dataesr/scientific-paragraphs-categorization huggingface dataset:
rerio), Northern pike (Esox lucius), rainbow trout (Oncorhynchus mykiss), Atlantic cod (Gadus morhua), and medaka (O. latipes). The phylogenetic tree is a representative tree of life. The presence or absence of each alternative exon (B, A, and C) is represented by a green check mark or a Red Cross mark, respectively. For each species, the location of the gene is specified in the table. (E) Relative expressions of the two lamp2a in eight tissues of rainbow trout were determined by RT-qPCR and normalized to 18S levels. Sets of data are expressed as means þ SD (n ¼ 3). Nonparametric statistical test pairwise Wilcoxon signed-rank test was performed (*P < 0. 05, **P < 0. 01, ***P < 0. 001). Chaperone-Mediated Autophagy in the Light of Evolution. doi:10. 1093/molbev/msaa127 MBE LAMP genes originated from the first and second rounds of vertebrate WGDs (VGD1 and VGD2) that occurred at the root of the vertebrate lineage.
2014 ). These observations are further supported by the recently reported analysis of the PhyloFish RNA-seq database-providing gene expression data from 23 different ray-finned fish species  (Pasquier et al. 2016 )-revealing the expression of a single lamp2a transcript in various salmonid species including rainbow trout but also brook trout (Salvelinus fontinalis), brown trout (Salmo trutta), grayling (Thymallus thymallus), European whitefish (Coregonus lavaretus), and American whitefish (Coregonus clupeaformis)  (Lescat et al. 2018). It is therefore tempting to speculate that loss of expression of the second lamp2a transcript has a common origin, probably in the ancestor of salmoniforms shortly after the SaGD. Collectively, these sets of data clearly demonstrate the existence, in a large variety of ray-finned fish species, of an active lamp2a splice form and therefore imply that CMA function might have appeared much earlier during evolution than initially thought.
Establishment of the lamp2a Knockout Medaka Line Using CRISPR/Cas9 Genome Editing Genome editing of the medaka lamp2a splice variant was performed by mean of the CRISPR/Cas9 method according to the protocol described by  Herpin et al. (2019). In details, identification of CRISPR/Cas target sites and design of oligonucleotides were performed by the use of the ZiFiT software (http://zifit. partners. org/ZiFiT/Disclaimer. aspx). For preparation of sgRNAs, the DR274 plasmid (Addgene No. 42250) was first linearized with Bsa1, electrophoresed in a 2% agarose gel, and purified. Pairs of complementary oligonucleotides were annealed (40 mM Tris-HCl [pH 8. 0], 20 mM MgCl 2 , and 50 mM NaCl buffer) by heating at 95 C for 2 min and then cooled down slowly to 25 C within 1 h. The doublestranded oligonucleotides were then ligated into the linearized pDR274 vector.
Different sgRNAs were designed to target several intronic sites spanning the lamp2a exon in order to create deletions specifically removing the whole 2a exon of the lamp2 gene. After linearization with Dra1 and Not1,  Lescat et al. doi:10. 1093/molbev/msaa127 MBE respectively, pDR274 and pCS2-nCas9n plasmids were used for generating either sgRNAs or Cas9 RNAs. The synthesized RNAs were then injected into one cell-staged embryos at the following concentrations: 25 ng/ml for each sgRNAs and 100 ng/ml for the Cas9 mRNA. CRISPR-positive fish were then screened for mutations using PCR primers flanking the site of deletion (supplementary data set S2, Supplementary Material online). Mutant fish used in this study have been outcrossed for at least four generations and always the phenotype observed correlated with the genotype.
Histology and Electron Microscopy For oil-red-O (ORO) and periodic acid Schiff (PAS) staining, liver tissue was frozen in OCT, sectioned, and stained with ORO at 0. 7% or PAS at 0. 5%. Quantification of area occupied by PAS staining and quantification of number and area occupied by lipid droplets (LD) were performed using ImageJ software. For electron microscopy, fresh liver sampled 16 h after the last meal (N ¼ 3/conditions) was fixed with 2. 5% glutaraldehyde in 0. 1 M phosphate buffer (pH 7. 4), and postfixed in 1% osmium tetroxide in phosphate buffer. Conditions of both the treatments of ultrathin sections and observations were already described in  Seiliez et al. (2016).
