human ceacam7  (R&D Systems)

Journal: Frontiers in Immunology

Article Title: Regulation of CEACAM Family Members by IBD-Associated Triggers in Intestinal Epithelial Cells, Their Correlation to Inflammation and Relevance to IBD Pathogenesis

doi: 10.3389/fimmu.2021.655960

Figure Lengend Snippet: CEACAMs gene expression in pediatric colonic mucosa of patients with Ulcerative colitis (UC) and Crohn’s disease (CD) and controls. Gene expression of (A) CEACAM1, (B) CEACAM3, (C) CEACAM5, (D) CEACAM6, (E) CEACAM7, (F) IL-8 as determined by RT-qPCR. (G) Spearman correlation on relative mRNA levels between IL-8 and CEACAM3 and CEACAM6 in pediatric colonic CD samples and of (H) IL-8 and CEACAM1 in pediatric colonic UC. n=9, Ulcerative Colitis (UC); n=12 Crohn’s Disease (CD); and n=6 controls. An ANOVA test with post-hoc analysis was used to analyze gene expression data and Spearman correlation test. *P < 0.05, **P < 0.01.

Article Snippet: Samples were run on Bolt 4–12% gradient Bis-Tris Plus Gels (cat nr. NW04122Box, Thermo Fisher Scientific) at 120 V for 1 h and transferred to a Immobilon PVDF membrane (Millipore) for 1 h. Membranes were washed in Tris Buffer Saline (TBS) and blocked for 1 h in 5% skimmed milk (0.5% Tween20 in TBS), followed by incubation overnight and rocking at 4°C with the primary antibodies [human CEACAM6/CD66c (Mouse IgG2A, Clone #439424, cat nr. MAB3934-SP, R&D-Biotechne); human CEACAM7 (Sheep IgG, cat nr. AF4478, R&D-Biotechne); β-actin (Mouse IgG2b, cat nr. 8H10D10, Cell Signaling Technology], diluted at 1:1,000 in 5% BSA/TBS-T.

Techniques: Expressing, Quantitative RT-PCR

Journal: Frontiers in Immunology

Article Title: Regulation of CEACAM Family Members by IBD-Associated Triggers in Intestinal Epithelial Cells, Their Correlation to Inflammation and Relevance to IBD Pathogenesis

doi: 10.3389/fimmu.2021.655960

Figure Lengend Snippet: CEACAMs gene expression in adult colonic mucosa of patients with Ulcerative colitis (UC) and Crohn’s disease (CD) with active and inactive disease and controls. Gene expression of (A) CEACAM1, (B) CEACAM3, (C) CEACAM5, (D) CEACAM6, (E) CEACAM7, (F) IL-8 as determined by RT q-PCR. (G) Spearman correlation on relative mRNA levels between IL-8 and CEACAM1 in adult CD Active samples and of (H) IL-8 and CEACAM7 in adult colonic UC. n=10–18, active Ulcerative Colitis (UC); n=10–19, inactive UC; n=7–11 active Crohn’s Disease (CD); n=10 inactive Crohn’s Disease and n=6–14 controls. An ANOVA test with post-hoc analysis was used to analyze gene expression data and Spearman correlation test. *P < 0.05, **P < 0.01, ***P < 0.001.

Article Snippet: Samples were run on Bolt 4–12% gradient Bis-Tris Plus Gels (cat nr. NW04122Box, Thermo Fisher Scientific) at 120 V for 1 h and transferred to a Immobilon PVDF membrane (Millipore) for 1 h. Membranes were washed in Tris Buffer Saline (TBS) and blocked for 1 h in 5% skimmed milk (0.5% Tween20 in TBS), followed by incubation overnight and rocking at 4°C with the primary antibodies [human CEACAM6/CD66c (Mouse IgG2A, Clone #439424, cat nr. MAB3934-SP, R&D-Biotechne); human CEACAM7 (Sheep IgG, cat nr. AF4478, R&D-Biotechne); β-actin (Mouse IgG2b, cat nr. 8H10D10, Cell Signaling Technology], diluted at 1:1,000 in 5% BSA/TBS-T.

