mann–whitney tests in graphpad prism Search Results


statsdirect software  (StatsDirect ltd)
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StatsDirect ltd statsdirect software
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statistics mann whitney non-parametric test  (RStudio)
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RStudio statistics mann whitney non-parametric test
Statistics Mann Whitney Non Parametric Test, supplied by RStudio, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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2017 n a agxt human tagged orf  (OriGene)
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Figure 1. Dysregulated glycine and oxalate metabolism in patients and mice with atherosclerosis (A) Schematic representation of glycine metabolic pathways. (B–E) Targeted metabolomics assessing the ratios of glycine to (B) serine, (C) threonine, (D) alanine, and (E) oxalate in serum from age- and sex- matched patients with or without sCAD (n = 24). (F) En face analysis of atherosclerotic lesions in male Apoe/ mice fed a standard diet (SD) or Western diet (WD) for 12 weeks (n = 5). (G–J) Targeted metabolomics assessing the ratios of glycine to (G) serine, (H) threonine, (I) alanine, and (J) oxalate in plasma from male Apoe/ mice fed a SD or WD for 12 weeks (n = 5). (K) Western blot analysis of <t>AGXT</t> protein abun- dance in livers from Apoe/ mice fed a SD or WD for 12 weeks (n = 5). Mann-Whitney U test for (B)-(E), (G), (H), and (K). Unpaired t test for (F), (I), and (J). Data are pre- sented as mean ± SEM. All points and p values are shown.
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inc software  (STATA Corporation)
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STATA Corporation inc software
Figure 1. Dysregulated glycine and oxalate metabolism in patients and mice with atherosclerosis (A) Schematic representation of glycine metabolic pathways. (B–E) Targeted metabolomics assessing the ratios of glycine to (B) serine, (C) threonine, (D) alanine, and (E) oxalate in serum from age- and sex- matched patients with or without sCAD (n = 24). (F) En face analysis of atherosclerotic lesions in male Apoe/ mice fed a standard diet (SD) or Western diet (WD) for 12 weeks (n = 5). (G–J) Targeted metabolomics assessing the ratios of glycine to (G) serine, (H) threonine, (I) alanine, and (J) oxalate in plasma from male Apoe/ mice fed a SD or WD for 12 weeks (n = 5). (K) Western blot analysis of <t>AGXT</t> protein abun- dance in livers from Apoe/ mice fed a SD or WD for 12 weeks (n = 5). Mann-Whitney U test for (B)-(E), (G), (H), and (K). Unpaired t test for (F), (I), and (J). Data are pre- sented as mean ± SEM. All points and p values are shown.
Inc Software, supplied by STATA Corporation, used in various techniques. Bioz Stars score: 99/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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kruskal-wallis rank sum test  (RStudio)
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RStudio kruskal-wallis rank sum test
Figure 1. Dysregulated glycine and oxalate metabolism in patients and mice with atherosclerosis (A) Schematic representation of glycine metabolic pathways. (B–E) Targeted metabolomics assessing the ratios of glycine to (B) serine, (C) threonine, (D) alanine, and (E) oxalate in serum from age- and sex- matched patients with or without sCAD (n = 24). (F) En face analysis of atherosclerotic lesions in male Apoe/ mice fed a standard diet (SD) or Western diet (WD) for 12 weeks (n = 5). (G–J) Targeted metabolomics assessing the ratios of glycine to (G) serine, (H) threonine, (I) alanine, and (J) oxalate in plasma from male Apoe/ mice fed a SD or WD for 12 weeks (n = 5). (K) Western blot analysis of <t>AGXT</t> protein abun- dance in livers from Apoe/ mice fed a SD or WD for 12 weeks (n = 5). Mann-Whitney U test for (B)-(E), (G), (H), and (K). Unpaired t test for (F), (I), and (J). Data are pre- sented as mean ± SEM. All points and p values are shown.
Kruskal Wallis Rank Sum Test, supplied by RStudio, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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il21  (Addgene inc)
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Addgene inc il21
Figure 1. <t>IL-21/IL-21R</t> signaling reduces murine L-GMPs in vivo (A) Il21R mRNA expression (qRT-PCR) in FACS-sorted L-GMPs from the BM of BL/6 AML mice 35 days after leukemia transplantation (n = 10 mice). Red bar indicates the mean. Each dot represents the mean of two technical replicates. (B) IL-21 levels in BM samples from AML mice (n = 11 mice) and naive controls (n = 5 mice). Each dot represents the mean of two technical replicates. Data are shown as mean ± SEM. Statistics: Student’s t test. (C) Fold change colony-forming units from FACS-sorted L-GMPs cultured in methylcellulose for 7 days in the presence or absence of 300 pg/mL rm-IL-21 (n = 6 mice/group). Each dot represents the mean of two technical replicates. Pooled data from two independent experiments are shown and displayed as mean ± SD. Statistics: Student’s t test. (D) Experimental setup: 5 3 104 MLL-AF9-GFP-transduced LSKs from the BM of BL/6 donors were injected intravenously into non-irradiated BL/6 and <t>Il21/</t>
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mann–whitney u-test  (STATCON GmbH)
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STATCON GmbH mann–whitney u-test
Figure 1. <t>IL-21/IL-21R</t> signaling reduces murine L-GMPs in vivo (A) Il21R mRNA expression (qRT-PCR) in FACS-sorted L-GMPs from the BM of BL/6 AML mice 35 days after leukemia transplantation (n = 10 mice). Red bar indicates the mean. Each dot represents the mean of two technical replicates. (B) IL-21 levels in BM samples from AML mice (n = 11 mice) and naive controls (n = 5 mice). Each dot represents the mean of two technical replicates. Data are shown as mean ± SEM. Statistics: Student’s t test. (C) Fold change colony-forming units from FACS-sorted L-GMPs cultured in methylcellulose for 7 days in the presence or absence of 300 pg/mL rm-IL-21 (n = 6 mice/group). Each dot represents the mean of two technical replicates. Pooled data from two independent experiments are shown and displayed as mean ± SD. Statistics: Student’s t test. (D) Experimental setup: 5 3 104 MLL-AF9-GFP-transduced LSKs from the BM of BL/6 donors were injected intravenously into non-irradiated BL/6 and <t>Il21/</t>
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r studio software  (RStudio)
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RStudio r studio software
Figure 1. <t>IL-21/IL-21R</t> signaling reduces murine L-GMPs in vivo (A) Il21R mRNA expression (qRT-PCR) in FACS-sorted L-GMPs from the BM of BL/6 AML mice 35 days after leukemia transplantation (n = 10 mice). Red bar indicates the mean. Each dot represents the mean of two technical replicates. (B) IL-21 levels in BM samples from AML mice (n = 11 mice) and naive controls (n = 5 mice). Each dot represents the mean of two technical replicates. Data are shown as mean ± SEM. Statistics: Student’s t test. (C) Fold change colony-forming units from FACS-sorted L-GMPs cultured in methylcellulose for 7 days in the presence or absence of 300 pg/mL rm-IL-21 (n = 6 mice/group). Each dot represents the mean of two technical replicates. Pooled data from two independent experiments are shown and displayed as mean ± SD. Statistics: Student’s t test. (D) Experimental setup: 5 3 104 MLL-AF9-GFP-transduced LSKs from the BM of BL/6 donors were injected intravenously into non-irradiated BL/6 and <t>Il21/</t>
R Studio Software, supplied by RStudio, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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fisher’s exact test  (MedCalc Software Ltd)
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MedCalc Software Ltd fisher’s exact test
Figure 1. <t>IL-21/IL-21R</t> signaling reduces murine L-GMPs in vivo (A) Il21R mRNA expression (qRT-PCR) in FACS-sorted L-GMPs from the BM of BL/6 AML mice 35 days after leukemia transplantation (n = 10 mice). Red bar indicates the mean. Each dot represents the mean of two technical replicates. (B) IL-21 levels in BM samples from AML mice (n = 11 mice) and naive controls (n = 5 mice). Each dot represents the mean of two technical replicates. Data are shown as mean ± SEM. Statistics: Student’s t test. (C) Fold change colony-forming units from FACS-sorted L-GMPs cultured in methylcellulose for 7 days in the presence or absence of 300 pg/mL rm-IL-21 (n = 6 mice/group). Each dot represents the mean of two technical replicates. Pooled data from two independent experiments are shown and displayed as mean ± SD. Statistics: Student’s t test. (D) Experimental setup: 5 3 104 MLL-AF9-GFP-transduced LSKs from the BM of BL/6 donors were injected intravenously into non-irradiated BL/6 and <t>Il21/</t>
Fisher’s Exact Test, supplied by MedCalc Software Ltd, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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mann whitney test  (Dotmatics Limited)
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Dotmatics Limited mann whitney test
Figure 1. <t>IL-21/IL-21R</t> signaling reduces murine L-GMPs in vivo (A) Il21R mRNA expression (qRT-PCR) in FACS-sorted L-GMPs from the BM of BL/6 AML mice 35 days after leukemia transplantation (n = 10 mice). Red bar indicates the mean. Each dot represents the mean of two technical replicates. (B) IL-21 levels in BM samples from AML mice (n = 11 mice) and naive controls (n = 5 mice). Each dot represents the mean of two technical replicates. Data are shown as mean ± SEM. Statistics: Student’s t test. (C) Fold change colony-forming units from FACS-sorted L-GMPs cultured in methylcellulose for 7 days in the presence or absence of 300 pg/mL rm-IL-21 (n = 6 mice/group). Each dot represents the mean of two technical replicates. Pooled data from two independent experiments are shown and displayed as mean ± SD. Statistics: Student’s t test. (D) Experimental setup: 5 3 104 MLL-AF9-GFP-transduced LSKs from the BM of BL/6 donors were injected intravenously into non-irradiated BL/6 and <t>Il21/</t>
Mann Whitney Test, supplied by Dotmatics Limited, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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algorithms synapsemech  (Addgene inc)
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Figure 3. The CTL synapse exhibits negative membrane curvatures at regions cleared of F-actin (A) Timelapse confocal images of live CTL F-actin distribution at the synapse during cognate tumor interaction in the DPA assay. Top: maximum intensity projection. Middle/bottom: <t>SynapseMech</t> analysis of CTL F-actin distribution and membrane curvature within the synapse. Displayed F-actin signal restricted to synapse region and intensity-thresholded. Scale bar, 5 mm. (B) Dynamics of area cleared from F-actin (gray shaded region) and average mean curvature of complete (dark gray) and F-actin-cleared (brown) synapse area for data shown in (A). (C) Time-averaged mean curvature of complete, F-actin-cleared, and F-actin-rich (outside of cleared space) synapse area. Boxplot shows median, IQR, and range for n = 6 cells. p values by matched one-way ANOVA test with prior Kolmogorov-Smirnov test for normality. (D) Surface tension of resting CTLs, tumor cells and tumor cytoplasts. Boxplots show median, IQR and range for n = 24 (CTL), n = 36 (tumor cells), and n = 19 (tumor cytoplasts). p values by Mann-Whitney test. (E) Timelapse confocal images of a live CTL interacting with a cognate tumor cytoplast in the DPA assay. t = 0 s: time of contact. Scale bar, 10 mm. (F) Main: dynamics of average mean synapse curvature of CTL interacting with cognate tumor cell or tumor cytoplast. Data are mean ± SEM (shaded area) for n = 7 (tumor cell) and n = 8 (tumor cytoplast). Inset: time-averaged mean synapse curvature of CTLs interacting with cognate tumor cells or tumor cytoplasts. p value by Mann-Whitney test.
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mann whitney two  (Xybion Inc)
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Xybion Inc mann whitney two
Figure 3. The CTL synapse exhibits negative membrane curvatures at regions cleared of F-actin (A) Timelapse confocal images of live CTL F-actin distribution at the synapse during cognate tumor interaction in the DPA assay. Top: maximum intensity projection. Middle/bottom: <t>SynapseMech</t> analysis of CTL F-actin distribution and membrane curvature within the synapse. Displayed F-actin signal restricted to synapse region and intensity-thresholded. Scale bar, 5 mm. (B) Dynamics of area cleared from F-actin (gray shaded region) and average mean curvature of complete (dark gray) and F-actin-cleared (brown) synapse area for data shown in (A). (C) Time-averaged mean curvature of complete, F-actin-cleared, and F-actin-rich (outside of cleared space) synapse area. Boxplot shows median, IQR, and range for n = 6 cells. p values by matched one-way ANOVA test with prior Kolmogorov-Smirnov test for normality. (D) Surface tension of resting CTLs, tumor cells and tumor cytoplasts. Boxplots show median, IQR and range for n = 24 (CTL), n = 36 (tumor cells), and n = 19 (tumor cytoplasts). p values by Mann-Whitney test. (E) Timelapse confocal images of a live CTL interacting with a cognate tumor cytoplast in the DPA assay. t = 0 s: time of contact. Scale bar, 10 mm. (F) Main: dynamics of average mean synapse curvature of CTL interacting with cognate tumor cell or tumor cytoplast. Data are mean ± SEM (shaded area) for n = 7 (tumor cell) and n = 8 (tumor cytoplast). Inset: time-averaged mean synapse curvature of CTLs interacting with cognate tumor cells or tumor cytoplasts. p value by Mann-Whitney test.
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Image Search Results


