDK8; breast cancer
15.  The sequence and de novo assembly of the giant panda genome 
Li, Ruiqiang | Fan, Wei | Tian, Geng | Zhu, Hongmei | He, Lin | Cai, Jing | Huang, Quanfei | Cai, Qingle | Li, Bo | Bai, Yinqi | Zhang, Zhihe | Zhang, Yaping | Wang, Wen | Li, Jun | Wei, Fuwen | Li, Heng | Jian, Min | Li, Jianwen | Zhang, Zhaolei | Nielsen, Rasmus | Li, Dawei | Gu, Wanjun | Yang, Zhentao | Xuan, Zhaoling | Ryder, Oliver A. | Leung, Frederick Chi-Ching | Zhou, Yan | Cao, Jianjun | Sun, Xiao | Fu, Yonggui | Fang, Xiaodong | Guo, Xiaosen | Wang, Bo | Hou, Rong | Shen, Fujun | Mu, Bo | Ni, Peixiang | Lin, Runmao | Qian, Wubin | Wang, Guodong | Yu, Chang | Nie, Wenhui | Wang, Jinhuan | Wu, Zhigang | Liang, Huiqing | Min, Jiumeng | Wu, Qi | Cheng, Shifeng | Ruan, Jue | Wang, Mingwei | Shi, Zhongbin | Wen, Ming | Liu, Binghang | Ren, Xiaoli | Zheng, Huisong | Dong, Dong | Cook, Kathleen | Shan, Gao | Zhang, Hao | Kosiol, Carolin | Xie, Xueying | Lu, Zuhong | Zheng, Hancheng | Li, Yingrui | Steiner, Cynthia C. | Lam, Tommy Tsan-Yuk | Lin, Siyuan | Zhang, Qinghui | Li, Guoqing | Tian, Jing | Gong, Timing | Liu, Hongde | Zhang, Dejin | Fang, Lin | Ye, Chen | Zhang, Juanbin | Hu, Wenbo | Xu, Anlong | Ren, Yuanyuan | Zhang, Guojie | Bruford, Michael W. | Li, Qibin | Ma, Lijia | Guo, Yiran | An, Na | Hu, Yujie | Zheng, Yang | Shi, Yongyong | Li, Zhiqiang | Liu, Qing | Chen, Yanling | Zhao, Jing | Qu, Ning | Zhao, Shancen | Tian, Feng | Wang, Xiaoling | Wang, Haiyin | Xu, Lizhi | Liu, Xiao | Vinar, Tomas | Wang, Yajun | Lam, Tak-Wah | Yiu, Siu-Ming | Liu, Shiping | Zhang, Hemin | Li, Desheng | Huang, Yan | Wang, Xia | Yang, Guohua | Jiang, Zhi | Wang, Junyi | Qin, Nan | Li, Li | Li, Jingxiang | Bolund, Lars | Kristiansen, Karsten | Wong, Gane Ka-Shu | Olson, Maynard | Zhang, Xiuqing | Li, Songgang | Yang, Huanming | Wang, Jian | Wang, Jun
Nature  2009;463(7279):311-317.
Using next-generation sequencing technology alone, we have successfully generated and assembled a draft sequence of the giant panda genome. The assembled contigs (2.25 gigabases (Gb)) cover approximately 94% of the whole genome, and the remaining gaps (0.05 Gb) seem to contain carnivore-specific repeats and tandem repeats. Comparisons with the dog and human showed that the panda genome has a lower divergence rate. The assessment of panda genes potentially underlying some of its unique traits indicated that its bamboo diet might be more dependent on its gut microbiome than its own genetic composition. We also identified more than 2.7 million heterozygous single nucleotide polymorphisms in the diploid genome. Our data and analyses provide a foundation for promoting mammalian genetic research, and demonstrate the feasibility for using next-generation sequencing technologies for accurate, cost-effective and rapid de novo assembly of large eukaryotic genomes.
