Journal: iScience

Article Title: In vivo reprogramming of NG2 glia improves bladder function after spinal cord injury

doi: 10.1016/j.isci.2026.114850

Figure Lengend Snippet: SOX2-mediated glial reprogramming in the adult mouse spinal cord (A) Experimental scheme. SOX2-expressing lentiviruses were injected into the adult spinal cord, and tissues were analyzed 4 weeks (wk) later using immunohistochemistry (IHC). (B) Quantification of SOX2-induced DCX + immature neurons (mean ± SEM, n = 3–4 mice per group). Notably, DCX + cells were nearly absent in the p75-2-only condition. (C) Confocal images of DCX + cells induced by either wild-type SOX2 (SOX2 WT ) or the phospho-mimetic mutant SOX2 (SOX2 S251E ). Scale bars, 20 μm (D) Experimental scheme for tracing the cellular origin of mutant SOX2-reprogrammed cells. Adult transgenic mice were treated with tamoxifen (Tam), followed by intraspinal viral injections, and analyzed 4 weeks later. (E) Confocal images and quantifications (mean ± SEM, n = 3 mice per group) showing that NG2 glia are the cellular origin of mutant SOX2-reprogrammed cells in the adult spinal cord. NG2 glia and astrocytes were lineage-traced using Pdgfra-CreER T2 ;R26R-tdT and Aldh1l1-CreER T2 ;R26R-tdT mice, respectively. Scale bar, 20 μm (F) Experimental scheme for a time-course analysis of mutant SOX2-mediated reprogramming. dpv, days post virus injection. (G) Quantification of mutant SOX2-induced ASCL1 + neural progenitors (mean ± SEM, n = 3 mice per group). (H) Confocal images showing induction of ASCL1 + neural progenitors by mutant SOX2. Scale bar, 20 μm (I) Experimental scheme for tracing the progeny of mutant SOX2-induced ASCL1 + progenitors in the adult spinal cord. wpv, weeks post virus injection. (J) Quantification of mutant SOX2-induced tdT + cells at 4 wpv (mean ± SEM, n = 5 mice). (K) Confocal images showing robust detection of DCX + tdT + cells in mutant SOX2-expressing spinal cord. Scale bar, 20 μm.

Article Snippet: Mouse: Pdgfra-CreER T2 , The Jackson Laboratory , JAX: 032770; RRID: IMSR_JAX: 032770.

Techniques: Expressing, Injection, Immunohistochemistry, Mutagenesis, Transgenic Assay, Virus

Journal: Circulation

Article Title: The Role of Venous Endothelial Cells in Developmental and Pathologic Angiogenesis

doi: 10.1161/CIRCULATIONAHA.121.054071

Figure Lengend Snippet: A. Schematic diagram showing a strategy for generation of Smad4iECKO;VECiDrereporter mice. Smad4 is deleted by inducible panEC specific Cre (Cdh5(PAC)CreERT2) while VECs is labeled with GFP using VEC-specific inducible Dre (Gm5127(BAC)DreERT2). B. Strategy for 4-OHT administration and analysis. C. Representative images showing the vascular phenotype (Isolectin B4 : red and GFP: green, respectively) of Smad4iECKO;VECiDrereporter (right) mice and WT littertmate(left, VECiDrereporter). White arrowheads indicate AVMs. D. Representative images showing the P7 retinal vasculature (Isolectin B4 : red, GFP: green) of Smad4iECKO;VECiDrereporter mice. White arrowheads indicate AV shunt. E. Representative images showing the P6 retinal vasculature (Isolectin B4 : red, GFP: green) of Smad4iECKO;VECiDrereporter mice. Note that vein starts to form abnormal branch (white dotted line) toward the nearby artery and ECs in the sprout are labeled with GFP. F. Confocal images showing EdU and IB4 staining in P7 retinal vasculature of Smad4iECKO mice. Note that ECs in AV shunt are highly proliferative. G. Representative images showing GM130 (cyan) and DAPI (green) immunostaining in P7 retinal vasculature (Isolectin B4 : red, GFP: green) of Smad4iECKO mice. Right panel represents endothelial nucleus and golgi in AV shunt. Note that endothelial golgi orientation in AV shunt is toward artery. H. Representative images showing the P7 retinal vasculature (Isolectin B4 : red, GFP: green) of Smad4iECKO;VECiDrereporter mice and WT littermate at low magnification. I. Quantification of GFP+ area in the P7 retinal vasculature of VECiDrereporter (blue) and Smad4iECKO;VECiDrereporter (orange) mice. (n=7, unpaired two-samples t-test) J. Representative images showing the vascular front area of P7 Smad4iECKO;VECiDrereporter mice (bottom) and WT littermate (top). (Isolectin B4 : purple, GFP: green). Green arrowheads indicate GFP+ TipECs. K. Quantification of GFP+ TipECs in the P7 retinal vasculature of VECiDrereporter (blue) and Smad4iECKO;VECiDrereporter (orange) mice. (n=8, unpaired two-samples t-test)