Figure 1. Dysregulated glycine and oxalate metabolism in patients and mice with atherosclerosis (A) Schematic representation of glycine metabolic pathways. (B–E) Targeted metabolomics assessing the ratios of glycine to (B) serine, (C) threonine, (D) alanine, and (E) oxalate in serum from age- and sex- matched patients with or without sCAD (n = 24). (F) En face analysis of atherosclerotic lesions in male Apoe/ mice fed a standard diet (SD) or Western diet (WD) for 12 weeks (n = 5). (G–J) Targeted metabolomics assessing the ratios of glycine to (G) serine, (H) threonine, (I) alanine, and (J) oxalate in plasma from male Apoe/ mice fed a SD or WD for 12 weeks (n = 5). (K) Western blot analysis of AGXT protein abun- dance in livers from Apoe/ mice fed a SD or WD for 12 weeks (n = 5). Mann-Whitney U test for (B)-(E), (G), (H), and (K). Unpaired t test for (F), (I), and (J). Data are pre- sented as mean ± SEM. All points and p values are shown.

Journal: Cell reports

Article Title: Dysregulated oxalate metabolism is a driver and therapeutic target in atherosclerosis.

doi: 10.1016/j.celrep.2021.109420

Figure Lengend Snippet: Figure 1. Dysregulated glycine and oxalate metabolism in patients and mice with atherosclerosis (A) Schematic representation of glycine metabolic pathways. (B–E) Targeted metabolomics assessing the ratios of glycine to (B) serine, (C) threonine, (D) alanine, and (E) oxalate in serum from age- and sex- matched patients with or without sCAD (n = 24). (F) En face analysis of atherosclerotic lesions in male Apoe/ mice fed a standard diet (SD) or Western diet (WD) for 12 weeks (n = 5). (G–J) Targeted metabolomics assessing the ratios of glycine to (G) serine, (H) threonine, (I) alanine, and (J) oxalate in plasma from male Apoe/ mice fed a SD or WD for 12 weeks (n = 5). (K) Western blot analysis of AGXT protein abun- dance in livers from Apoe/ mice fed a SD or WD for 12 weeks (n = 5). Mann-Whitney U test for (B)-(E), (G), (H), and (K). Unpaired t test for (F), (I), and (J). Data are pre- sented as mean ± SEM. All points and p values are shown.

Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Primer sequences for Agxt genotyping:, forward: 50-ACACCTCCACTGTCCTG TCC-30 Integrated DNA Technologies N/A Primer sequences for Agxt genotyping:, reverse: 50-GGTCAGATCTGCCTGCTA CC-30 Integrated DNA Technologies N/A Primer for Sanger sequencing:, 50-GCAGAGCTAGCTGGGAAATG-30 Integrated DNA Technologies N/A Primer sequences for Apoe genotyping:, forward: 50-GCCTAGCCGAGGGAGAG CCG-30 Integrated DNA Technologies N/A Primer sequences for Apoe genotyping:, reverse 1: 50-TGTGACTTGGGAGCTC TGCAGC-30 Integrated DNA Technologies N/A Primer sequences for Apoe genotyping:, reverse 2: 50-GCCGCCCCGACTGC ATCT-30 Integrated DNA Technologies N/A Primer sequences for qPCR, see Table S2 Integrated DNA Technologies N/A Recombinant DNA pAdDeltaF6 Jiandie Lin’s lab, University of Michigan; Guo et al., 2017 N/A pAAV2/8 Jiandie Lin’s lab, University of Michigan; Guo et al., 2017 N/A pAAV-TBG-GFP Jiandie Lin’s lab, University of Michigan; Guo et al., 2017 N/A pAAV-TBG-MCS Jiandie Lin’s lab, University of Michigan; Guo et al., 2017 N/A AGXT Human Tagged ORF Clone Origene CAT# RG212899 Software and algorithms ImageJ National Institutes of Health RRID:SCR_003070 https://imagej.nih.gov/ ij/download.html GraphPad Prism GraphPad Software Version 8.0 RRID:SCR_002798 https:// www.graphpad.com/scientific-software/ prism/ R statistics Hosted by Vienna, University of Economics and Business http://www.R-project.org/ RStudio RStudio Team (2020). https://www.rstudio.com/ RRID:SCR_000432 IBM SPSS software IBM Corporation Version 23.0; RRID:SCR_019096 https:// www.ibm.com/analytics/ spss-statistics-software FastQC Babraham Bioinformatics Version 0.11.8 RRID:SCR_014583 https:// www.bioinformatics.babraham.ac.uk/ projects/fastqc/ Trimmomatic Bolger et al., 2014 Version 0.35 RRID:SCR_011848 http:// www.usadellab.org/cms/? page=trimmomatic HISAT2 Kim et al., 2015 Version 2.1.0.13; RRID:SCR_015530 http:// daehwankimlab.github.io/hisat2/ download/ HTSeq Anders et al., 2015 Version 0.6.0; RRID:SCR_005514 https:// htseq.readthedocs.io/en/master/install. html (Continued on next page) Cell Reports 36, 109420, July 27, 2021 e3 Article ll OPEN ACCESS

Techniques: Western Blot, Clinical Proteomics, MANN-WHITNEY

Figure 2. AGXT deficiency exacerbates atherosclerosis in male Apoe/ mice (A) Western blot analysis confirming the loss of AGXT in livers from male Agxt//Apoe/ mice (n = 6). (B) Glycine/oxalate ratio in plasma from male Agxt//Apoe/ mice and their Agxt+/+/Apoe/

Journal: Cell reports

Article Title: Dysregulated oxalate metabolism is a driver and therapeutic target in atherosclerosis.

doi: 10.1016/j.celrep.2021.109420

Figure Lengend Snippet: Figure 2. AGXT deficiency exacerbates atherosclerosis in male Apoe/ mice (A) Western blot analysis confirming the loss of AGXT in livers from male Agxt//Apoe/ mice (n = 6). (B) Glycine/oxalate ratio in plasma from male Agxt//Apoe/ mice and their Agxt+/+/Apoe/

Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Primer sequences for Agxt genotyping:, forward: 50-ACACCTCCACTGTCCTG TCC-30 Integrated DNA Technologies N/A Primer sequences for Agxt genotyping:, reverse: 50-GGTCAGATCTGCCTGCTA CC-30 Integrated DNA Technologies N/A Primer for Sanger sequencing:, 50-GCAGAGCTAGCTGGGAAATG-30 Integrated DNA Technologies N/A Primer sequences for Apoe genotyping:, forward: 50-GCCTAGCCGAGGGAGAG CCG-30 Integrated DNA Technologies N/A Primer sequences for Apoe genotyping:, reverse 1: 50-TGTGACTTGGGAGCTC TGCAGC-30 Integrated DNA Technologies N/A Primer sequences for Apoe genotyping:, reverse 2: 50-GCCGCCCCGACTGC ATCT-30 Integrated DNA Technologies N/A Primer sequences for qPCR, see Table S2 Integrated DNA Technologies N/A Recombinant DNA pAdDeltaF6 Jiandie Lin’s lab, University of Michigan; Guo et al., 2017 N/A pAAV2/8 Jiandie Lin’s lab, University of Michigan; Guo et al., 2017 N/A pAAV-TBG-GFP Jiandie Lin’s lab, University of Michigan; Guo et al., 2017 N/A pAAV-TBG-MCS Jiandie Lin’s lab, University of Michigan; Guo et al., 2017 N/A AGXT Human Tagged ORF Clone Origene CAT# RG212899 Software and algorithms ImageJ National Institutes of Health RRID:SCR_003070 https://imagej.nih.gov/ ij/download.html GraphPad Prism GraphPad Software Version 8.0 RRID:SCR_002798 https:// www.graphpad.com/scientific-software/ prism/ R statistics Hosted by Vienna, University of Economics and Business http://www.R-project.org/ RStudio RStudio Team (2020). https://www.rstudio.com/ RRID:SCR_000432 IBM SPSS software IBM Corporation Version 23.0; RRID:SCR_019096 https:// www.ibm.com/analytics/ spss-statistics-software FastQC Babraham Bioinformatics Version 0.11.8 RRID:SCR_014583 https:// www.bioinformatics.babraham.ac.uk/ projects/fastqc/ Trimmomatic Bolger et al., 2014 Version 0.35 RRID:SCR_011848 http:// www.usadellab.org/cms/? page=trimmomatic HISAT2 Kim et al., 2015 Version 2.1.0.13; RRID:SCR_015530 http:// daehwankimlab.github.io/hisat2/ download/ HTSeq Anders et al., 2015 Version 0.6.0; RRID:SCR_005514 https:// htseq.readthedocs.io/en/master/install. html (Continued on next page) Cell Reports 36, 109420, July 27, 2021 e3 Article ll OPEN ACCESS

Techniques: Western Blot, Clinical Proteomics

Figure 3. Oxalate homeostasis is main- tained and atherosclerosis is unaltered in fe- male Agxt//Apoe/ mice (A) Western blot analysis confirming the loss of AGXT in livers from female Agxt//Apoe/ mice (n = 6). (B) GlGlycine/oxalate ratio in plasma from female Agxt//Apoe/ and Agxt+/+/Apoe/ mice (n = 10). (C–G) Female Agxt//Apoe/ and Agxt+/+/ Apoe/ mice were fed a WD for 12 weeks (n = 10): (C) plasma TC, (D) atherosclerosis in the aortic tree, (E) H&E staining, (F) ORO staining, and (G) Mac-2 immunohistochemistry of aortic sinus. Scale bars, 200 mm. Unpaired t test for (B), (D), (E), and (G). Mann- Whitney U test for (C) and (F). Data are presented as mean ± SEM. All points and p values are shown.

Journal: Cell reports

Article Title: Dysregulated oxalate metabolism is a driver and therapeutic target in atherosclerosis.

doi: 10.1016/j.celrep.2021.109420

Figure Lengend Snippet: Figure 3. Oxalate homeostasis is main- tained and atherosclerosis is unaltered in fe- male Agxt//Apoe/ mice (A) Western blot analysis confirming the loss of AGXT in livers from female Agxt//Apoe/ mice (n = 6). (B) GlGlycine/oxalate ratio in plasma from female Agxt//Apoe/ and Agxt+/+/Apoe/ mice (n = 10). (C–G) Female Agxt//Apoe/ and Agxt+/+/ Apoe/ mice were fed a WD for 12 weeks (n = 10): (C) plasma TC, (D) atherosclerosis in the aortic tree, (E) H&E staining, (F) ORO staining, and (G) Mac-2 immunohistochemistry of aortic sinus. Scale bars, 200 mm. Unpaired t test for (B), (D), (E), and (G). Mann- Whitney U test for (C) and (F). Data are presented as mean ± SEM. All points and p values are shown.

Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Primer sequences for Agxt genotyping:, forward: 50-ACACCTCCACTGTCCTG TCC-30 Integrated DNA Technologies N/A Primer sequences for Agxt genotyping:, reverse: 50-GGTCAGATCTGCCTGCTA CC-30 Integrated DNA Technologies N/A Primer for Sanger sequencing:, 50-GCAGAGCTAGCTGGGAAATG-30 Integrated DNA Technologies N/A Primer sequences for Apoe genotyping:, forward: 50-GCCTAGCCGAGGGAGAG CCG-30 Integrated DNA Technologies N/A Primer sequences for Apoe genotyping:, reverse 1: 50-TGTGACTTGGGAGCTC TGCAGC-30 Integrated DNA Technologies N/A Primer sequences for Apoe genotyping:, reverse 2: 50-GCCGCCCCGACTGC ATCT-30 Integrated DNA Technologies N/A Primer sequences for qPCR, see Table S2 Integrated DNA Technologies N/A Recombinant DNA pAdDeltaF6 Jiandie Lin’s lab, University of Michigan; Guo et al., 2017 N/A pAAV2/8 Jiandie Lin’s lab, University of Michigan; Guo et al., 2017 N/A pAAV-TBG-GFP Jiandie Lin’s lab, University of Michigan; Guo et al., 2017 N/A pAAV-TBG-MCS Jiandie Lin’s lab, University of Michigan; Guo et al., 2017 N/A AGXT Human Tagged ORF Clone Origene CAT# RG212899 Software and algorithms ImageJ National Institutes of Health RRID:SCR_003070 https://imagej.nih.gov/ ij/download.html GraphPad Prism GraphPad Software Version 8.0 RRID:SCR_002798 https:// www.graphpad.com/scientific-software/ prism/ R statistics Hosted by Vienna, University of Economics and Business http://www.R-project.org/ RStudio RStudio Team (2020). https://www.rstudio.com/ RRID:SCR_000432 IBM SPSS software IBM Corporation Version 23.0; RRID:SCR_019096 https:// www.ibm.com/analytics/ spss-statistics-software FastQC Babraham Bioinformatics Version 0.11.8 RRID:SCR_014583 https:// www.bioinformatics.babraham.ac.uk/ projects/fastqc/ Trimmomatic Bolger et al., 2014 Version 0.35 RRID:SCR_011848 http:// www.usadellab.org/cms/? page=trimmomatic HISAT2 Kim et al., 2015 Version 2.1.0.13; RRID:SCR_015530 http:// daehwankimlab.github.io/hisat2/ download/ HTSeq Anders et al., 2015 Version 0.6.0; RRID:SCR_005514 https:// htseq.readthedocs.io/en/master/install. html (Continued on next page) Cell Reports 36, 109420, July 27, 2021 e3 Article ll OPEN ACCESS

Techniques: Western Blot, Clinical Proteomics, Staining, Immunohistochemistry, MANN-WHITNEY

Figure 4. Dysregulated oxalate metabolism induces a pro-inflammatory response and CCL5 release (A–C) RNA sequencing of livers collected from male Agxt//Apoe/ and Agxt+/+/Apoe/ mice fed a WD for 12 weeks (n = 5): (A) volcano plot of DEGs (padj < 0.05, log2 fold change > 1) in male Agxt//Apoe/ versus Agxt+/+/Apoe/ mice (blue indicates downregulated; red indicates upregulated). (B) Kyoto Encyclopedia of Genes and Genomes (KEGG)-based pathway analysis. The significance of the enrichment was determined by a right-tailed Fisher’s exact test followed by a Benjamini-Hochberg multiple testing adjustment. (C) Heatmap-based representation of 50 DEGs linking inflammation, fibrogenesis, and the metabolism of lipids and sterols/steroids with atherosclerosis.

Journal: Cell reports

Article Title: Dysregulated oxalate metabolism is a driver and therapeutic target in atherosclerosis.

doi: 10.1016/j.celrep.2021.109420

Figure Lengend Snippet: Figure 4. Dysregulated oxalate metabolism induces a pro-inflammatory response and CCL5 release (A–C) RNA sequencing of livers collected from male Agxt//Apoe/ and Agxt+/+/Apoe/ mice fed a WD for 12 weeks (n = 5): (A) volcano plot of DEGs (padj < 0.05, log2 fold change > 1) in male Agxt//Apoe/ versus Agxt+/+/Apoe/ mice (blue indicates downregulated; red indicates upregulated). (B) Kyoto Encyclopedia of Genes and Genomes (KEGG)-based pathway analysis. The significance of the enrichment was determined by a right-tailed Fisher’s exact test followed by a Benjamini-Hochberg multiple testing adjustment. (C) Heatmap-based representation of 50 DEGs linking inflammation, fibrogenesis, and the metabolism of lipids and sterols/steroids with atherosclerosis.

Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Primer sequences for Agxt genotyping:, forward: 50-ACACCTCCACTGTCCTG TCC-30 Integrated DNA Technologies N/A Primer sequences for Agxt genotyping:, reverse: 50-GGTCAGATCTGCCTGCTA CC-30 Integrated DNA Technologies N/A Primer for Sanger sequencing:, 50-GCAGAGCTAGCTGGGAAATG-30 Integrated DNA Technologies N/A Primer sequences for Apoe genotyping:, forward: 50-GCCTAGCCGAGGGAGAG CCG-30 Integrated DNA Technologies N/A Primer sequences for Apoe genotyping:, reverse 1: 50-TGTGACTTGGGAGCTC TGCAGC-30 Integrated DNA Technologies N/A Primer sequences for Apoe genotyping:, reverse 2: 50-GCCGCCCCGACTGC ATCT-30 Integrated DNA Technologies N/A Primer sequences for qPCR, see Table S2 Integrated DNA Technologies N/A Recombinant DNA pAdDeltaF6 Jiandie Lin’s lab, University of Michigan; Guo et al., 2017 N/A pAAV2/8 Jiandie Lin’s lab, University of Michigan; Guo et al., 2017 N/A pAAV-TBG-GFP Jiandie Lin’s lab, University of Michigan; Guo et al., 2017 N/A pAAV-TBG-MCS Jiandie Lin’s lab, University of Michigan; Guo et al., 2017 N/A AGXT Human Tagged ORF Clone Origene CAT# RG212899 Software and algorithms ImageJ National Institutes of Health RRID:SCR_003070 https://imagej.nih.gov/ ij/download.html GraphPad Prism GraphPad Software Version 8.0 RRID:SCR_002798 https:// www.graphpad.com/scientific-software/ prism/ R statistics Hosted by Vienna, University of Economics and Business http://www.R-project.org/ RStudio RStudio Team (2020). https://www.rstudio.com/ RRID:SCR_000432 IBM SPSS software IBM Corporation Version 23.0; RRID:SCR_019096 https:// www.ibm.com/analytics/ spss-statistics-software FastQC Babraham Bioinformatics Version 0.11.8 RRID:SCR_014583 https:// www.bioinformatics.babraham.ac.uk/ projects/fastqc/ Trimmomatic Bolger et al., 2014 Version 0.35 RRID:SCR_011848 http:// www.usadellab.org/cms/? page=trimmomatic HISAT2 Kim et al., 2015 Version 2.1.0.13; RRID:SCR_015530 http:// daehwankimlab.github.io/hisat2/ download/ HTSeq Anders et al., 2015 Version 0.6.0; RRID:SCR_005514 https:// htseq.readthedocs.io/en/master/install. html (Continued on next page) Cell Reports 36, 109420, July 27, 2021 e3 Article ll OPEN ACCESS

Techniques: RNA Sequencing

Figure 5. Oxalate overload induces mito- chondrial dysfunction and overproduction of superoxide, leading to CCL5 release in macrophages (A) qPCR analyses of genes regulating redox ho- meostasis in livers from male Agxt//Apoe/

Journal: Cell reports

Article Title: Dysregulated oxalate metabolism is a driver and therapeutic target in atherosclerosis.

doi: 10.1016/j.celrep.2021.109420

Figure Lengend Snippet: Figure 5. Oxalate overload induces mito- chondrial dysfunction and overproduction of superoxide, leading to CCL5 release in macrophages (A) qPCR analyses of genes regulating redox ho- meostasis in livers from male Agxt//Apoe/

Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Primer sequences for Agxt genotyping:, forward: 50-ACACCTCCACTGTCCTG TCC-30 Integrated DNA Technologies N/A Primer sequences for Agxt genotyping:, reverse: 50-GGTCAGATCTGCCTGCTA CC-30 Integrated DNA Technologies N/A Primer for Sanger sequencing:, 50-GCAGAGCTAGCTGGGAAATG-30 Integrated DNA Technologies N/A Primer sequences for Apoe genotyping:, forward: 50-GCCTAGCCGAGGGAGAG CCG-30 Integrated DNA Technologies N/A Primer sequences for Apoe genotyping:, reverse 1: 50-TGTGACTTGGGAGCTC TGCAGC-30 Integrated DNA Technologies N/A Primer sequences for Apoe genotyping:, reverse 2: 50-GCCGCCCCGACTGC ATCT-30 Integrated DNA Technologies N/A Primer sequences for qPCR, see Table S2 Integrated DNA Technologies N/A Recombinant DNA pAdDeltaF6 Jiandie Lin’s lab, University of Michigan; Guo et al., 2017 N/A pAAV2/8 Jiandie Lin’s lab, University of Michigan; Guo et al., 2017 N/A pAAV-TBG-GFP Jiandie Lin’s lab, University of Michigan; Guo et al., 2017 N/A pAAV-TBG-MCS Jiandie Lin’s lab, University of Michigan; Guo et al., 2017 N/A AGXT Human Tagged ORF Clone Origene CAT# RG212899 Software and algorithms ImageJ National Institutes of Health RRID:SCR_003070 https://imagej.nih.gov/ ij/download.html GraphPad Prism GraphPad Software Version 8.0 RRID:SCR_002798 https:// www.graphpad.com/scientific-software/ prism/ R statistics Hosted by Vienna, University of Economics and Business http://www.R-project.org/ RStudio RStudio Team (2020). https://www.rstudio.com/ RRID:SCR_000432 IBM SPSS software IBM Corporation Version 23.0; RRID:SCR_019096 https:// www.ibm.com/analytics/ spss-statistics-software FastQC Babraham Bioinformatics Version 0.11.8 RRID:SCR_014583 https:// www.bioinformatics.babraham.ac.uk/ projects/fastqc/ Trimmomatic Bolger et al., 2014 Version 0.35 RRID:SCR_011848 http:// www.usadellab.org/cms/? page=trimmomatic HISAT2 Kim et al., 2015 Version 2.1.0.13; RRID:SCR_015530 http:// daehwankimlab.github.io/hisat2/ download/ HTSeq Anders et al., 2015 Version 0.6.0; RRID:SCR_005514 https:// htseq.readthedocs.io/en/master/install. html (Continued on next page) Cell Reports 36, 109420, July 27, 2021 e3 Article ll OPEN ACCESS

Techniques:

Figure 6. AAV-mediated overexpression of AGXT reduces oxidative stress and inflammation AAV-AGXT or AAV-GFP was injected into male Apoe/ mice and the mice were fed a WD for 12 weeks (n = 8). (A) Western blot analysis confirming the over- expression of AGXT in livers from mice treated with AAV-AGXT. (B) Plasma glycine/oxalate ratio. (C and D) qPCR analyses of genes regulating (C) redox homeostasis and (D) inflammatory re- sponses. Gene expression levels were normalized to 18S. (E and F) Plasma concentrations of (E) CCL2 and (F) CCL5. (G) DHE fluorescence and Mac-2 immunofluores- cence in the aortic sinuses. Scale bars, 50 mm. Unpaired t test for (B), (E), and (G), and Mann- Whitney U test for (F). Statistical differences in gene expression in (C) and (D) were tested using unpaired t test or Mann-Whitney U test, depending on normality tests. *P < 0.05, **P <0.01, ***P <0.001 versus mice injected with AAV-GFP. Data are presented as mean ± SEM. All points and p values are shown.

Journal: Cell reports

Article Title: Dysregulated oxalate metabolism is a driver and therapeutic target in atherosclerosis.

doi: 10.1016/j.celrep.2021.109420

Figure Lengend Snippet: Figure 6. AAV-mediated overexpression of AGXT reduces oxidative stress and inflammation AAV-AGXT or AAV-GFP was injected into male Apoe/ mice and the mice were fed a WD for 12 weeks (n = 8). (A) Western blot analysis confirming the over- expression of AGXT in livers from mice treated with AAV-AGXT. (B) Plasma glycine/oxalate ratio. (C and D) qPCR analyses of genes regulating (C) redox homeostasis and (D) inflammatory re- sponses. Gene expression levels were normalized to 18S. (E and F) Plasma concentrations of (E) CCL2 and (F) CCL5. (G) DHE fluorescence and Mac-2 immunofluores- cence in the aortic sinuses. Scale bars, 50 mm. Unpaired t test for (B), (E), and (G), and Mann- Whitney U test for (F). Statistical differences in gene expression in (C) and (D) were tested using unpaired t test or Mann-Whitney U test, depending on normality tests. *P < 0.05, **P <0.01, ***P <0.001 versus mice injected with AAV-GFP. Data are presented as mean ± SEM. All points and p values are shown.

Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Primer sequences for Agxt genotyping:, forward: 50-ACACCTCCACTGTCCTG TCC-30 Integrated DNA Technologies N/A Primer sequences for Agxt genotyping:, reverse: 50-GGTCAGATCTGCCTGCTA CC-30 Integrated DNA Technologies N/A Primer for Sanger sequencing:, 50-GCAGAGCTAGCTGGGAAATG-30 Integrated DNA Technologies N/A Primer sequences for Apoe genotyping:, forward: 50-GCCTAGCCGAGGGAGAG CCG-30 Integrated DNA Technologies N/A Primer sequences for Apoe genotyping:, reverse 1: 50-TGTGACTTGGGAGCTC TGCAGC-30 Integrated DNA Technologies N/A Primer sequences for Apoe genotyping:, reverse 2: 50-GCCGCCCCGACTGC ATCT-30 Integrated DNA Technologies N/A Primer sequences for qPCR, see Table S2 Integrated DNA Technologies N/A Recombinant DNA pAdDeltaF6 Jiandie Lin’s lab, University of Michigan; Guo et al., 2017 N/A pAAV2/8 Jiandie Lin’s lab, University of Michigan; Guo et al., 2017 N/A pAAV-TBG-GFP Jiandie Lin’s lab, University of Michigan; Guo et al., 2017 N/A pAAV-TBG-MCS Jiandie Lin’s lab, University of Michigan; Guo et al., 2017 N/A AGXT Human Tagged ORF Clone Origene CAT# RG212899 Software and algorithms ImageJ National Institutes of Health RRID:SCR_003070 https://imagej.nih.gov/ ij/download.html GraphPad Prism GraphPad Software Version 8.0 RRID:SCR_002798 https:// www.graphpad.com/scientific-software/ prism/ R statistics Hosted by Vienna, University of Economics and Business http://www.R-project.org/ RStudio RStudio Team (2020). https://www.rstudio.com/ RRID:SCR_000432 IBM SPSS software IBM Corporation Version 23.0; RRID:SCR_019096 https:// www.ibm.com/analytics/ spss-statistics-software FastQC Babraham Bioinformatics Version 0.11.8 RRID:SCR_014583 https:// www.bioinformatics.babraham.ac.uk/ projects/fastqc/ Trimmomatic Bolger et al., 2014 Version 0.35 RRID:SCR_011848 http:// www.usadellab.org/cms/? page=trimmomatic HISAT2 Kim et al., 2015 Version 2.1.0.13; RRID:SCR_015530 http:// daehwankimlab.github.io/hisat2/ download/ HTSeq Anders et al., 2015 Version 0.6.0; RRID:SCR_005514 https:// htseq.readthedocs.io/en/master/install. html (Continued on next page) Cell Reports 36, 109420, July 27, 2021 e3 Article ll OPEN ACCESS

