Journal: Physiological reports

Article Title: Expression of human carcinoembryonic antigen-related cell adhesion molecule 6 and alveolar progenitor cells in normal and injured lungs of transgenic mice.

doi: 10.14814/phy2.12657

Figure Lengend Snippet: Figure 1. Representative Western blot analyses of lung CEACAM6 in adult CEABAC mice compared to human infants. (A) Whole lung homogenate. 10 lg protein was loaded for each sample. Bands are observed at approximately 90, 70, and 50 kDa for most CEABAC and term human infant samples but not in wt mice. CEACAM6 in CEABAC mice consistently ran at slightly higher molecular weight than human lung CEACAM6, probably reflecting different amount of glycosylation. By scanning densitometry, total CEACAM6 signal is similar for CEABAC mice and human lungs. (B) Large aggregate surfactant fraction of BAL. 2 lg protein was loaded for mouse samples and 0.1 lg for a sample from an intubated premature human infant. A single 90 kDa band is observed for CEABAC samples and human surfactant but not wt mouse samples. (C) Supernatant of samples from B. 5 lg protein was loaded for mouse samples and 1 lg for the human. CEACAM6 was detected in the human specimen as previously reported (Chapin et al. 2012) but not in mouse samples.

Article Snippet: All blots were run with recombinant human CEACAM6 (R & D Systems Inc., Minneapolis, MN) and adult human lung homogenate as internal controls.

Techniques: Western Blot, Molecular Weight, Glycoproteomics

Journal: Physiological reports

Article Title: Expression of human carcinoembryonic antigen-related cell adhesion molecule 6 and alveolar progenitor cells in normal and injured lungs of transgenic mice.

doi: 10.14814/phy2.12657

Figure Lengend Snippet: Figure 2. Immunohistochemistry of CEACAM6 in cryosections of lung from adult wt and CEABAC mice. (A) Immunostaining of wt mouse lung. OTS-8 red staining shows alveolar type I cells, blue nuclei (DAPI) and absence of CEACAM6 (green). (b) Corresponding phase contrast image. (C) OTS-8 and CEACAM6 immunostaining of CEABAC lung. Relatively low intensity CEACAM6 staining (green) is observed in some alveoli and does not appear to co-localize with OTS-8 as a marker of type I cells. (D) Corresponding phase contrast image. (E) SP-B and CEACAM6 immunostaining (red) of CEABAC lung. Low intensity CEACAM6 staining (green) is observed in some alveoli and does not appear to co-localize with SP-B (red) as a marker of type II cells. (F) Corresponding phase contrast image. Insets in c and d show ~ twofold magnified views of CEACAM6+ alveolar regions (arrows). Images are representative of multiple sections from lungs examined. Bar = 20 lm.

Article Snippet: All blots were run with recombinant human CEACAM6 (R & D Systems Inc., Minneapolis, MN) and adult human lung homogenate as internal controls.

Techniques: Immunohistochemistry, Immunostaining, Staining, Marker

Journal: Physiological reports

Article Title: Expression of human carcinoembryonic antigen-related cell adhesion molecule 6 and alveolar progenitor cells in normal and injured lungs of transgenic mice.

doi: 10.14814/phy2.12657

Figure Lengend Snippet: Figure 4. Effect of bleomycin and saline on CEACAM6 content in lung tissue and lavage surfactant at 10 days. (A) Representative Western blots of lung homogenate. Equal amounts of total protein were loaded. Compared to No Treatment and Saline, increased total CEACAM6 signal is observed for lungs of bleomycin-treated animals. (B) CEACAM6 content of homogenate by scanning densitometry (mean SD). (C) Representative immunodot blot for large aggregate surfactant (with duplicate rows containing serial dilutions of equal amounts of PL/well). Increased signal is observed for bleomycin versus saline that is most evident at higher dilutions. (D) CEACAM6 content of surfactant by scanning densitometry. Data are mean SE, and n = 5; *P < 0.05 versus saline treatment.

