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1.  Increased expression of RRM2 by human papillomavirus E7 oncoprotein promotes angiogenesis in cervical cancer 
Wang, N | Zhan, T | Ke, T | Huang, X | Ke, D | Wang, Q | Li, H
British Journal of Cancer  2014;110(4):1034-1044.
Background:
The purpose of this study was to confirm that RRM2 as a novel target of HPVE7 involved in cervical cancer angiogenesis.
Methods:
Gene expression was analysed by RT-qPCR, western blot and immunohistochemistry in cervical cancer tissue and cell lines. Luciferase reporter assay was used to determine the activities of various RRM2 promoters. Secreted VEGF was measured by ELISA. RRM2-mediated capillary tube formation induced by HPVE7 in cervical cancer cells were evaluated using human umbilical vein endothelial cells in vitro. ROS induced by RRM2 in cercal cancer cells was confirmed by flow cytometry. The growth of cervical cancer cell overexpression RRM2 was examined by nude mouse xenograft.
Results:
RRM2 as a novel downstream target for HPVE7 was upregulated by it at the transcriptional level through the E7-pRb interaction and binding of E2F to the RRM2 promoter region. Immunohistochemical analysis showed that the level of RRM2 positively correlated with the HPVE7 level in human cervical cancer. Functionally, overexpression of RRM2 enhanced the expression of HIF-1α and VEGF via activation of the ERK1/2 signalling pathway in cervical cancer cells, and significantly associated with increased microvessel densities in cervical cancer tissues. In vitro, HPVE7 stimulated RRM2-dependent capillary tube formation by HUVECs, and RRM2-enhanced angiogenesis was VEGF dependent. RRM2-activated ERK1/2 pathway was mediated through production of ROS. In the xenograft mouse model, overexpression of RRM2 in cervical cancer cells enhanced tumour growth as well as microvessel densities.
Conclusion:
HPVE7 induces upregulation of RRM2, which then promotes cervical carcinogenesis via ROS-ERK1/2-HIF-1α-VEGF-induced angiogenesis. Thus, the inhibition of RRM2 activity may be a novel therapeutic strategy for human cervical cancer.
doi:10.1038/bjc.2013.817
PMCID: PMC3929894  PMID: 24423925
RRM2; HPVE7; ROS; angiogenesis; ERK1/2
3.  An injectable acoustic transmitter for juvenile salmon 
Scientific Reports  2015;5:8111.
Salmon recovery and the potential detrimental effects of dams on fish have been attracting national attention due to the environmental and economic implications. In recent years acoustic telemetry has been the primary method for studying salmon passage. However, the size of the existing transmitters limits the minimum size of fish that can be studied, introducing a bias to the study results. We developed the first acoustic fish transmitter that can be implanted by injection instead of surgery. The new injectable transmitter lasts four times longer and weighs 30% less than other transmitters. Because the new transmitter costs significantly less to use and may substantially reduce adverse effects of implantation and tag burden, it will allow for study of migration behavior and survival of species and sizes of fish that have never been studied before. The new technology will lead to critical information needed for salmon recovery and the development of fish-friendly hydroelectric systems.
doi:10.1038/srep08111
PMCID: PMC4309970  PMID: 25630763
4.  CtBP and Associated LSD1 Are Required for Transcriptional Activation by NeuroD1 in Gastrointestinal Endocrine Cells 
Molecular and Cellular Biology  2014;34(12):2308-2317.
Gene expression programs required for differentiation depend on both DNA-bound transcription factors and surrounding histone modifications. Expression of the basic helix-loop-helix (bHLH) protein NeuroD1 is restricted to endocrine cells in the gastrointestinal (GI) tract, where it is important for endocrine differentiation. RREB1 (RAS-responsive element binding protein 1), identified as a component of the CtBP corepressor complex, binds to nearby DNA elements to associate with NeuroD and potentiate transcription of a NeuroD1 target gene. Transcriptional activation by RREB1 depends on recruitment of CtBP with its associated proteins, including LSD1, through its PXDLS motifs. The mechanism of transcriptional activation by CtBP has not been previously characterized. Here we found that activation was dependent on the histone H3 lysine 9 (H3K9) demethylase activity of LSD1, which removes repressive methyl marks from dimethylated H3K9 (H3K9Me2), to facilitate subsequent H3K9 acetylation by the NeuroD1-associated histone acetyltransferase, P300/CBP-associated factor (PCAF). The secretin, β-glucokinase, insulin I, and insulin II genes, four known direct targets of NeuroD1 in intestinal and pancreatic endocrine cells, all show similar promoter occupancy by CtBP-associated proteins and PCAF, with acetylation of H3K9. This work may indicate a mechanism for selective regulation of transcription by CtBP and LSD1 involving their association with specific transcription factors and cofactors to drive tissue-specific transcription.
doi:10.1128/MCB.01600-13
PMCID: PMC4054299  PMID: 24732800
5.  Mitochondrial dysfunction-associated OPA1 cleavage contributes to muscle degeneration: preventative effect of hydroxytyrosol acetate 
Wang, X | Li, H | Zheng, A | Yang, L | Liu, J | Chen, C | Tang, Y | Zou, X | Li, Y | Long, J | Liu, J | Zhang, Y | Feng, Z
Cell Death & Disease  2014;5(11):e1521-.
