Journal: Oncogene

Article Title: Targeting chemoresistant colorectal cancer via systemic administration of a BMP7 variant

doi: 10.1038/s41388-019-1047-4

Figure Lengend Snippet: BMP7v treatment promotes CR-CSC differentiation. a Phase-contrast microscopy analysis of CD44v6 + CRC sphere cells treated with BMP7v at the indicated time points. One representative of CSC#1, 2, 4, 5, 7, and 23–26 is shown. The scale bar represents 20 µm. b Percentage of CK20 positive cells in CD44v6 + CR-CSCs treated with vehicle or BMP7v up to 21 days evaluated by immunofluorescence analysis. Data are expressed as mean ± SD of experiments performed in 15 CRC sphere cell lines (CSC#1–3, 5–7, 10,11, 14–16, 18, 25, 33, and 40). c Flow cytometry analysis of CD133/CD44v6 on CRC sphere cells treated with vehicle or BMP7v for 14 days. Data reported are mean ± SD of 15 CRC sphere cell lines analyzed (CSC#1–8, 10,11, 14–16, 18, and 25). d (left panels) Immunofluorescence analysis of CDX2 on CR-CSCs upon 14 days of BMP7v treatment. One representative of CSC# 3, 9, and 21 is shown. Nuclei were stained with Toto-3 (blue color). The scale bars represent 20 µm. (right panel) Percentage of CDX2 positive cells in CD44v6 + CR-CSCs treated with vehicle or BMP7v up to 14 days evaluated by immunofluorescence analysis. Data are expressed as mean ± SD of experiments performed in CSC# 3, 9, and 21. e Flow cytometry analysis of TOP-dGFP or CD44v6 in enriched CD44v6 + sphere cells treated with BMP7v up to 14 days. One representative experiment of CSC#1, 2, 4, 7, and 10 is shown. f Phase-contrast microscopy analysis of TOP-dGFP CRC sphere cells grown in matrigel drops and treated with vehicle, BMP7v or FBS for 14 days. One representative of CSC# 8, 9, and 11 is shown. The scale bar represents 100 µm. g Immunofluorescence analysis of E-cadherin, vimentin, and β-catenin (green color) in CD44v6 + CRC cells exposed to vehicle or BMP7v for 14 days. One representative experiment performed in cells as in e is shown. Nuclei were stained with Toto-3 (blue color). The scale bars represent 20 µm. h Migrating CD44v6 + and CD44v6 − cells treated with vehicle or BMP7v up to 48 h. Data are shown as mean ± SD of three independent experiments performed in five CRC sphere cell lines (CSC#1, 5, 7, 10, and 12). i Cell viability percentage of enriched CD44v6 + and CD44v6 − cells treated with vehicle or BMP7v up to 96 h. Data are shown as mean ± SD of different experiments performed in CSC#1, 2, 4, 7, and 10. j Cell cycle analysis in CD44v6 + CR-CSCs exposed to vehicle or BMP7v for 72 h. The data show percentage of cell number in sub-G0, G0/G1, S, and G2/M phases. Data are expressed as mean ± SD of three independent experiments performed in five different CRC sphere cell lines as in e . k Immunoblot analysis of PARP, cleaved PARP (cPARP), Caspase-3 (Casp-3), cleaved Caspase-3 (cCasp-3), Bcl-2, Bcl-xL in CD44v6 + , and CD44v6 − enriched cells treated as in e for 72 h. β-actin was used as loading control. One representative experiment performed in three different CRC sphere cell lines (CSC#1, 4, and 7)

Article Snippet: Following blocking with 3% bovine serum albumin (BSA) for 30 min, cells were exposed overnight at 4 °C to BMP7 (MAB3541, mouse, IgG2 b , R&D system), LGR5 (GPR49, rabbit, IgG, Abgent), CDX2 (MAB3665, mouse, IgG1, R&D Systems), CK20 (NCL-L-CK20, mouse, IgG2 k , Novocastra Leica), E-cadherin (#3195, rabbit, IgG, CST), vimentin (#5741, rabbit, IgG, CST), β-catenin (MAB1329, mouse, IgG2 b , R&D Systems), CD44v6 (BBA13, clone 2F10, mouse, IgG1, R&D system), BMPR1A (MAB2406, mouse, IgG2 b , R&D Systems), BMPR1B (MAB505, mouse, IgG2 a , R&D Systems) and, BMPR2 (MAB811, mouse, IgG2 b , R&D Systems) antibodies or isotype-matched controls (IMCs).

Techniques: Microscopy, Immunofluorescence, Flow Cytometry, Staining, Cell Cycle Assay, Western Blot, Control

Journal: PloS one

Article Title: Brg1 is required for Cdx2-mediated repression of Oct4 expression in mouse blastocysts.

doi: 10.1371/journal.pone.0010622

Figure Lengend Snippet: Figure 1. Brg1 is required for repression of Oct4 expression at the blastocyst stage. (A) qRT-PCR analysis of Brg1, Cdx2, and Oct4 transcripts in Brg1 KD 8-cell embryos, morulae, and blastocysts. Data were normalized to Ubtf (house keeping gene) and are relative to control embryos at each stage; black line = 1. Asterisk denotes significant difference between Brg1 KD and control blastocysts (p,0.05). (B) ICC analysis of Oct4 and Cdx2 expression in Brg1 KD 8-cell embryos, morulae, and blastocysts. Nuclei were counter stained with DAPI (blue). (C) Brg1 represses Oct4 expression in a dose dependent manner. One-cell embryos were injected with 0 mM (control), 0.1 mM, 1 mM, or 100 mM Brg1 siRNA and cultured to the blastocyst stage. Real-time qPCR was used to analyze the levels of Brg1 and Oct4 transcripts. Data were normalized to Ubtf and are relative to control blastocysts; dashed line = 1. doi:10.1371/journal.pone.0010622.g001

Article Snippet: CDX2 was detected by Western blot analysis using an affinity purified rabbit anti-CDX2 antibody (A300-692A, Bethyl Laboratories, Inc).

