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1.  Comprehensive molecular characterization of gastric adenocarcinoma 
Bass, Adam J. | Thorsson, Vesteinn | Shmulevich, Ilya | Reynolds, Sheila M. | Miller, Michael | Bernard, Brady | Hinoue, Toshinori | Laird, Peter W. | Curtis, Christina | Shen, Hui | Weisenberger, Daniel J. | Schultz, Nikolaus | Shen, Ronglai | Weinhold, Nils | Kelsen, David P. | Bowlby, Reanne | Chu, Andy | Kasaian, Katayoon | Mungall, Andrew J. | Robertson, A. Gordon | Sipahimalani, Payal | Cherniack, Andrew | Getz, Gad | Liu, Yingchun | Noble, Michael S. | Pedamallu, Chandra | Sougnez, Carrie | Taylor-Weiner, Amaro | Akbani, Rehan | Lee, Ju-Seog | Liu, Wenbin | Mills, Gordon B. | Yang, Da | Zhang, Wei | Pantazi, Angeliki | Parfenov, Michael | Gulley, Margaret | Piazuelo, M. Blanca | Schneider, Barbara G. | Kim, Jihun | Boussioutas, Alex | Sheth, Margi | Demchok, John A. | Rabkin, Charles S. | Willis, Joseph E. | Ng, Sam | Garman, Katherine | Beer, David G. | Pennathur, Arjun | Raphael, Benjamin J. | Wu, Hsin-Ta | Odze, Robert | Kim, Hark K. | Bowen, Jay | Leraas, Kristen M. | Lichtenberg, Tara M. | Weaver, Stephanie | McLellan, Michael | Wiznerowicz, Maciej | Sakai, Ryo | Getz, Gad | Sougnez, Carrie | Lawrence, Michael S. | Cibulskis, Kristian | Lichtenstein, Lee | Fisher, Sheila | Gabriel, Stacey B. | Lander, Eric S. | Ding, Li | Niu, Beifang | Ally, Adrian | Balasundaram, Miruna | Birol, Inanc | Bowlby, Reanne | Brooks, Denise | Butterfield, Yaron S. N. | Carlsen, Rebecca | Chu, Andy | Chu, Justin | Chuah, Eric | Chun, Hye-Jung E. | Clarke, Amanda | Dhalla, Noreen | Guin, Ranabir | Holt, Robert A. | Jones, Steven J.M. | Kasaian, Katayoon | Lee, Darlene | Li, Haiyan A. | Lim, Emilia | Ma, Yussanne | Marra, Marco A. | Mayo, Michael | Moore, Richard A. | Mungall, Andrew J. | Mungall, Karen L. | Nip, Ka Ming | Robertson, A. Gordon | Schein, Jacqueline E. | Sipahimalani, Payal | Tam, Angela | Thiessen, Nina | Beroukhim, Rameen | Carter, Scott L. | Cherniack, Andrew D. | Cho, Juok | Cibulskis, Kristian | DiCara, Daniel | Frazer, Scott | Fisher, Sheila | Gabriel, Stacey B. | Gehlenborg, Nils | Heiman, David I. | Jung, Joonil | Kim, Jaegil | Lander, Eric S. | Lawrence, Michael S. | Lichtenstein, Lee | Lin, Pei | Meyerson, Matthew | Ojesina, Akinyemi I. | Pedamallu, Chandra Sekhar | Saksena, Gordon | Schumacher, Steven E. | Sougnez, Carrie | Stojanov, Petar | Tabak, Barbara | Taylor-Weiner, Amaro | Voet, Doug | Rosenberg, Mara | Zack, Travis I. | Zhang, Hailei | Zou, Lihua | Protopopov, Alexei | Santoso, Netty | Parfenov, Michael | Lee, Semin | Zhang, Jianhua | Mahadeshwar, Harshad S. | Tang, Jiabin | Ren, Xiaojia | Seth, Sahil | Yang, Lixing | Xu, Andrew W. | Song, Xingzhi | Pantazi, Angeliki | Xi, Ruibin | Bristow, Christopher A. | Hadjipanayis, Angela | Seidman, Jonathan | Chin, Lynda | Park, Peter J. | Kucherlapati, Raju | Akbani, Rehan | Ling, Shiyun | Liu, Wenbin | Rao, Arvind | Weinstein, John N. | Kim, Sang-Bae | Lee, Ju-Seog | Lu, Yiling | Mills, Gordon | Laird, Peter W. | Hinoue, Toshinori | Weisenberger, Daniel J. | Bootwalla, Moiz S. | Lai, Phillip H. | Shen, Hui | Triche, Timothy | Van Den Berg, David J. | Baylin, Stephen B. | Herman, James G. | Getz, Gad | Chin, Lynda | Liu, Yingchun | Murray, Bradley A. | Noble, Michael S. | Askoy, B. Arman | Ciriello, Giovanni | Dresdner, Gideon | Gao, Jianjiong | Gross, Benjamin | Jacobsen, Anders | Lee, William | Ramirez, Ricardo | Sander, Chris | Schultz, Nikolaus | Senbabaoglu, Yasin | Sinha, Rileen | Sumer, S. Onur | Sun, Yichao | Weinhold, Nils | Thorsson, Vésteinn | Bernard, Brady | Iype, Lisa | Kramer, Roger W. | Kreisberg, Richard | Miller, Michael | Reynolds, Sheila M. | Rovira, Hector | Tasman, Natalie | Shmulevich, Ilya | Ng, Santa Cruz Sam | Haussler, David | Stuart, Josh M. | Akbani, Rehan | Ling, Shiyun | Liu, Wenbin | Rao, Arvind | Weinstein, John N. | Verhaak, Roeland G.W. | Mills, Gordon B. | Leiserson, Mark D. M. | Raphael, Benjamin J. | Wu, Hsin-Ta | Taylor, Barry S. | Black, Aaron D. | Bowen, Jay | Carney, Julie Ann | Gastier-Foster, Julie M. | Helsel, Carmen | Leraas, Kristen M. | Lichtenberg, Tara M. | McAllister, Cynthia | Ramirez, Nilsa C. | Tabler, Teresa R. | Wise, Lisa | Zmuda, Erik | Penny, Robert | Crain, Daniel | Gardner, Johanna | Lau, Kevin | Curely, Erin | Mallery, David | Morris, Scott | Paulauskis, Joseph | Shelton, Troy | Shelton, Candace | Sherman, Mark | Benz, Christopher | Lee, Jae-Hyuk | Fedosenko, Konstantin | Manikhas, Georgy | Potapova, Olga | Voronina, Olga | Belyaev, Smitry | Dolzhansky, Oleg | Rathmell, W. Kimryn | Brzezinski, Jakub | Ibbs, Matthew | Korski, Konstanty | Kycler, Witold | ŁaŸniak, Radoslaw | Leporowska, Ewa | Mackiewicz, Andrzej | Murawa, Dawid | Murawa, Pawel | Spychała, Arkadiusz | Suchorska, Wiktoria M. | Tatka, Honorata | Teresiak, Marek | Wiznerowicz, Maciej | Abdel-Misih, Raafat | Bennett, Joseph | Brown, Jennifer | Iacocca, Mary | Rabeno, Brenda | Kwon, Sun-Young | Penny, Robert | Gardner, Johanna | Kemkes, Ariane | Mallery, David | Morris, Scott | Shelton, Troy | Shelton, Candace | Curley, Erin | Alexopoulou, Iakovina | Engel, Jay | Bartlett, John | Albert, Monique | Park, Do-Youn | Dhir, Rajiv | Luketich, James | Landreneau, Rodney | Janjigian, Yelena Y. | Kelsen, David P. | Cho, Eunjung | Ladanyi, Marc | Tang, Laura | McCall, Shannon J. | Park, Young S. | Cheong, Jae-Ho | Ajani, Jaffer | Camargo, M. Constanza | Alonso, Shelley | Ayala, Brenda | Jensen, Mark A. | Pihl, Todd | Raman, Rohini | Walton, Jessica | Wan, Yunhu | Demchok, John A. | Eley, Greg | Mills Shaw, Kenna R. | Sheth, Margi | Tarnuzzer, Roy | Wang, Zhining | Yang, Liming | Zenklusen, Jean Claude | Davidsen, Tanja | Hutter, Carolyn M. | Sofia, Heidi J. | Burton, Robert | Chudamani, Sudha | Liu, Jia
Nature  2014;513(7517):202-209.
Gastric cancer is a leading cause of cancer deaths, but analysis of its molecular and clinical characteristics has been complicated by histological and aetiological heterogeneity. Here we describe a comprehensive molecular evaluation of 295 primary gastric adenocarcinomas as part of The Cancer Genome Atlas (TCGA) project. We propose a molecular classification dividing gastric cancer into four subtypes: tumours positive for Epstein–Barr virus, which display recurrent PIK3CA mutations, extreme DNA hypermethylation, and amplification of JAK2, CD274 (also known as PD-L1) and PDCD1LG2 (also knownasPD-L2); microsatellite unstable tumours, which show elevated mutation rates, including mutations of genes encoding targetable oncogenic signalling proteins; genomically stable tumours, which are enriched for the diffuse histological variant and mutations of RHOA or fusions involving RHO-family GTPase-activating proteins; and tumours with chromosomal instability, which show marked aneuploidy and focal amplification of receptor tyrosine kinases. Identification of these subtypes provides a roadmap for patient stratification and trials of targeted therapies.
doi:10.1038/nature13480
PMCID: PMC4170219  PMID: 25079317
2.  The Protective Effect of MicroRNA-320 on Left Ventricular Remodeling after Myocardial Ischemia-Reperfusion Injury in the Rat Model 
The primary objective of this study investigated the role of microRNA-320 (miR-320) on left ventricular remodeling in the rat model of myocardial ischemia-reperfusion (I/R) injury, and we intended to explore the myocardial mechanism of miR-320-mediated myocardium protection. We collected 120 male Wistar rats (240–280 g) in this study and then randomly divided them into three groups: (1) sham surgery group (sham group: n = 40); (2) ischemia-reperfusion model group (I/R group: n = 40); and (3) I/R model with antagomir-320 group (I/R + antagomir-320 group: n = 40). Value changes of heart function in transesophageal echocardiography were recorded at various time points (day 1, day 3, day 7, day 15 and day 30) after surgery in each group. Myocardial sections were stained with hematoxylin and eosin (H&E) and examined with optical microscope. The degree of myocardial fibrosis was assessed by Sirius Red staining. Terminal dUTP nick end-labeling (TUNEL) and qRT-PCR methods were used to measure the apoptosis rate and to determine the miR-320 expression levels in myocardial tissues. Transesophageal echocardiography showed that the values of left ventricular ejection fraction (LVEF), left ventricular fractional shortening (LVFS), left ventricular systolic pressure (LVSP) and ±dp/dtmax in the I/R group were obviously lower than those in the sham group, while the left ventricular end-diastolic pressure (LVEDP) value was higher than that in the sham group. The values of LVEF, LVFS, LVSP and ±dp/dtmax showed a gradual decrease in the I/R group, while the LVEDP value showed an up tendency along with the extension of reperfusion time. The H&E staining revealed that rat myocardial tissue in the I/R group presented extensive myocardial damage; for the I/R + antagomir-320 group, however, the degree of damage in myocardial cells was obviously better than that of the I/R group. The Sirius Red staining results showed that the degree of myocardial fibrosis in the I/R group was more severe along with the extension of the time of reperfusion. For the I/R + antagomir-320 group, the degree of myocardial fibrosis was less severe than that in the I/R group. Tissues samples in both the sham and I/R + antagomir-320 groups showed a lower apoptosis rate compared to I/R group. The qRT-PCR results indicated that miR-320 expression in the I/R group was significantly higher than that in both the sham and I/R + antagomir-320 groups. The expression level of miR-320 is significantly up-regulated in the rat model of myocardial I/R injury, and it may be implicated in the prevention of myocardial I/R injury-triggered left ventricular remodeling.
doi:10.3390/ijms151017442
PMCID: PMC4227171  PMID: 25268616
microRNA-320; myocardial ischemia-reperfusion injury; left ventricular remodeling
3.  Draft genome of the kiwifruit Actinidia chinensis 
Nature Communications  2013;4:2640.