Article Snippet: All blots were run with recombinant human CEACAM6 (R & D Systems Inc., Minneapolis, MN) and adult human lung homogenate as internal controls.

Techniques: Saline, Western Blot

Journal: Physiological reports

Article Title: Expression of human carcinoembryonic antigen-related cell adhesion molecule 6 and alveolar progenitor cells in normal and injured lungs of transgenic mice.

doi: 10.14814/phy2.12657

Figure Lengend Snippet: Figure 5. Immunohistochemistry for CEACAM6 expression in cryosections of lung from adult CEABAC mice treated with saline (A–C) or bleomycin (D–F) for 10 days. (A) OTS-8 staining in saline-instilled lung. (B) punctate CEACAM6 staining (green, arrow) of one cell that is immunopositive for OTS-8. (C) phase contrast image corresponding to A,B. (D) OTS-8 staining in bleomycin-instilled lung. (E) CEACAM6 staining of multiple cells that are either positive (e.g., arrows) or negative (e.g., arrowheads) for OTS-8. (F) phase contrast image corresponding to d and e; note thickened interstitium compared to saline (C), reflecting bleomycin injury. Bar = 20 lm. All fluorescence images were obtained at the same exposure settings.

Article Snippet: All blots were run with recombinant human CEACAM6 (R & D Systems Inc., Minneapolis, MN) and adult human lung homogenate as internal controls.

Techniques: Immunohistochemistry, Expressing, Saline, Staining

Journal: Physiological reports

Article Title: Expression of human carcinoembryonic antigen-related cell adhesion molecule 6 and alveolar progenitor cells in normal and injured lungs of transgenic mice.

doi: 10.14814/phy2.12657

Figure Lengend Snippet: Figure 6. CEACAM6 staining score for lungs after bleomycin instillation. A, score for cryosections of uninjured lungs (day 0) and at days 5–19 after bleomycin instillation (mean SE for 10 fields on each of 2 sections for lungs of 8 mice). (B) representative images for scores of 2, 3 and 4. The score is a composite of staining intensity and number of positive cells per field by a blinded observer. Maximal signal occurs by day 11; *P < 0.05 versus day 0. Results from a second experiment were similar.

Article Snippet: All blots were run with recombinant human CEACAM6 (R & D Systems Inc., Minneapolis, MN) and adult human lung homogenate as internal controls.

Techniques: Staining

Journal: Physiological reports

Article Title: Expression of human carcinoembryonic antigen-related cell adhesion molecule 6 and alveolar progenitor cells in normal and injured lungs of transgenic mice.

doi: 10.14814/phy2.12657

Figure Lengend Snippet: Figure 7. Increased CEACAM6 immunostaining in epithelial cells after bleomycin instillation. Panel A, A–C, co-localization of CEACAM6 and AQP5 in alveolar epithelium. By confocal imaging, some alveolar cells co-stain (yellow) for CEACAM6 (green) and AQP5 (red, e.g., arrows) and other CEACAM6+ cells are negative for AQP5 (e.g., arrow heads). Bar = 50 lm. Panel B, CEACAM6 and CC10 staining in airways of control (A,C) and bleomycin-treated (B,D) CEABAC mice. Most cells of control animals are CC10-positive with sparse CEACAM6 signal (A and C— higher power). After bleomycin (B,D) co-localization of CEACAM6 and CC10 is observed in airway epithelial cells (arrows) and CEACAM6 signal occurs basal to the epithelium. Bars = (A) 40 lm; (B) 100 lm; (C) and (D) 40 lm.

Article Snippet: All blots were run with recombinant human CEACAM6 (R & D Systems Inc., Minneapolis, MN) and adult human lung homogenate as internal controls.