Techniques: Expressing, Quantitative RT-PCR, Control, Staining, Injection, Cell Culture

Journal: PloS one

Article Title: Brg1 is required for Cdx2-mediated repression of Oct4 expression in mouse blastocysts.

doi: 10.1371/journal.pone.0010622

Figure Lengend Snippet: Figure 2. Expression and localization of Cdx2 and Oct4 in Brg1 KD blastocysts. (A) ICC analysis of Oct4 and Cdx2 in Brg1 KD and control blastocysts. In control blastocysts (a–d) Oct4 expression (green) is restricted to the ICM and is largely absent in the Cdx2-positive (red) trophectoderm. In contrast, in Brg1 KD blastocysts (e–h) Oct4 is widely expressed in both the ICM and cdx2-positive (yellow) trophectoderm. Arrowheads denote co- expression of Oct4 and Cdx2. Nuclei were counter stained with DAPI (blue). (B) Quantification of the average number of cells in Brg1 KD and control blastocysts expressing Oct4, Cdx2, and Oct4 & Cdx2 (double expression). Asterisks denote statistical significance (p,0.05) between Brg1 KD and control blastocysts. A total of 25 Brg1 KD blastocysts and 15 control blastocysts were analyzed. doi:10.1371/journal.pone.0010622.g002

Article Snippet: CDX2 was detected by Western blot analysis using an affinity purified rabbit anti-CDX2 antibody (A300-692A, Bethyl Laboratories, Inc).

Techniques: Expressing, Control, Staining

Journal: PloS one

Article Title: Brg1 is required for Cdx2-mediated repression of Oct4 expression in mouse blastocysts.

doi: 10.1371/journal.pone.0010622

Figure Lengend Snippet: Figure 5. Model for Brg1/Cdx2-mediated repression of Oct4 expression in trophectoderm. Schematic diagram of Oct4 regulation in blastocysts. In the trophectoderm Brg1 is recruited to the ARE of the Oct4 promoter via Cdx2. Once at the Oct4 promoter Brg1 and Cdx2 facilitate recruitment of additional co-repressors to repress transcription. doi:10.1371/journal.pone.0010622.g005

Article Snippet: CDX2 was detected by Western blot analysis using an affinity purified rabbit anti-CDX2 antibody (A300-692A, Bethyl Laboratories, Inc).

Techniques: Expressing

Journal: Genes & Development

Article Title: Enhancer, transcriptional, and cell fate plasticity precedes intestinal determination during endoderm development

doi: 10.1101/gad.318832.118

Figure Lengend Snippet: Shh Cre -mediated Cdx2 deletion results in anterior homeotic transformation. ( A ) The highest enriched sequences within region-specific enhancer clusters represent canonical TF motifs. ( B ) Trajectories of Eso/FS- and Int-specific TF transcripts in E12, E14, and E16 Eso/FS ( left ) and Int ( right ) endoderm. Log 2 transformed RPKM values, plotted over time, show increases and decreases in the different regions. ( C ) Intestinal regions (from the duodenum to the ileum) stained with PAS or with ATP4B or TRP63 antibody in E18 wild-type and Shh Cre ; Cdx2 Fl/Fl embryos. In the mutant duodenum and jejunum, gastric foveolar cells (recognized by a mauve apical rim; black arrows) extensively replace intestinal goblet cells (recognized by a purple cytoplasmic blob; black arrowheads). These contrasting features of wild-type and mutant jejunal villi are highlighted in the dashed boxes and magnified at the right . In addition, ATP4B + gastric parietal cells appear in the mutant jejunal (white arrowheads) but not duodenal crypts. The mutant ileum loses crypt–villus structure, including PAS + goblet cells, and is lined instead by TRP63 + stratified squamous mucosa (red arrows). Bars, 25 µm.

Article Snippet: CDX2 antibody-bound (Cell Signaling Technology) protein A and protein G magnetic Dynabeads (1:1 ratio; Thermo Fisher) were incubated overnight at 4°C with 2 μg of chromatin in 300 μL of ChIP buffer containing protease inhibitors and two carriers: 20 μL of recombinant human histone H2B solution (New England Biolabs, M2505S) and 1 μL of carrier RNA from the RNeasy microkit (Qiagen, 74004).