Techniques: Expressing

Journal: Frontiers in Immunology

Article Title: Regulation of CEACAM Family Members by IBD-Associated Triggers in Intestinal Epithelial Cells, Their Correlation to Inflammation and Relevance to IBD Pathogenesis

doi: 10.3389/fimmu.2021.655960

Figure Lengend Snippet: Relative gene expression profile in HT29 cells treated with 0.25% polysorbate 80 (p80) for 1, 2, 4, and 6 h for analysis on gene expression of (A) CEACAM1, (B) CEACAM3, (C) CEACAM5, (D) CEACAM6, (E) CEACAM7, (F) IL-8 as determined by RT-qPCR. Graphs are representative of two to four independent experiments with pooled triplicate samples and qRT-PCR assay performed in duplicates. β-actin was used as a housekeeping gene, and results are expressed as fold change. A Student t-test was used to analyze gene expression data. *P < 0.05, **P < 0.01 when compared to untreated cells.

Article Snippet: Samples were run on Bolt 4–12% gradient Bis-Tris Plus Gels (cat nr. NW04122Box, Thermo Fisher Scientific) at 120 V for 1 h and transferred to a Immobilon PVDF membrane (Millipore) for 1 h. Membranes were washed in Tris Buffer Saline (TBS) and blocked for 1 h in 5% skimmed milk (0.5% Tween20 in TBS), followed by incubation overnight and rocking at 4°C with the primary antibodies [human CEACAM6/CD66c (Mouse IgG2A, Clone #439424, cat nr. MAB3934-SP, R&D-Biotechne); human CEACAM7 (Sheep IgG, cat nr. AF4478, R&D-Biotechne); β-actin (Mouse IgG2b, cat nr. 8H10D10, Cell Signaling Technology], diluted at 1:1,000 in 5% BSA/TBS-T.

Techniques: Expressing, Quantitative RT-PCR

Journal: Frontiers in Immunology

Article Title: Regulation of CEACAM Family Members by IBD-Associated Triggers in Intestinal Epithelial Cells, Their Correlation to Inflammation and Relevance to IBD Pathogenesis

doi: 10.3389/fimmu.2021.655960

Figure Lengend Snippet: Relative gene expression profile in C2BBe1 cells co-cultured for 3 h with whole bacteria— Escherichia coli K12 (K12 E. coli ), Adhesion and Invasive E. coli (AIEC)-HM605, Salmonella typhimurium ( S. typh ) at a MOI 10:1 and untreated. After the 3 h co-culture, cells are washed in medium containing antibiotics followed by further culture in full culture media for 2 (T2), 5 (T5), and 13 (T13) h. Samples are collected at each time point for analysis on gene expression of (A) CEACAM1, (B) CEACAM3, (C) CEACAM5, (D) CEACAM6, (E) CEACAM7, (F) IL-8 as determined by RT q-PCR. Graphs are representative of two to four independent experiments with pooled triplicate samples and qRT-PCR assay performed in duplicates. β-actin was used as a housekeeping gene, and results are expressed as fold change. A Student t-test was used to analyze gene expression data. *P < 0.05, **P < 0.01, ***P < 0.001 when compared to untreated cells.

Article Snippet: Samples were run on Bolt 4–12% gradient Bis-Tris Plus Gels (cat nr. NW04122Box, Thermo Fisher Scientific) at 120 V for 1 h and transferred to a Immobilon PVDF membrane (Millipore) for 1 h. Membranes were washed in Tris Buffer Saline (TBS) and blocked for 1 h in 5% skimmed milk (0.5% Tween20 in TBS), followed by incubation overnight and rocking at 4°C with the primary antibodies [human CEACAM6/CD66c (Mouse IgG2A, Clone #439424, cat nr. MAB3934-SP, R&D-Biotechne); human CEACAM7 (Sheep IgG, cat nr. AF4478, R&D-Biotechne); β-actin (Mouse IgG2b, cat nr. 8H10D10, Cell Signaling Technology], diluted at 1:1,000 in 5% BSA/TBS-T.

