Journal: Poultry Science

Article Title: CSF3 enhances the innate immune responses to ALV-J infections via NF-κB and interferon pathways

doi: 10.1016/j.psj.2025.105648

Figure Lengend Snippet: CSF3 enhances the immune response to ALV-J via the NFκB signaling pathway. (A) Western blot analysis of p52, p100, phosphorylated p65 (p-p65), IκBα, and phosphorylated IκBα (p-IκBα) protein following CSF3 overexpression in DF-1 cells. (B) Quantification of protein levels in (A) based on relative grayscale values. (C) Western blot analysis of p52, p100, phosphorylated p65 (p-p65), IκBα, and phosphorylated IκBα (p-IκBα) protein following CSF3 knockdown in DF-1 cells. (D) Quantification of protein levels in (C) based on relative grayscale values. (D, E) RT-qPCR analysis of TNF-α, IL-1β , and IL-6 mRNA expression following CSF3 overexpression (D) or knockdown (E) in DF-1 cells. (F, G) ELISA measurement of TNF-α, IL-1β, and IL-6 protein levels following CSF3 overexpression (F) or knockdown (G) in DF-1 cells. (H, I) RT-qPCR analysis of TNF-α, IL-1β , and IL-6 mRNA expression following CSF3 overexpression (H) or knockdown (I) in CEF cells. (J, K) ELISA measurement of TNF-α, IL-1β, and IL-6 protein levels following CSF3 overexpression (J) or knockdown (K) in CEF cells. (L) Western blot analysis of STAT3, phosphorylated STAT3 (p-STAT3), env, β-actin, IκBα, p-IκBα, p-p65, p52, and p100 after STAT3 phosphorylation inhibition in CSF3-overexpressing DF-1 cells. (M) Quantification of protein levels in (L) based on relative grayscale values. Statistical significance was determined using a two-tailed unpaired Student’s t-test ( p < 0.05). * p < 0.05, **p < 0.01, *** p < 0.001.

Article Snippet: Mouse Anti-ALV-J envelope protein JE9 (kindly provided by Prof. Aijian Qin, Yangzhou University, Yangzhou, China), Rabbit Anti-STAT3 antibody (bs-1141R; Boss, China; 1:1000), Rabbit Anti-phospho-STAT3 (Ser727) antibody (bs-3429R; Boss, China; 1:1000), Rabbit Anti-NFκB2 antibody (10037P, Boss, China; 1:1000), Rabbit Anti-NFκB p-p65 (bs-0982R, Boss, China; 1:1000), Rabbit Anti-IKBα Rabbit (10268-1-AP, Proteintech, USA; 1:1000), Anti-p-IKBα (bs-2513R, Boss, China; 1:1000), and goat anti-rabbit IgG/HRP (bs13278), goat anti-mouse IgG/HRP (bs12478), goat Anti-Mouse IgG ( H + L ) FITC (bs10950) secondary antibody were purchased from Bioss (Beijing, China).

Techniques: Western Blot, Over Expression, Knockdown, Quantitative RT-PCR, Expressing, Enzyme-linked Immunosorbent Assay, Phospho-proteomics, Inhibition, Two Tailed Test

Journal: Cells

Article Title: Effects of Adipose-Derived Mesenchymal Stem Cell-Secretome on Pyroptosis of Laparoscopic Hepatic Ischemia Reperfusion Injury in a Porcine Model.

doi: 10.3390/cells14100722

Figure Lengend Snippet: Figure 6. ADSC-secretome inhibited the HMGB1/TLR4/NF-κB signaling pathway during HIRI. (a) Representative Western blot analysis of HMGB1, TLR4, p-NF-κBp65/NF-κBp65, and p-IκB. (b–e) The levels of HMGB1, TLR4, p-NF-κBp65/NF-κBp65, and p-IκB proteins were evaluated. The protein levels of HMGB1, p-NF-κBp65, and NF-κBp65 were adjusted against the respective levels of β-actin. For TLR4 and p-IκB, their protein expression levels were normalized to α-tubulin levels. (f–h) The gene expression analysis for HMGB1, TLR4, and NF-κBp65 was conducted. Three replicates were performed for each sample. The relative mRNA levels were standardized with reference to β-actin. The significance of results is indicated as ▲p < 0.05, ▲▲p < 0.01, when compared to the IRI group; * p < 0.01 and ** p < 0.01, relative to the DMEM group; # p < 0.05 and ## p < 0.01, in comparison to the ADSCs group.

Article Snippet: Following this, the membranes were incubated overnight with primary antibodies specific to β-actin (1:1000, Cell Signaling Technology, MA, USA), α-tubulin (1:10,000, Proteintech, IL, USA), IL-1β (1:500, ABmart, Shanghai, China), IL-18, caspase1, NLRP3, HMGB1, TLR4 (1:1000, WanLei Biologicals, Shenyang, China), apoptosis-associated speck-like protein containing a CARD (ASC), GSDMD-N (1:1000, Affinity Biosciences, FLA, USA), NF-κBp65 (1:2000, Proteintech, IL, USA), p-IκB (1:1000, Immunoway, TX, USA), p-NF-κBp65 (1:500, Santa Cruz Biotechnology, CA, USA), and IκB (1:1000, Immunoway, TX, USA).

Techniques: Western Blot, Expressing, Gene Expression, Comparison