dataesr/scientific-paragraphs-categorization huggingface dataset:
Cell Immunostaining and Flow Cytometry After treatment, cells were harvested using trypsin/EDTA before immunostaining. Cells were labeled using anti-pan HLA class I (anti-HLA-A, -B and -C; clone W6/32), anti-pan HLA class II (anti-HLA-DR, -DP, -DQ; clone L243), anti-MICA (clone AMO1; BamOmab, Tubingen, Germany), anti-HLA-E-APC (clone 3D12; Miltenyi Biotec, Paris, France) mouse IgG as primary antibodies and anti-mouse IgG + IgM (H + L)-FITC (Jackson Immunoresearch Laboratories, West Grove, PA, USA) as secondary antibodies. An isotype-matched IgG was used as the negative control. Fluorescence was measured by flow cytometry on 10,000 cells/sample using a BD FACSCanto TM II flow cytometer (Becton Dickinson, Satn Jose, CA, USA). Acquired data were analyzed with FlowJo software (Tree Star, Inc. , Ashland, OR, USA) and depicted in histograms plotting geometric mean of fluorescence intensity (GFI) on a four-decade logarithmic scale (x-axis) versus cell number (y-axis).
Data and Statistical Analysis Data are represented as means ± SD for replicates experiments. Statistical analyses were performed with Graphpad Prism ® Software (Graphpad Software, San Diego, CA, USA) by a Student's t-test, a parametric or Kruskal Wallis non-parametric analysis of variance test as appropriate. A p value < 0. 05 (*) was considered statistically significant. (**) indicates p < 0. 01 and (***) p < 0. 005.
Molecular Docking Protein preparation: The 3D structure of histone acetyltransferase p300 in complex with a co-crystallized inhibitor was downloaded from Protein Data Bank (PDB entry 5KJ2, rcsb. org) [22]. All implicit hydrogens (of the protein and the ligand) were added using GOLD 5. 6. 3 (CCDC, Cambridge, UK)  [23]. Water molecules that take part in at least three hydrogen bonds with the protein and/or the ligand, at least 2 of which are with the protein, were kept. Other water molecules were deleted. Hydrogen bonds were analyzed using LigandScout 4. 4 (Inteligand, Vienna, Austria)  [24]. Protein treatment was done with GOLD 5. 6. 3. The protein structure (in the absence of the ligand) was saved as a mol2 file. Ligand preparation: Three types of ligands for redocking were prepared as follows: • Co-crystallized extracted ligand from the complex structure with no changes in conformation and configuration: the ligand (after hydrogens were added) was saved as a mol2 file without the presence of the protein and any further modification.
• Energy-minimized reconstructed ligand: the structure of the ligand was rebuilt with ChemDraw Professional 16. 0  [25] , its 3D structure was then visualized from the SMILES code in the ligand-based view of LigandScout 4. 4, after which the energy minimization step was carried out as described above. The fully processed ligand structure was output and saved as a mol2 file. All test-set molecules (guttiferone J (1), xanthochymol (2), guttiferone F (3) and garcinol (4)) were downloaded from SciFinder in the sd file format. Their 2D structures were then converted into 3D, and energy minimization was carried out with LigandScout 4. 4 as previously described. All output ligands were saved separately as a mol2 file. Molecular docking: A rigid molecular docking procedure was carried out using GOLD 5.
Phytochemical Analysis Natural PPAPs are usually isolated from Garcinia (Clusiaceae) and Hypericum (Hypericaceae) genera  [26]. For the present study, PPAPs exhibiting a similar molecular scaffold to guttiferone J (1) were required. According to the exhaustive database created by Grossman and colleagues  [27] , on the 774 known naturally occurring PPAPs, 75 of them share the selected skeleton (Figure  1 ). Among them, garcinol (  4 ) is commercially available and was thus selected for biological assays. Moreover, to date, about 75% of PPAPs exhibiting the selected scaffold were mainly biosynthesized by Garcinia species. The genus Garcinia includes about 400 trees and shrubs growing in tropical and equatorial areas including Malaysia  [28, 29]. Therefore, the phytochemistry of various organs (i.