The kiwifruit (Actinidia chinensis) is an economically and nutritionally important fruit crop with remarkably high vitamin C content. Here we report the draft genome sequence of a heterozygous kiwifruit, assembled from ~140-fold next-generation sequencing data. The assembled genome has a total length of 616.1 Mb and contains 39,040 genes. Comparative genomic analysis reveals that the kiwifruit has undergone an ancient hexaploidization event (γ) shared by core eudicots and two more recent whole-genome duplication events. Both recent duplication events occurred after the divergence of kiwifruit from tomato and potato and have contributed to the neofunctionalization of genes involved in regulating important kiwifruit characteristics, such as fruit vitamin C, flavonoid and carotenoid metabolism. As the first sequenced species in the Ericales, the kiwifruit genome sequence provides a valuable resource not only for biological discovery and crop improvement but also for evolutionary and comparative genomics analysis, particularly in the asterid lineage.
The kiwifruit is an economically and nutritionally important fruit crop with high vitamin C content. Here, the authors report the draft genome sequence of a heterozygous kiwifruit and through comparative genomic analysis provide valuable insight into kiwifruit evolution.
doi:10.1038/ncomms3640
PMCID: PMC4089393  PMID: 24136039
4.  Whole-Exome Sequencing to Decipher the Genetic Heterogeneity of Hearing Loss in a Chinese Family with Deaf by Deaf Mating 
PLoS ONE  2014;9(10):e109178.
Inherited deafness has been shown to have high genetic heterogeneity. For many decades, linkage analysis and candidate gene approaches have been the main tools to elucidate the genetics of hearing loss. However, this associated study design is costly, time-consuming, and unsuitable for small families. This is mainly due to the inadequate numbers of available affected individuals, locus heterogeneity, and assortative mating. Exome sequencing has now become technically feasible and a cost-effective method for detection of disease variants underlying Mendelian disorders due to the recent advances in next-generation sequencing (NGS) technologies. In the present study, we have combined both the Deafness Gene Mutation Detection Array and exome sequencing to identify deafness causative variants in a large Chinese composite family with deaf by deaf mating. The simultaneous screening of the 9 common deafness mutations using the allele-specific PCR based universal array, resulted in the identification of the 1555A>G in the mitochondrial DNA (mtDNA) 12S rRNA in affected individuals in one branch of the family. We then subjected the mutation-negative cases to exome sequencing and identified novel causative variants in the MYH14 and WFS1 genes. This report confirms the effective use of a NGS technique to detect pathogenic mutations in affected individuals who were not candidates for classical genetic studies.
doi:10.1371/journal.pone.0109178
PMCID: PMC4188603  PMID: 25289672
5.  Ingredients of Huangqi decoction slow biliary fibrosis progression by inhibiting the activation of the transforming growth factor-beta signaling pathway 
Background
Huangqi decoction was first described in Prescriptions of the Bureau of Taiping People's Welfare Pharmacy in Song Dynasty (AD 1078), and it is an effective recipe that is usually used to treat consumptive disease, anorexia, and chronic liver diseases. Transforming growth factor beta 1 (TGFβ1) plays a key role in the progression of liver fibrosis, and Huangqi decoction and its ingredients (IHQD) markedly ameliorated hepatic fibrotic lesions induced by ligation of the common bile duct (BDL). However, the mechanism of IHQD on hepatic fibrotic lesions is not yet clear. The purpose of the present study is to elucidate the roles of TGFβ1 activation, Smad-signaling pathway, and extracellular signal-regulated kinase (ERK) in the pathogenesis of biliary fibrosis progression and the antifibrotic mechanism of IHQD.
Methods
A liver fibrosis model was induced by ligation of the common bile duct (BDL) in rats. Sham-operation was performed in control rats. The BDL rats were randomly divided into two groups: the BDL group and the IHQD group. IHQD was administrated intragastrically for 4 weeks. At the end of the fifth week after BDL, animals were sacrificed for sampling of blood serum and liver tissue. The effect of IHQD on the TGFβ1 signaling pathway was evaluated by western blotting and laser confocal microscopy.
Results
Decreased content of hepatic hydroxyproline and improved liver function and histopathology were observed in IHQD rats. Hepatocytes, cholangiocytes, and myofibroblasts in the cholestatic liver injury released TGFβ1, and activated TGFβ1 receptors can accelerate liver fibrosis. IHQD markedly inhibited the protein expression of TGFβ1, TGFβ1 receptors, Smad3, and p-ERK1/2 expression with no change of Smad7 expression.
Conclusion
IHQD exert significant therapeutic effects on BDL-induced fibrosis in rats through inhibition of the activation of TGFβ1-Smad3 and TGFβ1-ERK1/2 signaling pathways.
doi:10.1186/1472-6882-12-33
PMCID: PMC3419610  PMID: 22471627
Ingredients of Huangqi decoction; Cholestatic liver fibrosis; Transforming growth factor beta 1; Smad-signaling pathway, Extracellular signal-regulated kinase
6.  The impact of educational status on the clinical features of major depressive disorder among Chinese women 
Journal of Affective Disorders  2012;136(3):988-992.
Background
Years of education are inversely related to the prevalence of major depressive disorder (MDD), but the relationship between the clinical features of MDD and educational status is poorly understood. We investigated this in 1970 Chinese women with recurrent MDD identified in a clinical setting.
Methods
Clinical and demographic features were obtained from 1970 Han Chinese women with DSM-IV major depression between 30 and 60 years of age across China. Analysis of linear, logistic and multiple logistic regression models were used to determine the association between educational level and clinical features of MDD.
Results
Subjects with more years of education are more likely to have MDD, with an odds ratio of 1.14 for those with more than ten years. Low educational status is not associated with an increase in the number of episodes, nor with increased rates of co-morbidity with anxiety disorders. Education impacts differentially on the symptoms of depression: lower educational attainment is associated with more biological symptoms and increased suicidal ideation and plans to commit suicide.
Limitations
Findings may not generalize to males or to other patient populations. Since the threshold for treatment seeking differs as a function of education there may an ascertainment bias in the sample.
Conclusions
The relationship between symptoms of MDD and educational status in Chinese women is unexpectedly complex. Our findings are inconsistent with the simple hypothesis from European and US reports that low levels of educational attainment increase the risk and severity of MDD.
doi:10.1016/j.jad.2011.06.046
PMCID: PMC3314924  PMID: 21824664
Major depressive disorder; Education; Socio-economic status; Symptom
7.  Hierarchical Macro-meso-microporous ZSM-5 Zeolite Hollow Fibers With Highly Efficient Catalytic Cracking Capability 
Scientific Reports  2014;4:7276.