Article Snippet: The CreER T2 cassette construct was obtained from Addgene (pCAG-CreERT2) 7 and the DreER T2 cassette was subcloned from pCAG-NLS-HA-Dre construct (Addgene) 16 .

Techniques: Labeling, Staining, Immunostaining

Journal: Nature genetics

Article Title: Early TP53 Alterations Engage Environmental Exposures to Promote Gastric Premalignancy in an Integrative Mouse Model

doi: 10.1038/s41588-019-0574-9

Figure Lengend Snippet: (a) Schematic showing Lgr5-p53 experimental design and outcomes; percentages represent frequency of mice with antral dysplastic lesions. DCA/MNU treatment and tamoxifen injection schedule indicated on schematic below. (b) Representative H&E staining of gastric lesions from untreated and DCA/MNU treated Lgr5-p53 mice. Scale bar = 250μM. (c) Quantification of frequency of dysplasia in mice at 12 months in DCA/MNU treated Lgr5-p53 WT (n = 11) and Lgr5-p53 KO (n = 19) mice. This experiment was repeated once with similar results. (d) Mutation burden in the form of insertions and deletions (indels) in dysplastic gastric lesions from Lgr5-p53 WT (n = 3) and Lgr5-p53 KO (n = 10) mice using whole-exome sequencing. Data is presented as mean ± s.e.m.; two-sided Kolmogorov-smirnov p = 0.084. (e) Schematic showing Mist-p53;mtmg experimental design and outcomes; percentages represent frequency of dysplastic lesions found in the antrum or corpus of mice. Same DCA/MNU treatment and tamoxifen injection schedule as shown in (a) (f) Quantification of mice with antral or corpus dysplasia in DCA/MNU treated Mist-p53 WT (n = 11) and Mist-p53 KO (n = 15) mice at 12 months. This experiment was performed once. (g) Representative images from DCA/MNU-treated Mist-p53 WT and Mist-p53 KO mice. Dissection microscope images of lower esophagus and stomach flayed open along greater curvature; arrow indicates antral lesion (left panel). H&E staining and GFP immunohistochemistry of regions of dysplasia and antral polyps. Scale bar = 125μM.

Article Snippet: Lgr5 -EGFP-Ires-CreER T2 mice were backcrossed in C57BL/6J mice and subsequently SNP tested to ensure >97% pure background (Taconic).

Techniques: Injection, Staining, Mutagenesis, Sequencing, Dissection, Microscopy, Immunohistochemistry

Journal: Nature genetics

Article Title: Early TP53 Alterations Engage Environmental Exposures to Promote Gastric Premalignancy in an Integrative Mouse Model

doi: 10.1038/s41588-019-0574-9

Figure Lengend Snippet: (a) Recombination specific PCR of DNA extract from gastric lesions of Lgr5-p53 WT and Lgr5-p53 KO mice. Data presented as mean ± s.d of three technical replicates. (b) Schematic showing Lgr5-p53 LSL-R270H experimental design; DCA/MNU treatment and tamoxifen injection schedule during indicated duration depicted below. (c) Kaplan-Meier survival curve of Lgr5-p53 R270H experiments; table shows frequency of dysplasia in DCA/MNU treated mice of indicated genotype. (d) Schematic showing Lgr5-p53 KO experimental design; DCA alone, MNU alone, or DCA/MNU combination as well as tamoxifen injection schedule during indicated duration depicted below. Table shows frequency of dysplasia in Lgr5-p53 KO mice with indicated treatment after 12 months. *1/6 Cre-negative, tamoxifen-induced control mice treated with DCA/MNU developed dysplasia.