Techniques: Immunostaining, Imaging, Staining, Control

Journal: Physiological reports

Article Title: Expression of human carcinoembryonic antigen-related cell adhesion molecule 6 and alveolar progenitor cells in normal and injured lungs of transgenic mice.

doi: 10.14814/phy2.12657

Figure Lengend Snippet: Figure 8. CEACAM6 and EGFP fluorescence signal in control and bleomycin-treated CEABAC/CBG mice. In control animals (A–D), cuboidal type II cells (green) are evident along with 2 CEACAM6+ (red) cells, with one example of co-localization (arrow). In bleomycin-treated mice (E–H), there is increased CEACAM6 signal (red) in injured areas and altered cell shape: F versus B, shape of many EGFP+ cells is flattened after bleomycin (arrowheads) with one cell showing apical CEACAM6 and cytoplasmic EGFP signal (G—arrow and inset); Phase micrographs are shown in D and H. Bar = 40 lm. * alveolus with EGFP+ epithelium.

Article Snippet: All blots were run with recombinant human CEACAM6 (R & D Systems Inc., Minneapolis, MN) and adult human lung homogenate as internal controls.

Techniques: Control

Journal: Physiological reports

Article Title: Expression of human carcinoembryonic antigen-related cell adhesion molecule 6 and alveolar progenitor cells in normal and injured lungs of transgenic mice.

doi: 10.14814/phy2.12657

Figure Lengend Snippet: Figure 9. Representative scatter plots for sorting of lung cells. (A) Cells isolated from a bleomycin-treated CEABAC mouse. In this 2-way sort of live/single cells, 0.16% were CEACAM6+ and negative for CD11b, a lymphoid marker (box). 100,000 cells were recorded. B, Cells isolated from bleomycin-treated CEABAC/CBG mice. A 2-way sort provided populations of EGFP+ cells that were negative (square) or positive (ellipse) for CEACAM6. 1,000,000 cells were recorded.

Article Snippet: All blots were run with recombinant human CEACAM6 (R & D Systems Inc., Minneapolis, MN) and adult human lung homogenate as internal controls.

Techniques: Isolation, Marker

Journal: Physiological reports

Article Title: Expression of human carcinoembryonic antigen-related cell adhesion molecule 6 and alveolar progenitor cells in normal and injured lungs of transgenic mice.

doi: 10.14814/phy2.12657

Figure Lengend Snippet: Figure 10. Lung structure and CEACAM6 expression in mice exposed to hyperoxia. Representative images of newborn CEABAC mice exposed to room air (A,C) or 80% oxygen (B,D) for 7 days. Note the simplified septal morphology and increased airspace size in mice exposed to hyperoxia (B vs. A, bar = 10 lm). C versus D, CEACAM6 and AQP5 staining. In mice exposed to hyperoxia there was a 1.8-fold increase in CEACAM6+ cell (green, arrows and arrowheads) compared to normoxic mice. There were similar numbers of cells that expressed both CEACAM6 and AQP5 (arrows) in each group. Bar = 40 lm

Article Snippet: All blots were run with recombinant human CEACAM6 (R & D Systems Inc., Minneapolis, MN) and adult human lung homogenate as internal controls.

Techniques: Expressing, Staining

Journal: bioRxiv

Article Title: L-DOS47 enhances response to immunotherapy in pancreatic cancer tumor

doi: 10.1101/2023.08.28.555194

Figure Lengend Snippet: A . Percentage of CEACAM6-expressing cells analyzed by flow cytometry in KPC961 parental and KPC961 clone 1B6 cells. B . Ratio of extracellular acidification (ECAR) and oxygen consumption rate (OCR) profiles in KPC961 parental and clone 1B6 cells; C . Lactate production in KPC961 parental and clone 1B6 cells. D . Tumor growth as a function of inoculum size. KPC961-CEACAM6 cells were inoculated into the pancreas of B6.129 mice at 100000, 50000, and 20000 cells, and tumor volumes were measured by ultrasound. Ascites fluid was observed in two mice inoculated with 100,000 cells on days 24 and 29, two mice inoculated with 50,000 cells at day 29 and one mouse inoculated with 20,000 cells at day 27. E . Expression of CEACAM6, PD-L1 and H2Kb in KPC961-1B6 cells of inoculated orthotopic tumors by flow cytometry.