Zeolite fibers have attracted growing interest for a range of new applications because of their structural particularity while maintaining the intrinsic performances of the building blocks of zeolites. The fabrication of uniform zeolite fibers with tunable hierarchical porosity and further exploration of their catalytic potential are of great importance. Here, we present a versatile and facile method for the fabrication of hierarchical ZSM-5 zeolite fibers with macro-meso-microporosity by coaxial electrospinning. Due to the synergistic integration of the suitable acidity and the hierarchical porosity, high yield of propylene and excellent anti-coking stability were demonstrated on the as-prepared ZSM-5 hollow fibers in the catalytic cracking reaction of iso-butane. This work may also provide good model catalysts with uniform wall thickness and tunable porosity for studying a series of important catalytic reactions.
doi:10.1038/srep07276
PMCID: PMC4250908  PMID: 25450726
8.  Oleanolic Acid Suppresses Aerobic Glycolysis in Cancer Cells by Switching Pyruvate Kinase Type M Isoforms 
PLoS ONE  2014;9(3):e91606.
Warburg effect, one of the hallmarks for cancer cells, is characterized by metabolic switch from mitochondrial oxidative phosphorylation to aerobic glycolysis. In recent years, increased expression level of pyruvate kinase M2 (PKM2) has been found to be the culprit of enhanced aerobic glycolysis in cancer cells. However, there is no agent inhibiting aerobic glycolysis by targeting PKM2. In this study, we found that Oleanolic acid (OA) induced a switch from PKM2 to PKM1, and consistently, abrogated Warburg effect in cancer cells. Suppression of aerobic glycolysis by OA is mediated by PKM2/PKM1 switch. Furthermore, mTOR signaling was found to be inactivated in OA-treated cancer cells, and mTOR inhibition is required for the effect of OA on PKM2/PKM1 switch. Decreased expression of c-Myc-dependent hnRNPA1 and hnRNPA1 was responsible for OA-induced switch between PKM isoforms. Collectively, we identified that OA is an antitumor compound that suppresses aerobic glycolysis in cancer cells and there is potential that PKM2 may be developed as an important target in aerobic glycolysis pathway for developing novel anticancer agents.
doi:10.1371/journal.pone.0091606
PMCID: PMC3953484  PMID: 24626155
9.  SCFβ-TRCP-mediated degradation of NEDD4 inhibits tumorigenesis through modulating the PTEN/Akt signaling pathway 
Oncotarget  2014;5(4):1026-1037.
The HECT domain-containing ubiquitin E3 ligase NEDD4 is widely expressed in mammalian tissues and plays a crucial role in governing a wide spectrum of cellular processes including cell growth, tissue development and homeostasis. Recent reports have indicated that NEDD4 might facilitate tumorigenesis through targeted degradation of multiple tumor suppressor proteins including PTEN. However, the molecular mechanism by which NEDD4 stability is regulated has not been fully elucidated. Here we report that SCFβ-TRCP governs NEDD4 protein stability by targeting it for ubiquitination and subsequent degradation in a Casein Kinase-I (CKI) phosphorylation-dependent manner. Specifically, depletion of β-TRCP, or inactivation of CKI, stabilized NEDD4, leading to down-regulation of its ubiquitin target PTEN and subsequent activation of the mTOR/Akt oncogenic pathway. Furthermore, we found that CKIδ-mediated phosphorylation of Ser347 and Ser348 on NEDD4 promoted its interaction with SCFβ-TRCP for subsequent ubiquitination and degradation. As a result, compared to ectopic expression of wild-type NEDD4, introducing a non-degradable NEDD4 (S347A/S348A-NEDD4) promoted cancer cell growth and migration. Hence, our findings revealed the CKI/SCFβ-TRCP signaling axis as the upstream negative regulator of NEDD4, and further suggested that enhancing NEDD4 degradation, presumably with CKI or SCFβ-TRCP agonists, could be a promising strategy for treating human cancers.
PMCID: PMC4011580  PMID: 24657926
β-TRCP; NEDD4; degradation; PTEN; Akt; cancer; phosphorylation; ubiquitination; therapy
10.  The efficacy and safety of electroacupuncture for women with pure stress urinary incontinence: study protocol for a multicenter randomized controlled trial 
Trials  2013;14:315.
Background
Although available evidence relating to its effectiveness is weak, acupuncture is used as an alternative therapy for stress urinary incontinence. We report a protocol of a randomized controlled trial using electroacupuncture (the passing of a weak current between inserted acupuncture needles) to treat women with pure stress urinary incontinence.
Methods/Design
This is a large-scale multicenter subject-blinded randomized controlled trial. A total of 500 women with pure stress urinary incontinence will be randomly assigned to two groups: a treatment group and a control group. The treatment group will receive electroacupuncture with deep needling at acupuncture points BL33 and BL35. The control group will receive sham electroacupuncture with non-penetrating needling at sham locations for the acupuncture points of BL33 and BL35. Participants will be given three sessions a week for 6 weeks. A 24-week-long follow-up will be conducted. The primary outcome will be the change in amount of urine leakage at the sixth week from a baseline measured by a 1-h pad test. The secondary outcomes include: the 72-h incontinence episode frequency based on a 72-h bladder diary; the score of International Consultation on Incontinence Questionnaire-Urinary Incontinence Short Form; the degree of urinary incontinence based on a 72-h bladder diary; self-assessment of the therapeutic effect; weekly consumption of pads; application of other treatments for stress urinary incontinence; and subgroup analysis stratified by incontinence severity. The safety of electroacupuncture will also be assessed.
Discussion
This trial will help to identify whether electroacupuncture is effective for stress urinary incontinence, and, if so, whether it is a therapeutic effect rather than a placebo effect.