Techniques: Expressing, Cell Culture, Bacteria, Co-Culture Assay, Quantitative RT-PCR

Journal: Frontiers in Immunology

Article Title: Regulation of CEACAM Family Members by IBD-Associated Triggers in Intestinal Epithelial Cells, Their Correlation to Inflammation and Relevance to IBD Pathogenesis

doi: 10.3389/fimmu.2021.655960

Figure Lengend Snippet: CEACAM expression in intestinal epithelial cells cultured with bacterial conditioned medium with bacterial conditioned medium and SCFA.

Article Snippet: Samples were run on Bolt 4–12% gradient Bis-Tris Plus Gels (cat nr. NW04122Box, Thermo Fisher Scientific) at 120 V for 1 h and transferred to a Immobilon PVDF membrane (Millipore) for 1 h. Membranes were washed in Tris Buffer Saline (TBS) and blocked for 1 h in 5% skimmed milk (0.5% Tween20 in TBS), followed by incubation overnight and rocking at 4°C with the primary antibodies [human CEACAM6/CD66c (Mouse IgG2A, Clone #439424, cat nr. MAB3934-SP, R&D-Biotechne); human CEACAM7 (Sheep IgG, cat nr. AF4478, R&D-Biotechne); β-actin (Mouse IgG2b, cat nr. 8H10D10, Cell Signaling Technology], diluted at 1:1,000 in 5% BSA/TBS-T.

Techniques: Expressing, Cell Culture

Journal: Frontiers in Immunology

Article Title: Regulation of CEACAM Family Members by IBD-Associated Triggers in Intestinal Epithelial Cells, Their Correlation to Inflammation and Relevance to IBD Pathogenesis

doi: 10.3389/fimmu.2021.655960

Figure Lengend Snippet: C2BBe1 cells cultured for 3 h with the pro-inflammatory cytokine cocktail containing recombinant human (rh) TNF-α (20 ng/ml), rh IFN-γ (10 ng/ml), rh IL-1β (10 ng/ml), and untreated. After the 3 h co-culture (T0), cells are washed followed by further culture in full culture media for 2 (T2), 5 (T5), and 13 (T13) h. Samples are collected at T0, T2, T5, and T13 time points for analysis on gene and protein expression of (A, B) CEACAM1, (C, D) CEACAM3, (E, F) CEACAM5, (G, H) CEACAM6, (H, I) CEACAM7, and (J) IL-8 as determined by RT q-PCR, flow cytometry (CEACAM1, -3, -5), WB (CEACAM6 and 7). For gene expression analysis, graphs are representative of three to five independent experiments with pooled triplicate samples and RT-qPCR assay performed in duplicates. β-actin was used as a housekeeping gene, and results are expressed as fold change. For protein analysis with flow cytometry and WB, graphs are representative of two independent experiments with pooled duplicates/triplicates. In the flow cytometry graphs (B, D, F) white colums represent untreated samples and black columns represent cytokine cocktail treated samples collected at T5. A Student t-test was used to analyze gene and protein data. *P < 0.05, **P < 0.01, ***P < 0.001 when compared to untreated cells. UT, untreated; CK, cytokine cocktail treatment.

Article Snippet: Samples were run on Bolt 4–12% gradient Bis-Tris Plus Gels (cat nr. NW04122Box, Thermo Fisher Scientific) at 120 V for 1 h and transferred to a Immobilon PVDF membrane (Millipore) for 1 h. Membranes were washed in Tris Buffer Saline (TBS) and blocked for 1 h in 5% skimmed milk (0.5% Tween20 in TBS), followed by incubation overnight and rocking at 4°C with the primary antibodies [human CEACAM6/CD66c (Mouse IgG2A, Clone #439424, cat nr. MAB3934-SP, R&D-Biotechne); human CEACAM7 (Sheep IgG, cat nr. AF4478, R&D-Biotechne); β-actin (Mouse IgG2b, cat nr. 8H10D10, Cell Signaling Technology], diluted at 1:1,000 in 5% BSA/TBS-T.