Mitochondrial dysfunction contributes to the development of muscle disorders, including muscle wasting, muscle atrophy and degeneration. Despite the knowledge that oxidative stress closely interacts with mitochondrial dysfunction, the detailed mechanisms remain obscure. In this study, tert-butylhydroperoxide (t-BHP) was used to induce oxidative stress on differentiated C2C12 myotubes. t-BHP induced significant mitochondrial dysfunction in a time-dependent manner, accompanied by decreased myosin heavy chain (MyHC) expression at both the mRNA and protein levels. Consistently, endogenous reactive oxygen species (ROS) overproduction triggered by carbonyl cyanide 4-(trifluoromethoxy) phenylhydrazone (FCCP), a mitochondrial oxidative phosphorylation inhibitor, was accompanied by decreased membrane potential and decreased MyHC protein content. However, the free radical scavenger N-acetyl-L-cysteine (NAC) efficiently reduced the ROS level and restored MyHC content, suggesting a close association between ROS and MyHC expression. Meanwhile, we found that both t-BHP and FCCP promoted the cleavage of optic atrophy 1 (OPA1) from the long form into short form during the early stages. In addition, the ATPase family gene 3-like 2, a mitochondrial inner membrane protease, was also markedly increased. Moreover, OPA1 knockdown in myotubes was accompanied by decreased MyHC content, whereas NAC failed to prevent FCCP-induced MyHC decrease with OPA1 knockdown, suggesting that ROS might affect MyHC content by modulating OPA1 cleavage. In addition, hydroxytyrosol acetate (HT-AC), an important compound in virgin olive oil, could significantly prevent t-BHP-induced mitochondrial membrane potential and cell viability loss in myotubes. Specifically, HT-AC inhibited t-BHP-induced OPA1 cleavage and mitochondrial morphology changes, accompanied by improvement on mitochondrial oxygen consumption capacity, ATP productive potential and activities of mitochondrial complex I, II and V. Moreover, both t-BHP- and FCCP-induced MyHC decrease was sufficiently inhibited by HT-AC. Taken together, our data provide evidence indicating that mitochondrial dysfunction-associated OPA1 cleavage may contribute to muscle degeneration, and olive oil compounds could be effective nutrients for preventing the development of muscle disorders.
doi:10.1038/cddis.2014.473
PMCID: PMC4260731  PMID: 25393477
6.  THE ASSOCIATION BETWEEN RENAL FUNCTION BIOMARKERS AND SUBCLINICAL CARDIOVASCULAR MEASURES IN AFRICAN CARIBBEAN FAMILIES 
Ethnicity & disease  2013;23(4):492-498.
Background
Risk of cardiovascular disease (CVD) and mortality are increased in people with subclinical CVD. The impact of ethnicity and race on subclinical CVD is substantial. Previous studies assessed the heritability of several renal function biomarkers and their relationship with subclinical CVD among populations of European ancestries, but no such data is available in African ancestry populations.
Objective
Our aim was to investigate the relationships between renal function biomarkers and subclinical CVD among Afro-Caribbeans residing on the island of Tobago.
Design and Methods
402 participants aged 18 to 103 years from 7 large, multi-generation pedigrees (average family size: 50; range: 19 to 96; ~3500 relative pairs) were included in this study. Subclinical cardiovascular disease (SCVD) was assessed by brachial-ankle pulse wave velocity (baPWV) and carotid intima-media thickness (IMT). Serum cystatin C, creatinine, and eGFR based on the CKD-EPI (Chronic Kidney Disease Epidemiology Collaboration) equation were used to assess kidney function. The variance component approach, implemented in Sequential Oligogenic Linkage Analysis Routines (SOLAR), was used to assess heritability of these traits, and association with SCVD.
Results
Heritability of renal function biomarkers ranged from .19–.32 (all P< .001), and was highest for cystatin C (h2=.32, p<.0001). Serum cystatin C was independently associated with arterial stiffness (P=.04). This association was not found with other renal function biomarkers. No significant association between renal function and IMT was found.
Conclusion
Our data suggest that cystatin C is significantly heritable and associated with arterial stiffness among Afro-Caribbeans.
PMCID: PMC4059560  PMID: 24392614
7.  Sample size estimation for time-dependent ROC 
Statistics in medicine  2013;33(6):958-970.
