Journal: Frontiers in Immunology

Article Title: Promoting intestinal antimicrobial defense and microbiome symbiosis contributes to IL-22-mediated protection against alcoholic hepatitis in mice

doi: 10.3389/fimmu.2023.1289356

Figure Lengend Snippet: Primer sequences used for qPCR analysis.

Article Snippet: Cryostat sections of mouse ileum or liver tissues were incubated with anti-IL-22 (Genetex, Irvine, CA; Cat. #GTX18498), anti-F4/80 (BD Biosciences, Franklin Lakes, NJ; Cat. #565409), anti-MPO (Lifespan Biosciences, Seattle, WA; Cat. #LS-B6699), anti-DEFA5 (Elabscience, Houston, TX; ESAP13305), anti-ZO-1 (Millipore, Burlington, MA; Cat. #MABT11), anti-Ki67 antibody (Millipore; Cat. #AB9260), or anti-sodium hydrogen antiporter 3 (NHE3) antibody (Novus Biologicals, Centennial, CO; Cat. # NBP1-82574) followed by Alexa Fluor 594-conjugated IgG antibodies (Jackson ImmunoResearch Laboratories, West Grove, PA).

Techniques:

Journal: Frontiers in Immunology

Article Title: Promoting intestinal antimicrobial defense and microbiome symbiosis contributes to IL-22-mediated protection against alcoholic hepatitis in mice

doi: 10.3389/fimmu.2023.1289356

Figure Lengend Snippet: IL-22 stimulates IEC differentiation in a STAT3-depedent manner in mice and cultured organoids. WT mice were subjected to an 8-wk alcohol feeding, and IL-22 was i.p. injected at 1 mg/kg for the last 2 wk of feeding. (A) IF staining of ileal Ki67 (red) and nuclei (blue). Scale bar, 50 μm. (B) IF staining of ileal NHE3 (red) and nuclei (blue). Scale bar, 50 μm. (C) Organoid forming efficiency assessed by bright field imaging. Scale bar, 100 μm. Small intestinal organoids isolated from Stat3 fl/fl and Stat3 IEC-/- mice were treated with 100 ng/ml IL-22 for 10 h. (D) The mRNA levels of Nhe3 in organoids isolated from Stat3 fl/fl and Stat3 IEC-/- mice after IL-22 treatment. *** P < 0.001. (E) IF staining of NHE3 (red) in organoids after IL-22 treatment. Nuclei were stained by DAPI (blue). Scale bar, 20 μm.

Article Snippet: Cryostat sections of mouse ileum or liver tissues were incubated with anti-IL-22 (Genetex, Irvine, CA; Cat. #GTX18498), anti-F4/80 (BD Biosciences, Franklin Lakes, NJ; Cat. #565409), anti-MPO (Lifespan Biosciences, Seattle, WA; Cat. #LS-B6699), anti-DEFA5 (Elabscience, Houston, TX; ESAP13305), anti-ZO-1 (Millipore, Burlington, MA; Cat. #MABT11), anti-Ki67 antibody (Millipore; Cat. #AB9260), or anti-sodium hydrogen antiporter 3 (NHE3) antibody (Novus Biologicals, Centennial, CO; Cat. # NBP1-82574) followed by Alexa Fluor 594-conjugated IgG antibodies (Jackson ImmunoResearch Laboratories, West Grove, PA).

Techniques: Cell Culture, Injection, Staining, Imaging, Isolation

Journal: Frontiers in Immunology

Article Title: Promoting intestinal antimicrobial defense and microbiome symbiosis contributes to IL-22-mediated protection against alcoholic hepatitis in mice

doi: 10.3389/fimmu.2023.1289356

Figure Lengend Snippet: Schematic figure summarizing the major findings of the present study (created with BioRender.com). AMP, antimicrobial peptide; IEC, intestinal epithelial cells; NHE3, sodium-hydrogen exchanger 3; PAMP, pathogen-associated molecular pattern; STAT3, signal transducer and activator of transcription 3.