Techniques: Cell Culture, Recombinant, Co-Culture Assay, Expressing, Flow Cytometry, Quantitative RT-PCR

Journal: Frontiers in Immunology

Article Title: Regulation of CEACAM Family Members by IBD-Associated Triggers in Intestinal Epithelial Cells, Their Correlation to Inflammation and Relevance to IBD Pathogenesis

doi: 10.3389/fimmu.2021.655960

Figure Lengend Snippet: C2BBe1 cells were pretreated for 1 h with the IBD drugs—Tofacitinib (Tofa, 10 mM), Methylprednisolone (MetPred, 10 mM), Mercaptopurine (6-MP, 5 mM)—followed by culture for 3 h with the pro-inflammatory cytokine cocktail (Cytokine Mix), containing recombinant human (rh) TNF-α (20 ng/ml), rh IFN-γ (10 ng/ml), rh IL-1β (10 ng/ml), and untreated. After the 3 h co-culture, cells are washed in medium followed by further culture in full culture media containing respective IBD-drug for 2 (T2) and 13 (T13) h. Control wells (DMSO Control) are treated with 0.1% DMSO. Samples are collected at T2 and T13 time points for analysis on gene expression of (A) CEACAM1, (B) CEACAM3, (C) CEACAM5, (D) CEACAM6, (E) CEACAM7 as determined by RT q-PCR and (F) IL-8 determined by ELISA. Graphs are representative of two independent experiments with pooled triplicate samples and qRT-PCR assay performed in duplicates. β-actin was used as a housekeeping gene, and results are expressed as fold change. A Student t-test was used to analyze gene expression data. *P < 0.05, **P < 0.01, ***P < 0.001 when compared outlined treatment.

Article Snippet: Samples were run on Bolt 4–12% gradient Bis-Tris Plus Gels (cat nr. NW04122Box, Thermo Fisher Scientific) at 120 V for 1 h and transferred to a Immobilon PVDF membrane (Millipore) for 1 h. Membranes were washed in Tris Buffer Saline (TBS) and blocked for 1 h in 5% skimmed milk (0.5% Tween20 in TBS), followed by incubation overnight and rocking at 4°C with the primary antibodies [human CEACAM6/CD66c (Mouse IgG2A, Clone #439424, cat nr. MAB3934-SP, R&D-Biotechne); human CEACAM7 (Sheep IgG, cat nr. AF4478, R&D-Biotechne); β-actin (Mouse IgG2b, cat nr. 8H10D10, Cell Signaling Technology], diluted at 1:1,000 in 5% BSA/TBS-T.

Techniques: Recombinant, Co-Culture Assay, Expressing, Enzyme-linked Immunosorbent Assay, Quantitative RT-PCR

Journal: Journal of advanced research

Article Title: CEACAM7 expression contributes to early events of pancreatic cancer.

doi: 10.1016/j.jare.2023.02.013

Figure Lengend Snippet: Fig. 1. Expression profiling of CEACAM7 in normal and cancerous tissues: (a) CEACAM7 in normal tissues: CEACAM7 is cell adhesion molecule 7 (Gene ID: 1087). According to BioProject (PRJEB4337) entitled ‘‘HPA RNA-seq normal tissues”, the expression level of CEACAM7 at normal condition in various tissue of human was showcased. This data was retrieved from the NCBI BioProject database. In this BioProject database, RNA-seq analysis was performed on normal tissue samples from 95 human individuals. This expression profiling clearly depicted that CEACAM7 does not express in most of the tissue in normal condition expect colon tissue. (b) Pan cancer gene expression of CEACAM7: The gene expression profile across all tumor samples and paired normal tissues is indicated in the dot plot. Each dot represents the expression of samples. The expression data was scaled by log2(TPM + 1). Red: Probable high tumor expression of CEACAM7; Green: Normal cell expression of CEACAM7. Source: GEPIA. (For interpretation of the references to color in this figure legend, the reader is referred to the web version of this article.)

Article Snippet: To quantify the protein concentration of CECAM7 in various PDAC cell lines, commercially available R&D System, Human CEACAM7 DuoSet ELISA kit (Cat. No. DY4478-05, Lot: P187793) was used as per the manufacturer’s protocol.