In contrast to the usual ROC analysis with a contemporaneous reference standard, the time-dependent setting introduces the possibility that the reference standard refers to an event at a future time and may not be known for every patient due to censoring. The goal of this research is to determine the sample size required for a study design to address the question of the accuracy of a diagnostic test using the area under the curve in time-dependent ROC analysis. We adapt a previously published estimator of the time-dependent AUC which is a function of the expected conditional survival functions. This estimator accommodates censored data. The estimation of the required sample size is based on approximations of the expected conditional survival functions and their variances, derived under parametric assumptions of an exponential failure time and an exponential censoring time. We also consider different patient enrollment strategies. The proposed method can provide an adequate sample size to ensure that the test’s accuracy is estimated to a prespecified precision. We present results of a simulation study to assess the accuracy of the method and its robustness to departures from the parametric assumptions. We apply the proposed method to design of a study of PET as predictor of disease free survival in women undergoing therapy for cervical cancer.
doi:10.1002/sim.6005
PMCID: PMC4170805  PMID: 24123273
Censoring; Parametric survival model; Sample size estimation; Time-dependent AUC; Time-dependent ROC; Uniform accrual period
8.  Expression, Identification and Characterize of CD25-Binding Epitope Modified Human IL-2 in Pichia pastoris 
Indian Journal of Microbiology  2013;53(3):283-287.
Interleukin-2 (IL-2) plays important roles in variety of immune functions and it is widely used in the medication. But in recent years it was reported that vascular leak syndrome (VLS) was induced by IL-2. Evidences showed that the interaction of IL-2 and IL-2Rαβγ (CD25) caused VLS. Thus, this experiment modified the CD25-binding epitope in human IL-2 (hIL-2) to minimize the side effect of IL-2 in the medication. In this study, a recombinant human interleukin 2 (rhIL-2) was expressed in Pichia (P.) pastoris. An effective strategy was established to express rhIL-2 protein in 120 L scale and the optimal purification procedure was investigated. The purity of rhIL-2 in final product was about 98 % and the concentration of the rhIL-2 was 0.45 mg/mL. Bioactivity analysis showed that the purified rhIL-2 protein displayed high activity on proliferation of CTLL-2 cells and increased the ratio of CD4+/CD8+. It indicates that the target protein is expressed and the character of the rhIL-2 has high activity. This study provides a strategy for large-scale production of bioactive rhIL-2 protein using P. pastoris as an expression host.
doi:10.1007/s12088-013-0366-3
PMCID: PMC3689400  PMID: 24426123
IL-2; Expression; Purification; Immune function
9.  Integrated high-throughput analysis identifies Sp1 as a crucial determinant of p53-mediated apoptosis 
Cell Death and Differentiation  2014;21(9):1493-1502.
The restoration of p53 tumor suppressor function is a promising therapeutic strategy to combat cancer. However, the biological outcomes of p53 activation, ranging from the promotion of growth arrest to the induction of cell death, are hard to predict, which limits the clinical application of p53-based therapies. In the present study, we performed an integrated analysis of genome-wide short hairpin RNA screen and gene expression data and uncovered a previously unrecognized role of Sp1 as a central modulator of the transcriptional response induced by p53 that leads to robust induction of apoptosis. Sp1 is indispensable for the pro-apoptotic transcriptional repression by p53, but not for the induction of pro-apoptotic genes. Furthermore, the p53-dependent pro-apoptotic transcriptional repression required the co-binding of Sp1 to p53 target genes. Our results also highlight that Sp1 shares with p53 a common regulator, MDM2, which targets Sp1 for proteasomal degradation. This uncovers a new mechanism of the tight control of apoptosis in cells. Our study advances the understanding of the molecular basis of p53-mediated apoptosis and implicates Sp1 as one of its key modulators. We found that small molecules reactivating p53 can differentially modulate Sp1, thus providing insights into how to manipulate p53 response in a controlled way.
doi:10.1038/cdd.2014.69
PMCID: PMC4131181  PMID: 24971482
10.  An animal experimental study of porous magnesium scaffold degradation and osteogenesis 
Our objective was to observe the biodegradable and osteogenic properties of magnesium scaffolding under in vivo conditions. Twelve 6-month-old male New Zealand white rabbits were randomly divided into two groups. The chosen operation site was the femoral condyle on the right side. The experimental group was implanted with porous magnesium scaffolds, while the control group was implanted with hydroxyapatite scaffolds. X-ray and blood tests, which included serum magnesium, alanine aminotransferase (ALT), creatinine (CREA), and blood urea nitrogen (BUN) were performed serially at 1, 2, and 3 weeks, and 1, 2, and 3 months. All rabbits were killed 3 months postoperatively, and the heart, kidney, spleen, and liver were analyzed with hematoxylin and eosin (HE) staining. The bone samples were subjected to microcomputed tomography scanning (micro-CT) and hard tissue biopsy. SPSS 13.0 (USA) was used for data analysis, and values of P<0.05 were considered to be significant. Bubbles appeared in the X-ray of the experimental group after 2 weeks, whereas there was no gas in the control group. There were no statistical differences for the serum magnesium concentrations, ALT, BUN, and CREA between the two groups (P>0.05). All HE-stained slices were normal, which suggested good biocompatibility of the scaffold. Micro-CT showed that magnesium scaffolds degraded mainly from the outside to inside, and new bone was ingrown following the degradation of magnesium scaffolds. The hydroxyapatite scaffold was not degraded and had fewer osteoblasts scattered on its surface. There was a significant difference in the new bone formation and scaffold bioabsorption between the two groups (9.29±1.27 vs 1.40±0.49 and 7.80±0.50 vs 0.00±0.00 mm3, respectively; P<0.05). The magnesium scaffold performed well in degradation and osteogenesis, and is a promising material for orthopedics.