Article Snippet: Cryostat sections of mouse ileum or liver tissues were incubated with anti-IL-22 (Genetex, Irvine, CA; Cat. #GTX18498), anti-F4/80 (BD Biosciences, Franklin Lakes, NJ; Cat. #565409), anti-MPO (Lifespan Biosciences, Seattle, WA; Cat. #LS-B6699), anti-DEFA5 (Elabscience, Houston, TX; ESAP13305), anti-ZO-1 (Millipore, Burlington, MA; Cat. #MABT11), anti-Ki67 antibody (Millipore; Cat. #AB9260), or anti-sodium hydrogen antiporter 3 (NHE3) antibody (Novus Biologicals, Centennial, CO; Cat. # NBP1-82574) followed by Alexa Fluor 594-conjugated IgG antibodies (Jackson ImmunoResearch Laboratories, West Grove, PA).

Techniques:

Journal: American Journal of Translational Research

Article Title: Tempol treatment normalizes membrane expression of epithelial transport proteins in the kidney of salt-loaded hypertensive diabetic db/db mice

doi:

Figure Lengend Snippet: Sources of antibodies used in this study

Article Snippet: NHE3 , Novus Biologicals , NB110-81529.

Techniques:

Journal: Frontiers in Physiology

Article Title: Dipeptidyl Peptidase IV Inhibition Exerts Renoprotective Effects in Rats with Established Heart Failure

doi: 10.3389/fphys.2016.00293

Figure Lengend Snippet: Treatment with vildagliptin increases NHE3 phosphorylation at the PKA consensus site serine 552 in the renal cortex of HF rats. (A) Representative immunoblotting of renal cortical proteins probed with a phosphospecific NHE3 mAb that recognizes NHE3 only when it is phosphorylated at serine 552 (PS552-NHE3), total NHE3 mAb or anti-actin. (B) Graphical representation of the relative PS552-NHE3 and (C) total NHE3 levels. The relative abundance of PS552-NHE3 was quantitated by densitometry and expressed as a ratio of PS552-NHE3/total. The combined data from 4 experiments are represented. Values are means ± SEM. * p < 0.05 and *** p < 0.001 vs. Sham. ### p < 0.001 vs. HF.

Article Snippet: A mAb antibody against DPPIV (clone 5H8), a mouse mAb, clone 14D5, that recognizes NHE3 only when it is phosphorylated at serine 552 (Kocinsky et al., ) and polyclonal antibodies against the glucagon-like peptide-1 receptor (GLP-1R), cubilin, nephrin, and podocin were purchased from Santa Cruz Biotechnology (Santa Cruz, CA).

Techniques: Phospho-proteomics, Western Blot

Journal: American Journal of Physiology - Renal Physiology

Article Title: Furosemide reduces BK-αβ4-mediated K + secretion in mice on an alkaline high-K + diet

doi: 10.1152/ajprenal.00223.2018

Figure Lengend Snippet: Western blot and quantification of Na+/H+ exchanger 3 (NHE3) and vacuolar H+-ATPase (V-ATPase) expression of whole kidney (A) and medulla (B) of mice treated with HK + control (Ctrl) and HK + furosemide (Furo). *P < 0.05 vs. HK + Ctrl analyzed with Student’s t-test; n = 4/group.

Article Snippet: Primary antibodies included anti-BK-β4 (rabbit polyclonal, diluted 1:500; Alomone Laboratories, Jerusalem, Israel), anti-NHE3 (mouse monoclonal, diluted 1:200; Invitrogen, Carlsbad, CA), and anti-V-ATPase B1 (rabbit polyclonal, diluted 1:200; GeneTex, Irvine, CA).

Techniques: Western Blot, Expressing

Journal: bioRxiv

Article Title: Niche-inspired synthetic matrices for epithelial organoid culture

doi: 10.1101/806919

Figure Lengend Snippet: a , Bright-field images of 10-day old human enteroids growing in the synthetic ECM showing diverse phenotypes upon inducing differentiation. Scale bar: 200 μm. b-c , Immunostaining analysis of 10-day old organoids in the synthetic ECM and Matrigel showing proliferative cells (Ki67), Paneth cells (Lyz), Goblet cells (Muc2). Epithelial marker (E-cad), apical marker (NHE3) and basolateral markers (CD44v6, C-IV, and LMN). Scale bar: 500 μm.