Techniques: Transformation Assay, Staining, Mutagenesis

Journal: Genes & Development

Article Title: Enhancer, transcriptional, and cell fate plasticity precedes intestinal determination during endoderm development

doi: 10.1101/gad.318832.118

Figure Lengend Snippet: Time-sensitive activation of FG-specific transcriptional programs in the absence of CDX2. ( A ) Schema of Cdx2 deletions mediated by either Shh Cre (approximately E10) or treatment of Villin-Cre ER-T2 dams with tamoxifen at E13 or E15. Embryos deleted before E14 were assessed at E16, and those deleted at E15 were assessed at E18. CDX2 immunostains verify protein loss. ( B ) Unsupervised hierarchical clustering (Pearson coefficients) for the 2604 genes expressed differently (greater than fourfold, q < 0.05) in E16 wild-type tissues ( Supplemental Fig. S1D ) and E16 Cdx2 −/− Int after deletion at approximately E10 or E13. Transcriptomes in early, but not late, Cdx2 -deleted Int endoderm resemble those in FG epithelia. ( C ) mRNA differences in Cdx2 −/− Int after deletion at approximately E10 ( n = 3), E13, or E15 or in adult duodenum and ileum ( n = 2 each). Clusters are those with differential expression in wild-type E16 EPCAM + Eso/FS, HS, and Int cells (from Supplemental Fig. S1D ). For each FG cluster, colored violin plots represent the distributions of mRNA levels (colors represent the region, and white dots represent the median) in wild-type tissues, and gray violins show the distributions after Cdx2 deletion (knockout) at different times. Violin plots for Int genes are shown in Supplemental Figure S3B . The Mann-Whitney U -test was applied to determine the significance between Cdx2 knockout at different times. (n.s.) Not significant; (*) P < 0.05; (****) P < 0.0001. ( D – F ) IGV tracks of RNA-seq data showing native (wild-type FS, HS, and Int) and ectopic FG gene expression in Cdx2 −/− intestines after deletion at E10, E13, or E15. Numbers represent the scale for normalized read counts.

Article Snippet: CDX2 antibody-bound (Cell Signaling Technology) protein A and protein G magnetic Dynabeads (1:1 ratio; Thermo Fisher) were incubated overnight at 4°C with 2 μg of chromatin in 300 μL of ChIP buffer containing protease inhibitors and two carriers: 20 μL of recombinant human histone H2B solution (New England Biolabs, M2505S) and 1 μL of carrier RNA from the RNeasy microkit (Qiagen, 74004).

Techniques: Activation Assay, Quantitative Proteomics, Knock-Out, MANN-WHITNEY, RNA Sequencing, Gene Expression

Journal: Genes & Development

Article Title: Enhancer, transcriptional, and cell fate plasticity precedes intestinal determination during endoderm development

doi: 10.1101/gad.318832.118

Figure Lengend Snippet: CDX2 binding in developing Int endoderm and the consequences of late depletion. ( A ) PAS staining of intestinal regions after Cdx2 deletion at E13 or E15. Wild-type and mutant Ints were assessed at E18. Goblet cells (black arrowheads) were intact, and no region showed gastric foveolar differentiation (no cells were observed with apical rim staining). Bar, 25 μm. ( B ) Normalized ChIP-seq data for CDX2 at different developmental stages. Binding at enhancers from any regional cluster is minimal at E13; thereafter, it increases progressively and selectively at Int enhancers. ( C ) Lack of CDX2 binding at P1 FG enhancers (red- and gold-shaded areas). The blue-shaded box demarcates a CDX2-bound intestinal enhancer. Numbers represent the scales of ChIP or ATAC signals. ( D ) E13-specific CDX2-bound sites overlap minimally with FG-specific enhancer regions, implying the absence of direct CDX2 control over FG genes.

Article Snippet: CDX2 antibody-bound (Cell Signaling Technology) protein A and protein G magnetic Dynabeads (1:1 ratio; Thermo Fisher) were incubated overnight at 4°C with 2 μg of chromatin in 300 μL of ChIP buffer containing protease inhibitors and two carriers: 20 μL of recombinant human histone H2B solution (New England Biolabs, M2505S) and 1 μL of carrier RNA from the RNeasy microkit (Qiagen, 74004).

Techniques: Binding Assay, Staining, Mutagenesis, ChIP-sequencing, Control

Journal: Genes & Development

Article Title: Enhancer, transcriptional, and cell fate plasticity precedes intestinal determination during endoderm development

doi: 10.1101/gad.318832.118

Figure Lengend Snippet: Persistence of open chromatin at FG enhancers after early, but not late, CDX2 loss. ( A ) Mice were treated to induce Cdx2 deletion according to the schema in A, and isolated Int epithelium was assessed for accessible chromatin by ATAC. “Overlap” and HS enhancers, which are ordinarily open in wild-type embryos until E14 ( A), remained accessible after Cdx2 deletion at approximately E10 or E13 but were closed after Cdx2 deletion at E15 or in adult mice. Intestinal enhancers were significantly compromised after early, and less compromised after late, Cdx2 deletion. The IGV track shows a representative locus: Foxp4 . Shaded boxes denote Eso/FS- and HS-specific enhancers showing open chromatin after early Cdx2 deletion. ( B ) The fractions of genes up-regulated in E10 Cdx2 −/− intestines (gray) located 25 or 50 kb away from “overlapping” or HS enhancers where ATAC signals persist in E10 Cdx2 −/− intestines. ( C ) Model for the temporal relation of enhancer accessibility to tissue plasticity in wild-type endoderm ( top ), as inferred from Cdx2 −/− intestines ( bottom ). IGV tracks show original ATAC-seq peaks at representative FG- and Int-specific enhancers located far from the respective TSSs. ( D ) Model depicting midgut developmental potential as a function of temporally regulated waves of enhancer accessibility and region-specific mRNA expression. Low-level expression of FG genes is initially pervasive, while FG enhancers are accessible throughout the gut tube. Regional determination occurs after E14, when enhancer signatures become consolidated, gene expression domains become restricted, and regional anatomy approaches adult forms.

