dataesr/scientific-paragraphs-categorization huggingface dataset:
Another study, focusing specifically on the phylogeny of NATs in fungi, has also been published since then  [3]. In the present study, we expanded the previous survey by Glenn et al. [2]  by performing a comprehensive search of NAT-like sequences in the genomes of vertebrate species and compiled an up-to-date dataset of 77 vertebrate NAT sequences from 38 distinct species, among which 26 were identified for the first time (Figure  1 ). In contrast to the previous dataset of 55 vertebrate sequences in Glenn et al. [2] , we decided to exclude one lizard and three fish sequences from analysis (Anolis carolinensis NAT3, Oryzias latipes NAT3, Fugu rubripes NAT1, Tetraodon nigroviridis NAT1) because they aligned poorly with other vertebrate NATs and introduced too many gaps and alignment ambiguities.
Interestingly, codon 102 is also known to be involved in CoA binding and codon 272 is located in the C-terminus region of the protein. Moreover, two codon sites (173 and 191) were pinpointed to be under diversifying selection throughout simian NAT2 evolution. While the functional and/or structural significance of these two codons have not yet been explored, they are likely to be important in the function of the NAT2 protein and further investigations are warranted to define their potential relevance. In contrast to these positively selected codons, very low values of ω, suggesting rather strong functional constraints, were observed for the three sites of the catalytic triad: 0. 054, 0. 051 and 0. 050 for Cys68, His107, Asp122, respectively, in the vertebrate dataset. These three sites were all conserved in the entire set of NAT coding sequences considered, thereby confirming the importance of these three residues in the activation of the active site cysteine residue.
Interestingly, domain I was found to be more constrained than the other domains of the protein, consistent with the lower variability displayed by this region (mean Shannon entropy (H) = 0. 70) as compared to domain II (H = 0. 84) and domain III (H = 0. 93). Conversely, higher ω values were observed for the interdomain, 17residue insert and C-terminus regions of the protein. In particular, the C-terminal tail was shown to evolve under more relaxed selective constraints in the four datasets examined and many sites pinpointed to be under adaptive evolution were located in this region. This is consistent with the association of this region with different acetyl-CoA binding properties and substrate specificities.
Collection of vertebrate NAT sequences from genomic databases All available vertebrate genomes in various stages of draft sequencing, assembly, and annotation were searched for the presence of putative NAT genes by TBLA-STN searches in the National Center for Biotechnology Information (http://www. ncbi. nlm. nih. gov) and Ensembl (http://www. ensembl. org) databases, using the protein sequence of human NAT1 as the query and an E-value cut-off of 10 -5. Once we identified the first set of vertebrate sequences using the human probe, subsequent searches were conducted by using the sequences of representative species within a given taxonomic class or order to identify additional sequences in the same group of taxa. More specific searches were also performed using the NAT protein-coding sequence identified in one organism to look for additional NAT paralogs in its genome.
We discarded sequences with more than 200 missing nucleotides and only sequences with an identified ORF were kept in the final data set. Sequences from the Syrian hamster (Mesocricetus auratus) and the Chinese hamster (Cricetulus griseus) were obtained from GenBank (accession numbers: U05271 Mesocricetus auratus Nat1, L24912 Mesocricetus auratus Nat2, NM_001244413 Cricetulus griseus Nat2). Fine-scale synteny data around the NAT genes locus were retrieved from Genomicus v63. 01  [59]  and the UCSC genome browser  [60]. To name the NAT sequences identified in vertebrate species, we followed the official gene nomenclature for non-human NAT genes in eukaryotes and prokaryotes (http://mbg. duth. gr/non-humannatnomenclature/): names were given according to the descending identity between each reported NAT sequence and human NAT1.
Amplification, cloning and sequencing of NATs in primates Based on available DNAs, we selected a representative number of primate specimens belonging to 13 distinct genera, including 3 hominoids, 8 Old World monkeys and 2 New World monkeys, for which the NAT homologous sequences were determined: Pan paniscus (bonobo), Hylobates lar (white-handed gibbon), Nomascus gabriellae (yellow-cheeked gibbon, Macaca sylvanus (barbary macaque), Mandrillus sphinx (mandrill), Cercopithecus diana (diana monkey), Chlorocebus tantalus (tantalus monkey), Erythrocebus patas (patas monkey), Allenopithecus nigroviridis (allen's swamp monkey), Colobus guereza (guereza), Presbytis cristatus (silveredleaf monkey), Pithecia pithecia (white-faced saki), and Cebus apella (tufted capuchin). We designed amplification primers based on conserved sequences in the multiple alignments of database sequences retrieved from the genomes of Homo sapiens, Pan troglodytes, Gorilla gorilla, Pongo pygmaeus, Macaca mulatta, and Papio hamadryas.
