Fig 1: Nicotine activates the pyroptosis pathway in BMDMs by upregulating ROS production. (A, B) Western blot was used to detect the expression of TXNIP, NLRP3, ASC, caspase-1, and GSDMD. The average ratios were calculated based on gray intensity analysis (*P < 0.05, **P < 0.01, *** P < 0.001, **** P < 0.0001, n = 4). (C) IL-1ß and IL18 in the supernatant of BMDMs treated with or without nicotine were measured by ELISA (*P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001, n=5).
Fig 2: Protein levels of cleaved caspase-1 are upregulated and TXNIP and NLRP3 staining is strong in sepsis-induced myocardial injury in rats. (A) Protein levels of cleaved caspase-1 and caspase-1 were assessed by western blotting. (B) Relative levels of proteins were determined with GAPDH for normalization. (C) TXNIP and NLRP3 were stained using immunochemistry. **P<0.01 vs. Con. TXNIP, thioredoxin-interacting protein; NLRP3, NOD-like receptor pyrin domain containing 3; LPS, lipopolysaccharide; Con, control.
Fig 3: Downregulation of TXNIP reversed the cisplatin-induced ultramicroscopic changes in MCs, similar to that after downregulating NLRP3. (A) The morphologic changes of MCs after different treatments were evaluated under TEM. The red arrows indicate the ruptured cell membrane and local lighter-stained cytoplasm. Scale bars: 2 µm. (B) The morphological changes of MCs were observed by SEM. The red arrows indicate the withered tree-like cell membrane at the edge of the cell. Scale bars: 15 µm. Representative results of at least three repeated experiments are shown.
Fig 4: LR reduces oxidative stress through suppressing the TXNIP/NLRP3 pathway in LPS-treated cells. The lung epithelia cells of BEAS-2B were pretreated with different concentrations of LR (0, 20, 40 and 80 µM) for 24 h, followed by LPS exposure for 1 h. Then, all cells were harvested for the following research. (A) 2'-7'-Dichlorofluorescein (DCF) analysis was used to calculate ROS generation, and the representative images were displayed. The scale bar is 50 µm. (B) The quantification of ROS production was exhibited. (C) Western blot analysis of XO, TXNIP, LPRP3, apoptosis-associated speck-like protein containing a C-terminal caspase recruitment domain (ASC) and caspase-1 in LPS-treated cells in the presence or absence of LR. (D) p-p38, p-ERK1/2 and p-JNK levels were measured through western blot analysis. Data are represented as mean ± standard error of the mean of three independent experiments (n=6). +++P<0.001 vs. the Con group in the absence of any treatments. **P<0.01 and ***P<0.001 vs. the LPS groups. LR, linarin; LPS, lipopolysaccharide; XO, xanthine oxidase; ERK, extracellular signal-regulated kinase; JNK, c-Jun N-terminal kinase; ROS, reactive oxygen species; Con, control.
Fig 5: LR inactivates TXNIP/NLRP3 signaling pathway in the lung tissues of mice with LPS challenge. (A) xanthine oxidase (XO) and (B) TXNIP expression levels were evaluated through immunohistochemical analysis. The scale bar is 100 µm. (C) The immunoblotting analysis of XO and TXNIP. (D) Western blot analysis was conducted to determine NLRP3, apoptosis-associated speck-like protein containing a C-terminal caspase recruitment domain (ASC) and caspase-1 protein levels in the lung tissue samples of mice treated under various conditions. Data are represented as mean ± standard error of the mean of three independent experiments (n=6). +++P<0.001 vs. the Con group in the absence of any treatments. **P<0.01 and ***P<0.001 vs. the LPS groups. LR, linarin; LPS, lipopolysaccharide; XO, xanthine oxidase; Con, control; LC-LR, low concentration linarin; MC-LR, medium concentration linarin; HC-LR, high concentration linarin.
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