Article Snippet: Lgr5 -EGFP-Ires-CreER T2 mice were backcrossed in C57BL/6J mice and subsequently SNP tested to ensure >97% pure background (Taconic).

Techniques: Injection, Control

Journal: Nature genetics

Article Title: Early TP53 Alterations Engage Environmental Exposures to Promote Gastric Premalignancy in an Integrative Mouse Model

doi: 10.1038/s41588-019-0574-9

Figure Lengend Snippet: (a) Schematic showing isolation of dysplastic organoids from premalignant dys-Lgr5-p53 KO and dys-Lgr5-p53 WT gastric organoids from experiments shown in . Phase contrast images of Nutlin-3 treated dys-Lgr5-p53 KO and dys-Lgr5-p53 WT gastric organoids (b) Quantification of colony forming ability in ultra-low attachment culture of dys-Lgr5-p53 KO and dys-Lgr5-p53 WT gastric organoids. Data presented as mean ± s.d. from three biological replicates; p-values calculated by two-sided Student’s t-test. (c) Quantification of percent Lgr5+ cells in dys-Lgr5-p53 KO and dys-Lgr5-p53 WT gastric organoid culture as measured by GFP+. Data presented as mean ± s.d. from three biological replicates; p-values calculated by two-sided Student’s t-test. (d) Histopathological analysis of gastric dysplasia in dys-Lgr5-p53 KO and dys-Lgr5-p53 WT mice: H&E stain (top), Alcian blue stain (middle), anti-GFP/Lgr5 IHC (bottom) (e) Karyotype analysis of dys-Lgr5-p53 KO (n = 2 mice) and dys-Lgr5-p53 WT (n = 2 mice) gastric organoids; chromosome count of 10 cells per group. Representative images shown to the right; inserts are higher magnification images of chromosomes. Data presented as mean ± s.e.m.; p-values calculated by two-sided Student’s t-test. (f) Primary xenograft growth of dys-Lgr5-p53 KO (n = 10 mice) and dys-Lgr5-p53 WT (n = 10 mice) gastric organoids in nude mice

Article Snippet: Lgr5 -EGFP-Ires-CreER T2 mice were backcrossed in C57BL/6J mice and subsequently SNP tested to ensure >97% pure background (Taconic).

Techniques: Isolation, Staining

Journal: Nature genetics

Article Title: Early TP53 Alterations Engage Environmental Exposures to Promote Gastric Premalignancy in an Integrative Mouse Model

doi: 10.1038/s41588-019-0574-9

Figure Lengend Snippet: (a) Recombination specific PCR of DNA extract from premalignant dys-Lgr5-p53 WT and dys-Lgr5-p53 KO gastric organoids. Data presented as mean ± s.d of n = 3 technical replicates per group. (b) Quantification of proliferation by CellTiter-Glo and phase contrast images of nondysplastic and dysplastic Lgr5-p53 WT and Lgr5-p53 KO gastric organoids in the presence or absence of Nutlin-3 (30uM for 72 hours). Data presented as mean ± s.d. of n = 6 cell culture replicates. (c) Representative images of phase contrast and GFP-immunofluorescent images from dys-Lgr5-p53 KO and dys-Lgr5-p53 WT gastric organoid cultures. The experiment was repeated once with similar results. (d) Karyotype analysis of premalignant Lgr5-p53 KO (n = 2 mice), Lgr5-p53 WT (n = 2 mice), and Lgr5-p53 WT + AdenoCre gastric organoids; chromosome count of 10 cells per group. Data presented as mean ± s.d.; p-values calculated by Ordinary One-way ANOVA with Sidak’s multiple comparisons adjustment. (e) Copy number analysis of a patient specimen of a high-grade dysplastic Barrett’s metaplasia harboring TP53 R175H mutation and genome doubling. (f) Histopathology of primary tumor xenograft of Lgr5-p53 KO gastric organoid showing features of dysplasia. This experiment was repeated twice with similar results. (g) DAPI staining and GFP (Lgr5+) immunofluorescence of primary xenograft from Lgr5-p53 KO gastric organoids. This experiment was repeated twice with similar results. (h) Schematic of low-pass whole genome sequencing (LP-WGS) experiment of cultured and xenograft dys-Lgr5-p53 KO gastric organoids. Common and distinct broad somatic copy number alterations that were only found in xenograft tissue are displayed.