doi:10.1590/1414-431X20144009
PMCID: PMC4165299  PMID: 25098717
Biodegradable; Osteogenic; Magnesium scaffold
11.  Anthropometric measures and epithelial ovarian cancer risk among Chinese women: results from the Shanghai Women's Health Study 
British Journal of Cancer  2013;109(3):751-755.
Background:
Studies of anthropometric measures and ovarian cancer risk have predominantly included women of European descent with mixed findings.
Methods:
Data from the prospective Shanghai Women's Health Study (SWHS) were used to evaluate associations between anthropometric measures and risk of epithelial ovarian cancer (EOC). Hazard ratios (HRs) and 95% confidence intervals (CIs) were estimated by Cox proportional hazards regression.
Results:
A total of 152 EOC cases occurred among 70 258 women. Increasing quartiles of weight, hip circumference, and weight gain during adulthood were associated with significantly increased EOC risks. Body mass index (BMI) was also associated; overweight (25⩽BMI<29.99) and obese women (BMI⩾30.0) had significantly increased risks (HR: 1.49, 95% CI: 1.05, 2.13, and HR: 2.42, 95% CI: 1.37, 4.28, respectively). No significant associations were observed for height, waist circumference, waist-to-hip ratio (WHR), and waist-to-height ratio (WHER).
Conclusion:
Results from this large prospective study of Chinese women support the hypothesis that general adiposity contributes to the aetiology of ovarian cancer.
doi:10.1038/bjc.2013.384
PMCID: PMC3738128  PMID: 23860524
adiposity; obesity; body mass index; ovarian cancer; prospective cohort
12.  Identification and biochemical characterization of Laodelphax striatellus neutral ceramidase 
Insect molecular biology  2013;22(4):10.1111/imb.12028.
Ceramidases are a group of enzymes that catalyze hydrolysis of ceramides to generate fatty acid and sphingosine. In this study, we report the cloning and characterization of the rice small brown planthopper Laodelphax striatellus neutral ceramidase (nCDase), LsnCer. LsnCer was identified by sequencing the transcriptome of Laodelphax striatellus. LsnCer is a protein of 717 amino acids with a predicted molecular weight of 79.3 kDa. Similar to other known nCDases, LsnCer has a pH optimum at 8.0 and a temperature optimum at 37 °C for its in vitro activity. LsnCer activity is inhibited by Zn2+ significantly and Fe2+ slightly. LsnCer has broad substrate specificity with preference for ceramides with a medium acyl-chain or a mono unsaturated long acyl-chain. Infection with the rice strip virus (RSV) or treatment with insecticides significantly increased LsnCer mRNA expression and its enzymatic activity in L. striatellus. These results suggest that LsnCer is a bona fide nCDase that may have a role in adaption of L. striatellus to environmental stresses.
doi:10.1111/imb.12028
PMCID: PMC3879266  PMID: 23601004
nCDase; LsnCer; activity; mRNA; rive strip virus; insecticide
13.  Comparative analysis with collagen type II distinguishes cartilage oligomeric matrix protein as a primary TGFβ-responsive gene 
Summary
Objective
This study aims to investigate the regulation of expression of Cartilage oligomeric matrix protein (COMP), which is predominately expressed by chondrocytes and functions to organize the extracellular matrix. Mutations in COMP cause two skeletal dysplasias: pseudoachondroplasia and multiple epiphyseal dysplasia. The mechanism controlling COMP expression during chondrocyte differentiation is still poorly understood.
Design
Primary human bone marrow-derived stem cells were induced to differentiate into chondrocyte by pellet cultures. We then compared the temporal expression of COMP with the well-characterized cartilage-specific Type II collagen (Col2a1), and their response to transforming growth factor (TGF) β and Sox trio (Sox5, 6, and 9) stimulation.
Results
COMP and Col2a1 expression are differentially regulated by three distinct mechanisms. First, upregulation of COMP mRNA precedes Col2a1 by several days during chondrogenesis. Second, COMP expression is independent of high cell density but requires TGF-β1. Induction of COMP mRNA by TGF-β1 is detected within 2 h in the absence of protein synthesis and is blocked by specific inhibitors of the TGFβ signaling pathway; and therefore, COMP is a primary TFGβ-response gene. Lastly, while Col2a1 expression is intimately controlled by the Sox trio, overexpression of Sox trio fails to activate the COMP promoter.