doi:10.1038/nature08696
PMCID: PMC3951497  PMID: 20010809
16.  MiRNA-107 inhibits proliferation and migration by targeting CDK8 in breast cancer 
Background: MicroRNAs (miRNAs) are small, non-coding RNAs (18-25 nucleotides) that post-transcriptionally modulate gene expression by negatively regulating the stability or translational efficiency of their target mRNAs. The aim of this study was to investigate the expression pattern of microRNA-107 (miR-107) in human breast cancer, and its potential role in disease pathogenesis. Methods: Quantitative reverse transcription-polymerase chain reaction (qRT-PCR) was performed to determine the expression level of miR-107 in 30 breast cancer specimens and adjacent normal breast tissues. MTT and colony formation assays, transwell and wound healing test, cell cycle assays were conducted to explore the potential function of miR-107 in human MDA-MB-231 breast cancer cells. Luciferase reporter assays were employed to validate regulation of a putative target of miR-107. The effect of modulating miR-107 on endogenous levels of this target were subsequently confirmed via Western blotting. Results: miR-107 expression was relatively decreased in breast cancer specimens compared with adjacent normal tissues (P<0.01). Overexpression of miR-107 suppressed MDA-MB-231 cell proliferation and migration, meanwhile the cells were arrested at G0/G1 phase. Luciferase assays using a reporter carrying a putative miR-107 target site in the 3’, untranslated region (3’-UTR) of CDK8 revealed that miR-107 directly targets CDK8. Overexpression of miR-107 led to downregulation of CDK8 at the mRNA and protein level, as assessed by Western blotting. Conclusions: miR-107 may play an important role in breast cancer progression, which might negatively regulate the expression of CDK8 and inhibit the proliferation and migration of MDA-MB-231 cell line.
PMCID: PMC3902238  PMID: 24482686
Breast cancer; miR-107; CDK8
17.  MicroRNA-195-5p is a potential diagnostic and therapeutic target for breast cancer 
Oncology Reports  2014;31(3):1096-1102.
MicroRNAs (miRNAs) are a class of highly conserved, small endogenous single-strand non-coding RNAs. They are aberrantly expressed in the circulation and tissue of patients with cancer. Therefore, it has been suggested that they may act as key regulators of carcinogenesis. The aim of the present study was to examine the expression level of miR-195-5p in human breast cancer and its potential role in carcinogenesis. The expression level of miR-195-5p was measured in 40 breast cancer specimens and adjacent normal breast tissues by quantitative polymerase chain reaction (qPCR). Next, to explore the potential function of miR-195-5p, we used MDA-MB-231 human breast cancer cells and carried out MTT, colony formation, Transwell chamber migration and cell cycle assays. The dual-luciferase reporter assay was also performed to determine putative targets of miR-195-5p, which were validated using qPCR and western blot assays. We found that miR-195-5p expression was significantly decreased in the 40 breast cancer specimens when compared with that in the adjacent normal breast tissues (P<0.05). Overexpression of miR-195-5p inhibited cell proliferation, reduced cell colony formation, suppressed cell migration and caused an accumulation of cells in the G1 phase of the cell cycle. In the 3′-untranslated region (3′-UTR) of cyclin E1 (CCNE1), we found two putative target sites which may bind miR-195-5p, suggesting that CCNE1 is a direct target of miR-195-5p. Furthermore, through qPCR and western blot assays we showed that overexpression of miR-195-5p reduced CCNE1 mRNA and protein levels, respectively. Our study suggests that miR-195-5p may act as a tumor suppressor in breast cancer. Therefore, targeting of this miRNA may provide a novel strategy for the diagnosis and treatment of patients with this lethal disease.
doi:10.3892/or.2014.2971
PMCID: PMC3926672  PMID: 24402230
microRNA; miR-195-5p; breast cancer
18.  siRNA-mediated silencing of CDK8 inhibits proliferation and growth in breast cancer cells 
CDK8 is a cyclin-dependent kinase (CDK) member of the mediator complex that couples transcriptional regulators to the basal transcriptional machinery, and it has been investigated for possible tumor promoting functions. However, it is unclear whether CDK8 is involved in breast tumor cells growth. The aim of this study was to determine whether the suppression of CDK8 by small interfering RNA (siRNA) inhibits the growth of human breast cancer cell. Methods: CDK8-siRNA transfection was used to silencing the CDK8 gene in established breast cancer cell line, MDA-MB-231 and MCF-7, successful transfection being confirmed by Real-time PCR and could be shown by Western Blotting. CDK8 deletion caused significant decline in cell proliferation was observed in breast cancer cell lines as investigated by MTS assay, the number and size of the colonies formed were also significantly reduced in the absence of CDK8. Furthermore, transwell test were conducted to explore the migration of breast cancer cells. Moreover CDK8 gene knockdown arrested cell cycle. Results: CDK8 mRNA expression was reduced after transfection with CDK8-siRNA, and protein expression had a similar trend. Transfection of CDK8-siRNA suppressed breast cancer cells proliferation and migration; meanwhile the cells were arrested at G0/G1 phase. Conclusions: CDK8 plays an essential role in breast cancer progression, which might inhibit the proliferation and migration in breast cancer cells.