Cells were harvested for RNA isolation and qRT-PCR for the following transcripts: HLA-E, MICA, STAT1, SOCS1, CIITA pIV, CREB/CBP, p300 and HDAC3. Results shown are means of triplicate experiments, data were normalized using a housekeeping gene (RPLPO) and expressed as 2 -∆∆Ct using cells treated with diluent alone (DMSO, 1/1000) as reference. (B) Dose-dependent effect of PPAPs 2-4, HAT (A485, C646) and HDAC3 (SAHA) inhibitors on cell viability. ECs were cultured for 18 h with compounds (1-20 µM) or diluent in the presence of IFNγ (100 U/mL) for 18 h. Cell viability was assessed in triplicates using MTT staining and expressed as relative percentages calculated using diluent as reference (100%), * p < 0. 05, ** p < 0. 01 versus diluent. To explore further the properties of PPAPs, the following experiments were conducted using purified xanthochymol (2) and guttiferone F (3) in comparison to garcinol (4) and to two synthetic inhibitors of HAT CBP/p300 (C646 and A485) and HDAC (SAHA).
In contrast, IFNγ had no effect on MICB, GATA2 and SHP2 and decreased significantly the NKG2D ligands MICA, ULBP2 and ULBP3. In these conditions, PPAPs 2-4 significantly inhibited the regulatory effect of IFNγ by decreasing the transcript level for HLA class I, HLA-E, β2M and tapasin. TAP1 was selectively inhibited by 4 while 2-3 had no effect. PPAPs 2-4 further decreased MICA and MICB levels but had an opposite effect on ULBP2 and no effect on ULBP3. A significant inhibitory effect of STAT1 and SOCS1, which reflects a loss of IFNγ signal, was observed for 2 and 3 confirming the data obtained with the mixture GX before their purification. Next, target genes of STAT1, coding for factors involved in the transcriptional activation of MHC and HLA-E, CIITA, GATA2 and IRF9 were quantified. CIITA and GATA2 mRNA levels were dramatically reduced in the presence of PPAPs 2-4 consistent with an upstream dysfunction of IFNγ signaling.
IRF9 was lightly increased by 2 but reduced by 4, A485 and SAHA. No effect was found for Src homology region 2 (SH2)-containing protein tyrosine phosphatase 2 (SHP2) a protein tyrosine phosphatase involved in signal transduction by regulating several canonical pathways (MAPK and PI3K). Using Western blot experiments, we observed that PPAPs 2-4 reduce significantly the phosphorylation of STAT1 (Y701) and also reduce the acetylation of CBP (K1535)/p300 (K1499) suggesting hypoacetylation of CBP/p300 (Figure  4B, C ). Inhibition of HAT acetylation was also achieved using A485. Together these data may suggest that, similar to garcinol (4), PPAPs 1-3 display an inhibition of HAT activities that subsequently impairs IFNγ signaling and transcriptional activity required for MHC regulation. Finally, we used flow cytometry to assess whether inhibition of HAT in EC cultures may affect MHC protein expression.
Molecular Docking in the Binding Site of Histone Acetyltransferase p300 Biologically evaluated PPAPs 1-4 were also successively docked in the selective catalytic site of the histone acetyltransferase p300 previously described for A-485  [37]. Previously, the best redocked poses into the binding site (heavy atoms only) of the original structure and its energy-minimized conformation, as well as the rebuilt structure (created according to our protocol described in Materials and Methods) of the native ligand (HET code: 6TF, namely A-485) obtained with GOLD 5. 6. 3 deviated 0. 45 Å, 0. 73 Å and 1. 43 Å from the true crystal pose deposited in the Protein Data Bank, respectively, denoted that the docking procedure managed to correctly pose the ligand and could be employed for further investigation (Figure  S7 ).
As this amino acid is located at the center of the binding site, its strong interaction with the ligands could help to stabilize the protein-ligand complexes and is expected to impair the biological activity of the protein in a similar manner to that observed with the native ligand A-485. 5. 6. 3 deviated 0. 45 Å, 0. 73 Å and 1. 43 Å from the true crystal pose deposited in the Protein Data Bank, respectively, denoted that the docking procedure managed to correctly pose the ligand and could be employed for further investigation (Figure  S7 ). It was observed that the key interactions between the native ligand and several amino acid residues of the binding site were preserved in all redocked poses. Such interactions include a hydrogen bond between the A-485 ligand's methyl-urea moiety and the backbone carbonyl of Gln1455, as well as another hydrogen bond that involves the carbonyl group substituted at the C-4′ position on the oxazolidinedione structure of A-485 and the hydroxyl group of Ser1400.