Protein and codon sequence alignments After discarding a few very divergent sequences that aligned poorly (Anolis carolinensis NAT3, Oryzias latipes NAT3, Fugu rubripes NAT1, Tetraodon nigroviridis NAT1), the remaining set of NAT ORF sequences were translated using the ExPASy Translate tool (http://web. expasy. org/translate/) and the amino acid sequences were aligned using ClustalW  [61]  and Multalin  [62]. Both programs yielded similar results. The amino acid alignments were then converted to the corresponding alignments of codons using the PAL2NAL program  [63]. These nucleotide alignments were used as inputs for the construction of phylogenetic trees, the analysis of gene conversion and tests of positive selection.
Phylogenetic analysis Phylogenetic trees were built from two nucleotide alignments: the first one included the 77 vertebrate NAT sequences retrieved from genomic databases (referred to as the 'vertebrate dataset') and the second one was composed of 58 primate sequences, including both the 17 database sequences and the 41 newly generated ones (referred to as the 'primate dataset'). Phylogenetic relationships of NAT sequences were reconstructed using the maximum likelihood method implemented in PhyML v2. 4. 4  [64] , after determining the optimal model of sequence substitution with Modeltest 3. 04  [65]. For both datasets, a general time reversible model with a proportion of invariant sites and gamma distributed among-site rate variation (GTR + I + G) was used, as selected by the Akaike information criterion.
Detection of gene conversion Possible recombination and gene conversion events were detected using GeneConv version 1. 18  [66, 67]. The program tests for gene conversion by finding identical fragments between pairs of sequences in a nucleotide alignment. Global and pairwise P-values are calculated in order to assess the statistical significance of the observed fragment lengths. Global P-values are more conservative because they are based on the comparison of each fragment with all possible fragments for the entire alignment. GeneConv was run using a group structure defining a distinct group for each set of NAT coding sequences within a species so as to look only for withinspecies gene conversion events and adjust Bonferroni corrections All the default settings were used except for the mismatch penalty (set to 2) and the number of random permutations to compute global and pairwise P-values (set to 1,000,000).
Only gene conversion events with global permutation P-values ≤ 0. 01 were retained. In order to avoid confounding effects due to strong selection or recent mutation, GeneConv was also run using only silent sites. Intragenic recombination events were also detected using five other methods implemented in the RDP3 v. 3. 44 software package  [68] : two of these are phylogenetic methods, which infer recombination when different parts of the genome result in discordant topologies (RDP  [69]  and Bootscan  [70] ), while the other three are nucleotide substitution methods, which examine the sequences either for a significant clustering of substitutions or for a fit to an expected statistical distribution: MaxChi  [71] , Chimaera  [72] , and SiScan  [73]. Common settings for all methods were to consider sequences as linear, to require phylogenetic evidence, to polish breakpoints and to check alignment consistency.
Maximum likelihood tests of positive selection The ratio of nonsynonymous (amino acid altering) to synonymous (silent) substitution rates (dN/dS or ω) was used to evaluate the selective pressures acting on the NAT coding region, expecting dN/dS = 1 for neutrality, dN/dS < 1 for purifying selection, and dN/dS >1 for positive selection. The analysis was performed using the codeml tool of the PAML package version 4. 2b  [26, 74] , which allows pairs of nested models with and without positive selection to be tested in a likelihood ratio framework to determine if adaptive evolution has occurred. We used two pairs of site-evolution models to test whether some sites (codons) are under positive selection: (1) the M1a (Nearly Neutral) and M2a (Positive Selection) models and (2) the M7 (beta) and M8 models (beta & ω).