Article Snippet: CDX2 antibody-bound (Cell Signaling Technology) protein A and protein G magnetic Dynabeads (1:1 ratio; Thermo Fisher) were incubated overnight at 4°C with 2 μg of chromatin in 300 μL of ChIP buffer containing protease inhibitors and two carriers: 20 μL of recombinant human histone H2B solution (New England Biolabs, M2505S) and 1 μL of carrier RNA from the RNeasy microkit (Qiagen, 74004).

Techniques: Isolation, Expressing, Gene Expression

Journal: Frontiers in Pharmacology

Article Title: Rabeprazole suppressed gastric intestinal metaplasia through activation of GPX4-mediated ferroptosis

doi: 10.3389/fphar.2024.1409001

Figure Lengend Snippet: Rabeprazole suppressed gastric intestinal metaplasia. (A) HEK293 cells were transfected with reporter gene containing CDX2 promoter and renilla plasmid for 12 h, following by treatment with or without rabeprazole for another48 h, the CDX2 transactivation was determined and analyzed using two sample t-test. Data was exhibited as mean ± s.d of three independent experiments, *** p < 0.001. (B) After treatment with or without rabeprazole for 48 h in BGC823 and AGS cells, the whole RNA was extracted to detect CDX2 ( left panel ) and MUC2 ( right panel ) mRNA level, data was displayed as mean ± s.d of three independent experiments, one sample t-test was employed to determine the significance. **** p < 0.0001; (C) After cell adhesion overnight, BGC823 and AGS cells were starved with serum-free medium for 16 h and treated with or without rabeprazole (100uM) for another 48 h. The whole cell lysate was collected and detected indicated protein expression by immunoblotting, band intensity was analyzed and quantified using two sample t-test, Data was exhibited as mean ± s.d of three independent experiments, **** p < 0.0001.

Article Snippet: Antibody against CDX2 monoclonal antibody (60243-1-Ig, Proteintech), SLC7A11/xCT polyclonal antibody (26864-1-AP, Proteintech), GPX4 monoclonal antibody (67763-1-Ig, Proteintech), Lamin A/C recombinant antibody (81042-1-RR, Proteintech), CREB1 Monoclonal antibody (67927-1-Ig, Proteintech) and phospho-CREB1 (Ser133) Polyclonal antibody (28792-1-AP, Proteintech) were from Proteintech (Wuhan, China).

Techniques: Transfection, Plasmid Preparation, Expressing, Western Blot

Journal: Frontiers in Pharmacology

Article Title: Rabeprazole suppressed gastric intestinal metaplasia through activation of GPX4-mediated ferroptosis

doi: 10.3389/fphar.2024.1409001

Figure Lengend Snippet: Rabeprazole modulated gastric intestinal metaplasia in dependent of ferroptosis. (A) BGC823 cells were digested and re-seeded into a 24-well plate and grown to 80% confluence, after serum-sstarvation overnight, BGC823 cells were treated with or without rabeprazole (100uM) for 48 h, the ROS level was detected according to the manufacter’s instruction and captured under fluorescent microscope (upper panel) as well as FCM analysis (bottom panel), bar: 250 um. (B) BGC823 and AGS cells were cultured without FBS overnight and incubated with or without rabeprazole (100uM) for another 48 h, real-time PCR was utilized to test indicated genes expression, control group was normalized as 1, data was exhibited as mean ± s.d of three independent experiments, one sample t-test was used to determine the significance. ** p < 0.01, **** p < 0.0001. (C) BGC823 and AGS cells was treated as described in B, immunoblotting was utilized to analyze GPX4 and SLC7A11 protein expression. Control group was normalized as 1, data was exhibited as mean ± s.d of three independent experiments, one sample t-test was used to determine the significance. * p < 0.05, ** p < 0.01, **** p < 0.0001. (D) After serum-starved culture overnight, BGC823 cells were treated with Ferrostatin-1 (Fer-1, 10 uM) for 1 h, subsequently followed by treatment with rabeprazole for another 48 h. The total protein was harvested to test MUC2 and CDX2 expression by immunoblotting. the band intensity was measured and quantified by one-ANOVA with multiple comparisons, followed by Bonferroni post hoc test for significance, control group was normalized as 1, data was exhibited as mean ± s.d of three independent experiments, **** p < 0.0001. (E) CCK8 analysis of cell viability in BGC823 cells treated as indicated for 48 h, data was exhibited as mean ± s.d of five independent experiments, one-ANOVA with multiple comparisons, followed by Bonferroni post hoc test for significance, **** p < 0.0001.

Article Snippet: Antibody against CDX2 monoclonal antibody (60243-1-Ig, Proteintech), SLC7A11/xCT polyclonal antibody (26864-1-AP, Proteintech), GPX4 monoclonal antibody (67763-1-Ig, Proteintech), Lamin A/C recombinant antibody (81042-1-RR, Proteintech), CREB1 Monoclonal antibody (67927-1-Ig, Proteintech) and phospho-CREB1 (Ser133) Polyclonal antibody (28792-1-AP, Proteintech) were from Proteintech (Wuhan, China).