Techniques: Over Expression, Injection, Western Blot, Clinical Proteomics, Gene Expression, MANN-WHITNEY

Figure 7. AAV-mediated overexpression of AGXT reduces atherosclerosis independent of circulating cholesterol AAV-AGXT or AAV-GFP was injected into male Apoe/ mice and the mice were fed a WD for 12 weeks (n = 8). (A) Plasma TC. (B) Atherosclerosis in the aortic tree. (C–E) H&E staining (D), ORO staining (D), and Mac- 2 immunohistochemistry (E) of aortic sinus. Scale bars, 200 mm. (F) Proposed model of dysregulated oxalate metabolism as a driver and therapeutic target in atherosclerosis. Unpaired t test for (A)–(E). Data are presented as mean ± SEM. All points and p values are shown.

Journal: Cell reports

Article Title: Dysregulated oxalate metabolism is a driver and therapeutic target in atherosclerosis.

doi: 10.1016/j.celrep.2021.109420

Figure Lengend Snippet: Figure 7. AAV-mediated overexpression of AGXT reduces atherosclerosis independent of circulating cholesterol AAV-AGXT or AAV-GFP was injected into male Apoe/ mice and the mice were fed a WD for 12 weeks (n = 8). (A) Plasma TC. (B) Atherosclerosis in the aortic tree. (C–E) H&E staining (D), ORO staining (D), and Mac- 2 immunohistochemistry (E) of aortic sinus. Scale bars, 200 mm. (F) Proposed model of dysregulated oxalate metabolism as a driver and therapeutic target in atherosclerosis. Unpaired t test for (A)–(E). Data are presented as mean ± SEM. All points and p values are shown.

Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Primer sequences for Agxt genotyping:, forward: 50-ACACCTCCACTGTCCTG TCC-30 Integrated DNA Technologies N/A Primer sequences for Agxt genotyping:, reverse: 50-GGTCAGATCTGCCTGCTA CC-30 Integrated DNA Technologies N/A Primer for Sanger sequencing:, 50-GCAGAGCTAGCTGGGAAATG-30 Integrated DNA Technologies N/A Primer sequences for Apoe genotyping:, forward: 50-GCCTAGCCGAGGGAGAG CCG-30 Integrated DNA Technologies N/A Primer sequences for Apoe genotyping:, reverse 1: 50-TGTGACTTGGGAGCTC TGCAGC-30 Integrated DNA Technologies N/A Primer sequences for Apoe genotyping:, reverse 2: 50-GCCGCCCCGACTGC ATCT-30 Integrated DNA Technologies N/A Primer sequences for qPCR, see Table S2 Integrated DNA Technologies N/A Recombinant DNA pAdDeltaF6 Jiandie Lin’s lab, University of Michigan; Guo et al., 2017 N/A pAAV2/8 Jiandie Lin’s lab, University of Michigan; Guo et al., 2017 N/A pAAV-TBG-GFP Jiandie Lin’s lab, University of Michigan; Guo et al., 2017 N/A pAAV-TBG-MCS Jiandie Lin’s lab, University of Michigan; Guo et al., 2017 N/A AGXT Human Tagged ORF Clone Origene CAT# RG212899 Software and algorithms ImageJ National Institutes of Health RRID:SCR_003070 https://imagej.nih.gov/ ij/download.html GraphPad Prism GraphPad Software Version 8.0 RRID:SCR_002798 https:// www.graphpad.com/scientific-software/ prism/ R statistics Hosted by Vienna, University of Economics and Business http://www.R-project.org/ RStudio RStudio Team (2020). https://www.rstudio.com/ RRID:SCR_000432 IBM SPSS software IBM Corporation Version 23.0; RRID:SCR_019096 https:// www.ibm.com/analytics/ spss-statistics-software FastQC Babraham Bioinformatics Version 0.11.8 RRID:SCR_014583 https:// www.bioinformatics.babraham.ac.uk/ projects/fastqc/ Trimmomatic Bolger et al., 2014 Version 0.35 RRID:SCR_011848 http:// www.usadellab.org/cms/? page=trimmomatic HISAT2 Kim et al., 2015 Version 2.1.0.13; RRID:SCR_015530 http:// daehwankimlab.github.io/hisat2/ download/ HTSeq Anders et al., 2015 Version 0.6.0; RRID:SCR_005514 https:// htseq.readthedocs.io/en/master/install. html (Continued on next page) Cell Reports 36, 109420, July 27, 2021 e3 Article ll OPEN ACCESS

Techniques: Over Expression, Injection, Clinical Proteomics, Staining, Immunohistochemistry

Figure 1. IL-21/IL-21R signaling reduces murine L-GMPs in vivo (A) Il21R mRNA expression (qRT-PCR) in FACS-sorted L-GMPs from the BM of BL/6 AML mice 35 days after leukemia transplantation (n = 10 mice). Red bar indicates the mean. Each dot represents the mean of two technical replicates. (B) IL-21 levels in BM samples from AML mice (n = 11 mice) and naive controls (n = 5 mice). Each dot represents the mean of two technical replicates. Data are shown as mean ± SEM. Statistics: Student’s t test. (C) Fold change colony-forming units from FACS-sorted L-GMPs cultured in methylcellulose for 7 days in the presence or absence of 300 pg/mL rm-IL-21 (n = 6 mice/group). Each dot represents the mean of two technical replicates. Pooled data from two independent experiments are shown and displayed as mean ± SD. Statistics: Student’s t test. (D) Experimental setup: 5 3 104 MLL-AF9-GFP-transduced LSKs from the BM of BL/6 donors were injected intravenously into non-irradiated BL/6 and Il21/

Journal: Cell reports. Medicine

Article Title: IL-21/IL-21R signaling renders acute myeloid leukemia stem cells more susceptible to cytarabine treatment and CAR T cell therapy.

doi: 10.1016/j.xcrm.2024.101826

Figure Lengend Snippet: Figure 1. IL-21/IL-21R signaling reduces murine L-GMPs in vivo (A) Il21R mRNA expression (qRT-PCR) in FACS-sorted L-GMPs from the BM of BL/6 AML mice 35 days after leukemia transplantation (n = 10 mice). Red bar indicates the mean. Each dot represents the mean of two technical replicates. (B) IL-21 levels in BM samples from AML mice (n = 11 mice) and naive controls (n = 5 mice). Each dot represents the mean of two technical replicates. Data are shown as mean ± SEM. Statistics: Student’s t test. (C) Fold change colony-forming units from FACS-sorted L-GMPs cultured in methylcellulose for 7 days in the presence or absence of 300 pg/mL rm-IL-21 (n = 6 mice/group). Each dot represents the mean of two technical replicates. Pooled data from two independent experiments are shown and displayed as mean ± SD. Statistics: Student’s t test. (D) Experimental setup: 5 3 104 MLL-AF9-GFP-transduced LSKs from the BM of BL/6 donors were injected intravenously into non-irradiated BL/6 and Il21/

Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Experimental models: Organisms/strains Mouse: C57BL/6J Charles River MGI: 3028467 Il21R / (B6.129-Il21rtm1Kopf/J) MGI: 5435248 Il21 / (B6.129S-Il21tm1Lex/Mmucd) MGI: 4843320 Il21mCherry(B6.Cg-Tg(Il21-mCherry)1Wjl/Mmnc) MGI: 5478531 Recombinant DNA Plasmid pMDLg/pRRE Addgene #12259 Plasmid pRSV-Rev Addgene #12253 Plasmid pCMV-VSV-g Addgene #8454 CD19-CAR plasmid Addgene #200671 Software and algorithms FlowJoTM software v.10.6 TreeStar N/A GraphPad Prism software v9.0 GraphPad N/A ELDA Software Hu et al.21 http://bioinf.wehi.edu.au/software/elda/ fastqc v. 0.11.9 https://www.bioinformatics.babraham. ac.uk/projects/fastqc/ RSeQC v. 4.0.0 Wang et al.72 N/A HiSat2 v. 2.2.1 Kim et al.73 N/A FeatureCounts v. 2.0.1 Liao et al.74 N/A DESeq2 v.1.32.0 Love et al.75 N/A TopGo v. 2.44.0 https://bioconductor.org/packages/ release/bioc/html/topGO.html ClusterProfiler v. 4.0.2 Yu et al.76 N/A R 4.1.0 https://www.R-project.org/ INSPIRE software https://www.merckmillipore.com/ IDEAS software https://www.merckmillipore.com/ Kaluza analysis software Beckman Coulter Life Sciences N/A Other HALLMARK OXIDATIVE PHOSPHORYLATION gsea-msigdb.org Somervaille_UP Somervaille et al.26 N/A Somervaille_DOWN Somervaille et al.26 N/A STEMNESS_UP gsea-msigdb.org STEMNESS_ DOWN gsea-msigdb.org GAL_LEUKEMIC STEM CELL_DOWN gsea-msigdb.org BROWN_MYELOID_CELL_DEVELOPMENT_DN gsea-msigdb.org BROWN_MYELOID_CELL_DEVELOPMENT_UP gsea-msigdb.org HALLMARK_TNFA_SIGNALING_VIA_NFKB gsea-msigdb.org HSC Differentiation and Lineage-specific markers pathcards.genecards.org Wnt Signaling (beta-Catenin) pathcards.genecards.org MAPK-Erk Signaling pathcards.genecards.org POSITIVE REGULATION OF CELL CYCLE gsea-msigdb.org Respiratory electron transport_ATP synthesis pathcards.genecards.org FRIDMAN_SENESCENCE_UP gsea-msigdb.org TANG_SENESCENCE_TP53_TARGETS_DN gsea-msigdb.org