The innominate artery was formalin fixed, paraffin embedded, cut into serial 4-m sections, and stained with U. S. trichrome (UST; ref. 35). Luminal, lesion (neointima), and medial areas of atherosclerotic vessels were quantified by light microscopy with computerized planimetry (MCID Basic 7. 0 image analysis software; Imaging Research, London, ON, Canada) and, where appropriate, expressed as a percentage of the total area within the internal elastic laminae (IEL). The section with the maximum cross-sectional narrowing was chosen to represent each artery when calculating the mean area in a group. Empty spaces within the lesion (areas of extracellular lipid or cholesterol crystal accumulation; ref. 36)  were determined, and fibrous caps were counted  (37). Serial sections at the point of maximal cross-sectional narrowing were stained with picrosirius red (PSR) to show collagen content; quantification of picrosirius red staining was achieved using color deconvolution (Photoshop CS3 Extended, Adobe Systems, San Francisco, CA, USA) and expressed as a fraction of the total neointimal area, as described previously  (38).
Controls included a nonspecific IgG raised in the same species as the primary antibody. Immunoreactivity for ␣-SMA was quantified using color deconvolution (as described for collagen) and expressed as a fraction of the total neointimal area, as described previously  (38). Mac-2 and CD3 staining were quantified by manually counting nuclei in areas of immunoreactivity and expressing the data as number of cells per square millimeter of lesion. All image analysis was performed blind to genotype.
After washing, flow cytometric analyses were performed using BD FACS Calibur or LSRFortessa Cell Analyzer (BD Biosciences). Data analysis was performed using FlowJo 8. 2 software (TreeStar, Ashland, OR, USA). Fluorescent flow-check fluorospheres (Beckman and Coulter, High Wycombe, UK) were added to blood, BM, and peritoneal lavage samples prior to analysis in order to calculate the absolute number of cells. Antibodies used were as follows. Anti-CD11b-peridinin chlorophyll protein cyanin 5. 5 (PerCPCy5. 5) or -fluorescein isothiocyanate (FITC) (clone M1/70), anti-F4/80-PerCPCy5. 5 (clone BM8), anti-CD45-pacific blue (clone 30-F11), and anti-Ly6G-phycoerythrin-cyanine 7 (PECy7)  or -FITC (clone 1A8) were all from Biolegend (San Diego, CA, USA). Anti-CD115 phycoerythrin (PE; clone AFS98), anti-CD3-allophycocyanin (APC) or -PE (clone 17A2), and anti-CD45-PerCPCy5.
Acknowledgments The authors thank  Karine Devilliers , HerveBoutal,  Marie-Claire Nevers ,  Maud Goislard  (CEA) and  Marie-Francoise Drumare (INRA)  for expert technical assistance,  Farida Nato  ( Institut Pasteur ) for her gift of the F1 mAb, and  Mino Rajerison  ( Institut Pasteur, Madagascar ) for providing F1dipsticks.
Colony Formation Assay Cells growing in log phase were seeded at a density of 1000-2000 cells/well into 60 mm dishes in complete medium. After allowing the cells to adhere for 24 h, medium was replaced with complete medium containing ACCA at the indicated concentrations. Cells were allowed to grow for 3-4 weeks, with a medium change containing ACCA in complete medium, performed every fourth or fifth day. Colonies were then fixed and stained with Methylene blue-50% ethanol and counted. Individual assays were performed in triplicate with a total of three plates per data point.
However, and consistent with the data obtained above, on treatment of cells for 48 h. with increasing concentrations of ACA (25, 50, 100, 150 and 200 mM), tumor cell growth was strongly inhibited in a dose-dependent manner (Fig. 2B ). This suppressive effect of ACCA on tumor cells was most remarkable at the concentration of 200 mM, 150 mM and 50 mM for MCF-7 (54. 47%; P,0. 005), T47D (77. 47%; P,0. 005), and MDA-231 (64. 67%; P,0. 005) cells, respectively. In contrast, treatment of HBL100 immortal breast cell line, with increasing concentrations of ACCA (25, 50, 100, 150 and 200 mM), do not affect the growth of MDA-231, T47D and MCF-7 cells (Fig. 2B ). In total, these results suggest that ACCA acting through MCT1 selectively inhibits the growth of breast cancer cells in vitro. We next questioned whether the growth-inhibitory effect of breast cancer cell proliferation persists on removal of ACCA.