Medaka Primary Cells Caudal fins from wild-type and Lamp2a-Ko medaka (30 fish per group) were collected, washed in PBS containing 10 mg/ ml gentamycin and 250 lg/ml amphotericin B. Caudal fins were then minced and digested for 30 min at room temperature in ESM4 containing 2 mg/ml of collagenase (Sigma C2674). Then the mix was filtrated and centrifuged at 150 g for 10 min and the pellet was resuspended in ESM4 medium. Cells were cultivated for 48 h at 25 C before experimentation. Construction of the CMA Reporter Plasmid and Anti-lamp2a Morpholino Design The pKFERQ-PA-mCherry1 plasmid was constructed by inserting the oligonucleotide coding for the N-terminal sequence of the bovine ribonuclease A containing the KFERQ-CMA targeting motif (MKETAAAKFERQHMDSSTSAA) (accession number AAB35594) into the NheI (g/ctagc) and BamHI (g/gatcc) sites of the pPA-mCherry1-N1 vector (addgene), as previously reported in  Koga et al.
Acknowledgments We thank all members of the  Club d'Autophagie Bordelais (CAB)  for discussions and helpful suggestions. We also thank  C. Duret ,  A. Patinote ,  S. Gay , and  T. Nguyen  (from  INRA UR1037 LPGP, Rennes ),  M. Cluzeaud  (from  INRA-UPPA UMR1419 NuMeA, Saint P ee sur Nivelle ), and  X. Pinson  and  S. Dutertre  (from  Microscopy Rennes Imaging Center, MRic ) for their technical assistance. This research was supported by the  INRA "Animal Physiology and Livestock Systems" Division , the  French National Research Agency  ( ANR-17-CE20-0033  " Fish-and-Chap "), and the  Doctoral School of Exact Sciences  and their Applications (ED211) of the UPPA. N. R. -H received fellowship from the  MICIUN  ( BES-2015-074654 ).
Correlation of lymphatic marker expression with MB subgroups. We analyzed the basal expression of markers of lymphatic vessel development  21, 22  as a function of MB subgroups-in the most aggressive subgroups (SHH, Group 4, Group 3)-in the Cavalli  23  database of the R2: Genomics Analysis and Visualization Platform (http://r2. amc. nl). Early markers of lymphatic vessel development (i. e. , LYVE1, VEGFR3, PROX1) are also lymphatic endothelial cell (LEC) specification markers and VEGFC, a late marker in the development of lymphatic vessels, is rather a marker of lymphatic network expansion  22. LYVE1, VEGFR3 and PROX1 were significantly more expressed in the SHH group (moderate to intermediate prognosis) than in more aggressive MB subgroups (groups 3 and 4): p < 0. 001. Conversely, the SHH tumors presented a lower expression of VEGFC: p < 0.
Both VEGFR3  46  and PROX1  47, 48  mRNAs are highly expressed in the developing cerebellum (https://www. proteinatlas. org/ENSG00000037280-FLT4/ tissue/cerebellum), respectively in the Purkinje cells and the external granule layer, and both mRNAs regulate the neuronal development at early postnatal stages. The high level of VEGFR3 and PROX1 mRNAs might thus be related not only to lymphatic, but also to neuronal development and interestingly, specific neural progenitors from the CNS have recently been shown to promote growth and metastasis of tumors of different origin  49. VEGFC, a late gene in the lymphangiogenesis process, essential to the lymphatic system expansion  22  , presented lower expression in the patients from SHH subgroup when compared to more aggressive subgroups. Patient database analysis and immunohistochemistry experiments demonstrated that lymphatic gene expression (VEGFC, LYVE1, PDPN) is a function of the metastasis status.
The cells were counted by the ImageJ (NIH) software. RNA isolation for reverse transcription and real-time PCR. Protein extraction and immunoblot assays. Daoy and HD-MB03 cells were cultured to sub-confluence, then lysed on ice with 1X Laemmli buffer (6 mM Tris-Cl pH 6. 8, 2% SDS, 10% glycerol, 5% β-mercaptoethanol). The lysates were sonicated for 30 s and protein amount was determined by the Pierce™ BCA Protein Assay Kit assay (Thermo Fisher). Sample proteins were reduced by heating at 95 °C for 5 min with 5% β-mercaptoethanol. 50 μg of proteins were loaded on 10% polyacrylamide gels and SDS-PAGE was performed using the Mini-Protean® Tetra Vertical Electrophoresis Cell (Biorad, Marnes-la-Coquette, FRANCE). The proteins were transferred onto PVDF membranes in Tris-Glycine buffer (25 mM Tris, 192 mM glycine, pH 8.