Article Snippet: Lgr5 -EGFP-Ires-CreER T2 mice were backcrossed in C57BL/6J mice and subsequently SNP tested to ensure >97% pure background (Taconic).

Techniques: Cell Culture, Mutagenesis, Histopathology, Staining, Immunofluorescence, Sequencing

Journal: Nature genetics

Article Title: Early TP53 Alterations Engage Environmental Exposures to Promote Gastric Premalignancy in an Integrative Mouse Model

doi: 10.1038/s41588-019-0574-9

Figure Lengend Snippet: (a) Schematic showing methodology for gene expression profiling of p53 KO (n = 2), p53 WT (n = 2), Lgr5-p53 KO (n = 2), Lgr5-p53 WT (n = 2), dys-Lgr5-p53 KO (n = 6) and dys-Lgr5-p53 WT (n = 6) gastric organoids. (b) Volcano plot of gene-set enrichment pathways in premalignant dLgr5-p53 KO (n = 5 organoids from 2 mice) versus dLgr5-p53 WT (n = 6 organoids from 2 mice) gastric organoids plotted as normalized enrichment scores (NES) by -log10(p-value), where p-values for gene set enrichment scores were determined by randomly permuting genes (n=100,000 permutations) (left). Individual inflammation gene sets are described with corresponding Normalized Enrichment Scores (NES). Gene-set enrichment plot of inflammatory response pathway ; associated heat map of scoring genes in gene-set (right). (c) Scatter plot of NES of gene-sets in dys-Lgr5-p53 KO /dys-Lgr5-p53 WT (y-axis) versus nondysplastic Lgr5-p53 KO /Lgr5-p53 WT (x-axis) gastric organoids; Select pathways downregulated in both comparisons are indicated in blue; upregulated in both comparisons indicated in red; and only upregulated in dys-Lgr5-p53 KO comparison indicated in orange. (d) Immunoblot showing protein levels of WNT target Sox9, total β-catenin, and vinculin (loading control) in p53 WT , p53 KO , Lgr5-p53 WT , Lgr5-p53 KO , dys-Lgr5-p53 WT , dys-Lgr5-p53 KO gastric organoids. This experiment was repeated once. (e) Histopathology of nondysplastic gastric antrum of untreated Lgr5-p53 WT and Lgr5-p53 KO mice and dysplastic gastric lesions from treated dys-Lgr5-p53 WT and dys-Lgr5-p53 KO mice; H&E staining (top panel) and Sox9 immunohistochemistry (bottom panel) of gastric antrum. Scale bar = 250μM

Article Snippet: Lgr5 -EGFP-Ires-CreER T2 mice were backcrossed in C57BL/6J mice and subsequently SNP tested to ensure >97% pure background (Taconic).