Conclusion
COMP and Col2a1 expression are regulated differently during chondrogenesis. COMP is a primary response gene of TGFβ and its fast induction during chondrogenesis suggests that COMP is suitable for rapidly accessing the chondrogenic potential of stem cells.
doi:10.1016/j.joca.2011.07.011
PMCID: PMC4098880  PMID: 21843649
Chondrocyte differentiation; Chondrogenesis; TGFβ; Sox9
14.  Nanogranular TiN-ZrO2 intermediate layer induced improvement of isolation and grain size of FePt thin films 
Scientific Reports  2014;4:5607.
The effects of TiN-ZrO2 intermediate layer on the microstructures and magnetic properties of FePt films were investigated. The TiN-ZrO2 intermediate layer was granular consisting of grains of solid solution of Ti(Zr)ON segregated by amorphous ZrO2. By doping ZrO2 into TiN intermediate layer, the FePt grains became better isolated from each other and the FePt grain size was reduced. For 20 vol. % ZrO2 doping into TiN, the grain size decreased dramatically from 11. 2 nm to 6. 4 nm, and good perpendicular anisotropy was achieved simultaneously. For the FePt 4nm-SiO2 35 vol. % -C 20 vol. % films grown on top of the TiN-ZrO2 20 vol. % intermediate layer, well isolated FePt (001) granular films with coercivity higher than 18. 1 kOe and an average size as small as 6. 4 nm were achieved.
doi:10.1038/srep05607
PMCID: PMC4085629  PMID: 25001593
15.  Cruciferous vegetables consumption and the risk of female lung cancer: a prospective study and a meta-analysis 
Annals of Oncology  2013;24(7):1918-1924.
Background
Epidemiological studies evaluating the association between cruciferous vegetables (CVs) intake and female lung cancer risk have produced inconsistent results.
Patients and methods
This study followed 74 914 Chinese women aged 40–70 years who participated in the Shanghai Women's Health Study. CV intake was assessed through a validated food-frequency questionnaire (FFQ) at baseline and reassessed during follow-up. Hazard ratios (HRs) and 95% confidence interval (CIs) were estimated by using Cox proportional hazards models. Furthermore, we carried out a meta-analysis of all observational studies until December 2011.
Results
After excluding the first 2 years of follow-up, 417 women developed lung cancer over a mean of 11.1 years of follow-up. An inverse association of borderline statistical significance was observed between CV consumption and female lung cancer risk, with HR for the highest compared with the lowest quartiles of 0.73 (95% CI 0.54–1.00, P trend = 0.1607). The association was strengthened in analyses restricting to never smokers, with the corresponding HR of 0.59 (95% CI 0.40–0.87, P trend = 0.0510). The finding of an inverse association between CV intake and lung cancer risk in women was supported by our meta-analysis of 10 included studies.
Conclusions
Our study suggests that CV consumption may reduce the risk of lung cancer in women, particularly among never smokers.
doi:10.1093/annonc/mdt119
PMCID: PMC3690909  PMID: 23553059
cruciferous vegetable; lung cancer; meta-analysis; prospective study; women
16.  The androgen receptor mediates antiapoptotic function in myometrial cells 
Cell Death & Disease  2014;5(7):e1338-.
During pregnancy, myometrial phenotype is programmed into three characteristic stages referred to as the early proliferative, the midterm hypertrophic, and the late contractile stage. Increased myometrial growth in the early and midterm of pregnancy involves a complex process of cell proliferation, antiapoptosis and differentiation. We have previously demonstrated that the androgen receptor (AR) is required for myometrial cell proliferation by modulating IGF-1 signaling during early pregnancy. Here, we report that AR also exerts its antiapoptotic function in human myometrial cells. Enhanced AR expression protects, whereas AR silencing sensitizes myometrial cells to both intrinsic and extrinsic apoptotic stimuli. AR agonist inhibits, whereas AR antagonist induces myometrial cells to undergo apoptotic cell death. Gene microarray analysis confirms that the central functions of AR in myometrial cells are to regulate cell cycling and apoptosis through three major gene groups involving the epidermal growth factor (EGF) signaling, RNA splicing and DNA repair processes. AR mediates its antiapoptotic function through two distinct pathways. In the receptor-dependent pathway, AR is required for the expression of several protein factors within the EGF signaling pathway. Through the PI3K/Akt pathway, AR enhances the expression of the antiapoptotic protein Mcl-1. In the ligand-dependent pathway, AR agonist triggers the activation of Src kinase, which in turn phosphorylates STAT3 to increase Mcl-1 expression. We conclude from these results that the AR signaling exerts antiapoptotic function in myometrial cells, further supporting its key role in programming of myometrial phenotype.
doi:10.1038/cddis.2014.303
PMCID: PMC4123094  PMID: 25032861
17.  miR-17 extends mouse lifespan by inhibiting senescence signaling mediated by MKP7 
Cell Death & Disease  2014;5(7):e1355-.