MDA-231 cells were treated with medium alone or containing 50 mM of ACCA for 48 h. , and proliferation was assessed by MTT assay (d0). Our data show that, consistent with our previous results, ACCA significantly inhibited tumor cell proliferation (d0, OD control, 0. 235; OD ACCA, 0. 084, 62. 38% inhibition, P,0. 001). Plating medium was removed and replaced with growth medium containing 10% of FBS, and cell growth was monitored for a further 4 days. Our data show that the suppressive effects of ACCA at day 4 (d4) were similar to that observed at d0, (d0, OD control, 1. 775; OD ACCA, 0. 580, 67. 33% inhibition, P,0. 001). These results clearly demonstrate that the inhibitory effect of ACCA on cell proliferation persists, even on removal of the drug.
As shown in Fig. 5 , ACCA decreased Bcl-2 protein level (1. 8-, 2. 4-, and 2. 6-fold, respectively) whereas Bax protein expression were elevated (3. 4-, 2. 4, and 3. 2-fold, respectively) in MCF-7, T47D and MDA-231 breast cancer cells, respectively. In contrast, the levels of the various pro-and antiapoptotic proteins did not change significantly in HBL-100 cells following treatment with. ACCA (data not shown). These studies suggest that ACCA increases the ratio of of pro-to antiapoptotic proteins, thereby tipping the balance of cancer cells from survival to programmed cell death.
A shown in Fig. 6B , the average tumor volume in mice treated with ACCA was significantly lower compared to vehicle-treated control mice. At 32 days after tumor cell injection, MDA-231 cells gave rise to small tumors as compared to vehicle-treated control mice (195612 mm 3 vs 1262 mm 3 ) indicating that ACCA reduced tumour growth by ,94%. Although, treatment of mice receiving MDA-231 cells with ACCA at 8 days resulted in a less dramatic decrease in tumor growth, the difference was still evident (Fig. 6B ). Of note, animals remained well throughout the entire experiment and no weight loss was observed upon treatment, suggesting that ACCA was well tolerated at this dose regimen (data not shown).
The capacity of ACCA to efficiently inhibit the growth of wild-type and mutant p53 breast carcinoma cells, supports the possibility that ACCA may prove efficacious in the treatment of human breast and other cancers. Additionally, it is important to note that the cell viability and proliferation in immortal normal human breast epithelial cells were not significantly altered by ACCA at concentrations ranging from 25 to 200 uM, which significantly induced cell death in breast carcinoma cells. Furthermore, our results showed that ACCA significantly inhibited both migration, invasion and in vivo tumor growth of MDA-231 cells. These data together with previous studies demonstrating similar anti-proliferative effects of a-cyano-4-hydroxycinnamate through inhibition of lactate trans-porters in other cellular contexts  [13] [14] [15] , suggest that ACCA may possess potential therapeutic activity against breast cancer cells, without significantly affecting viability of normal cells.
Although previous reports have shown induction of apoptosis in several cancer cell lines  [13] [14] [15] [16] , our data are the first to highlight mechanisms by which ACCA induces apoptosis in breast cancer cells. Our investigation also revealed that although Bax expression is regulated positively by wild-type p53  [31] , ACCA is able to induce Bax irrespective of p53 status in breast cancer cells, suggesting that alternative pathways can be involved in Bax up-regulation after treatment with ACCA. Our results add to the growing evidence that has been accumulated over the past few years supporting the existence of p53-independent cell death induced by chemotherapeutic drugs  [32] [33] [34] [35]. At present, it unknown whether ACCA directly or indirectly induces Bax expression in breast cancer cells.
Previous investigators have performed such quantitative assessment of histopathological injury  [37]. 2. 12. Statistical Analysis. The results were expressed as means ± S. E. All data were done with the Statistical Package for Social Sciences (SPSS 17. 0 for windows). The results were analyzed using one way analysis of variance (ANOVA) followed by Duncan's test for comparison between different treatment groups. Statistical significance was set at  ≤ 0. 05.
Acknowledgments The authors thank the  Academy of Scientific Research & Technology (ASRT, Egypt)  and  IMRCP UMR 5623 ,  Faculty of Pharmacy, University Paul Sabatier (France) , for supporting this study within the  IMHOTEP program  (Project no. 109/EGY/FR 8-02 ). Also, the authors are grateful to  Professor Dr. Adel Mohamed Bakeer Kholoussy ,  Professor of Pathology, Faculty of Veterinary Medicine, Cairo University , for reading the histopathological sections.