Techniques: Microscopy, Cell Culture, Incubation, Real-time Polymerase Chain Reaction, Expressing, Control, Western Blot

Journal: Frontiers in Pharmacology

Article Title: Rabeprazole suppressed gastric intestinal metaplasia through activation of GPX4-mediated ferroptosis

doi: 10.3389/fphar.2024.1409001

Figure Lengend Snippet: Ferroptosis was decreased in patients with IM. (A) IHC was performed to detect GPX4 and SLC7A11 expression as well as CDX2 expression in a set of healthy control and patients with IM, the quantitation of GPX4 and SLC7A11 expression were performed upon IHC score, ** p < 0.01, **** p < 0.0001. (B) IF experiment was employed to determine GPX4 and SLC7A11 expression in a set of healthy control and patients with IM, the quantitation of GPX4 and SLC7A11 expression were performed upon mean fluorescence intensity (MFI) analysis, ** p < 0.01, **** p < 0.0001.

Article Snippet: Antibody against CDX2 monoclonal antibody (60243-1-Ig, Proteintech), SLC7A11/xCT polyclonal antibody (26864-1-AP, Proteintech), GPX4 monoclonal antibody (67763-1-Ig, Proteintech), Lamin A/C recombinant antibody (81042-1-RR, Proteintech), CREB1 Monoclonal antibody (67927-1-Ig, Proteintech) and phospho-CREB1 (Ser133) Polyclonal antibody (28792-1-AP, Proteintech) were from Proteintech (Wuhan, China).

Techniques: Expressing, Control, Quantitation Assay, Fluorescence

Journal: Molecular Medicine Reports

Article Title: Overexpression of caudal type homeobox transcription factor 2 inhibits the growth of the MGC-803 human gastric cancer cell line in vivo

doi: 10.3892/mmr.2015.3413

Figure Lengend Snippet: Sequences of the primers used for reverse transcription semi-quantitative polymerase chain reaction.

Article Snippet: Specific rabbit anti-human polyclonal antibodies to CDX2 (#12306), c-Myc (#5605), Skp2 (#2652), Bax (#5023), Bcl-2 (#2827), cyclin D1 (#2978), survivin (#2808) and GAPDH (#2118) were provided by Cell Signaling Technology, Inc. (Beverly, MA, USA) .

Techniques: Reverse Transcription, Sequencing

Journal: Molecular Medicine Reports

Article Title: Overexpression of caudal type homeobox transcription factor 2 inhibits the growth of the MGC-803 human gastric cancer cell line in vivo

doi: 10.3892/mmr.2015.3413

Figure Lengend Snippet: Determination of lentivirus titers. The viral titers of recombinant lentivirus transfected into 293T cells were determined by end point dilution, through counting the numbers of fluorescent green-labeled cells under a fluorescence microscope (magnification, x100). The viral dilution factor was 1:1,000. LV-GFP, lentivirus-green fluorescent protein; NC, negative control; CDX2, caudal type homeobox transcription factor 2.

Article Snippet: Specific rabbit anti-human polyclonal antibodies to CDX2 (#12306), c-Myc (#5605), Skp2 (#2652), Bax (#5023), Bcl-2 (#2827), cyclin D1 (#2978), survivin (#2808) and GAPDH (#2118) were provided by Cell Signaling Technology, Inc. (Beverly, MA, USA) .

Techniques: Recombinant, Transfection, Labeling, Fluorescence, Microscopy, Negative Control

Journal: Molecular Medicine Reports

Article Title: Overexpression of caudal type homeobox transcription factor 2 inhibits the growth of the MGC-803 human gastric cancer cell line in vivo

doi: 10.3892/mmr.2015.3413

Figure Lengend Snippet: Overexpression of CDX2 inhibits tumor growth and induces tumor cell apoptosis in LV-GFP-CDX2-treated mice. (A) Relative tumor volume growth curve revealed significant growth tendencies in the PBS and LV-GFP-NC groups, while the MGC-803 tumor growth in the LV-GFP-CDX2 group was markedly inhibited. (B) Tumor volumes in the LV-GFP-CDX2 groupe were smaller compared with those in the control group 14 days after tumor injection ( * P<0.05). (C) Tumor cell apoptosis was assessed using a TUNEL assay and HE staining, revealing that the MGC-803 tumor cells in the LV-GFP-CDX2 group had higher levels of apoptosis compared with the LV-GFP-NC and PBS groups (magnification, x400). CDX2, vaudal type homeobox transcription factor 2; LV-GFP, lentivirus-green fluoresent protein; PBS, phosphate-buffered saline; NC, negative control; TV, tumor volume; RTV, relative TV; TUNEL, terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling; HE, hematoxylin and eosin.

Article Snippet: Specific rabbit anti-human polyclonal antibodies to CDX2 (#12306), c-Myc (#5605), Skp2 (#2652), Bax (#5023), Bcl-2 (#2827), cyclin D1 (#2978), survivin (#2808) and GAPDH (#2118) were provided by Cell Signaling Technology, Inc. (Beverly, MA, USA) .