Trial Registration
Clinical Trials.gov NCT01784172
doi:10.1186/1745-6215-14-315
PMCID: PMC3850726  PMID: 24079823
Electroacupuncture; Pure stress urinary incontinence; Efficacy; RCT; Study protocol
11.  Prevalence and Correlates of Discomfort and Acceptability of Acupuncture among Outpatients in Chinese Acupuncture and Moxibustion Departments: A Cross-Sectional Study 
Objective. This study aims to give a profile of discomfort and acceptability of acupuncture, including the prevalence and association with demographic and acupuncture-related factors. Methods. A cross-sectional study was conducted in Beijing, China. Outpatients of acupuncture and moxibustion departments were recruited using purposive sampling. 925 subjects were interviewed with an anonymous questionnaire. Multinomial and binary logistic regression were used to analyze factors affecting discomfort and acceptability of acupuncture. Results. The average VAS value of 925 subjects' acupuncture discomfort was 2.66 ± 2.02, within the range of mild discomfort. Acupuncture was easily accepted by 81.1% of the subjects. Results of logistic regression were as follows: (1) subjects with a better knowledge of acupuncture, or a greater fear of pain or needles, experienced more “moderate to severe discomfort” and showed a decreased acupuncture acceptance (P < 0.001 or P < 0.01); (2) Acupuncture with less discomfort or implemented by a more qualified doctor was easy to be accepted (P < 0.001); (3) subjects aged 20–29 preferred to report “moderate to severe discomfort” while those aged 40–59 preferred to report “slight discomfort” (P < 0.001). Conclusion. Acupuncture is an acceptable therapy with less discomfort, which can be greatly affected by fear of pain or needles, age, knowledge of acupuncture, and professional title of acupuncturist.
doi:10.1155/2013/715480
PMCID: PMC3707286  PMID: 23864895
12.  The efficacy and safety study of electro-acupuncture for severe chronic functional constipation: study protocol for a multicenter, randomized, controlled trial 
Trials  2013;14:176.
Background
Previous research has shown that electro-acupuncture (EA) may be effective for functional constipation. We report a protocol for a randomized controlled trial using EA to confirm the efficacy and safety for severe chronic functional constipation.
Methods/design
This is a randomized, controlled trial. A total of 1,034 patients will be randomly allocated into the EA group (n=517) and the sham EA group (n=517). The EA group receives needling at ST25, SP14 and ST37 and the sham EA group receives needling at sham ST25, SP14 and ST37. The primary outcome measure is the changed number of weekly average complete spontaneous bowel movements(CSBMs) during 8 weeks of treatment, compared with baseline. The secondary outcome measures are: 1) the proportion of participants having three or more CSBMs on average per week; 2) the changed number of weekly average CSBMs during weeks 9 to 20; 3) the changed number of weekly average spontaneous bowel movements during 8 weeks of treatment; 4) stool consistency; 5) degree of difficulty in defecation; 6) patient assessment of constipation quality of life questionnaire (PAC-QOL); 7) incidence of adverse events; and 8) usage of medicine for constipation.
Discussion
This trial will evaluate the efficacy and safety of EA for severe chronic functional constipation.
Trial registration
Protocol Registration System of ClinicalTrial.gov, NCT01726504
doi:10.1186/1745-6215-14-176
PMCID: PMC3706209  PMID: 23768191
Chronic functional constipation; Electro-acupuncture; Multicenter RCT; Efficacy; Safety
13.  Genetic Polymorphisms of Phase II Metabolic Enzymes and Lung Cancer Susceptibility in a Population of Central South China 
Disease Markers  2006;22(3):141-152.
A case-control study was conducted for analyzing the genetic polymorphisms of phase II metabolic enzymes in 97 patients with lung cancer and 197 healthy subjects from Han ethnic group of Hunan Province located in Central South China. The results showed that the frequencies of glutathione S-transferase (GST) M1-null (GSTM1-) or GSTT1-null (GSTT1-) genotype alone, or combined form of both in lung cancer patients were significantly higher than those of the controls. Genotypes of combining GSTP1 mutant/GSTM1(-) or GSTP1 mutant/GSTT1(-) led to high risk of lung cancer. Individuals carrying any two or all three of GSTM1(-), GSTT1(-) and GSTP1 mutant genotypes have a distinctly increased risk of lung cancer when compared to those with GSTM1 present (GSTM1+: GSTM1+/+ or GSTM1+/−), GSTT1 present (GSTT1+: GSTT1+/+ or GSTT1+/−) and GSTP1 wild genotypes. Furthermore, individuals possessing combined genotypes of N-acetyltransferase 2 (NAT2) rapid acetylator, GSTP1 mutant and both GSTT1(-) and GSTM1(-) have a remarkably higher lung cancer risk than those carrying combined NAT2 slow acetylator genotype, GSTP1 wild genotype and both GSTT1(+) and GSTM1(+) genotypes. All these findings suggest that the genetic polymorphisms of phase II metabolic enzymes affect the susceptibility of lung cancer in the Han ethnic group of Central South China.
doi:10.1155/2006/436497
PMCID: PMC3851385  PMID: 16788248
Genetic polymorphism; GST; NAT2; lung cancer; susceptibility
14.  Autoantibodies in Chinese patients with chronic hepatitis B: Prevalence and clinical associations 
AIM: To investigate the prevalence of autoantibodies and their associations with clinical features in Chinese patients with chronic hepatitis B (CHB).
METHODS: A total of 325 Chinese patients with CHB were enrolled in this retrospective, hospital-based study. Patients with chronic hepatitis C (CHC), autoimmune hepatitis (AIH), or primary biliary cirrhosis (PBC) were included, with healthy donors acting as controls. A panel of autoantibodies that serologically define AIH and PBC was tested by indirect immunofluorescence assay and line immunoassay. The AIH-related autoantibody profile included homogeneous anti-nuclear antibodies (ANA-H), smooth-muscle antibodies, anti-liver kidney microsome type 1, anti-liver cytosolic antigen type 1, and anti-soluble liver antigen/liver pancreas; the PBC-related antibodies were characterized by ANA-nuclear dots/membranous rim-like, anti-mitochondrial antibodies-M2 (AMA-M2), anti-BPO (recombinant antigen targeted by AMA-M2), anti-Sp100, anti-promyelocytic leukemia protein (anti-PML), and anti-gp210. The dichotomization of clustering was used to unequivocally designate the AIH or PBC profiles for each case. Anti-Ro52 antibodies were also tested.
RESULTS: The prevalence of any autoantibody in CHB amounted to 58.2%, which was similar to the 66.2% prevalence in CHC, significantly higher than the 6.7% in the healthy controls (P < 0.001), and lower than the 100% found in AIH and PBC (P = 0.004 and P < 0.001, respectively). There were more anti-PML and anti-gp210 antibodies among the CHB patients than the CHC patients (11.1% vs 0%, P = 0.003; 12.6% vs 0%, P < 0.001, respectively). The prevalence and titer of AMA, anti-BPO, anti-PML, and anti-gp210 were higher in PBC than in those with CHB. Among the CHB patients, the prevalence of ANA, especially ANA-H, was significantly lower in patients with compensated and decompensated cirrhosis compared with patients without cirrhosis. Thirty-eight cases of hepatocellular carcinoma (HCC) in CHB showed a significant difference compared with non-HCC patients in the prevalence of anti-PML (0% vs 12.5%, P = 0.013). Dichotomization of the autoantibodies revealed that the PBC profile was more prevalent in patients with CHB than in those with CHC, and that it was strongly correlated with both compensated and decompensated cirrhosis. In contrast, the prevalence of the AIH profile was significantly higher in non-cirrhosis patients with CHB than in those with compensated cirrhosis (18.5% vs 8.2%, P = 0.039). Moreover, the AIH profile was also closely associated with hepatitis B e-antigen positivity.
