Journal: Journal of Advanced Research

Article Title: Tussilagone attenuated cigarette smoke-induced chronic obstructive pulmonary disease through regulating Nrf2 and NF-κB/NLRP3 inflammasome via directly targeting cysteine 434 of KEAP1

doi: 10.1016/j.jare.2025.07.019

Figure Lengend Snippet: Discovery of TUS as a potent therapeutic agent for COPD. (A) High-throughput screening for CXCL1 inhibitory activity of compounds from FF. RAW 264.7 macrophages were treated with 1 μg/mL LPS and 20 μM diverse compounds from FF for 24 h, and the level of CXCL1 was detected with ELISA kit. (B) Structures of sesquiterpenoids from FF. (C) The H&E or Masson’s trichrome staining images of lung tissues of mice in different groups. (D) The protein levels of iNOS, COX-2, and IL-1β in the homogenate of lung tissues of mice in different groups. (E-F) The levels of MUC5AC and SP-D in the homogenate of lung tissues of mice in different groups. (G) The percentage of neutrophil in white blood cells of mice in different groups. (H-I) The levels of TNF-α and IL-1β in BALF of mice in different groups. The results were expressed as mean ± SD (n = 4). * p < 0.05, ** p < 0.01, *** p < 0.001 treated vs CS group.

Article Snippet: The primary antibodies for KEAP1 (10503-2-AP, 1:5000), Nrf2 (16396-1-AP, 1:2000), glutamate-cystine ligase, modifier subunit (GCLM, 14241-1-AP, 1:2000), cyclooxygenase-2 (COX-2, 27308-1-AP, 1:1000), IL-1β (16806-1-AP, 1:500), β-actin (20536-1-AP, 1:30000), inducible Nitric Oxide Synthase (iNOS, 22226-1-AP, 1:2000), HA (51064-2-AP, 1:2000), inhibitor of NF-κB (IκB, 10268-1-AP, 1:2000), Caspase-1 (22915-1-AP, 1:2000), NLRP3 (27458-1-AP, 1:2000), α-tubulin (11224-1-AP, 1:10000), and the secondary antibodies HRP-conjugated goat anti-mouse IgG (66031-1-Ig, 1:10000) and HRP-conjugated goat anti-rabbit IgG (66031-2-Ig, 1:10000) were purchased from Proteintech Group (Wuhan, China).

Techniques: High Throughput Screening Assay, Activity Assay, Enzyme-linked Immunosorbent Assay, Staining

Journal: Journal of Advanced Research

Article Title: Tussilagone attenuated cigarette smoke-induced chronic obstructive pulmonary disease through regulating Nrf2 and NF-κB/NLRP3 inflammasome via directly targeting cysteine 434 of KEAP1

doi: 10.1016/j.jare.2025.07.019

Figure Lengend Snippet: TUS inhibited NF-κB by activating Nrf2. (A) The protein levels of p -NF-κB, IκB, iNOS, and COX-2 in the RAW 264.7 cells treated with 1 μg/mL LPS and TUS at 2.5, 5, 10, and 20 μM. (B) The regulation of TUS on p -NF-κB levels in the cytoplasm and nucleus. RAW 264.7 cells treated with 1 μg/mL LPS and TUS at 10 and 20 μM for 8 h, and the protein level of p -NF-κB in the cytoplasm and nucleus were detected using Western blot. (C) Immunofluorescence indicated that TUS inhibited p -NF-κB translocation into the nucleus. RAW 264.7 cells were treated with 1 μg/mL LPS and TUS at 10 and 20 μM for 8 h, followed by incubation with p -NF-κB primary antibody and Alex 594 secondary antibody. The immunofluorescence of p -NF-κB was observed and imaged. (D-E) The regulatory effects of TUS on protein levels of p -NF-κB, IκB, iNOS, and COX-2 in the WT and Nrf2-silenced RAW 264.7 cells. WT and Nrf2-silenced RAW 264.7 cells were treated with 10 μM TUS, and the protein levels of p -NF-κB, iNOS, and COX-2 were detected using Western blot. (F) The regulatory effects of TUS on protein levels of iNOS and COX-2 in the WT or C434A mutant RAW 264.7 cells. WT and C434A mutant RAW 264.7 cells were treated with 10 μM TUS, and the protein levels of p -NF-κB, iNOS, and COX-2 were detected using Western blot. The results were expressed as mean ± SD (n = 3). * p < 0.05, ** p < 0.01, *** p < 0.001 treated vs LPS group.