Techniques: Expressing, RNA Sequencing, Gene Expression

Journal: Journal of advanced research

Article Title: CEACAM7 expression contributes to early events of pancreatic cancer.

doi: 10.1016/j.jare.2023.02.013

Figure Lengend Snippet: Fig. 2. (a) Differential gene expression box plot analysis of CEACAM7: GEPIA box plot analysis to observe significantly higher mRNA expression of CEACAM7 in PAAD followed by STAD. The red colored box is tumor tissue dataset and blue colored box is normal tissue dataset. Red * denoted significant data. The method for differential analysis is a one tailed unpaired student t-test, using disease state (tumor or normal) as a variable for calculating differential expression. The level of significance is p-value cutoff 0.01. (b) Protein expression level of CEACAM7: CPTAC analysis to observe mass-spectrometry-based proteomic characterization of CEACAM7 in various cancers. The statistics of each cancer indication are based on the Z-values, which denote standard deviations (SD) from the median value of all samples. The statistical significance p-value of normal vs Pancreatic adenocarcinoma (PAAD): 3.47e-04, normal vs colon cancer: 6.34e-18, normal vs Uterine corpus endometrial carcinoma (UCEC): 6.99e-02, normal vs Head and neck carcinoma (H&N Cancer): 4.03e-14. (For interpretation of the references to color in this figure legend, the reader is referred to the web version of this article.)

Article Snippet: To quantify the protein concentration of CECAM7 in various PDAC cell lines, commercially available R&D System, Human CEACAM7 DuoSet ELISA kit (Cat. No. DY4478-05, Lot: P187793) was used as per the manufacturer’s protocol.

Techniques: Gene Expression, Expressing, One-tailed Test, Quantitative Proteomics, Mass Spectrometry

Journal: Journal of advanced research

Article Title: CEACAM7 expression contributes to early events of pancreatic cancer.

doi: 10.1016/j.jare.2023.02.013

Figure Lengend Snippet: Fig. 3. Patients’ survival and gene correlation plots of CEACAM7: (a) This survival plot was generated by Kaplan-Meier Plotter (Hazard Ratio (HR): 1.56, Logrank p = 0.036, total no. of patients: 261); the cutoff value that was used in the analysis is 743 (ranging from 154 to 1426). This analysis was done based on mRNA expression. (b) Correlation graph of CEACAM7 and S100A4, which demonstrates the strong positive association of both of genes in pancreatic cancer. (c) Comparison of z-score gene-based expression of CEACAM7 and S100A4in pancreatic cancer.

Article Snippet: To quantify the protein concentration of CECAM7 in various PDAC cell lines, commercially available R&D System, Human CEACAM7 DuoSet ELISA kit (Cat. No. DY4478-05, Lot: P187793) was used as per the manufacturer’s protocol.

Techniques: Generated, Expressing, Comparison, Gene Expression

Journal: Journal of advanced research

Article Title: CEACAM7 expression contributes to early events of pancreatic cancer.

doi: 10.1016/j.jare.2023.02.013

Figure Lengend Snippet: Fig. 5. CEACAM7 positioning in pancreas: RNA expression of CEACAM7 in pancreas tissues. The single cell type clusters identified in this tissue were visualized by a UMAP plot.

Article Snippet: To quantify the protein concentration of CECAM7 in various PDAC cell lines, commercially available R&D System, Human CEACAM7 DuoSet ELISA kit (Cat. No. DY4478-05, Lot: P187793) was used as per the manufacturer’s protocol.

Techniques: RNA Expression

Journal: Journal of advanced research

Article Title: CEACAM7 expression contributes to early events of pancreatic cancer.

doi: 10.1016/j.jare.2023.02.013

Figure Lengend Snippet: Fig. 4. Volcano graph and pathways enrichment analyses of CEACAM7 and associated genes: (a) Co-expression genes of CEACAM7 in pancreatic cancers (LinkedOmics: LinkFinder). The significantly correlated genes with CEACAM7 were analyzed by Spearman test. Green and red dots represented the negative and positive correlations with CEACAM7, respectively. (b) Pathway enrichment analysis of CEACAM7 and associated genes by using LinkInterpreter. Blue and orange bar graphs represent pathway enrichment of positively and negatively associated genes with CEACAM7, respectively. (For interpretation of the references to color in this figure legend, the reader is referred to the web version of this article.)