Techniques: In Vivo, Expressing, Quantitative RT-PCR, Transplantation Assay, Cell Culture, Injection, Irradiation

Figure 3. CD4+ T cell-derived IL-21 reduces murine LSCs in vivo (A and B) MLL-AF9 AML was induced in Il21mcherry reporter mice. Frequencies of mcherry-Il21+ CD4+ T cells (A) and CD4+ mcherry-Il21+ cells (B) were determined by flow cytometry 16 and 33 days after leukemia transplantation, in BM of AML and naive Il21mcherry mice (n = 4 mice/group). (C) Experimental setup: 5 3 106 CD4+ T cells were FACS-sorted from the spleens of CD45.1 mice and injected intravenously into two out of four experimental groups (non-irradiated BL/6 and Il21/ recipients) one day prior to leukemia transplantation. One day after, 5 3 104 MLL-AF9-GFP-transduced LSKs from the BM of BL/6 donors were injected intravenously into all four experimental groups (BL/6 AML, Il21/ AML, BL/6 AML + CD4+, and Il21/ AML + CD4+, respectively). Mice were sacrificed 35 days after leukemia transplantation and BM and spleen were analyzed (n = 4 mice/group). (D–G) Spleen size (D), number of MLL-AF9-GFP+ leukemic cells in BM (E) and in peripheral blood (F), and number of L-GMPs (G) in the BM of BL/6 AML, Il21/

Journal: Cell reports. Medicine

Article Title: IL-21/IL-21R signaling renders acute myeloid leukemia stem cells more susceptible to cytarabine treatment and CAR T cell therapy.

doi: 10.1016/j.xcrm.2024.101826

Figure Lengend Snippet: Figure 3. CD4+ T cell-derived IL-21 reduces murine LSCs in vivo (A and B) MLL-AF9 AML was induced in Il21mcherry reporter mice. Frequencies of mcherry-Il21+ CD4+ T cells (A) and CD4+ mcherry-Il21+ cells (B) were determined by flow cytometry 16 and 33 days after leukemia transplantation, in BM of AML and naive Il21mcherry mice (n = 4 mice/group). (C) Experimental setup: 5 3 106 CD4+ T cells were FACS-sorted from the spleens of CD45.1 mice and injected intravenously into two out of four experimental groups (non-irradiated BL/6 and Il21/ recipients) one day prior to leukemia transplantation. One day after, 5 3 104 MLL-AF9-GFP-transduced LSKs from the BM of BL/6 donors were injected intravenously into all four experimental groups (BL/6 AML, Il21/ AML, BL/6 AML + CD4+, and Il21/ AML + CD4+, respectively). Mice were sacrificed 35 days after leukemia transplantation and BM and spleen were analyzed (n = 4 mice/group). (D–G) Spleen size (D), number of MLL-AF9-GFP+ leukemic cells in BM (E) and in peripheral blood (F), and number of L-GMPs (G) in the BM of BL/6 AML, Il21/

Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Experimental models: Organisms/strains Mouse: C57BL/6J Charles River MGI: 3028467 Il21R / (B6.129-Il21rtm1Kopf/J) MGI: 5435248 Il21 / (B6.129S-Il21tm1Lex/Mmucd) MGI: 4843320 Il21mCherry(B6.Cg-Tg(Il21-mCherry)1Wjl/Mmnc) MGI: 5478531 Recombinant DNA Plasmid pMDLg/pRRE Addgene #12259 Plasmid pRSV-Rev Addgene #12253 Plasmid pCMV-VSV-g Addgene #8454 CD19-CAR plasmid Addgene #200671 Software and algorithms FlowJoTM software v.10.6 TreeStar N/A GraphPad Prism software v9.0 GraphPad N/A ELDA Software Hu et al.21 http://bioinf.wehi.edu.au/software/elda/ fastqc v. 0.11.9 https://www.bioinformatics.babraham. ac.uk/projects/fastqc/ RSeQC v. 4.0.0 Wang et al.72 N/A HiSat2 v. 2.2.1 Kim et al.73 N/A FeatureCounts v. 2.0.1 Liao et al.74 N/A DESeq2 v.1.32.0 Love et al.75 N/A TopGo v. 2.44.0 https://bioconductor.org/packages/ release/bioc/html/topGO.html ClusterProfiler v. 4.0.2 Yu et al.76 N/A R 4.1.0 https://www.R-project.org/ INSPIRE software https://www.merckmillipore.com/ IDEAS software https://www.merckmillipore.com/ Kaluza analysis software Beckman Coulter Life Sciences N/A Other HALLMARK OXIDATIVE PHOSPHORYLATION gsea-msigdb.org Somervaille_UP Somervaille et al.26 N/A Somervaille_DOWN Somervaille et al.26 N/A STEMNESS_UP gsea-msigdb.org STEMNESS_ DOWN gsea-msigdb.org GAL_LEUKEMIC STEM CELL_DOWN gsea-msigdb.org BROWN_MYELOID_CELL_DEVELOPMENT_DN gsea-msigdb.org BROWN_MYELOID_CELL_DEVELOPMENT_UP gsea-msigdb.org HALLMARK_TNFA_SIGNALING_VIA_NFKB gsea-msigdb.org HSC Differentiation and Lineage-specific markers pathcards.genecards.org Wnt Signaling (beta-Catenin) pathcards.genecards.org MAPK-Erk Signaling pathcards.genecards.org POSITIVE REGULATION OF CELL CYCLE gsea-msigdb.org Respiratory electron transport_ATP synthesis pathcards.genecards.org FRIDMAN_SENESCENCE_UP gsea-msigdb.org TANG_SENESCENCE_TP53_TARGETS_DN gsea-msigdb.org

Techniques: Derivative Assay, In Vivo, Cytometry, Transplantation Assay, Injection, Irradiation

Figure 4. IL-21 is an independent positive prognostic marker for OS in AML (A) IL-21 levels in serum samples from newly diagnosed patients with AML (n = 193) and age-matched healthy controls (n = 10). Each dot represents the mean of two technical replicates. Red bars indicate the mean. Statistics: Mann-Whitney test. (B) Kaplan-Meier survival curves of the AML patients’ cohort (n = 193) divided into two groups at the IL-21 serum levels (sIL-21) threshold of 29 pg/mL. Statistics: log rank test. (C) Correlation of patients’ age with sIL-21 levels. Statistics: Pearson r test. (D) sIL-21 of patients in the different risk groups. Red bars indicate the mean. Statistics: one-way ANOVA. (E) Multivariate analysis for sIL-21 adjusted for BM infiltration, leukocyte counts, percentage of peripheral blood blasts, age, and risk group. Statistics: multiple Cox regression. (F and G) Two publicly available microarray datasets were analyzed for IL21 mRNA expression levels and their association with prognosis. Statistics: log rank test. (F) Valk dataset, accession number GSE6891, sIL-21 threshold 6.91 RPKM. (G) Metzeler dataset, accession number GSE12417, sIL-21 threshold 7.54 RPKM. (H–K) Enrichment plots depicting significantly enriched gene sets in patients with high levels of IL-21. HSC differentiation and lineage-specific markers (H), positive regulation of stem cell proliferation (I), Fridman-senescence-DN (J), Stemness-UP (K). Gene sets were derived from the Metzeler dataset, accession number GSE12417. Normalized enrichment score (NES) and p value are indicated for each plot. *p < 0.05; **p < 0.01; ***p < 0.001; ****p < 0.0001. Abbreviations: OS, overall survival; yr, years; fav., favorable; int., intermediate; adv., adverse; HR, hazard ratio; CI, confidence interval; PB, peripheral blood; RPKM, reads per kilobase per million mapped reads; NES, normalized enrichment score. See also Figure S5.

Journal: Cell reports. Medicine

Article Title: IL-21/IL-21R signaling renders acute myeloid leukemia stem cells more susceptible to cytarabine treatment and CAR T cell therapy.

doi: 10.1016/j.xcrm.2024.101826

Figure Lengend Snippet: Figure 4. IL-21 is an independent positive prognostic marker for OS in AML (A) IL-21 levels in serum samples from newly diagnosed patients with AML (n = 193) and age-matched healthy controls (n = 10). Each dot represents the mean of two technical replicates. Red bars indicate the mean. Statistics: Mann-Whitney test. (B) Kaplan-Meier survival curves of the AML patients’ cohort (n = 193) divided into two groups at the IL-21 serum levels (sIL-21) threshold of 29 pg/mL. Statistics: log rank test. (C) Correlation of patients’ age with sIL-21 levels. Statistics: Pearson r test. (D) sIL-21 of patients in the different risk groups. Red bars indicate the mean. Statistics: one-way ANOVA. (E) Multivariate analysis for sIL-21 adjusted for BM infiltration, leukocyte counts, percentage of peripheral blood blasts, age, and risk group. Statistics: multiple Cox regression. (F and G) Two publicly available microarray datasets were analyzed for IL21 mRNA expression levels and their association with prognosis. Statistics: log rank test. (F) Valk dataset, accession number GSE6891, sIL-21 threshold 6.91 RPKM. (G) Metzeler dataset, accession number GSE12417, sIL-21 threshold 7.54 RPKM. (H–K) Enrichment plots depicting significantly enriched gene sets in patients with high levels of IL-21. HSC differentiation and lineage-specific markers (H), positive regulation of stem cell proliferation (I), Fridman-senescence-DN (J), Stemness-UP (K). Gene sets were derived from the Metzeler dataset, accession number GSE12417. Normalized enrichment score (NES) and p value are indicated for each plot. *p < 0.05; **p < 0.01; ***p < 0.001; ****p < 0.0001. Abbreviations: OS, overall survival; yr, years; fav., favorable; int., intermediate; adv., adverse; HR, hazard ratio; CI, confidence interval; PB, peripheral blood; RPKM, reads per kilobase per million mapped reads; NES, normalized enrichment score. See also Figure S5.

Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Experimental models: Organisms/strains Mouse: C57BL/6J Charles River MGI: 3028467 Il21R / (B6.129-Il21rtm1Kopf/J) MGI: 5435248 Il21 / (B6.129S-Il21tm1Lex/Mmucd) MGI: 4843320 Il21mCherry(B6.Cg-Tg(Il21-mCherry)1Wjl/Mmnc) MGI: 5478531 Recombinant DNA Plasmid pMDLg/pRRE Addgene #12259 Plasmid pRSV-Rev Addgene #12253 Plasmid pCMV-VSV-g Addgene #8454 CD19-CAR plasmid Addgene #200671 Software and algorithms FlowJoTM software v.10.6 TreeStar N/A GraphPad Prism software v9.0 GraphPad N/A ELDA Software Hu et al.21 http://bioinf.wehi.edu.au/software/elda/ fastqc v. 0.11.9 https://www.bioinformatics.babraham. ac.uk/projects/fastqc/ RSeQC v. 4.0.0 Wang et al.72 N/A HiSat2 v. 2.2.1 Kim et al.73 N/A FeatureCounts v. 2.0.1 Liao et al.74 N/A DESeq2 v.1.32.0 Love et al.75 N/A TopGo v. 2.44.0 https://bioconductor.org/packages/ release/bioc/html/topGO.html ClusterProfiler v. 4.0.2 Yu et al.76 N/A R 4.1.0 https://www.R-project.org/ INSPIRE software https://www.merckmillipore.com/ IDEAS software https://www.merckmillipore.com/ Kaluza analysis software Beckman Coulter Life Sciences N/A Other HALLMARK OXIDATIVE PHOSPHORYLATION gsea-msigdb.org Somervaille_UP Somervaille et al.26 N/A Somervaille_DOWN Somervaille et al.26 N/A STEMNESS_UP gsea-msigdb.org STEMNESS_ DOWN gsea-msigdb.org GAL_LEUKEMIC STEM CELL_DOWN gsea-msigdb.org BROWN_MYELOID_CELL_DEVELOPMENT_DN gsea-msigdb.org BROWN_MYELOID_CELL_DEVELOPMENT_UP gsea-msigdb.org HALLMARK_TNFA_SIGNALING_VIA_NFKB gsea-msigdb.org HSC Differentiation and Lineage-specific markers pathcards.genecards.org Wnt Signaling (beta-Catenin) pathcards.genecards.org MAPK-Erk Signaling pathcards.genecards.org POSITIVE REGULATION OF CELL CYCLE gsea-msigdb.org Respiratory electron transport_ATP synthesis pathcards.genecards.org FRIDMAN_SENESCENCE_UP gsea-msigdb.org TANG_SENESCENCE_TP53_TARGETS_DN gsea-msigdb.org

Techniques: Marker, MANN-WHITNEY, Microarray, Expressing, Derivative Assay

Figure 5. IL-21 is produced by CD4+ T cells and reduces cell growth and colony-forming capacity of AML stem and progenitor cells in vitro (A) IL21 mRNA expression (qRT-PCR) in FACS-sorted CD4+ T cells (n = 32), CD8+ T cells (n = 27), and CD45dimSSClolinCD34+ LSPCs (n = 13) from newly diagnosed patients with AML. Red bars indicate the mean. Statistics: one-way ANOVA.

Journal: Cell reports. Medicine

Article Title: IL-21/IL-21R signaling renders acute myeloid leukemia stem cells more susceptible to cytarabine treatment and CAR T cell therapy.

doi: 10.1016/j.xcrm.2024.101826

Figure Lengend Snippet: Figure 5. IL-21 is produced by CD4+ T cells and reduces cell growth and colony-forming capacity of AML stem and progenitor cells in vitro (A) IL21 mRNA expression (qRT-PCR) in FACS-sorted CD4+ T cells (n = 32), CD8+ T cells (n = 27), and CD45dimSSClolinCD34+ LSPCs (n = 13) from newly diagnosed patients with AML. Red bars indicate the mean. Statistics: one-way ANOVA.

Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Experimental models: Organisms/strains Mouse: C57BL/6J Charles River MGI: 3028467 Il21R / (B6.129-Il21rtm1Kopf/J) MGI: 5435248 Il21 / (B6.129S-Il21tm1Lex/Mmucd) MGI: 4843320 Il21mCherry(B6.Cg-Tg(Il21-mCherry)1Wjl/Mmnc) MGI: 5478531 Recombinant DNA Plasmid pMDLg/pRRE Addgene #12259 Plasmid pRSV-Rev Addgene #12253 Plasmid pCMV-VSV-g Addgene #8454 CD19-CAR plasmid Addgene #200671 Software and algorithms FlowJoTM software v.10.6 TreeStar N/A GraphPad Prism software v9.0 GraphPad N/A ELDA Software Hu et al.21 http://bioinf.wehi.edu.au/software/elda/ fastqc v. 0.11.9 https://www.bioinformatics.babraham. ac.uk/projects/fastqc/ RSeQC v. 4.0.0 Wang et al.72 N/A HiSat2 v. 2.2.1 Kim et al.73 N/A FeatureCounts v. 2.0.1 Liao et al.74 N/A DESeq2 v.1.32.0 Love et al.75 N/A TopGo v. 2.44.0 https://bioconductor.org/packages/ release/bioc/html/topGO.html ClusterProfiler v. 4.0.2 Yu et al.76 N/A R 4.1.0 https://www.R-project.org/ INSPIRE software https://www.merckmillipore.com/ IDEAS software https://www.merckmillipore.com/ Kaluza analysis software Beckman Coulter Life Sciences N/A Other HALLMARK OXIDATIVE PHOSPHORYLATION gsea-msigdb.org Somervaille_UP Somervaille et al.26 N/A Somervaille_DOWN Somervaille et al.26 N/A STEMNESS_UP gsea-msigdb.org STEMNESS_ DOWN gsea-msigdb.org GAL_LEUKEMIC STEM CELL_DOWN gsea-msigdb.org BROWN_MYELOID_CELL_DEVELOPMENT_DN gsea-msigdb.org BROWN_MYELOID_CELL_DEVELOPMENT_UP gsea-msigdb.org HALLMARK_TNFA_SIGNALING_VIA_NFKB gsea-msigdb.org HSC Differentiation and Lineage-specific markers pathcards.genecards.org Wnt Signaling (beta-Catenin) pathcards.genecards.org MAPK-Erk Signaling pathcards.genecards.org POSITIVE REGULATION OF CELL CYCLE gsea-msigdb.org Respiratory electron transport_ATP synthesis pathcards.genecards.org FRIDMAN_SENESCENCE_UP gsea-msigdb.org TANG_SENESCENCE_TP53_TARGETS_DN gsea-msigdb.org

Techniques: Produced, In Vitro, Expressing, Quantitative RT-PCR

Figure 6. IL-21/IL-21R signaling promotes the sensitivity of AML LSCs to cytarabine treatment (A) Kaplan-Meier survival curves of patients with AML (n = 110) who received intensive chemotherapy (‘‘7 + 3’’ regimen) as a first-line treatment divided into two groups based on complete remission (CR) achievement. Statistics: log rank test. (B) Kaplan-Meier survival curves of the AML patient cohort that received intensive chemotherapy (n = 110) divided into three groups at the sIL-21 threshold of 29 and 72 pg/mL. Statistics: log rank test. (C) sIL-21 levels in patients who achieved CR (n = 80) versus patients who did not achieve CR (n = 30). Red bars indicate the mean. Statistics: Mann-Whitney test. (D) CR rates in patients with high, intermediate, and low sIL-21 levels. Statistics: chi-square test. (E) LSPCs colonies were enumerated after two weeks of culture in methylcellulose in the presence or absence of 100 pg/mL rh-IL-21, 1 nM cytarabine (AraC), or combination. Each dot represents the mean of three technical replicates. Different colors indicate different patients (n = 11). Data are shown as mean ± SD. Statistics: one-way ANOVA followed by Tukey’s multiple comparisons test. (F and G) LSPCs colonies were enumerated after two weeks of culture in methylcellulose in the presence or absence of 100 pg/mL rh-IL-21, anti-CD70 CAR T cells (1:5 E:T ratio), or combination. Two patients are shown in each graph (each dot represents a technical replicate). Statistics: one-way ANOVA followed by Tukey’s multiple comparisons test. *p < 0.05; **p < 0.01; ***p < 0.001; ****p < 0.0001. Abbreviations: ChTh, chemotherapy; OS, overall survival; CR, complete remission; CFU, colony-forming units, AraC, cytosine arabinoside (or cytarabine). See also Figure S7.