Article Snippet: The primary antibodies for KEAP1 (10503-2-AP, 1:5000), Nrf2 (16396-1-AP, 1:2000), glutamate-cystine ligase, modifier subunit (GCLM, 14241-1-AP, 1:2000), cyclooxygenase-2 (COX-2, 27308-1-AP, 1:1000), IL-1β (16806-1-AP, 1:500), β-actin (20536-1-AP, 1:30000), inducible Nitric Oxide Synthase (iNOS, 22226-1-AP, 1:2000), HA (51064-2-AP, 1:2000), inhibitor of NF-κB (IκB, 10268-1-AP, 1:2000), Caspase-1 (22915-1-AP, 1:2000), NLRP3 (27458-1-AP, 1:2000), α-tubulin (11224-1-AP, 1:10000), and the secondary antibodies HRP-conjugated goat anti-mouse IgG (66031-1-Ig, 1:10000) and HRP-conjugated goat anti-rabbit IgG (66031-2-Ig, 1:10000) were purchased from Proteintech Group (Wuhan, China).

Techniques: Western Blot, Immunofluorescence, Translocation Assay, Incubation, Mutagenesis

Journal: Journal of Advanced Research

Article Title: Tussilagone attenuated cigarette smoke-induced chronic obstructive pulmonary disease through regulating Nrf2 and NF-κB/NLRP3 inflammasome via directly targeting cysteine 434 of KEAP1

doi: 10.1016/j.jare.2025.07.019

Figure Lengend Snippet: TUS inhibited lung inflammation by activating Nrf2. (A) The images of lung tissues stained by H&E in indicated groups. (B-E) The levels of TNF-α and IL-1β in BALF of different groups. (F) The protein levels of Nrf2, iNOS, p -NF-κB, and IL-1β in the homogenate of lung tissues in indicated groups. (G) The mRNA levels of NF-κB, iNOS, NLRP3, COX-2, TNF-α, and IL-1β in the homogenate of lung tissues in indicated groups. The results were expressed as mean ± SD (n = 3). * p < 0.05, ** p < 0.01, *** p < 0.001 treated vs LPS group.

Article Snippet: The primary antibodies for KEAP1 (10503-2-AP, 1:5000), Nrf2 (16396-1-AP, 1:2000), glutamate-cystine ligase, modifier subunit (GCLM, 14241-1-AP, 1:2000), cyclooxygenase-2 (COX-2, 27308-1-AP, 1:1000), IL-1β (16806-1-AP, 1:500), β-actin (20536-1-AP, 1:30000), inducible Nitric Oxide Synthase (iNOS, 22226-1-AP, 1:2000), HA (51064-2-AP, 1:2000), inhibitor of NF-κB (IκB, 10268-1-AP, 1:2000), Caspase-1 (22915-1-AP, 1:2000), NLRP3 (27458-1-AP, 1:2000), α-tubulin (11224-1-AP, 1:10000), and the secondary antibodies HRP-conjugated goat anti-mouse IgG (66031-1-Ig, 1:10000) and HRP-conjugated goat anti-rabbit IgG (66031-2-Ig, 1:10000) were purchased from Proteintech Group (Wuhan, China).

Techniques: Staining

Journal: Clinical and Experimental Pharmacology & Physiology

Article Title: Mechanisms of IL‐17A Neutralisation in Alleviating Renal Fibrosis and Inflammation in Spontaneously Hypertensive Rats

doi: 10.1111/1440-1681.70116

Figure Lengend Snippet: Effects of IL‐17A neutralisation on macrophage polarisation in SHR renal tissues. (A) Representative IHC staining of M1 macrophage markers (iNOS and CD86) with quantitative analysis of their positive areas. (B) Representative IHC staining of M2 macrophage markers (Arg‐1 and CD163) with quantitative analysis of their positive areas. (C) Representative immunoblots and relative expression levels of iNOS, CD86, Arg‐1, and CD163 proteins. (D) Proportion of CD86 + and CD163 + cells among CD68 + macrophages. (E) mRNA expression levels of iNOS, CD86, Arg‐1, and CD163. Data are presented as mean ± SD ( n = 6).

Article Snippet: Sections were then incubated overnight at 4°C with primary antibodies against: E‐cadherin (Boster, China), Collagen III (Boster, China), inducible nitric oxide synthase (iNOS) (Boster, China), CD86 (Boster, China), arginase‐1 (Arg‐1) (Boster, China), CD163 (Boster, China), α‐smooth muscle actin (α‐SMA) (Cell Signalling Technology, USA).

Techniques: Immunohistochemistry, Western Blot, Expressing