V. minor, also known as the lesser periwinkle, is an evergreen herbaceous shrub belonging to the Apocynaceae family. In addition to the above mentioned pharmaceutically important MIA vincamine, this plant produces more than 50 indole alkaloids, including t less known interesting molecules such as lochnericine, akuammicine, vincadifformine, minovincinine  [7, 8]  and picrinine  [9]. To date, publicly available V. minor genomic resources are restricted to the 454 based-(V_minor_v1) and illumina based transcriptomes (V_minor_v3) from the PhytoMetaSyn database (https://bioinformatics. tugraz. at/phytometasyn/). Furthermore, knowledge on MIA biosynthetic pathways in V. minor is lacking and mainly results from homology-based identification of genes with its relative C. roseus sharing high genetic and metabolic similarities  [8, 10].
In our continuing effort to elucidate and engineer MIA biosynthesis, we present here a comprehensive consensus transcriptome resource for V. minor comprising eight different tissue types and experimental conditions. This new V. minor reference transcriptome displays improved BUSCO scores to the previously available 454 based (V_minor_v1) and illumina based assemblies (V_minor_v3) from the PhytoMetaSyn database (https://bioinformatics. tugraz. at/phytometasyn/). This improved resource was thus used to rationally predict candidate genes for all known steps of the V. minor MIA pathway as illustrated by the functional validation of a tabersonine/vincadifformine 16-O-methyltransferase (16OMT) isoform. Finally, a gene co-expression analysis was performed to predict new MIA biosynthetic candidate genes to deepen our knowledge on the MIA metabolism in V.
RNA-Sequencing RNA was extracted from two biological replicates for AR, YL, OL and YS, using TRIzol (Thermo Fisher Scientific, Illkirch-Graffenstaden, France) as previously described  [3]. The RNA was subsequently used to prepare libraries with the Illumina TruSeq RNA Library Prep Kit v2. Libraries were sequenced in paired-end mode (2 × 100 bp) by Eurofins Genomics (Les Ulis, France) using the Illumina HiSeq2500 technology. Sequencing data was deposited as project accession number PRJEB40906. EdgeR R package was used to process RNA-seq data. Replicate homogeneity was checked using a multidimensional scaling analysis (Figure  S1 ). It clearly indicated that biological replicates were consistent. The biological coefficient of variation was 0. 4 (common dispersion = 0. 168531), in the range of typical variation for human data.
Generation of a Consensus Transcriptome Reads were trimmed using FastP  [69]  version 0. 20 with default settings and used to de novo assemble individual transcriptomes using Bridger V2014-12-01  [70]. In order to construct a high quality reference transcriptome suitable for gene expression analysis and biosynthesis gene discovery, all assemblies were merged. Sequencing reads were pseudo aligned onto the merged assembly and counted with Salmon v0. 14. 1  [71]  with bias correction (-biasCorrect) in the variational Bayesian optimized (-vbo) mode to establish the abundance estimates as transcripts per million (TPM). In our compaction procedure we discarded weakly expressed transcripts (>TPM5) and applied a clustering algorithm MMSeqs2  [72]  using the following parameters: -mode 1, -c 0. 8. Assemblies were evaluated with BUSCO  [73]  version 3.
0. 2 compared to existing V. minor transcriptomes (Vinca_minor_v1 and Vinca_minor_v3) obtained from the Phytometasyn database (https://bioinformatics. tugraz. at/phytometasyn/). Annotation of the consensus transcriptome was achieved with the Trinotate pipeline  [74]  version 3. 0. 1 that uses Blastx and BlastP of TransDecoder  [75]  V5. 3. 0 predicted ORFs against Uniprot, and hmmscan  [76]  against the PFAM database (https://pfam. xfam. org/).
Functional Analysis of the Consensus Transcriptome Differentially expressed genes (DEGs) were identified by fitting a generalized linear model to each transcript using the edgeR R package  [77]. Genes were considered to be differentially expressed if the p-value of the exact test on the binomial fit was below 0. 01. In this analysis, genes with a log fold change above 2 were considered to be upregulated and genes with a log fold change below -2 were considered to be downregulated. Overlap in DEG sets between different comparisons were visualized by the UpSetR  [78]  package v1. 4. 0. Uniprot description keywords were retrieved from the transcriptome annotation and used in the enrichment tests by comparing effectives to a hypergeometric distribution with the phyper function in R  [79]. MIA gene conservation was determined through blastn  [80]  analysis of the V.
UPLC-MS Analyses UPLC-MS analyses were performed according to  [65]. The selected ion monitoring mode was used to collect data for the following compounds: tabersonine (m/z 337), retention time (RT) = 12. 36 min (RT) = 11. 3 min; 16-hydroxytabersonine (m/z 353), RT = 7. 9 min; 16-methoxytabersonine (m/z 367), RT = 12. 04 min; 19-hydroxytabersonine (m/z 353), RT = 6. 8 min; vincadifformine (m/z 339), RT = 11. 16 min; lochnericine (m/z 353), RT = 11. 2 min; 16-methoxylochnericine (m/z 383), RT = 11. 4; Ervinceine (m/z 369) and RT = 11. 5 min in Figure  7. Ervinceine was quantified as vincadifformine equivalent.