CONCLUSION: ANA-H could be an indicator of early-stage CHB. Dichotomizing the autoantibody profiles revealed that the PBC profile is strongly associated with cirrhosis in CHB.
doi:10.3748/wjg.v21.i1.283
PMCID: PMC4284347  PMID: 25574103
Autoantibodies; Chronic hepatitis B; Autoimmune hepatitis; Primary biliary cirrhosis; Cirrhosis; Hepatocellular carcinoma
15.  Non-alcoholic fatty liver disease associated with gallstones in females rather than males: a longitudinal cohort study in Chinese urban population 
BMC Gastroenterology  2014;14(1):213.
Background
Whether non-alcoholic fatty liver disease (NAFLD) is a risk factor for gallstones remains uncertain. Few longitudinal or cohort studies have been used to identify this relationship. The aim of this study was to confirm the association between NAFLD and gallstones in a longitudinal cohort of urban dwellers in China.
Methods
To elucidate the association between NAFLD and gallstones, we fitted a generalized estimating equation (GEE) model in a large-scale longitudinal cohort over 6 years, which included 11,200 participants with at least three regular health check-ups.
Results
A total of 498 cases of gallstones occurred during the 6-year follow-up, which resulted in a total incidence density of 12.73 per 1000 person-years (498/39, 135.5 person-years). The GEE analyses confirmed and clarified the association between NAFLD and gallstones (relative risk (RR) = 1.2381, 95% confidence interval (CI) = 1.003–1.528, P = 0.047) after adjusting for other potential confounding factors, especially in females (RR = 1.707, 95% CI = 1.245–2.341, P = 0.001).
Conclusions
NAFLD is associated with gallstones in an urban Chinese population from the middle to upper socioeconomic strata. Moreover, this association is more strongly apparent in females than in males. Further cohort studies must be conducted to confirm this association in the general population.
Electronic supplementary material
The online version of this article (doi:10.1186/s12876-014-0213-y) contains supplementary material, which is available to authorized users.
doi:10.1186/s12876-014-0213-y
PMCID: PMC4273434  PMID: 25496394
Gallstones; Non-alcoholic fatty liver disease (NAFLD); Longitudinal cohort study; Generalized estimated equation (GEE)
16.  Cocirculation of Three Hemagglutinin and Two Neuraminidase Subtypes of Avian Influenza Viruses in Huzhou, China, April 2013: Implication for the Origin of the Novel H7N9 Virus 
Journal of Virology  2014;88(11):6506-6511.
We detected three avian influenza hemagglutinin (HA) subtypes (H7, H9, and H5) and two neuraminidase (NA) subtypes (N9 and N2), as well as H7N9-related H9N9 reassortant intermediates, cocirculating among poultry in Huzhou, China, during April 2013. The results of our study reveal not only that Huzhou is one of the geographic origins of the novel H7N9 virus but also that cocirculation poses a potential threat to humans in the future.
doi:10.1128/JVI.03319-13
PMCID: PMC4093899  PMID: 24623437
17.  Isolation and Characterization of Marine Brevibacillus sp. S-1 Collected from South China Sea and a Novel Antitumor Peptide Produced by the Strain 
PLoS ONE  2014;9(11):e111270.
A Gram-positive, rod-shaped bacterium, designated as S-1, was isolated from a marine sediment sample collected from South China Sea. Phylogenetic analysis based on 16S rRNA gene sequence showed that S-1 belongs to the genus Brevibacillus. A novel cytotoxic peptide was isolated from the fermentation broth of the marine-derived bacterium Brevibacillus sp. S-1, using ion-exchange chromatography and reverse-phase HPLC chromatography. The molecular weight of this peptide was determined as 1570 Da by MALDI-TOF mass spectrometry, and its structure was proposed as a cyclic peptide elucidated by MALDI-TOF/TOF mass spectrometry and de novo sequencing. 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assay showed that this peptide exhibited cytotoxicity against BEL-7402 human hepatocellular carcinoma cells, RKO human colon carcinoma cells, A549 human lung carcinoma cells, U251 human glioma cells and MCF-7 human breast carcinoma cells. Additionally, SBP exhibited low cytotoxicity against HFL1 human normal fibroblast lung cells. The result suggested that the cytotoxic effect of the peptide is specific to tumor cells.
doi:10.1371/journal.pone.0111270
PMCID: PMC4220994  PMID: 25372839
18.  Dental Follicle Cells Rescue the Regenerative Capacity of Periodontal Ligament Stem Cells in an Inflammatory Microenvironment 
PLoS ONE  2014;9(10):e108752.
Aims
Periodontal ligament stem cells (PDLSCs) are one of the best candidates for periodontal regeneration. Their function could be impaired in periodontitis microenvironment. Dental follicle cells (DFCs), serving as precursor cells and mesenchymal stem cells, have intimate connection with PDLSCs. However, it is still unknown whether DFCs could provide a favorable microenvironment to improve the proliferation and differentiation capacity of PDLSCs from healthy subjects (HPDLSCs) and patients diagnosed with periodontitis (PPDLSCs).
Methods
HPDLSCs, PPDLSCs and DFCs were harvested and identified using microscopic and flow cytometric analysis. Then, the coculture systems of DFCs/HPDLSCs and DFCs/PPDLSCs were established with 0.4 µm transwell, in which all the detection indexs were obtained from HPDLSCs and PPDLSCs. The expression of stemness-associated genes was detected by real-time PCR, and the proliferation ability was assessed using colony formation and cell cycle assays. The osteogenic differentiation capacity was evaluated by real-time PCR, western blot, ALP activity, Alizarin Red S staining and calcium level analysis, while the adipogenic differentiation capacity was determined by real-time PCR and Oil Red O staining. The cell sheet formation in vitro was observed by HE staining and SEM, and the implantation effect in vivo was evaluated using HE staining and Masson’s trichrome staining.
