e coli s 17  (ATCC)

Journal: Nature Communications

Article Title: Pancreatic glycoprotein 2 is a first line of defense for mucosal protection in intestinal inflammation

doi: 10.1038/s41467-021-21277-2

Figure Lengend Snippet: a Fecal total IgA concentration in WT and Gp2 –/– mice, as determined by ELISA ( n = 5). N.S. not significant (two-tailed unpaired t -test). Data are presented as mean values ± SEM. b 16S rRNA gene sequencing of fecal bacteria. Phylum levels of bacteria are shown. c qPCR analysis of mucosal bacteria from WT ( n = 9) and Gp2 –/– ( n = 8) mice were examined. * p < 0.05., N.S. indicates not significant (two-tailed unpaired t -test). Data are presented as mean values ± SEM. d Survival ratio and body weight change of WT and Gp2 –/– mice with acute colitis ( n = 12/group). * p < 0.05, ** p < 0.01 (two-tailed unpaired t -test). Data are presented as mean values ± SEM. e Colon length and representative pictures of colon are shown, intact; WT ( n = 3), KO ( n = 4), and colitis; WT, KO ( n = 12). *** p < 0.0001 (two-tailed unpaired t -test). Data are presented as mean values ± SEM. f Hematoxylin and eosin staining of colon at day 8 of DSS treatment are shown. Scale bars: 100 μm. Data are representative of three independent experiments. g Flow cytometry analysis of infiltrated neutrophils is shown. Representative data are shown. h Immunohistochemical analysis of colitis (DSS 2%, day 8) colon with luminal contents of WT and Gp2 –/– mice. Bacteria (red), MUC2 (green), DAPI (blue). Scale bars: upper panels, 100 μm; lower panels, 20 μm. Data are representative of three independent experiments. i Serum anti- E. coli IgM, intact ( n = 4), and colitis ( n = 5), IgA, intact WT ( n = 6), KO ( n = 10), and colitis WT ( n = 15), KO ( n = 11), IgG intact WT ( n = 12), KO ( n = 4), and colitis WT ( n = 11), KO ( n = 7) in intact and colitis WT and Gp2 –/– mice were analyzed by ELISA (Kruskal-Wallis test followed by Mann–Whitney U test). Data are presented as mean values ± SEM. j Body weight changes after the induction of acute TNBS-induced colitis in WT and Gp2 –/– mice. *** p < 0.01 (two-tailed unpaired t -test). Data are presented as mean values ± SEM. k Immunohistochemical analysis of colon and luminal contents during TNBS-induced colitis in Gp2 Panc and control mice. EUB338 bacteria (red), MUC2 (green), DAPI (blue). Scale bars: upper panels, 100 μm; lower panels, 20 μm. Data are representative of two independent experiments. Source data are provided as a Source Data file.

Article Snippet: As previously described, E. coli strain S-17 were freeze-dried by Techno Suruga Laboratory.

Techniques: Concentration Assay, Enzyme-linked Immunosorbent Assay, Two Tailed Test, Sequencing, Bacteria, Staining, Flow Cytometry, Immunohistochemical staining, MANN-WHITNEY, Control

Journal: Nature Communications

Article Title: Pancreatic glycoprotein 2 is a first line of defense for mucosal protection in intestinal inflammation

doi: 10.1038/s41467-021-21277-2

Figure Lengend Snippet: a Luminal contents of colon were stained with GP2 (red) and EUB338 (blue in left bottom and green in right bottom) were shown. Data are representative of three independent experiments. b Fecal unbound GP2 in WT and Gp2 –/– mice with or without acute colitis were measured by ELISA (WT, n = 8; Gp2 –/– , n = 4). N.S. indicates not significant (two-tailed unpaired t -test). Data are presented as mean values ± SEM. c Percentage of rGP2-bound fecal bacteria isolated from Gp2 –/– mice, PBS and rGP2 Intact n = 7, PBS colitis ( n = 7), rGP2 colitis ( n = 6). * p = 0.017 (two-tailed unpaired t -test). Data are presented as mean values ± SEM. d Representative flow cytometry analysis of fecal bacteria isolated from WT and Gp2 –/– mice with or without colitis were shown. e E. coli number of non-selected (rGP2 MACS −) and selected by MACS (rGP2 MACS +) fecal bacteria ( n = 9 /group). N.D.: not detected. Data are presented as mean values ± SEM. Source data are provided as a Source Data file.

Article Snippet: As previously described, E. coli strain S-17 were freeze-dried by Techno Suruga Laboratory.

Techniques: Staining, Enzyme-linked Immunosorbent Assay, Two Tailed Test, Bacteria, Isolation, Flow Cytometry

Journal: Nature Communications

Article Title: Pancreatic glycoprotein 2 is a first line of defense for mucosal protection in intestinal inflammation

doi: 10.1038/s41467-021-21277-2

Figure Lengend Snippet: a Flow cytometry of E. coli strain S-17-GFP bound with or without rGP2 were performed. Representative data of three independent experiments were shown. *** p < 0.0001 (two-tailed unpaired t -test). Data are presented as mean values ± SEM. b CFU assay of S-17-GFP incubated with recombinant GP2 ( n = 4). N.S. indicates not significant (two-tailed unpaired t -test). c Approximately 1 × 10 6 CFU of WT S-17 cells and FimH-deficient S-17 cells (ΔFimH) were incubated with 1 μg of mouse or human GP2, and FACS analysis was conducted with anti-GP2 antibody. Representative data of three independent experiments are shown. d , e Immunohistochemical analysis of colon specimens isolated from ligated colon of S-17-GFP, pre-incubated with PBS or recombinant GP2 were shown. Scale bars, 50 μm (left panel) and 10 μm (right panels). f CFU assay of S-17-GFP injected looped colon (2% DSS day 6), n = 5 /group were shown. * p = 0.017 (two-tailed unpaired t -test). Data are presented as mean values ± SEM. g The amount of anti-GP2 IgG in the luminal contents of intact mice ( n = 5), DSS-treated mice ( n = 6), mice systemically administered GP2-bound heat-killed (70 °C, 30 min) S-17 cells ( n = 4), and GP2 knockout (GP2KO) mice immunized with rmGP2 and complete Freund’s adjuvant (CFA) as positive controls was determined by ELISA on day 27 after immunization ( n = 4). *** p < 0.01 (one-way ANOVA). Data are presented as mean values ± SEM. h Neutralizing assay for the binding of rGP2 to S-17 cells. Medium (Tris-HCL). Luminal contents from anti-GP2 IgG antibody–negative (intact) or -positive (DSS-treated) mice were pre-incubated with GP2 and added to S-17 cells. The GP2-bound SYTO9 + S-17 population is shown. Data are representative of two independent experiments. Source data are provided as a Source Data file.

Article Snippet: As previously described, E. coli strain S-17 were freeze-dried by Techno Suruga Laboratory.

Techniques: Flow Cytometry, Two Tailed Test, Colony-forming Unit Assay, Incubation, Recombinant, Immunohistochemical staining, Isolation, Injection, Knock-Out, Adjuvant, Enzyme-linked Immunosorbent Assay, Neutralizing Assay, Binding Assay