Co-Expression Network For each possible gene pair in the consensus transcriptome, the pairwise Pearson's correlation coefficient was calculated and subsequently ranked to determine the highest reciprocal ranks (HRR) as described previously  [86, 87]. The best co-expressed genes were captured for each known MIA pathway gene if their HRR < 100. The resulting associations were visualized using the igraph package in R  [88]  and communities of the closest co-expressed genes were determined using a fast greedy algorithm  [89].
Next, we aimed at constructing a high-quality reference transcriptome suitable for gene expression analysis and discovery of MIA biosynthetic genes. We thus combined the individual transcriptomes resulting in a merged resource of 896,265 transcripts and a complete BUSCO score of 99. 2% (all non compacted, Figure  2 ). However, this large number of transcripts is far from the closely related species C. roseus whose genome was predicted to contain 34,363 genes  [93]. Therefore, we aimed to decrease the final merged assembly size by removing redundant sequences without losing biologically relevant transcripts. In our downstream compaction procedures, we discarded weakly expressed genes (TPM<5) and applied a sequence clustering algorithm (MMSeqs2)  [72]. Finally, this procedure allowed a reduction of the merged transcriptome (896,265 transcripts) to a final consensus transcriptome of 41,823 transcripts (all compacted) and an acceptable complete BUSCO score of 90.
All candidates display nucleotide sequence similarity (> 70%) to the C. roseus 16OMT. The deduced protein sequences of all four candidates maintained Omethyltransferase domains (PF00891. 15, Figure  S12 ), as determined by hidden-Markov comparison against the PFAM database. Interestingly, the relative expression levels (as transcripts per million; TPM) of each candidate, obtained by mapping the reads from each tissue type to the consensus transcriptomes (Figure  6A ), revealed that YS2_comp61_seq0 display the highest expression in all the organs tested with a maximum in the young aerial parts of the plant (YL and YS, > 1300 TPM). This high level of expression was confirmed by quantitative RT-PCR providing additional evidence of high expression in YL (Figure  6B ). By contrast, the three remaining candidates were barely detectable in old leaves and roots.
Such formation was observed for the Vm16OMT-YFP N /Vm16OMT-YFP C combination while no BiFC complexes were reformed with the Vm16OMT-YFP N /YFP C -Vm16OMT combination. These results thus suggest that Vm16OMT is engaged in homodimer or homomultimer formation through a head-to-head association. This was in agreement with the presence of a dimerization domain (PF08100. 8, Figure  S12 ) located at the N-terminal end of plant Omethyltransferases  [99]. Interestingly, we also noticed that Vm16OMT-BiFC complexes were localized exclusively in the cytosol and not in the nucleus as revealed by the lack of superimposition of the BiFC YFP signal with that of the nucleus CFP marker (Figure  9M-P ). This nuclear exclusion probably results from the increase of Vm16OMT size due to homodimer formation (around 80 kDa) that may exceed nuclear pore exclusion size that is around 60 kDa  [100].
Gene Co-Expression Analysis Finally, after validating our V. minor consensus transcriptome for homology-based identification of MIA genes, we took advantage of the multiple samples from the dataset to initiate the prediction of candidate genes catalyzing unknown MIA biosynthetic steps. Usually, genes sharing similar functions or involved in similar pathways display similar expression profiles  [64]. Such similarities can be exploited in a "Guilt-By-Association" analysis to guide the identification of additional MIA pathway genes in the dataset  [101]. For example, the downstream tabersonine pathway genes TEX and TAT were identified in C. roseus by searching candidates that were highly co-expressed with T16H  [23, 24]. Additionally, a co-expression analysis was used to short list candidates for subsequent O-methyltransferase activity assays towards identifying the 10-hydroxycamptothecin Omethyltransferase (Ca10OMT) in C.
minor consensus transcriptome against elucidated C. roseus MIA genes, MIA transporters and MIA transcription factors. GenBank accession numbers of C. roseus genes are displayed in brackets. Table  S2 : Transcripts that were co-expressed with MIA genes listed and arranged in communities that were determined by a fast greedy algorithm. Transcripts were annotated using blastx against the UniProt database. Table  S3 : Primers used in this study.