Article Snippet: Cells were labeled with the following antibodies:(CEACAM6-FITC at 10 μg/mL, Sino Biological 10823-R408R; H2Kb-Pac Blue at 0.5 mg/mL, BioLegend 116514; CD45-BV605 at 0.2 mg/mL, BioLegend 103155; and PD-L1 – PE at 0.2 mg/mL, Invitrogen 12-5982-82) in FACS buffer for 20 minutes at 4°C in the dark.

Techniques: Expressing, Flow Cytometry

Journal: European journal of clinical investigation

Article Title: The influence of hypoxia-mediated CEACAM6 upregulation on epithelial cell and macrophage response in the context of gastric cancer.

doi: 10.1111/eci.14352

Figure Lengend Snippet: FIGURE 1 Hypoxia induces CEACAM6 in gastric cancer. (A) Western blotting of whole cell lysates of AGS and MKN45 cells kept at normoxia (N) or hypoxia (=H, 3% O2 for 12 h) showing levels of HIF1α and CEACAM6. α-tubulin was used as a loading control. Bar graphs showed a significant increase in CEACAM6 levels in AGS and MKN45. (B) A representative (n = 3) immunofluorescence micrograph of human metastatic gastric cancer biopsy tissue sample showing the status of HIF1α (green) and CEACAM6 (red). Bar graphs showing significant changes in HIF1α and CEACAM6 mean fluorescence intensities. Nuclei were stained with DAPI (blue). Tissues were sectioned at 5 μm thickness. Images were captured using 20X objective, and scale bars were set at 50 μm. (C) A representative western blot image showing CEACAM6 protein levels in hypoxia and/or normoxia-treated empty vector-transfected as well as CEACAM6 stably-transfected cells. Summary graph (n = 3) confirmed a significant increase of CEACAM6 compared to empty vector-transfected AGS cells kept at normoxia. Graphs = mean ± sem. Statistical significance was determined using the Student's t-test (n = 3). *p < .05; ***p < .001.

Article Snippet: For suppression studies, control siRNA (Santa Cruz Biotechnology, Dallas, Texas, USA) and CEACAM6 siRNA (Santa Cruz Biotechnology) were transfected into AGS cells using Lipofectamine 3000.

Techniques: Western Blot, Control, Immunofluorescence, Fluorescence, Staining, Plasmid Preparation, Transfection, Stable Transfection

Journal: European journal of clinical investigation

Article Title: The influence of hypoxia-mediated CEACAM6 upregulation on epithelial cell and macrophage response in the context of gastric cancer.

doi: 10.1111/eci.14352

Figure Lengend Snippet: FIGURE 3 Gastric cancer cell-macrophage crosstalk alters macrophage polarization status via CEACAM6. (A) Immune association analysis from TIMER2.0 showed that in STAD high CEACAM6 and macrophage infiltration caused lower cumulative survival, compared to lower CEACAM6 expression and low macrophage infiltration. (B) Representative (n = 3) immunofluorescence micrographs showed M2 macrophage infiltration in metastatic gastric biopsy tissue. (C) Representative (n = 3) flow cytometry analysis of PBMC-derived macrophages treated with supernatants from normoxic or hypoxic CEACAM6 or empty vector-overexpressing epithelial cells showed a downregulation of the M1(CD68+CD64+) phenotype in the presence of hypoxia and CEACAM6. H, hypoxia; N, normoxia.

Article Snippet: For suppression studies, control siRNA (Santa Cruz Biotechnology, Dallas, Texas, USA) and CEACAM6 siRNA (Santa Cruz Biotechnology) were transfected into AGS cells using Lipofectamine 3000.