Techniques: Over Expression, Control, Injection, TUNEL Assay, Staining, Saline, Negative Control, End Labeling

Journal: Molecular Medicine Reports

Article Title: Overexpression of caudal type homeobox transcription factor 2 inhibits the growth of the MGC-803 human gastric cancer cell line in vivo

doi: 10.3892/mmr.2015.3413

Figure Lengend Snippet: Overexpression of CDX2 mRNA and protein in the LV-GFP-CDX2 group. (A) Reverse transcription semi-quantitative polymerase chain reaction analysis of CDX2 and GAPDH in the MGC-803 tumor tissues from the LV-GFP-CDX2, LV-GFP-NC and PBS groups. M, 500 bp marker. (B) mRNA expression levels of CDX2 were measured in the three groups, normalized to GAPDH and presented as the mean ± standard error of the mean (n=8 in each group). (C) Western blot analysis of the protein expression levels of CDX2 and GAPDH in the MGC-803 tumor tissues from the three groups. (D) Protein expression levels of CDX2 were measured in the three groups, normalized to GAPDH and presented as the mean ± standard error of the mean (n=8 in each group). Lanes: 1, LV-GFP-CDX2 group; 2, LV-GFP-NC group; 3, PBS group, GAPDH: internal control mRNA and protein. * P<0.05 compared with LV-GFP-NC and PBS group, using analysis of variance and Student-Newman-Keuls analyses. CDX2, caudal type homeobox transcription factor 2; LV-GFP, lentivirus-green fluoresent protein; PBS, phosphate-buffered saline; NC, negative control.

Article Snippet: Specific rabbit anti-human polyclonal antibodies to CDX2 (#12306), c-Myc (#5605), Skp2 (#2652), Bax (#5023), Bcl-2 (#2827), cyclin D1 (#2978), survivin (#2808) and GAPDH (#2118) were provided by Cell Signaling Technology, Inc. (Beverly, MA, USA) .

Techniques: Over Expression, Reverse Transcription, Real-time Polymerase Chain Reaction, Marker, Expressing, Western Blot, Control, Saline, Negative Control

Journal: Molecular Medicine Reports

Article Title: Overexpression of caudal type homeobox transcription factor 2 inhibits the growth of the MGC-803 human gastric cancer cell line in vivo

doi: 10.3892/mmr.2015.3413

Figure Lengend Snippet: Overexpression of CDX2 induces downregulation in the mRNA expression levels of c-Myc, Skp2, Bcl-2, cyclin D1 and survivin and upregulation in the expression of Bax. (A) Reverse transcription semi-quantitative polymerase chain reaction analysis of c-Myc, Skp2, Bcl-2, cyclin D1, survivin, Bax and GAPDH in the MGC-803 tumor tissues from the LV-GFP-CDX2, 2, LV-GFP-NC and PBS groups. M, 500 bp marker (B) mRNA expression levels of c-Myc, Skp2, Bcl-2, cyclinD1, survivin and Bax were measured in the three groups, normalized to those of GAPDH and presented as the mean ± standard error of the mean (n=8 in each group). 1, LV-GFP-CDX2 group; 2, LV-GFP-NC group; 3, PBS group; GAPDH: internal control. * P<0.05, compared with the LV-GFP-NC and PBS groups, using analysis of variance and Student-Newman-Keuls analyses. Bcl-2, B-cell lymphoma 2; Bax, Bcl-2-associated X protein; Skp2, S-phase kinase-associated protein 2; CDX2, caudal type homeobox transcription factor 2; LV-GFP, lentivirus-green fluoresent protein; PBS, phosphate-buffered saline.

Article Snippet: Specific rabbit anti-human polyclonal antibodies to CDX2 (#12306), c-Myc (#5605), Skp2 (#2652), Bax (#5023), Bcl-2 (#2827), cyclin D1 (#2978), survivin (#2808) and GAPDH (#2118) were provided by Cell Signaling Technology, Inc. (Beverly, MA, USA) .

Techniques: Over Expression, Expressing, Reverse Transcription, Real-time Polymerase Chain Reaction, Marker, Control, Saline

Journal: Molecular Medicine Reports

Article Title: Overexpression of caudal type homeobox transcription factor 2 inhibits the growth of the MGC-803 human gastric cancer cell line in vivo

doi: 10.3892/mmr.2015.3413

Figure Lengend Snippet: Overexpression of CDX2 induces downregulation of the protein expression levels of c-Myc, Skp2, Bcl-2, cyclinD1 and survivin and upregulation of Bax. (A) Western blot analysis of c-Myc, Skp2, Bcl-2, cyclin D1, survivin, Bax and GAPDH in the MGC-803 tumor tissue from the LV-GFP-CDX2, LV-GFP-NC and PBS groups. (B) Protein expression levels of c-Myc, Skp2, Bcl-2, cyclin D1, survivin and Bax were measured in the three groups, normalized to those of GAPDH and expressed as the mean ± standard error of the mean (n=8 in each group). 1, LV-GFP-CDX2 group; 2, LV-GFP-NC group; 3, PBS group; GAPDH: internal control. * P<0.05, compared with the LV-GFP-NC and PBS group, using analysis of variance and Student-Newman-Keuls analyses. Bcl-2, B-cell lymphoma 2; Bax, Bcl-2-associated X protein; Skp2, S-phase kinase-associated protein 2; CDX2, caudal type homeobox transcription factor 2; LV-GFP, lentivirus-green fluoresenct protein; PBS, phosphate-buffered saline.

