anti il 22 neutralizing antibody  (R&D Systems)

Journal: Frontiers in Immunology

Article Title: Bacteroides fragilis strain ZY-312 facilitates colonic mucosa regeneration in colitis via motivating STAT3 signaling pathway induced by IL-22 from ILC3 secretion

doi: 10.3389/fimmu.2023.1156762

Figure Lengend Snippet: B.fragilis promoted IL-22 production, but not IL-1β and TNF-α mediated classical proinflammatory pathway in DSS-induced colitis. (A) ELISA analysis of the inflammatory factors including IL-22, IL-6, IL-23, IL-1β, TNF-α, IFN-γ, IL-17A, and IL-10 from colon tissue in DSS-induced colitis. (B) ELISA analysis of IL-22, and IL-6 from serum in DSS-induced colitis. (C) ELISA analysis of IL-22, IL-6, IL-23, IL-1β, TNF-α, IFN-γ, IL-17A, and IL-10 in supernatant from co-culture model including B. fragilis strain ZY-312 and CLP after lipopolysaccharide (LPS,1ug/ml) stimulation for 12 hours in vitro . Data are presented as Mean ± SEM (A–C) . Statistical analysis was performed using one-way ANOVA. * p < 0.05,** p < 0.01, *** p < 0.005, **** p < 0.001.ND indicates undetectable. NS indicates no significance.

Article Snippet: B. fragilis (10 4 CFU/well) and murine TNF-α (60 ng/mL; Peprotech, Rocky Hill, NJ, USA) were added independently or simultaneously to the culture medium for 24 h. Murine IL-22 (5 ng/ml, Peprotech), anti-IL-22 neutralizing antibody (0.1 μg/mL, AF582; RD, Minneapolis, MN, USA) , and Stattic (15 μM; Selleck, Houston, TX, USA) ( ) were also added to organoids for 24 hours.

Techniques: Enzyme-linked Immunosorbent Assay, Co-Culture Assay, In Vitro

Journal: Frontiers in Immunology

Article Title: Bacteroides fragilis strain ZY-312 facilitates colonic mucosa regeneration in colitis via motivating STAT3 signaling pathway induced by IL-22 from ILC3 secretion

doi: 10.3389/fimmu.2023.1156762

Figure Lengend Snippet: B. fragilis up-regulated IL-22/pSTAT3 pathway to facilitate colonic enterocytes proliferation and mucus secretion, inhibit apoptosis, and impact gut microbiota. (A, B) Percent of weight loss (A) and DAI (B) were monitored daily starting from DSS administration. Control/ IL-22 -/- group (N=4), DSS/ IL-22 -/- group (N=7), DSS+ZY-312/ IL-22 -/- group (N=8), DSS+ZY-312/ IL-22 +/+ group (N=7). N, number of mice. (C) Representative colon morphology. (D) Statistical analysis of colon length. (E) H&E staining of colon tissues (Scale bars, 200 μm). (F) Statistical analysis of HAI. (G) qPCR analysis of Caspase-3 and PCNA in colon tissue. (H) Immunochemistry analysis of pSTAT3 and Ki-67, alcian blue analysis of mucus from IL-22 +/+ and IL-22 -/- mice (Scale bars, 200 μm). (I) Tunel assay analysis of colon tissue from IL-22 -/- mice (Scale bars, 200 μm). (J) Western Blot analysis of pSTAT3, STAT3, PCNA, cleaved Caspase-3, and Caspase-3 from colonic epithelial cells in IL-22 -/- mice. (K) Theβdiversity analysis of microbiota in Stat3 △IEC mice between groups. (L) LEfSe analysis of differential bacteria in feces between the DSS group and ZY-312 group with different levels meeting a significant LDA threshold value of >3.5 in IL-22 -/- mice. NF.22KO indicates Control/IL-22 -/- group. DF.22KO indicates DSS/IL-22 -/- group. ZF.22KO indicates DSS+ZY-312/IL-22 -/- group. Data are presented as Mean ± SEM (A, B, D, F, G) . The P value was calculated using two-way ANOVA (A, B) . Statistical analysis was performed using one-way ANOVA (D, F, G) . * p < 0.05,** p < 0.01, *** p < 0.005, **** p < 0.001. ND indicates undetectable. NS indicates no significance.