Techniques: Gene Expression, Comparison, Western Blot, Control, Histopathology, Staining, Immunohistochemistry

Journal: Nature genetics

Article Title: Early TP53 Alterations Engage Environmental Exposures to Promote Gastric Premalignancy in an Integrative Mouse Model

doi: 10.1038/s41588-019-0574-9

Figure Lengend Snippet: (a) mRNA expression of Trp53 in p53 KO , p53 WT , Lgr5-p53 KO , Lgr5-p53 WT , dys-Lgr5-p53 KO and dys-Lgr5-p53 WT gastric organoids. Data presented as mean ± s.d. of n = 2 (nondysplastic) and n = 3 (dysplastic) cell culture replicates. (b) mRNA expression levels of Csf3 , Cxcl10 , and Ccl5 in p53 KO , p53 WT , Lgr5-p53 KO , Lgr5-p53 WT , dys-Lgr5-p53 KO and dys-Lgr5-p53 WT gastric organoids by RT-PCR. Data presented as mean ± s.d. of n = 3 cell culture replicates. (c) Volcano plot of differential expressed gene-sets in p53 KO gastric organoids plotted as normalized enrichment scores (NES) by -log10(p-value), where p-values for gene set enrichment scores were determined by randomly permuting genes (n=100,000 permutations). Individual inflammation gene sets are described with corresponding NES. (d) Scatter plot of single sample GSEA (ssGSEA) of interferon pathway and Ccl5 mRNA expression in gastric cancer cell lines annotated by TP53 mutation status (n = 26 for TP53 mutant and n = 10 for TP53 wildtype). Gray shaded region represents 95% confidence interval of the linear regression fit. Pearson’s correlation value R = 0.6 and p-value computed based on a two-sided t-distribution. (e) Schematic of proposed vicious feedback cycle in gastric premalignancy: chronic inflammation and carcinogen exposure selects for p53 mutant gastric cells, which in turn stimulate inflammatory cytokines, particularly Csf3, Cxcl10, Ccl5

Article Snippet: Lgr5 -EGFP-Ires-CreER T2 mice were backcrossed in C57BL/6J mice and subsequently SNP tested to ensure >97% pure background (Taconic).

Techniques: Expressing, Cell Culture, Reverse Transcription Polymerase Chain Reaction, Mutagenesis

Journal: Nature genetics

Article Title: Early TP53 Alterations Engage Environmental Exposures to Promote Gastric Premalignancy in an Integrative Mouse Model

doi: 10.1038/s41588-019-0574-9

Figure Lengend Snippet: (a) WNT-reporter activity in adherent culture of dys-Lgr5-p53 KO and dys-Lgr5-p53 WT gastric cells following transient transfection. Data presented as mean ± s.d. of n = 4 cell culture replicates; p-value calculated by two-sided Mann Whitney test. (b) Phase contrast images of dys-Lgr5-p53 KO and dys-Lgr5-p53 WT gastric organoids two days following the first and second passage in WNT independent media (DMEM + 10%FBS). This experiment was repeated twice with similar results. (c) Gene network of significantly mutated genes in dysplastic gastric lesions from DCA/MNU-treated Lgr5-p53 WT mice (n = 8) using GeNets (adjusted p-value = 0.00065 using Bonferroni correction) (Li et al., 2018).

Article Snippet: Lgr5 -EGFP-Ires-CreER T2 mice were backcrossed in C57BL/6J mice and subsequently SNP tested to ensure >97% pure background (Taconic).

Techniques: Activity Assay, Transfection, Cell Culture, MANN-WHITNEY

Journal: Nature genetics

Article Title: Early TP53 Alterations Engage Environmental Exposures to Promote Gastric Premalignancy in an Integrative Mouse Model