Here we show that transgenic expression of miR-17 extends lifespan and inhibits cellular senescence. We propose that miR-17 acts as a critical regulator of cellular senescence and tumorigenesis. We demonstrate that miR-17 targets both ADCY5 and IRS1, upregulating the downstream signals MKP7, FoxO3, LC3B, and HIF1α, and downregulating mTOR, c-myc, cyclin D1, and JNK. Silencing either ADCY5 or IRS1 promoted autophagy and repressed cellular senescence and apoptosis. Repression of ADCY5 by miR-17 translocated membrane-bound RGS2 into the nucleus, promoting interactions of RGS2 with HIF1α and the MKP7 promoter, enhancing MKP7 transcription. ADCY5 repression by miR-17 also facilitated the translocation of EGFR and MKP7 from membrane into cytoplasmic and mitochondrial fractions. Importantly, we found that MKP7 inhibited senescence by dephosphorylating PRAS40 at Thr246 and mTOR at Ser2248, facilitating the interaction and loss of function of both molecules. Thus, the oncogenic miR-17 also acts pleiotropically to inhibit cellular senescence and extend longevity.
doi:10.1038/cddis.2014.305
PMCID: PMC4123096  PMID: 25077541
18.  Performance of KCl:Eu2+ storage phosphor dosimeters for low dose measurements 
Physics in medicine and biology  2013;58(12):4357-4366.
Recent research has demonstrated that europium doped potassium chloride (KCl:Eu2+) storage phosphor material has the potential to become the physical foundation of a novel and reusable dosimetry system using either film-like devices or devices similar to thermoluminescent dosimeter (TLD) chips. The purposes of this work are to quantify the performance of KCl:Eu2+ prototype dosimeters for low dose measurements and to demonstrate how it can be incorporated into clinical application for in vivo peripheral dose measurements. Pellet-style KCl:Eu2+ dosimeters, 6 mm in diameter, and 1 mm thick, were fabricated in-house for this study. The dosimeters were read using a laboratory photostimulated luminescence detection system. KCl:Eu2+ prototype storage phosphor dosimeter was capable of measuring a dose-to-water as low as 0.01 cGy from a 6 MV photon beam with a signal-to-noise ratio greater than 6. A pre-readout thermal annealing procedure enabled the dosimeter to be read within an hour post irradiation. After receiving large accumulated doses (~10 kGy), the dosimeters retained linear response in the low dose region with only a 20 percent loss of sensitivity comparing to a fresh sample (zero Gy history). The energy-dependence encountered during low dose peripheral measurements could be accounted for via a single point outside-field calibration per each beam quality. With further development the KCl:Eu2+− based dosimeter could become a versatile and durable dosimetry tool with large dynamic range (sub-cGy to 100 Gy).
doi:10.1088/0031-9155/58/12/4357
PMCID: PMC3774152  PMID: 23735856
radiation therapy dosimetry; dosimeter; storage phosphor
19.  Prospective evaluation of type 2 diabetes mellitus on the risk of primary liver cancer in Chinese men and women 
Annals of Oncology  2013;24(6):1679-1685.
Background
No prospective study has investigated the relationship between type 2 diabetes mellitus (T2DM) and the risk of primary liver cancer (PLC) in mainland China, and little is known about the effect of diabetes duration on PLC risk.
Design
Data from two population-based cohorts (the Shanghai Men's Health Study, SMHS, 2002–2006 and the Shanghai Women's Health Study, SWHS, 1996–2000) were thus used to assess the associations among T2DM, diabetes duration and PLC risk in Chinese population.
Results
During follow-up through 2009, 344 incident PLC cases were identified among 60 183 men and 73 105 women. T2DM is significantly associated with the increased risk of PLC in both men [hazard ratio (HR) = 1.63, 95% confidence interval (CI) 1.06–2.51] and women (HR = 1.64, 95% CI 1.03–2.61). The highest risk of incident liver cancer was observed in the first 5 years after diabetes diagnosis, and decreased substantially with the prolonged diabetes duration (Ptrend < 0.001). No synergistic interaction in the development of PLC was found between diabetes and other known risk factors.
Conclusions
T2DM is associated with the increased risk of subsequent liver cancer within 5 years after diagnosis in Chinese population, suggesting that hyperinsulinaemia rather than hyperglycaemia is more likely to be a primary mediator for this association.
doi:10.1093/annonc/mdt017
PMCID: PMC3660077  PMID: 23406734
China; cohort study; primary liver cancer; type 2 diabetes
20.  IRF4 is a novel mediator for neuronal survival in ischaemic stroke 
Cell Death and Differentiation  2014;21(6):888-903.