Article Snippet: B. fragilis (10 4 CFU/well) and murine TNF-α (60 ng/mL; Peprotech, Rocky Hill, NJ, USA) were added independently or simultaneously to the culture medium for 24 h. Murine IL-22 (5 ng/ml, Peprotech), anti-IL-22 neutralizing antibody (0.1 μg/mL, AF582; RD, Minneapolis, MN, USA) , and Stattic (15 μM; Selleck, Houston, TX, USA) ( ) were also added to organoids for 24 hours.

Techniques: Staining, TUNEL Assay, Western Blot

Journal: Frontiers in Immunology

Article Title: Bacteroides fragilis strain ZY-312 facilitates colonic mucosa regeneration in colitis via motivating STAT3 signaling pathway induced by IL-22 from ILC3 secretion

doi: 10.3389/fimmu.2023.1156762

Figure Lengend Snippet: B. fragilis possibly promoted CLP-derived ILC3 to secrete IL-22 in DSS-induced colitis. (A) The heatmap analysis of inflammatory factors from colon tissue through protein microarray detection. Control group (N=2), DSS group (N=3), DSS+ZY-312 group (N=3). N, number of mice. Red represents up-regulation, and blue represents down-regulation. (B) The level of inflammatory factors including IL-7, IL-1α, GM-CSF, and IFN-γ showed significant differences between the DSS+ZY-312 group (N=3) and the DSS group (N=3). IL-7 (p value=0.0078), IL-1α (p value=0.0038), GM-CSF (p value=0.0173), IFN-γ (p value=0.0386), Red represents up-regulation, blue represents down-regulation. (C) The PPI (protein-protein interaction) analysis of DEGs (differentially expressed genes) between the DSS group and DSS+ZY-312 group. Network nodes represent proteins, edges represent protein-protein associations, colored nodes: query proteins and the first shell of interactors, white nodes: the second shell of interactors, and empty nodes: proteins of unknown 3D structure, contrary to a known 3D structure. (D) IL-22 secreting ILC3 (labeled with FVS-CD45+Lineage-RORγt+IL-22+) and CD4+T cells (labeled with FVS-CD45+Lineage+CD4+IL-22+) from CLP in C57BL/6 mice were analyzed by cell flow cytometry, and statistical analysis of cell percentage was showed (right panel). Control group (N=3), DSS group (N=13), DSS+ZY-312 group (N=11). N, number of mice. (E) Cell flow cytometry analysis of IL-22 secreting ILC3 in IL-22 +/+ mice. Control group/ IL-22 +/+ (N=3), DSS group/ IL-22 +/+ (N=3), DSS+ B. fragilis group/ IL-22 +/+ (N=3). N, number of mice. Data are presented as Mean ± SEM. Statistical analysis was performed using one-way ANOVA (D, E) . * p < 0.05,** p < 0.01, *** p < 0.005, NS indicates no significance.

Article Snippet: B. fragilis (10 4 CFU/well) and murine TNF-α (60 ng/mL; Peprotech, Rocky Hill, NJ, USA) were added independently or simultaneously to the culture medium for 24 h. Murine IL-22 (5 ng/ml, Peprotech), anti-IL-22 neutralizing antibody (0.1 μg/mL, AF582; RD, Minneapolis, MN, USA) , and Stattic (15 μM; Selleck, Houston, TX, USA) ( ) were also added to organoids for 24 hours.

Techniques: Derivative Assay, Microarray, Labeling, Flow Cytometry

Journal: Frontiers in Immunology

Article Title: Bacteroides fragilis strain ZY-312 facilitates colonic mucosa regeneration in colitis via motivating STAT3 signaling pathway induced by IL-22 from ILC3 secretion

doi: 10.3389/fimmu.2023.1156762

Figure Lengend Snippet: B. fragilis facilitates colonic mucosa regeneration in colitis via motivating the STAT3 pathway induced by IL-22 from ILC3 secretion. B. fragilis promoted CLP-derived ILC3 to secrete IL-22, then motivated the STAT3 signaling pathway, hence promoting colonic mucosa proliferation and mucus secretion, inhibiting apoptosis, and altering gut microbiota in DSS-induced colitis.

Article Snippet: B. fragilis (10 4 CFU/well) and murine TNF-α (60 ng/mL; Peprotech, Rocky Hill, NJ, USA) were added independently or simultaneously to the culture medium for 24 h. Murine IL-22 (5 ng/ml, Peprotech), anti-IL-22 neutralizing antibody (0.1 μg/mL, AF582; RD, Minneapolis, MN, USA) , and Stattic (15 μM; Selleck, Houston, TX, USA) ( ) were also added to organoids for 24 hours.

Techniques: Derivative Assay