doi: 10.1038/s41588-019-0574-9

Figure Lengend Snippet: (a) Volcano plot of differentially expressed genes in dys-Lgr5-p53 KO (n = 6, three cell culture replicates from 2 different mice) relative to dys-Lgr5-p53 WT (n = 6, three cell culture replicates from 2 different mice) gastric organoids show as log fold-change (logFC, x-axis) by -log 10 (p-value), where p-values were estimated from empirical-Bayes moderated t-statistics. Dots colored red have FDR < 0.1, where FDR-adjusted p-values are estimated using the Benjamini-Hochberg method. (b) mRNA expression of CDKN2A in p53 WT , p53 KO , Lgr5-p53 WT , Lgr5-p53 KO , dys-Lgr5-p53 WT and dys-Lgr5-p53 KO gastric organoids (c) Immunoblot showing protein levels of p53, p16, and vinculin (loading control) in dys-Lgr5-p53 WT expressing either a control sgRNA or Trp53 sgRNA. This experiment was repeated once. (d) Localization and expression levels of p16 (red) in dysplastic lesions of DCA/MNU-treated Lgr5-p53 WT and Lgr5-p53 KO mice. Nuclear staining by DAPI (blue). Scale bar = 125μM (e) Localization and expression levels of p16 (red) in dysplastic lesions of DCA/MNU-treated Mist-p53 WT and Mist-p53 KO mice. GFP+ represents conditionally induced Mist+ cells in indicated genetic mice. Nuclear staining by DAPI (blue). Scale bar = 125μM (f) Oncoprint from cbioportal showing alterations in CDKN2A and TP53 as well as CDKN2A mRNA expression levels in the CIN-subtype of human gastric (n = 223) and esophageal (n = 74) adenocarcinomas from TCGA. Deep deletions (solid blue); missense mutations in the COSMIC repository (solid green); nonsense or frameshift mutations (black); promoter methylation (magenta). CDKN2A mRNA expression from Z-scores of −3 to 3.

Article Snippet: Lgr5 -EGFP-Ires-CreER T2 mice were backcrossed in C57BL/6J mice and subsequently SNP tested to ensure >97% pure background (Taconic).

Techniques: Cell Culture, Expressing, Western Blot, Control, Staining, Methylation

Journal: Nature genetics

Article Title: Early TP53 Alterations Engage Environmental Exposures to Promote Gastric Premalignancy in an Integrative Mouse Model

doi: 10.1038/s41588-019-0574-9

Figure Lengend Snippet: (a) Immunoblot showing protein levels of p53, p16, and vinculin (loading control) in dys-Lgr5-p53 WT control or Trp53/Cdkn2a DKO (top panel) and dys-Lgr5-p53 KO control or p16 KO gastric organoids. This experiment was repeated once. (b) Quantification and representative dissection microscope images of dys-Lgr5-p53 WT control or Trp53/Cdkn2a DKO gastric organoid colonies formed under ultra-low attachment culture conditions in WRN media after 21 days. (c) Quantification and representative dissection microscope images of dys-Lgr5-p53 KO control or Cdkn2a DKO gastric organoid colonies formed under low-attachment culture conditions in indicated media after 7 days. Data presented as mean ± s.d. of three culture replicates; p-values calculated by two-sided Student’s t-test. (d) Primary xenograft growth of dys-Lgr5-p53 WT gastric organoids expressing either a control sgRNA, Trp53 sgRNA, Cdkn2a sgRNA, or Trp53 / Cdkn2a sgRNA (DKO) in nude mice (n=6 per group). ( e) Immunoblot showing protein levels of phospho-CHK1, total-CHK1, phospho-RPA32, p53, p16, and vinculin (loading control) in dys-Lgr5-p53 WT expressing either a control sgRNA, Trp53 sgRNA, Cdkn2a sgRNA, or Trp53 / Cdkn2a sgRNA (DKO). This experiment was repeated once. (f) Heatmap showing relative sensitivity of CHEK1/2 inhibitor AZD7762 in 30 human gastric cancer cell lines. Solid red rectangles indicate cell-lines with co-disruption of TP53 / CDKN2A ; p-value = 0.0051, FDR = 0 (Top panel); Box plot showing quantiles of sensitivity to CHEK1/2 inhibitor AZD77622 in 30 human gastric cancer cell lines (black dots) stratified into four molecular groups: CDKN2A WT /TP53 WT (n = 8), CDKN2A altered /TP53 WT (n = 1), CDKN2A WT /TP53 altered (n = 15), and CDKN2A altered /TP53 altered (n = 6). Drug response was measured as Area Under the Curve (AUC). Differences in sensitivity were determined by two-sided Student’s t-test with *p-value <0.05 and **p-value < 0.01 (Bottom Panel) (g) Dose-response curve of isogenic CP-A cell lines expressing either control sgRNA, p53 sg#1, or p53 sg#2 to indicated concentrations of Prexasertib. IC 50 scores are displayed; Data presented as mean ± s.d. of six culture replicates at each indicated dose; Comparison of Fits, p value = 0.011 based on differences in IC 50 (right panel). Immunoblot phospho-CHK1, phospho-ATR, total-ATR, and B-actin loading control in CP-A control or p53 KO BE cells treated with indicated concentrations of Prexasertib (left panel). This experiment was repeated once. (h) Dose-response curve of isogenic dys-Lgr5-p53 WT -control and dys-Lgr5-p53 WT -DKO gastric organoids to indicated concentrations of Prexasertib (Top). Comparison of Fits, p = 0.0001 based on bottom value. Dose-response curve of isogenic dys-Lgr5-p53 KO -control and dys-Lgr5-p53 KO -sg CDKN2A gastric organoids to indicated concentrations of Prexasertib. Comparison of Fits, p = 0.0047 based on IC 50 . Data presented as mean ± s.d. of six culture replicates at each indicated dose (i) Representative phase contrast images of dys-Lgr5-p53 WT -control, dys-Lgr5-p53 WT -DKO, dys-Lgr5-p53 KO -control, dys-Lgr5-p53 WT -p16 KO gastric organoids treated for 36 hours with DMSO or indicated concentration of CHK1/2 inhibitor Prexasertib. This experiment was repeated once. (j) Quantification of organoid size from each group in experiment (F). Data presented as mean ± s.d. of seven technical replicates; p-values calculated by two-sided Student’s t-test. (k) Schematic of functional and therapeutic role of CDKN2A and TP53 alterations in gastric premalignancy