Results
PPDLSCs had a greater proliferation capability but lower osteogenic and adipogenic potential than HPDLSCs. DFCs enhanced the proliferation and osteogenic/adipogenic differentiation of HPDLSCs and PPDLSCs to different degrees. Moreover, coculture with DFCs increased cell layers and extracellular matrix of HPDLSCs/PPDLSCs cell sheets in vitro and improved periodontal regeneration by HPDLSCs/PPDLSCs in vivo.
Conclusions
Our data suggest that the function of PPDLSCs could be damaged in the periodontitis microenvironment. DFCs appear to enhance the self-renewal and multi-differentiation capacity of both HPDLSCs and PPDLSCs, which indicates that DFCs could provide a beneficial microenvironment for periodontal regeneration using PDLSCs.
doi:10.1371/journal.pone.0108752
PMCID: PMC4183515  PMID: 25275580
19.  Extensive Recombination Due to Heteroduplexes Generates Large Amounts of Artificial Gene Fragments during PCR 
PLoS ONE  2014;9(9):e106658.
Artificial recombinants can be generated during PCR when more than two genetically distinct templates coexist in a single PCR reaction. These recombinant amplicons can lead to the false interpretation of genetic diversity and incorrect identification of biological phenotypes that do not exist in vivo. We investigated how recombination between 2 or 35 genetically distinct HIV-1 genomes was affected by different PCR conditions using the parallel allele-specific sequencing (PASS) assay and the next generation sequencing method. In a standard PCR condition, about 40% of amplicons in a PCR reaction were recombinants. The high recombination frequency could be significantly reduced if the number of amplicons in a PCR reaction was below a threshold of 1013–1014 using low thermal cycles, fewer input templates, and longer extension time. Heteroduplexes (each DNA strand from a distinct template) were present at a large proportion in the PCR products when more thermal cycles, more templates, and shorter extension time were used. Importantly, the majority of recombinants were identified in heteroduplexes, indicating that the recombinants were mainly generated through heteroduplexes. Since prematurely terminated extension fragments can form heteroduplexes by annealing to different templates during PCR amplification, recombination has a better chance to occur with samples containing different genomes when the number of amplicons accumulate over the threshold. New technologies are warranted to accurately characterize complex quasispecies gene populations.
doi:10.1371/journal.pone.0106658
PMCID: PMC4161356  PMID: 25211143
20.  Aplysin Sensitizes Cancer Cells to TRAIL by Suppressing P38 MAPK/Survivin Pathway 
Marine Drugs  2014;12(9):5072-5088.
TNF-related apoptosis-inducing ligand (TRAIL) is a tumor-selective apoptosis inducer and has been shown to be promising for treating various types of cancers. However, the application of TRAIL is greatly impeded by the resistance of cancer cells to its action. Studies show that overexpression of some critical pro-survival proteins, such as survivin, is responsible for TRAIL resistance. In this study, we found that Aplysin, a brominated compound from marine organisms, was able to restore the sensitivity of cancer cells to TRAIL both in vitro and in vivo. Aplysin was found to enhance the tumor-suppressing capacity of TRAIL on several TRAIL-resistant cancer cell lines. TRAIL-induced apoptosis was also potentiated in A549 and MCF7 cells treated with Aplysin. Survivin downregulation was identified as a mechanism by which Aplysin-mediated TRAIL sensitization of cancer cells. Furthermore, the activation of p38 MAPK was revealed in Aplysin-treated cancer cells, and its inhibitor SB203580 was able to abrogate the promoting effect of Aplysin on the response of cancer cells to TRAIL action, as evidenced by restored survivin expression, elevated cell survival and reduced apoptotic rates. In conclusion, we provided evidence that Aplysin acts as a sensitizer for TRAIL and its effect on p38 MAPK/survivin pathway may partially account for this activity. Considering its low cytotoxicity to normal cells, Aplysin may be a promising agent for cancer treatment in combination with TRAIL.
doi:10.3390/md12095072
PMCID: PMC4178493  PMID: 25257790
aplysin; TRAIL; p38 MAPK; survivin
21.  Individual Differences in Impulsivity Predict Head Motion during Magnetic Resonance Imaging 
PLoS ONE  2014;9(8):e104989.
Magnetic resonance imaging (MRI) provides valuable data for understanding the human mind and brain disorders, but in-scanner head motion introduces systematic and spurious biases. For example, differences in MRI measures (e.g., network strength, white matter integrity) between patient and control groups may be due to the differences in their head motion. To determine whether head motion is an important variable in itself, or just simply a confounding variable, we explored individual differences in psychological traits that may predispose some people to move more than others during an MRI scan. In the first two studies, we demonstrated in both children (N  =  245) and adults (N  =  581) that head motion, estimated from resting-state functional MRI and diffusion tensor imaging, was reliably correlated with impulsivity scores. Further, the difference in head motion between children with attention deficit hyperactivity disorder (ADHD) and typically developing children was largely due to differences in impulsivity. Finally, in the third study, we confirmed the observation that the regression approach, which aims to deal with motion issues by regressing out motion in the group analysis, would underestimate the effect of interest. Taken together, the present findings provide empirical evidence that links in-scanner head motion to psychological traits.
doi:10.1371/journal.pone.0104989
PMCID: PMC4141798  PMID: 25148416
22.  Development of an Acellular Tumor Extracellular Matrix as a Three-Dimensional Scaffold for Tumor Engineering 
PLoS ONE  2014;9(7):e103672.