Neuroprotection following ischaemic stroke is driven by the interplay between regulatory transcription factors and endogenous protective factors. IRF4, a member of the interferon regulatory factor (IRF) family, is implicated in the survival of tumour cells. However, its role in the survival of normal cells including neurons remains elusive. Using genetic approaches, we established a central role for IRF4 in protection against ischaemia/reperfusion (I/R)-induced neuronal death. IRF4 was expressed in neurons, and induced by ischaemic stroke. Neuron-specific IRF4 transgenic (IRF4-TG) mice exhibited reduced infarct lesions, and this effect was reversed in IRF4-knockout mice. Notably, we revealed that IRF4 rescues neurons from I/R-induced death both in vivo and in vitro. Integrative transcriptional and cell survival analyses showed that IRF4 functions mechanistically as a transcription activator of serum response factor (SRF) crucial to salvage neurons during stroke. Indeed, the expression of SRF and SRF-dependent molecules was significantly upregulated upon IRF4 overexpression and conversely inhibited upon IRF4 ablation. Similar results were observed in oxygen glucose deprivation (OGD)-treated primary cortical neurons. Furthermore, we identified the IRF4-binding site in the promoter region of the SRF gene essential for its transcription. To verify the IRF4–SRF axis in vivo, we generated neuron-specific SRF knockout mice, in which SRF exerted profound cerebroprotective effects similar to those of IRF4. More importantly, the phenotype observed in IRF4-TG mice was completely reversed by SRF ablation. Thus, we have shown that the IRF4–SRF axis is a novel signalling pathway critical for neuronal survival in the setting of ischaemic stroke.
doi:10.1038/cdd.2014.9
PMCID: PMC4013523  PMID: 24510125
IRF4; SRF; ischaemic stroke; neuronal survival
21.  Taxonomy and multi-gene phylogeny of Datronia(Polyporales, Basidiomycota) 
Taxonomic and phylogenetic studies on Datronia were carried out. Phylogeny based on ITS, nLSU and RBP2 regions revealed that Datronia in current sense includes species belonging to three distantly related clades in polypores. The Datronia in a restricted sense is proposed for the clade including the type species D. mollis and D. stereoides. Neodatronia gen. nov. was proposed for two new resupinate species, N. gaoligongensis and N. sinensis. Species of Neodatronia differ from Datronia s.s. by their resupinate basidiomes, moderately to frequently branched skeletal hyphae in subiculum. Datroniella gen. nov., typified by D. scutellata was proposed for species in the other clade. Four new species of Datroniella, D. melanocarpa, D. subtropica, D. tibetica and D. tropica, were identified. Species of Datroniella differ from Datronia s.s. by their moderately to frequently branched skeletal hyphae in context and absence of dendrohyphidia. While, differentiate from Neodatronia by their small pileate, effused-reflexed or rarely resupinate basidiomes and absence of dendrohyphidia. Illustrated descriptions of the new species and two new genera are provided. The main morphological differences between Datronia, Datroniella, Neodatronia and related genera are discussed, identification keys to related genera and species in each genus are provided.
doi:10.3767/003158514X681828
PMCID: PMC4150076  PMID: 25264389
ITS; nLSU; Polyporaceae; RPB2; wood-inhabiting fungi
22.  Targeting Bax interaction sites reveals that only homo-oligomerization sites are essential for its activation 
Peng, R | Tong, J-S | Li, H | Yue, B | Zou, F | Yu, J | Zhang, L
Cell Death and Differentiation  2013;20(5):744-754.
Bax is a proapoptotic Bcl-2 family member that has a central role in the initiation of mitochondria-dependent apoptosis. However, the mechanism of Bax activation during apoptosis remains unsettled. It is believed that the activation of Bax is mediated by either dissociation from prosurvival Bcl-2 family members, or direct association with BH3-only members. Several interaction sites on Bax that mediate its interactions with other Bcl-2 family members, as well as its proapoptotic activity, have been identified in previous studies by other groups. To rigorously investigate the functional role of these interaction sites, we knocked in their respective mutants using HCT116 colon cancer cells, in which apoptosis induced by several stimuli is strictly Bax-dependent. Bax-mediated apoptosis was intact upon knock-in (KI) of K21E and D33A, which were shown to block the interaction of Bax with BH3-only activators. Apoptosis was partially reduced by KI of D68R, which impairs the interaction of Bax with prosurvival members, and S184V, a constitutively mitochondria-targeting mutant. In contrast, apoptosis was largely suppressed by KI of L70A/D71A, which blocks homo-oligomerization of Bax and its binding to prosurvival Bcl-2 family proteins. Collectively, our results suggest that the activation of endogenous Bax in HCT116 cells is dependent on its homo-oligomerization sites, but not those previously shown to interact with BH3-only activators or prosurvival proteins only. We therefore postulate that critical interaction sites yet to be identified, or mechanisms other than protein-protein interactions, need to be pursued to delineate the mechanism of Bax activation during apoptosis.