Journal: Cell reports. Medicine

Article Title: IL-21/IL-21R signaling renders acute myeloid leukemia stem cells more susceptible to cytarabine treatment and CAR T cell therapy.

doi: 10.1016/j.xcrm.2024.101826

Figure Lengend Snippet: Figure 6. IL-21/IL-21R signaling promotes the sensitivity of AML LSCs to cytarabine treatment (A) Kaplan-Meier survival curves of patients with AML (n = 110) who received intensive chemotherapy (‘‘7 + 3’’ regimen) as a first-line treatment divided into two groups based on complete remission (CR) achievement. Statistics: log rank test. (B) Kaplan-Meier survival curves of the AML patient cohort that received intensive chemotherapy (n = 110) divided into three groups at the sIL-21 threshold of 29 and 72 pg/mL. Statistics: log rank test. (C) sIL-21 levels in patients who achieved CR (n = 80) versus patients who did not achieve CR (n = 30). Red bars indicate the mean. Statistics: Mann-Whitney test. (D) CR rates in patients with high, intermediate, and low sIL-21 levels. Statistics: chi-square test. (E) LSPCs colonies were enumerated after two weeks of culture in methylcellulose in the presence or absence of 100 pg/mL rh-IL-21, 1 nM cytarabine (AraC), or combination. Each dot represents the mean of three technical replicates. Different colors indicate different patients (n = 11). Data are shown as mean ± SD. Statistics: one-way ANOVA followed by Tukey’s multiple comparisons test. (F and G) LSPCs colonies were enumerated after two weeks of culture in methylcellulose in the presence or absence of 100 pg/mL rh-IL-21, anti-CD70 CAR T cells (1:5 E:T ratio), or combination. Two patients are shown in each graph (each dot represents a technical replicate). Statistics: one-way ANOVA followed by Tukey’s multiple comparisons test. *p < 0.05; **p < 0.01; ***p < 0.001; ****p < 0.0001. Abbreviations: ChTh, chemotherapy; OS, overall survival; CR, complete remission; CFU, colony-forming units, AraC, cytosine arabinoside (or cytarabine). See also Figure S7.

Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Experimental models: Organisms/strains Mouse: C57BL/6J Charles River MGI: 3028467 Il21R / (B6.129-Il21rtm1Kopf/J) MGI: 5435248 Il21 / (B6.129S-Il21tm1Lex/Mmucd) MGI: 4843320 Il21mCherry(B6.Cg-Tg(Il21-mCherry)1Wjl/Mmnc) MGI: 5478531 Recombinant DNA Plasmid pMDLg/pRRE Addgene #12259 Plasmid pRSV-Rev Addgene #12253 Plasmid pCMV-VSV-g Addgene #8454 CD19-CAR plasmid Addgene #200671 Software and algorithms FlowJoTM software v.10.6 TreeStar N/A GraphPad Prism software v9.0 GraphPad N/A ELDA Software Hu et al.21 http://bioinf.wehi.edu.au/software/elda/ fastqc v. 0.11.9 https://www.bioinformatics.babraham. ac.uk/projects/fastqc/ RSeQC v. 4.0.0 Wang et al.72 N/A HiSat2 v. 2.2.1 Kim et al.73 N/A FeatureCounts v. 2.0.1 Liao et al.74 N/A DESeq2 v.1.32.0 Love et al.75 N/A TopGo v. 2.44.0 https://bioconductor.org/packages/ release/bioc/html/topGO.html ClusterProfiler v. 4.0.2 Yu et al.76 N/A R 4.1.0 https://www.R-project.org/ INSPIRE software https://www.merckmillipore.com/ IDEAS software https://www.merckmillipore.com/ Kaluza analysis software Beckman Coulter Life Sciences N/A Other HALLMARK OXIDATIVE PHOSPHORYLATION gsea-msigdb.org Somervaille_UP Somervaille et al.26 N/A Somervaille_DOWN Somervaille et al.26 N/A STEMNESS_UP gsea-msigdb.org STEMNESS_ DOWN gsea-msigdb.org GAL_LEUKEMIC STEM CELL_DOWN gsea-msigdb.org BROWN_MYELOID_CELL_DEVELOPMENT_DN gsea-msigdb.org BROWN_MYELOID_CELL_DEVELOPMENT_UP gsea-msigdb.org HALLMARK_TNFA_SIGNALING_VIA_NFKB gsea-msigdb.org HSC Differentiation and Lineage-specific markers pathcards.genecards.org Wnt Signaling (beta-Catenin) pathcards.genecards.org MAPK-Erk Signaling pathcards.genecards.org POSITIVE REGULATION OF CELL CYCLE gsea-msigdb.org Respiratory electron transport_ATP synthesis pathcards.genecards.org FRIDMAN_SENESCENCE_UP gsea-msigdb.org TANG_SENESCENCE_TP53_TARGETS_DN gsea-msigdb.org

Techniques: MANN-WHITNEY

Figure 3. The CTL synapse exhibits negative membrane curvatures at regions cleared of F-actin (A) Timelapse confocal images of live CTL F-actin distribution at the synapse during cognate tumor interaction in the DPA assay. Top: maximum intensity projection. Middle/bottom: SynapseMech analysis of CTL F-actin distribution and membrane curvature within the synapse. Displayed F-actin signal restricted to synapse region and intensity-thresholded. Scale bar, 5 mm. (B) Dynamics of area cleared from F-actin (gray shaded region) and average mean curvature of complete (dark gray) and F-actin-cleared (brown) synapse area for data shown in (A). (C) Time-averaged mean curvature of complete, F-actin-cleared, and F-actin-rich (outside of cleared space) synapse area. Boxplot shows median, IQR, and range for n = 6 cells. p values by matched one-way ANOVA test with prior Kolmogorov-Smirnov test for normality. (D) Surface tension of resting CTLs, tumor cells and tumor cytoplasts. Boxplots show median, IQR and range for n = 24 (CTL), n = 36 (tumor cells), and n = 19 (tumor cytoplasts). p values by Mann-Whitney test. (E) Timelapse confocal images of a live CTL interacting with a cognate tumor cytoplast in the DPA assay. t = 0 s: time of contact. Scale bar, 10 mm. (F) Main: dynamics of average mean synapse curvature of CTL interacting with cognate tumor cell or tumor cytoplast. Data are mean ± SEM (shaded area) for n = 7 (tumor cell) and n = 8 (tumor cytoplast). Inset: time-averaged mean synapse curvature of CTLs interacting with cognate tumor cells or tumor cytoplasts. p value by Mann-Whitney test.

Journal: Developmental cell

Article Title: T cell cytoskeletal forces shape synapse topography for targeted lysis via membrane curvature bias of perforin.

doi: 10.1016/j.devcel.2022.08.012

Figure Lengend Snippet: Figure 3. The CTL synapse exhibits negative membrane curvatures at regions cleared of F-actin (A) Timelapse confocal images of live CTL F-actin distribution at the synapse during cognate tumor interaction in the DPA assay. Top: maximum intensity projection. Middle/bottom: SynapseMech analysis of CTL F-actin distribution and membrane curvature within the synapse. Displayed F-actin signal restricted to synapse region and intensity-thresholded. Scale bar, 5 mm. (B) Dynamics of area cleared from F-actin (gray shaded region) and average mean curvature of complete (dark gray) and F-actin-cleared (brown) synapse area for data shown in (A). (C) Time-averaged mean curvature of complete, F-actin-cleared, and F-actin-rich (outside of cleared space) synapse area. Boxplot shows median, IQR, and range for n = 6 cells. p values by matched one-way ANOVA test with prior Kolmogorov-Smirnov test for normality. (D) Surface tension of resting CTLs, tumor cells and tumor cytoplasts. Boxplots show median, IQR and range for n = 24 (CTL), n = 36 (tumor cells), and n = 19 (tumor cytoplasts). p values by Mann-Whitney test. (E) Timelapse confocal images of a live CTL interacting with a cognate tumor cytoplast in the DPA assay. t = 0 s: time of contact. Scale bar, 10 mm. (F) Main: dynamics of average mean synapse curvature of CTL interacting with cognate tumor cell or tumor cytoplast. Data are mean ± SEM (shaded area) for n = 7 (tumor cell) and n = 8 (tumor cytoplast). Inset: time-averaged mean synapse curvature of CTLs interacting with cognate tumor cells or tumor cytoplasts. p value by Mann-Whitney test.

Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Experimental models: Cell lines EL4 mouse lymphoma cells ATCC Cat. #: TIB-39 WM266-4 (Elkin1 KO) human melanoma cells Patkunarajah et al. (2020) N/A Experimental models: Organisms/strains Mouse: C57BL/6 OT-1 Gift from E. Deenick (Garvan Institute of Medical Research) N/A Mouse: C57BL/6 OT-1 x Lifeact-EGFP Galeano Niño et al. (2020) N/A Mouse: C57BL/6 OT-1 x TdTomato Galeano Niño et al. (2020); Muzumdar et al. (2007) N/A Human: primary CTLs transduced with 1G4 TCR Gift from J. Goyette (University of New South Wales) N/A Recombinant DNA Myosin IIA-GFP plasmid Addgene RRID: Addgene_38297 Software and algorithms SynapseMech This paper [Github]: https://github.com/BiroLab/ SynapseMech; https://doi.org/10.5281/ zenodo.7014229 FIJI LimeSeg plugin Machado et al. (2019) https://imagej.net/plugins/limeseg Imaris v8.4.1 Bitplane https://imaris.oxinst.com Prism v8 Graphpad https://graphpad.com Matlab R2021a Mathworks https://mathworks.com Other AccuCount Blank Particles Spherotech Cat. #: ACBP-70-10 Sensoplate Microplate, 96 well, PS, F-bottom Greiner Cat. #: 655892 35 mm Dish | No. 1.5 Coverslip | 14 mm Glass Diameter | Uncoated Mattek Cat. #: P35G-1.5-14-C

Techniques: Membrane, MANN-WHITNEY