Article Snippet: Immunostaining for cell-specific markers was done using the following primary antibodies, diluted in blocking solution; goat anti-E-cadherin (R&D, AF748, 1:200), goat anti-hDPPIV/CD26 (Invitrogen, AF1180, 1:400), rabbit anti-Col IV (Abcam, Ab6586, 1:200), rabbit anti-Lysozyme (Dako, A0099, 1:200), rabbit anti-LMN (Abcam, Ab11575, 1:200), rabbit anti-Ki67, (Abcam, Ab15580, 1:200 or ab16667, 1:100), rabbit anti-NHE3/SCL9A3 (Novus, NBP-82574), mouse anti-Villin (Santa Cruz, SC-58897, 1:50), mouse anti-CD44-v6 (Abcam, Ab78960, 1:200), mouse anti-Muc2 (Santa Cruz, SC515032 1:200), mouse anti-EpCAM (Abcam, ab7504 1:200).

Techniques: Immunostaining, Marker

Journal: bioRxiv

Article Title: Niche-inspired synthetic matrices for epithelial organoid culture

doi: 10.1101/806919

Figure Lengend Snippet: a-b , Immunostaining analysis of 10-day old organoids differentiated in the synthetic ECM and Matrigel showing proliferative cells (Ki67), Paneth cells (Lyz), epithelial marker (E-cad), apical markers (NHE3, Villin) and basolateral markers (CD44v6, C-IV, and LMN). Scale bar: 50 μm.

Article Snippet: Immunostaining for cell-specific markers was done using the following primary antibodies, diluted in blocking solution; goat anti-E-cadherin (R&D, AF748, 1:200), goat anti-hDPPIV/CD26 (Invitrogen, AF1180, 1:400), rabbit anti-Col IV (Abcam, Ab6586, 1:200), rabbit anti-Lysozyme (Dako, A0099, 1:200), rabbit anti-LMN (Abcam, Ab11575, 1:200), rabbit anti-Ki67, (Abcam, Ab15580, 1:200 or ab16667, 1:100), rabbit anti-NHE3/SCL9A3 (Novus, NBP-82574), mouse anti-Villin (Santa Cruz, SC-58897, 1:50), mouse anti-CD44-v6 (Abcam, Ab78960, 1:200), mouse anti-Muc2 (Santa Cruz, SC515032 1:200), mouse anti-EpCAM (Abcam, ab7504 1:200).

Techniques: Immunostaining, Marker

Journal: bioRxiv

Article Title: Niche-inspired synthetic matrices for epithelial organoid culture

doi: 10.1101/806919

Figure Lengend Snippet: Immunostaining analysis of 10-day old organoids emerging in the synthetic matrix and Matrigel showing proliferative cells (Edu), and correct localization of apical markers (F-actin, NHE3 and DPP4).

Article Snippet: Immunostaining for cell-specific markers was done using the following primary antibodies, diluted in blocking solution; goat anti-E-cadherin (R&D, AF748, 1:200), goat anti-hDPPIV/CD26 (Invitrogen, AF1180, 1:400), rabbit anti-Col IV (Abcam, Ab6586, 1:200), rabbit anti-Lysozyme (Dako, A0099, 1:200), rabbit anti-LMN (Abcam, Ab11575, 1:200), rabbit anti-Ki67, (Abcam, Ab15580, 1:200 or ab16667, 1:100), rabbit anti-NHE3/SCL9A3 (Novus, NBP-82574), mouse anti-Villin (Santa Cruz, SC-58897, 1:50), mouse anti-CD44-v6 (Abcam, Ab78960, 1:200), mouse anti-Muc2 (Santa Cruz, SC515032 1:200), mouse anti-EpCAM (Abcam, ab7504 1:200).

Techniques: Immunostaining