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: Bioactive Materials

Article Title: Sustained oxygen-releasing hydrogel implants enhance flap regeneration by promoting mitochondrial biogenesis under mild hypoxia

doi: 10.1016/j.bioactmat.2025.04.010

Figure Lengend Snippet: Anti-inflammatory and Antioxidant Effects Mediated by p65 Downregulation and SOD Upregulation Following cOMP-GelMA Treatment. (A) Representative immunohistochemistry images showing p65 (NF-κB subunit, upper row) and SOD expression (lower row). Scale bar = 50 μm. (B, C) Quantification of (B) p65-positive and (C) SOD-positive staining, n = 4–5 per group. (D, E) Expression levels of (D) p65 and (E) SOD were assessed by qPCR, n = 5 per group. (F) Schematic summary of the PGC-1α signaling pathway, highlighting the roles of p65 and SOD in reducing inflammation and oxidative stress. (G, H, I) Expression levels of pro-inflammatory cytokines as assessed by qPCR: (G) Il1b, (H) Il6, and (I) Tnfa, n = 5 per group. (J) Quantification of 8-OHdG-positive cells, n = 4 per group. (K) Representative immunofluorescence images of 8-OHdG (oxidative DNA damage marker) and DAPI staining. Scale bar = 100 μm. Data are presented as means ± SD, and statistical significance was determined using one-way ANOVA. ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001, ∗∗∗∗ p < 0.0001.

Article Snippet: Following deparaffinization, antigen retrieval, and blocking the tissue sections were incubated with primary antibodies, including vascular endothelial growth factor (VEGF; 1:250, sc-57496, Santa Cruz Biotechnology, Inc., Texas, USA), myeloperoxidase (MPO; 1:250, ab188211, Abcam, Cambridge, UK), CD31 (1:100, ab28364, Abcam), alpha-smooth muscle actin (α-SMA; 1:100, 14395-1-AP, Proteintech Group Inc., Rosemont, USA), Nuclear Respiratory Factor 1 (NRF-1, 1:100, ab175932, Abcam), and PGC-1α (1:100, 66369-1-lg, Proteintech Group Inc.), Hypoxia Inducible Factor 1 Alpha (HIF-1a, 1:400, 48085S, Cell signaling, Massachusetts, USA), Cluster of Differentiation 68(CD68, 1:250, MCA341R, Bio-rad, California, USA), Phospho-nuclear factor kappa-light-chain-enhancer of activated B cells p65 (p-NFκB p65, 1:100, sc-136548, Santa Cruz Biotechnology), 8-hydroxy-2′-deoxyguanosine (8-OhdG, 1:200, sc-393871, Santa Cruz Biotechnology) and Superoxide dismutase 2, mitochondrial (SOD2, 1:50, sc-133134, Santa Cruz Biotechnology).

Techniques: Immunohistochemistry, Expressing, Staining, Immunofluorescence, Marker

Journal: Journal of Advanced Research

Article Title: NAT10 promotes osteoclastogenesis in inflammatory bone loss by catalyzing Fos mRNA ac4C modification and upregulating MAPK signaling pathway

doi: 10.1016/j.jare.2024.07.031

Figure Lengend Snippet: Inhibition of NAT10 represses RANKL-activated MAPK signaling pathway. (A) Western blotting and quantification analysis of p-NFκB p65 and NFκB p65 in DMSO or Remodelin (20 μM)-pretreated RAW264.7 cells after RANKL stimulation at the designated times (n = 4). (B) Western blotting and quantification analysis of p-ERK1/2, ERK1/2, p-JNK, JNK, p-p38 and p38 in DMSO or Remodelin (20 μM)-pretreated RAW264.7 cells after RANKL stimulation at the designated times (n = 4). (C) Western blotting and quantification analysis of p-ERK1/2, ERK1/2, p-JNK, and JNK in DMSO or Remodelin (20 μM)-pretreated BMDMs after RANKL stimulation at the designated times (n = 4). (D) Western blotting and quantification analysis of p-ERK1/2, ERK1/2, p-JNK, and JNK in shNC and sh Nat10 RAW264.7 cells after RANKL stimulation at the designated times (n = 4). (E) The SRE-Luc and AP-1-Luc activities in DMSO or Remodelin (20 μM)-pretreated RAW264.7 cells after RANKL stimulation for 1 h (n = 4). Data were shown in the form of mean ± SD, two-way ANOVA was performed with Bonferroni’s multiple comparisons test, and Mann-Whitney U test was performed for comparisons between two groups. * p < 0.05, ** p < 0.01, *** p < 0.001 and **** p < 0.0001.

Article Snippet: Primary antibodies against β-actin, p-NFκB p65, NFκB p65, p-ERK1/2, ERK1/2, p-JNK, JNK, p-p38, and p38 were from Cell Signaling Technology, NAT10 was from Abcam (Cambridge, UK), ACP5, c-Jun, and c-Fos were from Proteintech (Wuhan, China), and NFATc1 was from Santa Cruz (CA, USA).

Techniques: Inhibition, Western Blot, MANN-WHITNEY