PMCID: PMC3885463  PMID: 24427329
Breast cancer; CDK8; siRNA
19.  MiRNA-497 regulates cell growth and invasion by targeting cyclin E1 in breast cancer 
Background
MicroRNAs are a class of endogenous single strand non-coding RNAs that are involved in many important physiological and pathological processes. The purpose of this study was to examine the expression levels of miR-497 in human breast cancer and its function in MDA-MB-231 breast cancer cells.
Methods
Quantitative polymerase chain reaction was used to measure the expression levels of miR-497 in 40 breast cancer specimens and adjacent normal breast tissues. MTT assays, colony formation assays, wound healing assays, transwell assays and cell cycle assays were used to explore the potential function of miR-497 in MDA-MB-231 breast cancer cells. Dual-luciferase reporter assays were performed to analyze the regulation of putative target of miR-497, and western blot assays were used to validate the dual-luciferase results.
Results
The expression of miR-497 in breast cancer specimens was lower than adjacent normal tissues (P < 0.05). Overexpression of miR-497 inhibited cellular growth, suppressed cellular migration and invasion, and caused a G1 arrest. Dual-luciferase reporter assays showed that miR-497 binds the 3′-untranslated region (3′-UTR) of cyclin E1, suggesting that cyclin E1 is a direct target of miR-497. Western blot assays confirmed that overexpression of miR-497 reduced cyclin E1 protein levels.
Conclusions
MiR-497 may act as a tumor suppressor gene in breast cancer. Inhibited cellular growth, suppressed cellular migration and invasion, and G1 cell cycle arrest were observed upon overexpression of miR-497 in cells, possibly by targeting cyclin E1. These results indicate miR-497 could be considered a therapeutic target for the development of treatment for breast cancer.
doi:10.1186/1475-2867-13-95
PMCID: PMC3853026  PMID: 24112607
MiR-497; Breast cancer; Cyclin E1
20.  Clinical significance of NOB1 expression in breast infiltrating ductal carcinoma 
Background: NIN/RPN Binding protein 1 homologue (NOBp1), encoded by NOB1 gene, was reported to play an essential role in the oncogenesis and prognosis of carcinomas. We conducted a study to reveal its expression and clinical significance in breast infiltrating ductal carcinoma. Methods: To explore the relationship between NOB1 expression and the clinical TNM (cTNM), 162 patients who undergone surgery were involved in the study. Compared to healthy tissues, abnormal localization and higher level of NOB1 in tumor cells was observed by Immunohistochemistry staining. Real-time PCR and western-blotting verified the up-regulation of NOB1 in carcinoma individuals. Results: A significant correlation between high level of NOB1 and the T stage, lymph node metastasis and cTNM was shown. Furthermore, patients with higher level of NOB1 predicted a declined overall survival (OS). Notably, multivariate analyses by Cox’s proportional hazard model revealed that expression of NOB1 was an independent prognostic factor in breast infiltrating ductal carcinoma. Conclusions: In summary, our present study clarify that the aberrant expression of NOB1 in breast infiltrating ductal carcinoma is possibly involved with tumorigenesis and development, and the NOB1 protein could act as a potential biomarker for prognosis assessment of breast infiltrating ductal carcinoma. Related mechanism is worthy of further investigation.
PMCID: PMC3796236  PMID: 24133592
Breast cancer; NOB1 protein; immunohistochemistry; tissue microarray
21.  Histone H2B lysine 120 monoubiquitination is required for embryonic stem cell differentiation 
Cell Research  2012;22(9):1402-1405.
doi:10.1038/cr.2012.114
PMCID: PMC3434344  PMID: 22847742
22.  ‘Expansion in-situ’ concept as a new technique for expanding skin and soft tissue 
Techniques for expanding skin and soft tissue are widely used to repair damaged areas since they facilitate the provision of new, additional skin tissue with similar quality, texture and color to that surrounding the defective area. Conventional expansion techniques involve placing expanders under the normal skin adjacent to a lesion. However, these techniques may involve additional incisions, complications with blood supply and ‘dog-ear’ deformities and may result in a low utilization rate of the expanded tissue. When reconstructing small defects that may not be sutured directly, these shortcomings, particularly the requirement to make additional incisions, limit the application of conventional techniques. The current study presents a novel approach to expansion called the ‘expansion in-situ’ technique. In this technique, the lesion is used as the center for expansion and expanders of optimal size are implanted under the lesion and surrounding normal soft tissue. Following expansion, the damaged area is excised directly. In order to avoid poor healing of the incision made during expander implantation, the overlapping suturing of both cut sides is conducted. This enlarges the contact area of both sides of the incision, thereby avoiding incision dehiscence and increasing wound healing during the expansion process. Between August 2006 and July 2011, the expansion in-situ technique was applied in 10 cases involving either nevus excision or scar removal. All 10 cases were treated successfully. Five of the cases were followed up over 1–3 years. The ‘expansion in-situ’ technique is likely to be useful for avoiding additional incisions and improving the utilization rate of expanded skin flaps.