Techniques: Expressing, Immunofluorescence, Flow Cytometry, Derivative Assay, Plasmid Preparation

Journal: Nature Communications

Article Title: A drug-free cardiovascular stent functionalized with tailored collagen supports in-situ healing of vascular tissues

doi: 10.1038/s41467-024-44902-2

Figure Lengend Snippet: a Representative rhodamine-conjugated phalloidin (TRITC-Phalloidin, red) and 4′,6-diamidino-2-phenylindole (DAPI, blue) fluorescence images of HUVECs showing the (rhCol III/PDA-PEI) n coatings loaded with different amounts of rhCol III all encouraged HUVECs compared with control PLA group. Scale bars, 200 μm. Quantification of ( b , c ) cell number and ( d , e ) cell viability of HUVECs cultured on uncoated and (rhCol III/PDA-PEI) n -coated PLA ( n = 1, 2, and 4) sheets after 1, 3, 5, and 7 days of culture ( n = 5 independent samples). f Volcano plot showing differentially expressed genes in the (rhCol III/PDA-PEI) 2 group compared to the PLA control group. Downregulated and upregulated genes are colored blue and red, respectively, at significantly differentially expressed thresholds │log 2 FC│ > 1.2 and p < 0.05. g Gene Ontology (GO) analysis of differentially expressed genes in PLA versus (rhCol III/PDA-PEI) 2 . h Heatmap of the differentially expressed genes in the PLA and (rhCol III/PDA-PEI) 2 groups. i Circular visualization of the results of gene-annotation enrichment analysis. j Quantification of the expression of representative genes in PLA versus (rhCol III/PDA-PEI) 2 , validated by qRT-PCR arrays. The value in the PLA group was normalized to that in the (rhCol III/PDA-PEI) 2 group ( n = 3 independent samples). k Representative immunofluorescence staining of CCL5 (green), GATA3 (green), XBP1 (green), and CEACAM6 (green) of HUVECs on 3 days of culture. Scale bar, 50 µm. l Schematic diagram of the potential three signaling pathways involved in the regulation of HUVEC behavior induced by the (rhCol III/PDA-PEI) 2 coating, including PI3K/AKT, mTOR, and MAPK. Two-way ANOVA with Tukey’s multiple comparisons was used for the comparisons in ( b )–( e ) and ( j ). Two-sided Student’s t test with multiple testing corrections was used in ( f ) and ( g ). The data are presented as the mean ± SD ( p values < 0.05 were considered statistically significant).

Article Snippet: The primary antibodies used in this study included mouse monoclonal XBP1 (Cat. No.: sc-8015, Clone: F-4, Santa Cruz Biotechnology, USA, 1:50), mouse polyclonal CCL5 (Cat. No.: sc-365826, Clone: A-4, Santa Cruz Biotechnology, USA, 1:50), mouse monoclonal CEACAM6 (Cat. No.: sc-59899, Clone: 9A6, Santa Cruz Biotechnology, USA, 1:50), rabbit monoclonal GATA3 (Cat. No.: ab199428, Clone: EPR16651, Abcam, USA, 1:500), rabbit polyclonal F4/80 (Cat. No.: 29414-1-AP, Proteintech, China, 1:100), rabbit monoclonal CD68 (Cat. No.: ab283654, Clone: EPR23917-164, Abcam, USA, 1:100), rabbit polyclonal CD86 (Cat. No.: bs-1035R, Biosynthesis Biotechnology co., ltd, USA, 1:200), rabbit monoclonal CD206 (Cat. No.: 24595, Clone: E6T5J, Cell Signaling Technology, USA, 1:200), mouse monoclonal α-SMA (Cat. No.: ab7817, Clone: 1A4, Abcam, USA, 1:200), and rabbit monoclonal MMP2 (Cat. No.: 10373-2-AP, Clone: SB13a, Proteintech, USA, 1:200), mouse monoclonal CD31 (Cat. No.: ab9498, Clone: JC/70A, Abcam, USA, 1:200), and rabbit polyclonal eNOS (Cat. No.: ab5589, Abcam, USA, 1:100).

Techniques: Fluorescence, Control, Cell Culture, Expressing, Quantitative RT-PCR, Immunofluorescence, Staining, Protein-Protein interactions