For each phase diagram, different weight ratios of oil and mixed surfactant/ cosurfactant were investigated: 10:0, 9:1, 8:2, 7:3, 6:4, 5:5, 4:6, 3:7, 2:8, 1:9, and 0:10 (w/w). Slow titration with the aqueous phase was performed for each weight ratio of oil and mixed surfactant/cosurfactant, and visual observation was carried out for a clear transparent W/O microemulsion. Once stable transparent formulations were obtained, simple tests of nondispersibility in water and conductance tests were conducted to confirm that they were W/O microemulsions. In addition, the W/O microemulsions were observed under polarizing light and found to be nonbirefringent, as expected from their isotropic nature. The physical state of the microemulsion was marked using the ProSim Ternary Diagram computer program (ProSim, Labège, France), on a pseudo-three-component phase diagram, with one axis representing the aqueous phase, another representing oil, and the third axis representing the mixture of surfactant and cosurfactant at fixed weight ratios.
Statistical analysis Each dilution of emulsion was tested in six wells, and each experiment was realized in triplicate. The mean values for each concentration were analyzed by a one-way analysis of variance (ANOVA), followed by Dunnett's test. 30, 31 The level of significance was fixed at 0. 05. Statistical analysis was performed using SigmaStat 2. 0 software (Systat Software Inc, San Jose, CA, USA).
The ability of the program Macromodel (from Schrödinger Inc. ) to predict the nonpolar interactions between host 4 and benzene derivatives was finally evaluated by means of docking simulations. Measured binding free energies were plotted as a function of the simulated inclusion enthalpies (Figure  6 ), defined as the enthalpy differences between the inclusion compounds and the free species. A significant linear correlation was obtained, confirming that the space filling of host cavity plays a dominant role within such series of complexes, and that the applied force field was reasonable. All guests fitted completely into the cavity of host 4 without steric hindrance. The slope of the experimental Gibbs free energy versus the simulated interaction enthalpy was still inferior to unity (0. 69), which indicated that the host-guest interactions were partially compensated by reduced solvation and loss of freedom of the host-guest system.
Acknowledgements We acknowledge support from  UCEIV , which participates in the  Institut de Recherche en ENvironnement Industriel (IRENI)  financed by the  Communauté Urbaine de Dunkerque , the  Région Nord Pas-de-Calais , the  Ministère de l'Enseignement Supérieur et de la Recherche , the  CNRS  and  European Regional Development Fund (ERDF).
ACKNOWLEDGEMENTS IAEA: For us the fellowship ( ZAI/07011 ) awarded under the  Technical Cooperation  Project  ZAI/5/016  between the  Democratic Republic of the Congo (DRC)  and the  International Atomic Energy Agency (IAEA)  through the  CGEA/CREN-K. General Atomic Energy Commission. Regional Center of Nuclear Studies of Kinshasa  ( GAEC / RCNS-K ). For we have given permission to leave the country (DRC) to do the some months internship in France. Research Institute for Development (RID)  / Mont-.
Potentiodynamic polarization Polarization measurements were carried out in a conventional three-electrode electrolytic cell. Saturated calomel electrode (SCE) and platinum electrode were used as reference and auxiliary electrodes respectively. The working electrode is in the form of a disc from carbon steel of the surface 0. 32 cm 2. These electrodes are connected to Voltalab PGZ 100 piloted by ordinate associated to "Volta Master 4" software. The scan rate was 1 mV/s started from an initial potential of -700 to -100 mV/SCE. Before recording each curve, a stabilization time of 30 min was allowed, which was proved to be satisfactory to attain a stable value for E cor. All experiments were repeated three times at temperature desired ±1°C. Corrosion current densities were obtained from the polarization curves by linear extrapolation of the Tafel curves.
Methods and Findings: A systematic search of the literature was conducted to identify all studies published between 1960 and February 2013, in which patients were enrolled and treated with DP. Principal investigators were approached and invited to share individual patient data with the WorldWide Antimalarial Resistance Network (WWARN). Data were pooled using a standardised methodology. Univariable and multivariable risk factors for parasite recrudescence were identified using a Cox's regression model with shared frailty across the study sites. Twenty-four published and two unpublished studies (n = 7,072 patients) were included in the analysis. After correcting for reinfection by parasite genotyping, Kaplan-Meier survival estimates were 97. 7% (95% CI 97. 3%-98. 1%) at day 42 and 97. 2% (95% CI 96.