Article Snippet: Specific rabbit anti-human polyclonal antibodies to CDX2 (#12306), c-Myc (#5605), Skp2 (#2652), Bax (#5023), Bcl-2 (#2827), cyclin D1 (#2978), survivin (#2808) and GAPDH (#2118) were provided by Cell Signaling Technology, Inc. (Beverly, MA, USA) .

Techniques: Over Expression, Expressing, Western Blot, Control, Saline

Journal: Life Science Alliance

Article Title: Laminin is the ECM niche for trophoblast stem cells

doi: 10.26508/lsa.201900515

Figure Lengend Snippet: (A) Expression of undifferentiated TSC marker CDX2 in trophectoderm cells in Lamc1 EQ/+ and Lamc1 EQ/EQ blastocysts. Bar, 50 μm. (B) Loss of integrin α7x2β1 binding to the basement membrane in E5.5 Lamc1 EQ/EQ embryos. Magenta, anti-laminin α1 antibody binding; green, recombinant integrin α7x2β1 binding; white, area double-positive for magenta and green signals. Bar, 50 μm. (C) Morphologies of E5.5 control and Lamc1 EQ/EQ embryos visualized by immunofluorescence. Cyan, laminin α1; magenta, CDX2; green, OCT4. ExE cells are enclosed by dotted lines. The arrow indicates CDX2-positive cells detached from the laminin-positive basement membrane. Bar, 50 μm. (D) Quantification of CDX2-positive and OCT4-positive cells in control WT and Lamc1 EQ/+ and Lamc1 EQ/EQ E5.5 egg cylinders. Data represent means ± SD ( n = 24 and 6 for WT or Lamc1 EQ/+ and Lamc1 EQ/EQ , respectively). *** P < 0.001, significant difference by Welch’s t test. (E) Diagram illustrating the dependence of TSCs on laminin–integrin interactions in the mouse conceptus. In addition to FGF4 and nodal from the epiblast, laminin also acts on TSCs as an ECM niche through binding to integrin receptors. The inset shows the region illustrated in the main figure. The diagram is based on that in .

Article Snippet: The following antibodies and reagents were obtained commercially: rat anti-laminin-γ1 mAb (Millipore); rabbit anti-laminin pAb, BSA, and heparin (Sigma-Aldrich); mouse anti-CDX2 mAb (Biocare); rabbit anti-OCT4 pAb (Santa Cruz Biotechnology); rat anti-integrin αV mAb (RMV-7), hamster anti-integrin β1 mAb (Ha2/5), and hamster anti-integrin β3 mAb (2C9.G2) (BD Biosciences); Alexa 488–conjugated goat antirabbit IgG, Alexa 546–conjugated goat antirat IgG, Alexa 405–conjugated goat antirabbit IgG, Alexa 488–conjugated goat antirat IgG, and Alexa 546–conjugated goat antimouse IgG (Invitrogen); bovine types I and IV collagens (Nippi Inc.); recombinant human FGF4 (Peprotech); and PermaFluor (Thermo Shandon).

Techniques: Expressing, Marker, Binding Assay, Membrane, Recombinant, Control, Immunofluorescence

Journal: Life Science Alliance

Article Title: Laminin is the ECM niche for trophoblast stem cells

doi: 10.26508/lsa.201900515

Figure Lengend Snippet: Morphologies of E5.5 control and Lamc1 EQ/EQ embryos. Cyan, laminin α1; magenta, CDX2; green, OCT4. ExE cells are enclosed by dotted lines. Bar, 50 μm.

Article Snippet: The following antibodies and reagents were obtained commercially: rat anti-laminin-γ1 mAb (Millipore); rabbit anti-laminin pAb, BSA, and heparin (Sigma-Aldrich); mouse anti-CDX2 mAb (Biocare); rabbit anti-OCT4 pAb (Santa Cruz Biotechnology); rat anti-integrin αV mAb (RMV-7), hamster anti-integrin β1 mAb (Ha2/5), and hamster anti-integrin β3 mAb (2C9.G2) (BD Biosciences); Alexa 488–conjugated goat antirabbit IgG, Alexa 546–conjugated goat antirat IgG, Alexa 405–conjugated goat antirabbit IgG, Alexa 488–conjugated goat antirat IgG, and Alexa 546–conjugated goat antimouse IgG (Invitrogen); bovine types I and IV collagens (Nippi Inc.); recombinant human FGF4 (Peprotech); and PermaFluor (Thermo Shandon).

Techniques: Control

Journal: Developmental Cell

Article Title: Maternal DNA Methylation Regulates Early Trophoblast Development

doi: 10.1016/j.devcel.2015.12.027

Figure Lengend Snippet: Scml2 Is Controlled by DNA Methylation and Affects SynT Formation and Cell Adhesion (A) RT-qPCR of Dnmt3a mKO EPCs confirms that Scml2 is controlled by oocyte methylation. (B) Methylation analysis by Sequenom MassARRAY in E7.5 male EPCs, confirming the DMR at an intragenic TSS of Scml2 . Each data point may include more than one CpG from the amplicon, as indicated on the x axis. (C) RT-qPCR analysis of TSCs grown in FGF+ (TSC conditions) or FGF− (differentiation conditions) medium for 6 days, with or without Scml2 overexpression. (D) Expression of Syna is reduced in Dnmt3a mKO EPCs, whereas markers of SynT-II Synb and Cebpa are unaffected. (E) E-cadherin staining of two independent Scml2 knockout clones from TKO TSCs shows a rescue of the morphological alterations seen in TKO TSCs. (F) Scml2 KO on TKO TSCs also rescues the defect in cell adhesion to cell culture wells in the absence of laminin. (G) RT-qPCR analysis of TKO Scml2 KO clones shows maintained expression of the TSC marker Cdx2 ; the expression of genes involved in cell adhesion is not rescued upon Scml2 deletion. Error bars represent SD. ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001; t test comparing WT and Dnmt3a mKO EPCs (A and D) or Scml2 -expressing TSCs versus vector control (C), or ANOVA with post hoc tests comparing Ctrl with DHet/DKO (B) or TKO TSC lines with WT TSCs. See also Figure S6 .