Tumor engineering is defined as the construction of three-dimensional (3D) tumors in vitro with tissue engineering approaches. The present 3D scaffolds for tumor engineering have several limitations in terms of structure and function. To get an ideal 3D scaffold for tumor culture, A549 human pulmonary adenocarcinoma cells were implanted into immunodeficient mice to establish xenotransplatation models. Tumors were retrieved at 30-day implantation and sliced into sheets. They were subsequently decellularized by four procedures. Two decellularization methods, Tris-Trypsin-Triton multi-step treatment and sodium dodecyl sulfate (SDS) treatment, achieved complete cellular removal and thus were chosen for evaluation of histological and biochemical properties. Native tumor tissues were used as controls. Human breast cancer MCF-7 cells were cultured onto the two 3D scaffolds for further cell growth and growth factor secretion investigations, with the two-dimensional (2D) culture and cells cultured onto the Matrigel scaffolds used as controls. Results showed that Tris-Trypsin-Triton multi-step treated tumor sheets had well-preserved extracellular matrix structures and components. Their porosity was increased but elastic modulus was decreased compared with the native tumor samples. They supported MCF-7 cell repopulation and proliferation, as well as expression of growth factors. When cultured within the Tris-Trypsin-Triton treated scaffold, A549 cells and human colorectal adenocarcinoma cells (SW-480) had similar behaviors to MCF-7 cells, but human esophageal squamous cell carcinoma cells (KYSE-510) had a relatively slow cell repopulation rate. This study provides evidence that Tris-Trypsin-Triton treated acellular tumor extracellular matrices are promising 3D scaffolds with ideal spatial arrangement, biomechanical properties and biocompatibility for improved modeling of 3D tumor microenvironments.
doi:10.1371/journal.pone.0103672
PMCID: PMC4114977  PMID: 25072252
23.  Tumor suppressor p53 negatively regulates glycolysis stimulated by hypoxia through its target RRAD 
Oncotarget  2014;5(14):5535-5546.
Cancer cells display enhanced glycolysis to meet their energetic and biosynthetic demands even under normal oxygen concentrations. Recent studies have revealed that tumor suppressor p53 represses glycolysis under normoxia as a novel mechanism for tumor suppression. As the common microenvironmental stress for tumors, hypoxia drives the metabolic switch from the oxidative phosphorylation to glycolysis, which is crucial for survival and proliferation of cancer cells under hypoxia. The p53's role and mechanism in regulating glycolysis under hypoxia is poorly understood. Here, we found that p53 represses hypoxia-stimulated glycolysis in cancer cells through RRAD, a newly-identified p53 target. RRAD expression is frequently decreased in lung cancer. Ectopic expression of RRAD greatly reduces glycolysis whereas knockdown of RRAD promotes glycolysis in lung cancer cells. Furthermore, RRAD represses glycolysis mainly through inhibition of GLUT1 translocation to the plasma membrane. Under hypoxic conditions, p53 induces RRAD, which in turn inhibits the translocation of GLUT1 and represses glycolysis in lung cancer cells. Blocking RRAD by siRNA greatly abolishes p53's function in repressing glycolysis under hypoxia. Taken together, our results revealed an important role and mechanism of p53 in antagonizing the stimulating effect of hypoxia on glycolysis, which contributes to p53's function in tumor suppression.
PMCID: PMC4170611  PMID: 25114038
p53; glycolysis; RRAD; hypoxia; lung cancer
24.  Neuroprotective Effects of Jitai Tablet, a Traditional Chinese Medicine, on the MPTP-Induced Acute Model of Parkinson's Disease: Involvement of the Dopamine System 
Jitai tablet (JTT) is a traditional Chinese medicine used to treat neuropsychiatric disorders. We previously demonstrated that JTT treatment led to increased level of dopamine transporter (DAT) in the striatum, thus indicating that JTT might have therapeutic potential for Parkinson's disease (PD), which is characterized by dysregulated dopamine (DA) transmission and decreased striatal DAT expression. The aim of this study was to investigate the neuroprotective effect of JTT on MPTP-induced PD mice. Using locomotor activity test and rotarod test, we evaluated the effects of JTT (0.50, 0.15, or 0.05 g/kg) on MPTP-induced behavioral impairments. Tyrosine hydroxylase TH-positive neurons in the substantia nigra and DAT and dopamine D2 receptor (D2R) levels in the striatum were detected by immunohistochemical staining and/or autoradiography. Levels of DA and its metabolites were determined by HPLC. In MPTP-treated mice, behavioral impairments were alleviated by JTT treatment. Moreover, JTT protected against impairment of TH-positive neurons and attenuated the MPTP-induced decreases in DAT and D2R. Finally, high dose of JTT (0.50 g/kg) inhibited the MPTP-induced increase in DA metabolism rate. Taken together, results from our present study provide evidence that JTT offers neuroprotective effects against the neurotoxicity of MPTP and thus might be a potential treatment for PD.
doi:10.1155/2014/542383
PMCID: PMC3996930  PMID: 24799940
25.  Polymorphisms in XPD Gene Could Predict Clinical Outcome of Platinum-Based Chemotherapy for Non-Small Cell Lung Cancer Patients: A Meta-Analysis of 24 Studies 
PLoS ONE  2013;8(11):e79864.
Objective
Xeroderma pigmentosum group D (XPD) is an essential gene involved in the nucleotide excision repair (NER) pathway. Two commonly studied single nucleotide polymorphisms (SNPs) of XPD (Lys751Gln, A>C, rs13181; Asp312Asn, G>A, rs1799793) are implicated in the modulation of DNA repair capacity, thus related to the responses to platinum-based chemotherapy. Here we performed a meta-analysis to better evaluate the association between the two XPD SNPs and clinical outcome of platinum-based chemotherapy in non-small cell lung cancer (NSCLC) patients.
Methods
A comprehensive search of PubMed database was conducted to identify relevant articles. Primary outcomes included objective response (i.e., complete response + partial response vs. stable disease + progressive disease), progression-free survival (PFS) and overall survival (OS). The pooled and 95% confidence intervals (CIs) of ORs (odds ratios) and HRs (hazard ratios) were estimated using the fixed or random effect model.
Results
Twenty-four studies were eligible according to the inclusion criteria. None of the XPD Lys751Gln/Asp312Asn polymorphisms was associated with objective response, PFS or OS in NSCLC patients treated with platinum drugs. However, in stratified analysis by ethnicity, the XPD Lys751Gln (A>C) polymorphism was not significantly associated with increased response in Caucasians (OR = 1.35, 95%CI = 1.0–1.83, P = 0.122 for heterogeneity) but was associated with decreased PFS in Asians (HR = 1.39, 95%CI = 1.07–1.81, P = 0.879 for heterogeneity). Furthermore, a statistically significant difference existed in the estimates of effect between the two ethnicities (P = 0.014 for TR; P<0.001 for PFS).
Conclusions
XPD Lys751Gln (A>C) may have inverse predictive and prognostic role in platinum-based treatment of NSCLC according to different ethnicities. Further studies are needed to validate our findings.
doi:10.1371/journal.pone.0079864
PMCID: PMC3829883  PMID: 24260311

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