The gating strategy of these experiments is described in Supplementary Fig. S7. The data were collected with the BD FACSDiva software application. Percentages of CD146-labeled isolated cells were calculated with FlowJo® software version 18. 0 (Flexera, Hamburg, GERMANY). Immunocytochemistry. Immunofluorescence studies were performed on medulloblastoma cell lines. Cells were plated (10 5 Daoy, 1. 5*10 5 HD-MB03) into 12-well plates with 14 mm coverslips first coat with 0. 1 mg/ml poly-L-Lysine (Sigma-Aldrich). After 24 h the cells were fixed in 4% PFA, 25 min. After two PBS washing, cells were stained 1 h at room temperature with primary antibody (1:100 dilution) in blocking solution (3% BSA, 0. 1% saponine, PBS). After washing, cells were incubated with fluorochrome-conjugated (AlexaFluor 488, and AlexaFluor 594) secondary antibodies for 1 h at room temperature.
Nuclei were visualized with 1 μg/ml DAPI (4, 6-diamidino-2-phenylindole) (Sigma-Aldrich). Images were acquired with a 63× (1. 4 numerical aperture) oil immersion objective and Zeiss LSM 870 confocal laser-scanning microscope. Images were processed using FIJI software. Enzyme-linked immunosorbent assay (ELISA). Cells were seeded in 12-well plates in triplicates (50,000 cells/well for Daoy or ONS-76 cells, 100,000 cells/well for HD-MB03, D458Med or D341Med). The cells were grown for 48 h, at 37 °C, in a humidified atmosphere with 5% CO 2. The supernatants were then recovered, and the cells of each well were counted for data normalization. VEGFC was quantified from supernatants with the R&D Systems Human VEGFC Duoset ELISA kit (Minneapolis, MN, USA), according to the manufacturer's recommendations.
Data are presented as secreted VEGFC in pg/ml/10 6 cells/48 h. Pseudo-vessel formation assay. Cells were deprived of serum overnight. They were then seeded (75,000 Daoy cells, 200,000 ONS-76 cells or 300,000 HD-MB03, D458Med or D341Med cells per well) in 24-well plates coated with Corning® Matrigel® Matrix, hESC-Qualified (VWR, Fontenay-sous-Bois, FRANCE). The plates were then incubated at 37 °C, 5% CO 2 for 6-30 h. Representative pictures were taken with the Evos XL Core Cell Imaging system at low magnification (4X). Subcutaneous xenografts. This study was conducted in compliance with the National Charter on the ethics of animal experimentation. Our experiments were approved by the "Comité National Institutionnel d'Éthique pour l'Animal de Laboratoire (CIEPAL)" (reference: NCE/2017-383). 10 6 Daoy cells or 0.
Burel-Vandenbos) evaluated marker expression. Technical resources. Dr. Steven C. Clifford (Newcastle-upon-Tyne, UK) provided us with a databank of 250 samples of MB, with transcriptomic and clinical data, including overall and progression-free survival  29. We also compiled and analyzed MB data from the "R2: Genomics Analysis and Visualization Platform" (http://r2. amc. nl) for lymphangiogenesis gene expression. Statistics and reproducibility. Results were expressed in the text as mean ± SEM of at least three independent experiments. Unless otherwise stated, statistical analyses were performed using one-way or two-way ANOVA tests with Dunnett's multiple comparison tests. For two independent groups, Mann-Whitney analyses were performed. Results were considered significant when p-value <0. 05.
For in vitro experiments, 3 to 5 samples were used to assess a biological effect. Most authors consider that replicating three times the same experiment, taking care that cell culture conditions do not change is enough to accurately describe a biological phenomenon. We increased the number of samples for qPCR experiments, where variability is higher. For in vivo experiments, each group of animals is composed of 8-10 individuals. Our experiments did not require blind tests. No data were excluded.
Acknowledgements Financial support: This work is supported by funding from the  Fondation Flavien, Monaco  ( Principality of Monaco ), e-mail: tsunamiflavien@gmail. com, from the  Fondation ARC, Villejuif (FRANCE) , post-doctoral supporting grant  n°PDF20161205242 , from The  Ligue Nationale contre le Cancer  ( Equipe Labellisée 2019 ), from the  Fondation BMS ,  Rueil-Malmaison  ( FRANCE) , e-mail: fondation@bms. com, from the  H2020 Ther-aLymph   Grant  project  ID: 874708.
Powder and solution EPR spectra Single-crystal EPR data are not always easily available, nor is their analysis always straightforward. We were therefore interested to assess whether we could derive useful as to the magnetic structures of ferromagnetic Cu II 3 pyrazolates from powder or frozen solution data. Accordingly, EPR spectra were collected from powders and frozen CH 2 Cl 2 solutions of 1-4. Powder spectra of complexes 1, 2 and 4 exhibited characteristic zfs spectra, attributed to their S = 3/2 ground states. Preliminary fits with a simple giant-spin model yielded excellent results assuming collinear g and D, with solid-state g av values for 1-3 (2. 121, 2. 164, 2. 115, respectively, Table  7 ) following closely the ones determined from magnetic susceptibility data, even reproducing the higher g-value of 2 (Table  2 ).