doi:10.1038/cdd.2013.4
PMCID: PMC3619242  PMID: 23392123
Bax; apoptosis; mitochondria; Bcl-2 family; caspases
23.  Model-Based Meta-Analysis for Quantifying Paclitaxel Dose Response in Cancer Patients 
Lu, D | Joshi, A | Li, H | Zhang, N | Ren, M M | Gao, Y | Wada, R | Jin, J Y
Model-based meta-analysis of dose response is a sophisticated method to guide dose and regimen selection. In this report, the effects of paclitaxel dose and regimen (weekly or every 3 weeks) on the efficacy and safety in cancer patients were quantified by model-based meta-analysis of 29 monotherapy trials. Logistic regression models were developed to assess the relationship between dose and objective response rate or neutropenia rate. Survival models were developed to assess the relationship between dose and overall survival or progression-free survival. Paclitaxel efficacy (e.g., objective response rate, median overall survival, and progression-free survival) is correlated with average dose per week (mg/m2/week), whereas safety (e.g., neutropenia rate) is correlated with dose per administration (mg/m2). Weekly paclitaxel regimen at 65–80 mg/m2 is supported to have comparable to better efficacy and lower neutropenia incidence than an every-3-week regimen at 175 mg/m2.
doi:10.1038/psp.2014.14
PMCID: PMC4055787  PMID: 24850445
24.  Empirical Selection of Informative Microsatellite Markers within Co-ancestry Pig Populations Is Required for Improving the Individual Assignment Efficiency 
The Lanyu is a miniature pig breed indigenous to Lanyu Island, Taiwan. It is distantly related to Asian and European pig breeds. It has been inbred to generate two breeds and crossed with Landrace and Duroc to produce two hybrids for laboratory use. Selecting sets of informative genetic markers to track the genetic qualities of laboratory animals and stud stock is an important function of genetic databases. For more than two decades, Lanyu derived breeds of common ancestry and crossbreeds have been used to examine the effectiveness of genetic marker selection and optimal approaches for individual assignment. In this paper, these pigs and the following breeds: Berkshire, Duroc, Landrace and Yorkshire, Meishan and Taoyuan, TLRI Black Pig No. 1, and Kaohsiung Animal Propagation Station Black pig are studied to build a genetic reference database. Nineteen microsatellite markers (loci) provide information on genetic variation and differentiation among studied breeds. High differentiation index (FST) and Cavalli-Sforza chord distances give genetic differentiation among breeds, including Lanyu’s inbred populations. Inbreeding values (FIS) show that Lanyu and its derived inbred breeds have significant loss of heterozygosity. Individual assignment testing of 352 animals was done with different numbers of microsatellite markers in this study. The testing assigned 99% of the animals successfully into their correct reference populations based on 9 to 14 markers ranking D-scores, allelic number, expected heterozygosity (HE) or FST, respectively. All miss-assigned individuals came from close lineage Lanyu breeds. To improve individual assignment among close lineage breeds, microsatellite markers selected from Lanyu populations with high polymorphic, heterozygosity, FST and D-scores were used. Only 6 to 8 markers ranking HE, FST or allelic number were required to obtain 99% assignment accuracy. This result suggests empirical examination of assignment-error rates is required if discernible levels of co-ancestry exist. In the reference group, optimum assignment accuracy was achievable achieved through a combination of different markers by ranking the heterozygosity, FST and allelic number of close lineage populations.
doi:10.5713/ajas.2013.13365
PMCID: PMC4093195  PMID: 25049996
Microsatellite Markers; Pigs; Assignment Test
25.  Transcriptome Analysis from Low Cell Numbers: Two RNA-Amplification Approaches 
Laser capture microdissection (LCM) permits the precise isolation of small populations of cells from complex tissue. Without the need for purification, the limited quantities of total RNA in LCM samples can be amplified for global gene expression profiling by RNA-Seq. In this study, two RNA amplification methods were evaluated. As input, 10, 30 and 100 human metastatic melanoma cells (c8161) were harvested by LCM after transplantation into the chick embryonic neural crest microenvironment. For comparison, 10 to 300 cultured c8161 cells were processed in identical manner. The amplified cDNA samples generated with either the SMARTer Ultra Low RNA Kit for Illumina Sequencing (Clontech) or the Ovation RNA-Seq System V2 kit (NuGEN) were fragmented prior to Illumina TruSeq library construction. As references, libraries were prepared from 10 ng of amplified and 1 μg of unamplified total RNA isolated from ∼7 million cultured cells. We show that both kits can be used to perform quantitative transcriptome analysis with as few as 10 cells. The cDNA yields obtained with the NuGEN kit were significantly higher than the Clontech yields. The number of expressed genes (>0 FPKM) increased with higher cell numbers, particularly for the culture-derived samples. Gene length and transcript abundance positively affected correlation with the unamplified reference for both the LCM and culture-derived samples. Considerably different expression profiles were observed for the samples amplified with the NuGEN versus Clontech methods which may be attributed to the different chemistries. Therefore, caution is advised when directly comparing inter-kit data sets.
PMCID: PMC4162268

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