Article Snippet: To quantify the protein concentration of CECAM7 in various PDAC cell lines, commercially available R&D System, Human CEACAM7 DuoSet ELISA kit (Cat. No. DY4478-05, Lot: P187793) was used as per the manufacturer’s protocol.

Techniques: Expressing

Journal: Journal of advanced research

Article Title: CEACAM7 expression contributes to early events of pancreatic cancer.

doi: 10.1016/j.jare.2023.02.013

Figure Lengend Snippet: Fig. 6. Transcriptomic and translational expression of CEACAM7: (a) mRNA expression analysis: Gene expression of CEACAM7 in progressive PDAC cell lines with respect to HPNE, a normal pancreatic cell line. n = 3; p-value *<= 0.05 and ***<=0.001. (b) Protein expression: Quantification of CEACAM7 protein expression in progressive PDAC cell lines was demonstrated using ELISA. n = 3; p-value *<= 0.05, ** <=0.01 and *** <=0.001. (c) Representative image of western blot recognition pattern of CEACAM7 in lysates PDAC cell lines (n = 3).

Article Snippet: To quantify the protein concentration of CECAM7 in various PDAC cell lines, commercially available R&D System, Human CEACAM7 DuoSet ELISA kit (Cat. No. DY4478-05, Lot: P187793) was used as per the manufacturer’s protocol.

Techniques: Expressing, Gene Expression, Enzyme-linked Immunosorbent Assay, Western Blot

Journal: Journal of advanced research

Article Title: CEACAM7 expression contributes to early events of pancreatic cancer.

doi: 10.1016/j.jare.2023.02.013

Figure Lengend Snippet: Fig. 7. Confocal microscopy of CEACAM7 in normal pancreas and PDAC cell line panels: Protein expression analysis of CEACAM7 in progressive PDAC cell lines with respect to HPNE, a normal pancreatic cell line. Cells were processed for CEACAM7 immunostaining and confocal microscopy. Original Magnifications 40.

Article Snippet: To quantify the protein concentration of CECAM7 in various PDAC cell lines, commercially available R&D System, Human CEACAM7 DuoSet ELISA kit (Cat. No. DY4478-05, Lot: P187793) was used as per the manufacturer’s protocol.

Techniques: Confocal Microscopy, Expressing, Immunostaining

Journal: Journal of advanced research

Article Title: CEACAM7 expression contributes to early events of pancreatic cancer.

doi: 10.1016/j.jare.2023.02.013

Figure Lengend Snippet: Fig. 8. IHC staining of commercially available Tumor Micro Arrays (TMAs) and its Mean Composite Scoring (MSC): Representative images of the tissues stained for CEACAM7 expression, depicted in color brown. Figure a, b, c, & d were 40 zoomed images of inset images (5–7.5 magnification full core of images) by using Case Viewer 2.4 software (3DHistech Ltd., Budapest, Hungary). In this IHC image (a) normal tissue with no staining; (b) well differentiated (high coverage and intense staining is present in duct); (c) moderately (high coverage but less intense staining is present in duct); and (d) poorly (low coverage and low intense staining is present in deformed duct) are the differentiated cores of PDAC samples, respectively. (e) Mean Composite Scoring of CEACAM7: Bar diagram of Mean Composite Score (MSC) of CEACAM7 expressions in various stages of pancreatitis, PDAC, and normal tissue cores.

Article Snippet: To quantify the protein concentration of CECAM7 in various PDAC cell lines, commercially available R&D System, Human CEACAM7 DuoSet ELISA kit (Cat. No. DY4478-05, Lot: P187793) was used as per the manufacturer’s protocol.

Techniques: Immunohistochemistry, Staining, Expressing, Software