Acknowledgments The authors thank  Dr. Yiannis Sanakis  for preliminary measurements and helpful discussions,  Dr. David I. Kreiger  for the single crystal of 4, and  Prof. Philippe Turek  for support throughout this study. This project has received funding from the  European Union 's  Horizon 2020 research and innovation programme  under the  Marie Sklodowska-Curie  grant agreement No  746060  (project " CHIRALQUBIT "), by the  Fondation pour la Recherche en Chimie (Strasbourg)  under the  Innovation grant agreement PTu-FRC-0003  (project " MiSSTri ") and the  National Science Foundation  ( CHE-1213683 ).
MA was supported by  Directorate-General for Research and Innovation of the European Commission (RIsk-based Screening for Cervical Cancer , grant number  847845 );  Belgian Foundation Against Cancer  ( IHUVAC  project) and from the  VALCOR  project. LB's research unit has received unrestricted research grants from  Merck ,  Sharpe & Dohme  and  GlaxoSmithKline. MG received travel support and honoraria from  Merck ,  Sharpe & Dohme  to be a speaker. DK, PB, MP, CB, DR, and  EW  declare no competing interests. Where authors are identified as personnel of the  International Agency for Research on Cancer or WHO , the authors alone are responsible for the views expressed in this Article and they do not necessarily represent the decisions, policy or views of the  International Agency for Research on Cancer or WHO.
Patients and Study Designs As part of the original studies, all patients provided written informed consent, and the trials were approved by all relevant review bodies. All trials were conducted in accordance with the principles of the Declaration of Helsinki and the International Conference on Harmonisation Good Clinical Practice guidelines. The popPK and exposure-response analyses included data (clinical cutoff date: January 8, 2019) from 4 clinical studies: 3 monotherapy studies -PAVO (MMY1004; ClinicalTrials. gov Identifier: NCT02519452) Part 2, MMY1008 (NCT03242889), COLUMBA (MMY3012; NCT03277105) -and 1 combination study, PLEIADES (MMY2040; NCT03412565). In all studies, subcutaneous DARA was administered by injection over 3 to 5 minutes at alternating left/right abdominal sites. Additional details on the study designs of each of these studies are presented in Table  1  and Supplementary Methods.
PopPK Analyses Analyses were conducted on the combined PKevaluable data set from studies in which subcutaneous DARA was administered to a total of 487 patients (PAVO Part 2, n = 25; MMY1008, n = 6; COLUMBA, n = 257; PLEIADES, n = 199), and intravenous DARA was administered to a total of 255 patients. The subcutaneous DARA data were described based on the previously developed final structural and covariate model from intravenous DARA, with an additional absorption phase, as no major differences in systemic PK characteristics and covariates were expected between intravenous and subcutaneous administration. 30 he base popPK model for intravenous DARA included a 2-compartment structure with parallel linear and Michaelis-Menten nonlinear elimination pathways. The linear clearance represents the nonspecific clearance for immunoglobulin G (IgG), and the Michaelis-Menten nonlinear elimination represents the saturable target-mediated clearance.
The developed subcutaneous DARA monotherapy popPK model was used to estimate individual PK parameters for combination therapies (D-VMP, D-Rd, and D-VRd) in the PLEIADES study through a maximum a posteriori probability Bayesian approach. 31 erum concentration-time data were used for nonlinear mixed-effects modeling (NONMEM; version 7. 2; ICON plc, Dublin, Ireland). The first-order conditional estimation method was used for continuous dependent variables. The software R (version 3. 4. 1 or higher) was used for any data management, postprocessing of the NONMEM run, simulations, and all other analyses. The package Xpose (version 4. 6. 1) was used for model evaluation. The intrinsic factors explored as covariates in the pooled population PK analysis were body weight, age, sex, race, baseline creatinine clearance, baseline albumin, alanine aminotransferase, alkaline phosphatase, and hepatic dysfunction categories using the National Cancer Institute criteria (based on aspartate aminotransferase and total bilirubin).
