Journal: Frontiers in Immunology

Article Title: Adiponectin Enhances B-Cell Proliferation and Differentiation via Activation of Akt1/STAT3 and Exacerbates Collagen-Induced Arthritis

doi: 10.3389/fimmu.2021.626310

Figure Lengend Snippet: The expression and activation of Akt1, ERK1/2, Egr-1, and STAT3 in B cells exposed to AD. (A) Purified mouse splenic B cells were incubated with 10 μg/ml AD for different time points (0, 5, 10, 20, 30 min), and the levels of t-Akt1, p-Akt1 Thr308 , p-Akt1 Ser473 , t-ERK1/2, p-ERK1/2, and β-actin were examined by the Western blot. (B) Purified mouse splenic B cells were incubated with 10 μg/ml AD for different time points (0, 3, 6, 12, and 24 h), and the levels of Egr-1, t-STAT3, p-STAT3, and β-actin were detected by the Western blot. Results from one representative experiment out of three are shown. Representative images are shown. Data are presented as means ± SE ( n = 3 in each time points). ** p < 0.01 vs. 0 min time point or 0 h time point.

Article Snippet: Akt1 siRNA (siAkt1), STAT3 siRNA (siSTAT3), control siRNA (siCTR), and fluorescein-conjugated control siRNA (FITC-siCTR) were purchased from Cell Signaling Technology.

Techniques: Expressing, Activation Assay, Purification, Incubation, Western Blot

Journal: Frontiers in Immunology

Article Title: Adiponectin Enhances B-Cell Proliferation and Differentiation via Activation of Akt1/STAT3 and Exacerbates Collagen-Induced Arthritis

doi: 10.3389/fimmu.2021.626310

Figure Lengend Snippet: The roles of PI3K, Akt1, and STAT3 activation in B-cell proliferation and differentiation in response to AD. Purified mouse splenic B cells were transfected with siPI3K, siAkt1, siSTAT3, or siCTR followed by AD stimulation (10 μg/ml) for different time points, and then the protein expression as well as B-cell proliferation and differentiation were evaluated. (A) The levels of PI3K, t-Akt1, p-Akt1 Ser473 , t-STAT3, p-STAT3, Blimp-1, and β-actin were examined by the Western blot. (B–E) The BrdU-positive rate (B,D) and CD138-positive rate (C,E) were detected by flow cytometry. Results from one representative experiment out of three are shown. Representative images are shown. Data are presented as means ± SE ( n = 3 in each group). ** p < 0.01 vs. siCTR + AD group.

Article Snippet: Akt1 siRNA (siAkt1), STAT3 siRNA (siSTAT3), control siRNA (siCTR), and fluorescein-conjugated control siRNA (FITC-siCTR) were purchased from Cell Signaling Technology.

Techniques: Activation Assay, Purification, Transfection, Expressing, Western Blot, Flow Cytometry

Journal: PLoS Pathogens

Article Title: Inflammatory monocytes provide a niche for Salmonella expansion in the lumen of the inflamed intestine

doi: 10.1371/journal.ppat.1007847

Figure Lengend Snippet: (A-C) Effects of Sm pretreatment and STm infection on C57BL/6J mice (n = 3–7 per group). Samples were harvested on day 4 after infection. Data show mean with SEM (A and B) or mean with SEM and individual data points (C), and were analyzed by one-way ANOVA with Sidak post hoc test. (A) Relative fold induction of Ccl2 expression in cecal tissues, normalized to Gapdh . (B) Percentage of IM among total gated cells from cecal tissues. (C) Number of STm CFU recovered from cecal contents, normalized to weight. (D-E) Four-day time course tracking IM recruitment into cecal tissues from C57BL/6J mice (n = 4–9 per group) treated with Sm prior to inoculation with STm. Mice treated with Sm prior to inoculation with PBS (uninfected, UI) were used as a control. Data show mean with SEM (D) or mean with SEM and individual data points (E), and were analyzed by one-way ANOVA with Fisher’s LSD post hoc test. (D) Percentage of IM among total gated cells from cecal tissues. (E) Number of STm CFU recovered from cecal contents, normalized to weight. (F) Percentage of IM among live cells in cecal contents from C57BL/6J mice (n = 6 per group) treated with Sm prior to inoculation with PBS (uninfected, UI) or STm. Cecal contents were collected on day 4 after inoculation. Data show mean with SEM and were analyzed by Student’s t -test. (G) Level of iNOS expressed by IM and neutrophils purified and pooled from cecal tissues of C57BL/6J mice (n = 5–6 per group) treated with Sm prior to inoculation with STm. Ceca were harvested on day 4 after inoculation. β-Actin was used as a loading control for Western blotting. Data are representative of two independent experiments. Also see .

