Journal: bioRxiv

Article Title: Transferrin-dependent crosstalk between the intestinal tract and commensal microbes contributes for immune tolerance

doi: 10.1101/2020.03.02.972281

Figure Lengend Snippet: THP-1 cells were stimulated in presence or absence of transferrin by LPS for 8 h. Total and phosphorylated TAK1, IKKα, IκBα, and p65 of the myD88-dependent pathway and TBK1 and IRF3 of the myD88-independent pathway in THP-1 cells were analyzed by Western blotting, respectively. Corresponding quantifications are shown on right (six panels). β-actin was used as loading control. Data represent means ± SD of five independent experiments, * p < 0.05, ** p < 0.01 by one-way ANOVA with Dunnett’s post-hoc test. Tf: transferrin.

Article Snippet: The primary antibodies used included anti-phospho-transforming growth factor-β (TGF-β)-activating kinase 1 (TAK1) (1:1□000 dilution, AF4379-100ul, Affinity Biosciences, USA), anti-total-TAK1 (1:1□000 dilution, AF4679-50ul, Affinity Biosciences, USA), anti-phospho-inhibitory subunit of nuclear factor κB (NF-κB) (IκB) kinase α (IKKα) (1:1□000 dilution, C84E11, Cell Signaling Technology, USA), anti-total-IKKα (1:1□000 dilution, ab32041-40ul, Abcam, USA), anti-phospho-IκBα (1:1□000 dilution, AF2002-100, Affinity Biosciences, USA), anti-total-IκBα (1:2□000 dilution, 4814T, Cell Signaling Technology, USA), anti-phospho-NF-κB p65 (1:1□000 dilution, AF2006-100, Affinity Biosciences, USA), anti-total-NF-κB p65 (1:1□000 dilution, 6956T, Cell Signaling Technology, USA), anti-phospho-TRAF-associated NF-κB activator (TANK)-binding kinase 1 (TBK1) (1:1□000 dilution, AF8190-50ul, Affinity Biosciences, USA), anti-total-TBK1 (1:1□000 dilution, DF7026-50, Affinity Biosciences, USA), anti-phospho-interferon regulatory factor 3 (IRF3) (1:1□000 dilution, 29047S, Cell Signaling Technology, USA), anti-total-IRF3 (1:1□000 dilution, ab50772, Abcam, USA), anti-phospho-c-JUN N-terminal kinase (JNK) (1:1□000 dilution, 9255S, Cell Signaling Technology, USA), anti-phospho-p38 (1:1□000 dilution, 4511S, Cell Signaling Technology, USA), anti-ALDH1A2 (1:1□000, DF4422, Affinity Biosciences, USA), anti-CCL22 (1:1□000, DF7781, Affinity Biosciences, USA), anti-TGF-β1 (1:1□000, 50698-T48, Sino Biological, China), anti-IL-10 (1:1□000, GTX130513, GeneTex, USA), anti-toll-like receptor 4 (TLR4) (1:1□000, AF7017, Affinity Biosciences, China), anti-transferrin (1:5□000, ab82411, Abcam, USA), anti-GAPDH (1:3□000, T0004-50, Affinity Biosciences, USA) and anti-β-actin (1:3□000, T0022-50, Affinity Biosciences, USA).

Techniques: Western Blot, Control