Exposure-Response Analyses The exposure-response analysis for subcutaneous DARA monotherapy included data from PAVO Part 2, MMY1008, and COLUMBA; the exposure-response analysis for subcutaneous DARA in combination therapies was based on data from PLEIADES. Responders were defined as patients who achieved partial response or better based on International Myeloma Working Group response criteria. 32 The exposure-efficacy analysis for ORR was evaluated graphically and compared between subcutaneous and intravenous DARA. The exposure-response analysis for safety was explored for selected adverse events (AEs), including overall serious AEs (SAEs), overall grade ≥3 treatment-emergent AEs (TEAEs), neutropenia, and IRRs. Peak DARA concentrations after the first dose and overall peak concentrations (except for IRRs) were investigated for their potential relationship with AEs by exposure quartiles and evaluated graphically.
Patient Demographics and Disease Characteristics The popPK data set comprised 5159 measurable PK samples from 742 patients, with 487 patients in the subcutaneous DARA group (monotherapy, n = 288; combination therapy, n = 199) and 255 patients in the intravenous DARA monotherapy group. Data from 3 patients with no measurable DARA concentrations were excluded from the popPK analysis. Patient baseline demographics and disease covariates are summarized in Table  2. Observed PK of Subcutaneous DARA Subcutaneous DARA (1800 mg) monotherapy and combination therapies achieved similar concentrations over time.
Both centers propose the same follow-up standards, questionnaires, laboratory tests, and physical exams. Medical directors meet every 3 months at the scientific board of the SHCS to discuss protocols and best medical practice. Every 6 months, at the patient visit, physicians of both centers fill in the SHCS case report forms that cover all of the social and medical outpatient assessments. Both centers also collect biannual, physicianassisted, self-reported adherence assessments  [17]. Both centers follow their patients more frequently during the first year of treatment according to the International AIDS Society guidelines. Data collected during each single additional visit, including CD4 count and HIV viral load, are transferred to the SHCS data base. The start date of the study was 2004, the date of start of the medication adherence program in Lausanne.
Data Analysis Baseline was defined as the date of cART initiation. Baseline sociodemographic and clinical variables were described according to center using percentages or median and interquartile ranges (IQRs) as appropriate. Several methods have been used to measure retention in care including measurement of CD4 count and viral load, missed medical visits, and gaps in care  [24, 25]. All laboratory and medical visits are captured and stored systematically in the available SHCS database. After the first year of treatment, visits are conducted on a semiannual basis and include a blood draw 7 to 14 days earlier  [26, 27]. Gaps in care were defined as gaps in blood draws lasting more than 6 or 12 months in duration. We used gaps in blood draws rather than gaps in medical visits because blood draws occur more systematically than medical visits.
The length of the gap in care is the number of weeks between patients' last blood draw and their return to care, stop date, or censoring date. Stop date was defined as the date of the last visit (eg, before the patient was transferred to another clinic). The censoring date was the administrative closure of the database, which was on December 31, 2012. The incidence of gaps, length of the gaps, and time until the first gap in care were also calculated. The percentage of patients with gaps was compared using χ 2 test. The median length of the gap and the median time until the first gap in care were compared using Wilcoxon rank-sum test. Individuals in the SHCS are asked biannually about missed doses of cART in the last 4 weeks (daily, more than once a week, once a week, once every second week, once a month, never) through the SHCS physician-assisted-self-reported adherence questionnaire (SHCS-AQ).
On the one hand, it is important to note that both ID centers work similarly in terms of medical activity. Medical interns attend medical school in Geneva or Lausanne, are trained in ID service in either center, and rotate every year. Second, several methodological approaches were used to adjust for differences in patient characteristics to attempt to balance the 2 centers as in a controlled trial. Based on the data collected through the SHCS, we tried to cover all crucial sociodemographic, clinical, and therapeutic variables. The stability of the intervention effect over all models, except the ISR-weighted model, suggests the results are quite robust. The ISR-weighted models consider the intervention group to be the standard population rather than the entire population from both centers, so they answer a different question than the other models.
However, we acknowledge we cannot completely exclude the possibility that fraction 1 may have contained a small fraction of stored RNPs, that we did not characterise in this study. Accordingly, we pooled the RNA fractions representing the NP-(fractions 2-5) and P-populations (fractions 6-10) and subjected them to RNA-seq according to protocols described in the "Methods" section. A principal component analysis (PCA) of the RNA-seq data (Figure  4a ) revealed that the component 1 (PC1; 28%) clustered the NP and P samples and that the NP samples were less variable in both components (PC2; 21%). Although a correlation of gene expression values showed that there was no profoundly obvious global difference between NP and P datasets (Figure  4b ). Out of all 25,574 mRNAs transcribed from autosomes and chromosome X; 11,511 transcripts (with a value of fragments per kilobase per million reads mapped (FPKM) > 0.