Article Snippet: Expression of iNOS and β-actin was detected with anti-NOS2 antibody (1:1,000; BioLegend Cat#690902; RRID: AB_2629826) and anti-β-actin antibody (1:1,000; Cell Signaling Technology Cat#4967S; RRID: AB_330288), respectively.

Techniques: Infection, Expressing, Purification, Western Blot

Journal: PLoS Pathogens

Article Title: Inflammatory monocytes provide a niche for Salmonella expansion in the lumen of the inflamed intestine

doi: 10.1371/journal.ppat.1007847

Figure Lengend Snippet: (A) Relative fold induction of Nos2 expression in cecal tissues from C57BL/6J (WT) and Ccr2 -/- mice (n = 5–7 per group) treated with Sm prior to inoculation with PBS (uninfected, UI) or STm. Ceca were harvested on day 4 after inoculation. Relative Nos2 expression was normalized to Gapdh . Data show mean with SEM and were analyzed by one-way ANOVA with Sidak post hoc test. (B) Relative level of iNOS expressed in cecal tissues from C57BL/6J (WT) and Ccr2 -/- mice (n = 6–9 per group) treated with Sm prior to inoculation with PBS (uninfected, UI) or STm. Ceca were harvested on day 4 after inoculation. β-Actin was used as a loading control for Western blotting. Data show mean with SEM and were analyzed by one-way ANOVA with Sidak post hoc test. (C) Concentration of nitrate in cecal mucus from C57BL/6J (WT) and Ccr2 -/- mice (n = 5–7 per group) left untreated or treated with Sm prior to inoculation with PBS or STm. Ceca were harvested on day 4 after inoculation. Data show mean with SEM and individual data points, and were analyzed by one-way ANOVA with Sidak post hoc test. Also see .

Article Snippet: Expression of iNOS and β-actin was detected with anti-NOS2 antibody (1:1,000; BioLegend Cat#690902; RRID: AB_2629826) and anti-β-actin antibody (1:1,000; Cell Signaling Technology Cat#4967S; RRID: AB_330288), respectively.

Techniques: Expressing, Western Blot, Concentration Assay

Journal: PLoS Pathogens

Article Title: Inflammatory monocytes provide a niche for Salmonella expansion in the lumen of the inflamed intestine

doi: 10.1371/journal.ppat.1007847

Figure Lengend Snippet: We postulate that IM contribute both directly and indirectly to the generation of host- derived nitrate used by STm for growth in the lumen of the inflamed intestine. IM may contribute directly to the generation of host-derived nitrate used by STm for growth in the lumen of the inflamed intestine by expressing Nos2 and indirectly by potentiating Nos2 expression by other cells such as IEC. See the text for details.

Article Snippet: Expression of iNOS and β-actin was detected with anti-NOS2 antibody (1:1,000; BioLegend Cat#690902; RRID: AB_2629826) and anti-β-actin antibody (1:1,000; Cell Signaling Technology Cat#4967S; RRID: AB_330288), respectively.

Techniques: Derivative Assay, Expressing

Journal: Oncogene

Article Title: A causal link from ALK to hexokinase II overexpression and hyperactive glycolysis in EML4-ALK-positive lung cancer

doi: 10.1038/onc.2016.150

Figure Lengend Snippet: ( A ) The effects of ALK knockdown in H2228 and H3122 or ALK ectopic expression in A549 on levels of HIF1α mRNA were analyzed by qPCR. ( B ) H2228 and H3122 cells were treated with vehicle, ceritinib (200 nM), LY294002 (10 μM), MK2206 (4 μM), or rapamycin (20 nM) for 8 hours. HIF1α protein and mRNA levels in control and treated cells were analyzed by immunoblotting and qPCR, respectively. ( C ) AKT1/2 expression in H2228 and H3122 cells was knocked down simultaneously by siRNA targeting both isoforms. The effects on HIF1α, HK2 and other proteins as well as HIF1α mRNA were analyzed as in ( B ). In all panels, qPCR results were normalized to 18S rRNA and presented as values relative to the corresponding control cells (defined as 1 fold).

Article Snippet: AKT1/2 siRNA were purchased from Cell Signaling.

Techniques: Expressing, Western Blot