It is therefore judicious to clearly define the system as the trihydrate of atorvastatin calcium, or if other trihydrates are discovered "trihydrate of atorvastatin calcium form I". Nonetheless, so far only one fully solved crystal structure with atorvastatin, a solvate containing ethylene glycol, has been described in the Cambridge Structural Database (IZOQIZ)  (Adhikari et al. , 2016). If a hydrate is heated up, two different melting behaviors can be encountered, either incongruent melting or congruent melting as indicated in Figure  2  for a trihydrate. Congruent melting in the right-hand panel of Figure  2  means that the entire trihydrate forms a liquid with the same composition as that of the trihydrate and such a transition is basically equivalent to fusion of a pure compound. Incongruent melting means that only part of the trihydrate forms a liquid that will be richer in water, therefore creating a "leftover" solid containing more API (active pharmaceutical ingredient) in comparison to the trihydrate.
TEMPERATURE-RESOLVED SECOND HARMONIC GENERATION (TR-SHG) An Insight X3 single laser with automated dispersion compensation (Spectra-Physics, Santa Clara, USA) and a TCS SP8 MP confocal microscope (Leica Microsystems, Wetzlar, Germany) were used to carry out confocal microscopy as well as two-photon microscopy on the samples. The laser delivers over 2. 54 W of average power at 900 nm and is tunable from 680 nm to 1300 nm. The pulse rate is 80 MHz and the temporal width at the output of the cavity is around 120 fs (< 100fs between 850 nm and 1050 nm). The laser is controlled by LASX Leica software. Two Leica hybrid descanned detectors (HyD) were used to record images. Long working distance dry Leica objectives (HC PL Fluotar 5X NA 0. 15, HC PL Fluotar 10X NA 0. 3 or HC PL APO 20X NA 0. 4 CS2) or oil-immersion Leica objective (HC PL APO 40X NA 1.
These regimen modifications may contribute to development of macrolide resistance  [5]  or suboptimal outcomes. There are very limited published data to support management decisions even for the most frequent AEs in MAC-PD treatment, and management of drug toxicity is poorly addressed in current guidelines. To know the opinions of clinicians with expertise treating MAC-PD, we designed a survey on AE management in MAC-PD, using the SurveyMonkey online tool (http:// www. surveymonkey. com). The survey presented common AEs in MAC-PD treatment (hepatotoxicity, ocular toxicity, gastrointestinal upset, tinnitus, and ototoxicity) and a choice of management strategies; the AEs were selected by a during a face-to-face meeting at the European Respiratory Society conference (Paris, September 2018). Experts were selected from the Nontuberculous Mycobacteria Network European Trials Group (NTM-NET) (www.
Another great concern is that the transfer of valuable knowledge from generation to generation mainly practiced by word of mouth, might be lost forever due to the lack of a proper written system. Indeed, while some medicinal plants were frequently cited, a lot of herbal species were used for medicinal purposes by only a quite limited number of people within the communities, such as Calamus palustris (4 reported), Engelhardtia serrata (5 reported). Although local people are aware of this vertical transmission, it is still declining quickly and they have no idea about how to resolve this inextricable problem. Therefore, it is crucial that the results of this project will provide them with feedback in various ways (documents and talks for example), as a thankful return for their participation in the study, and to allow the spread and sustainability of those skills and traditions in the community as well as in the whole population and eventually at an international level (publications, databases).
Conservation of medicinal plants: an important issue All medicinal plants are checked for their presence in the Vietnam Red Data Book (Plant section) any plants cited in the endangered or vulnerable group does not mean that there is no cause for concern. For example, although not mentioned as an endangered species, Dacrycarpus imbricatus has been decreasing in population according to IUCN and is even listed as a protected species in the Vietnam Red Data Book. Local people confirmed that it was hard to find big trees in the surrounding areas. Additionally, most of the medicinal plants (85. 5% of total) were collected from natural vegetation. However, there is no special protection or care given to these plants. Therefore, medicinal plants might be too damaged to satisfy the demands of local people who do not much pay attention to conservation issues.
Conclusion The present study recorded 34 traditional medicinal plants used by K'Ho-Cil people. These results help to preserve the plant lore and contribute basic data for phytochemical and pharmacological studies on potential medicinal plants as well as provide precious information for the Vietnamese pharmacopeia. For example, the information on the number of medicinal plant species distributed in the families in the present study along with data on phytochemical active compounds obtainined from literature surveys could serve as a signpost for future drug development with the Asteraceae and Fabaceae families containing the most medicinal plants. Among the medicinal plants cited, several species are widely used for the treatment of various health problems by not only local people but also other minorities elsewhere.