Article Snippet: Lgr5 -EGFP-Ires-CreER T2 mice were backcrossed in C57BL/6J mice and subsequently SNP tested to ensure >97% pure background (Taconic).

Techniques: Western Blot, Control, Dissection, Microscopy, Expressing, Disruption, Comparison, Concentration Assay, Functional Assay

Journal: Nature genetics

Article Title: Early TP53 Alterations Engage Environmental Exposures to Promote Gastric Premalignancy in an Integrative Mouse Model

doi: 10.1038/s41588-019-0574-9

Figure Lengend Snippet: (a) Dose-response curves of CHK1 inhibitor AZD7762 of two gastric cancer cell lines wildtype for either TP53 or CDKN2A and two gastric cancer cell lines with co-alteration of TP53 and CDKN2A . (b) Relative dependency of 23 gastric cancer cell lines to CHEK1, CHEK2, and WEE1 shRNA-mediated knockdown as shown by CERES dependency score. Black rectangles indicate gastric cancer cell lines with simultaneous disruption of TP53 and CDKN2A . (c) Dose-response curve of KE39 (TP53 MUT , CDKN2A WT ) and GSU (TP53 MUT , CDKN2A MUT ) gastric cancer cell lines to Prexasertib. Data presented as mean ± s.d. of n = 6 cell culture replicates at each dose; p-value calculated by Comparison of Fits based on differences in IC 50 . (d) Dose-response curve of KE39 (TP53 MUT , CDKN2A WT ) and GSU (TP53 MUT , CDKN2A MUT ) gastric cancer cell lines to WEE1 inhibitor AZD1775. Data presented as mean ± s.d. of n = 6 cell culture replicates at each dose; p-value calculated by Comparison of Fits based on differences in IC 50 . (e) Quantification by manual counting of number of organoids in experiment from - . (f) Quantification of organoid size and number from dys-Lgr5-p53 KO -control and dys-Lgr5-p53 WT -p16 KO gastric organoids treated for 36 hours with DMSO or indicated concentration of ATR inhibitor AZD6738 or WEE1 inhibitor AZD1775. Data presented as mean ± s.d. of n = 6-8 technical replicates at each dose (g) Representative phase contrast images from experiment (f)

Article Snippet: Lgr5 -EGFP-Ires-CreER T2 mice were backcrossed in C57BL/6J mice and subsequently SNP tested to ensure >97% pure background (Taconic).

Techniques: shRNA, Knockdown, Disruption, Cell Culture, Comparison, Control, Concentration Assay