Article Snippet: Paraffin sections were deparaffinized with Histo-Clear and dehydrated through an ethanol series, followed either by standard H&E staining or antigen retrieval by boiling slides for 30 min (in 1 mM EDTA, 0.05% Tween 20, pH 8) and cooling at room temperature for 20 min. After blocking with 1% BSA overnight, sections were incubated with a rabbit monoclonal anti-CDX2 antibody (EPR2764Y, Novus Biologicals, 1:250 dilution) for 2 hr.

Techniques: DNA Methylation Assay, Quantitative RT-PCR, Methylation, Amplification, Over Expression, Expressing, Staining, Knock-Out, Clone Assay, Cell Culture, Marker, Plasmid Preparation, Control

Journal: Diagnostic Pathology

Article Title: Colon adenoma and adenocarcinoma with clear cell components - two case reports

doi: 10.1186/s13000-019-0819-z

Figure Lengend Snippet: Tubular adenoma with clear cell change. The striking tubule structures of the clear cells are accompanied by conventional tubular adenoma cells at low magnification with HE staining ( a ). The boundary between the clear cell and conventional components at high magnification with HE staining ( b ). The clear cell component is negative for PAS ( c ) and alcian blue staining ( d ). Both components are positive for CDX2 staining ( e ). The localization of CEA (f) expression is diffusely cytoplasmic for the clear cell component, and luminal cell apical for the conventional one. Ki67 labeling ( g ) is slightly lower in the clear cell component. d-g represent immunohistochemistry. Ultrastructural examination ( h ) of the boundary between the clear cell area (left) and the conventional adenoma (right) at low magnification is shown and multiple cytoplasmic lipid-like vacuoles surround the nuclei in the clear cells ( i )

Article Snippet: CDX2 , AMT28 , Novus Biologicals, Newcastle, UK , 1:50 , pH 9.0.

Techniques: Staining, Expressing, Labeling, Immunohistochemistry

Journal: Diagnostic Pathology

Article Title: Colon adenoma and adenocarcinoma with clear cell components - two case reports

doi: 10.1186/s13000-019-0819-z

Figure Lengend Snippet: Clear cell adenocarcinoma. Low magnification ( a ) and high magnification of the clear cell ( b ) and conventional components ( c ) with HE staining. The boundary ( d ) between the clear cell and conventional components at high magnification with HE staining. The clear cells are negative for PAS ( e ) and alcian blue staining ( f ), whereas both components of the tumor are positive for CDX2 ( g ). The localization of CEA ( h ) expression is diffusely cytoplasmic for the clear cell component and luminal cell apical for the conventional component. CD10 ( i ) and adipophilin ( j ) expression is confined to the clear component and Ki67 labeling ( k ) is higher in the clear cell component. g-k represent immunohistochemistry. Immunoelectron microscopy analysis ( l ) at low (left) and high magnification (right) reveals multiple cytoplasmic lipid-like vacuoles in clear cells that are negative for adipophilin

Article Snippet: CDX2 , AMT28 , Novus Biologicals, Newcastle, UK , 1:50 , pH 9.0.

Techniques: Staining, Expressing, Labeling, Immunohistochemistry, Immuno-Electron Microscopy

Journal: Diagnostic Pathology

Article Title: Colon adenoma and adenocarcinoma with clear cell components - two case reports

doi: 10.1186/s13000-019-0819-z

Figure Lengend Snippet: Antibody information

Article Snippet: CDX2 , AMT28 , Novus Biologicals, Newcastle, UK , 1:50 , pH 9.0.

Techniques:

Journal: PLoS ONE

Article Title: Dynamics of DNA Methylation during Early Development of the Preimplantation Bovine Embryo

doi: 10.1371/journal.pone.0066230

Figure Lengend Snippet: Nuclei of all cells were labeled with propidium iodide (red). Panels represent merged images for all three fluorescent labels (A), 5-methylcytosine and PI (B), CDX2 and PI (C), CDX2 and 5-methylcytosine (D) and control antibodies for anti-5methylcytosine and anti-CDX2 as well as PI (E). The scale bar is 20 µm.

Article Snippet: Embryos were labeled for anti-methylcytosine as described above, washed six times and transferred to a solution of antibody against CDX2 (affinity purified mouse monoclonal antibody against CDX-2, ready to use solution; BioGenex, San Ramon, CA, USA) for 1 h. Embryos were washed three times and transferred to 10 μg/ml Alexa Fluor 350 labeled goat anti-mouse IgG diluted in DPBS containing 0.05% (v/v) Tween 20 and 0.01% (w/v) BSA for 1 h. Embryos were then washed, labeled with PI, mounted on slides and examined as described above.

Techniques: Labeling, Control