doi:10.3892/etm.2013.1269
PMCID: PMC3820849  PMID: 24223661
soft tissue expansion; incision dehiscence; expansion in-situ; additional incision; cicatrix
24.  MiRNA-26b inhibits proliferation by targeting PTGS2 in breast cancer 
Background
MicroRNAs (miRNAs) are small, non-coding RNAs (20–24 nucleotides) that post-transcriptionally modulate gene expression by negatively regulating the stability or translational efficiency of their target mRNAs. The aim of this study was to investigate the expression pattern of microRNA-26b (miR-26b) in human breast cancer, and its potential role in disease pathogenesis.
Methods
Quantitative reverse transcription-polymerase chain reaction (qRT-PCR) was performed to determine the expression level of miR-26b in 38 breast cancer specimens and adjacent normal breast tissues. MTT assays were conducted to explore the impact of miR-26b overexpression on the proliferation of human MDA-MB-231 breast cancer cells. Luciferase reporter assays were employed to validate regulation of a putative target of miR-26b. The effect of modulating miR-26b on endogenous levels of this target were subsequently confirmed via qRT-PCR and Western blot.
Results
MiR-26b expression was relatively decreased in breast cancer specimens compared with adjacent normal tissues (P<0.01). Overexpression of miR-26b suppressed MDA-MB-231 cell growth. Luciferase assays using a reporter carrying a putative miR-26b target site in the 3' untranslated region of PTGS2 revealed that miR-26b directly targets PTGS2. Overexpression of miR-26b led to downregulation of PTGS2 at the mRNA and protein level, as assessed by qRT-PCR and Western blot. Targeted knockdown of PTGS2 by siRNA significantly inhibited the proliferation of MDA-MB-231 breast cancer cells.
Conclusions
MiR-26b may act as a tumor suppressor in breast cancer. The overexpression of miR-26b inhibits cellular growth by targeting PTGS2, suggesting its use as a potential therapeutic target for breast cancer.
doi:10.1186/1475-2867-13-7
PMCID: PMC3599806  PMID: 23374284
MiR-26b; Proliferation; PTGS2; Breast cancer
25.  Frontalis Muscle Flap Suspension for the Correction of Congenital Blepharoptosis in Early Age Children 
PLoS ONE  2013;8(1):e53185.
Background
We aimed to report our successful use of frontalis muscle flap suspension for the correction of congenital blepharoptosis in early age children.
Methods
This retrospective study included 61 early age children (41 boys, 20 girls) with an average age of 6 years (range, 3–10 years) with congenital blepharoptosis who received surgery during the period from March 2007 to January 2011. There were 39 cases of unilateral blepharoptosis and 22 cases of bilateral blepharoptosis, thus a total of 83 eyes were affected. If patient had bilateral blepharoptosis, both eyes were operated on in the same surgery. Patients were followed for 3 months to 5 years. The procedure was performed without complications in all cases.
Results
The postoperative healing grade was good in 81 eyes (97.6%); the correction of blepharoptosis was satisfactory, the double eyelid folds were natural and aesthetic, the eyelid position and the curvature were ideal, and the eyes were bilaterally symmetrical. The postoperative healing grade was fair in 2 eyes (2.4%); blepharoptosis was improved compared with that before surgery. At discharge, lagophthalmos was noted in 10 eyes of which 4 cases resolved by the last follow-up. The remaining 6 cases were mild. Eleven eyes received reoperation for residual ptosis after the first surgery. The curvature of the palpebral margin was not natural in 4 eyes. These unnatural curvature possibly was caused by an excessively low lateral fixation point or postoperative avulsion.
Conclusion
Frontalis muscle flap suspension under general anesthesia for the correction of congenital blepharoptosis in early age children can achieve good surgical results.
doi:10.1371/journal.pone.0053185
PMCID: PMC3538777  PMID: 23308158

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