Journal: Immunity, Inflammation and Disease

Article Title: Kaempferitrin Attenuates Lipopolysaccharide‐Induced Cardiac Dysfunction Through Suppression of the NF‐κB/NLRP3 Signaling Pathway

doi: 10.1002/iid3.70323

Figure Lengend Snippet: Kae inhibited LPS‐induced cardiomyocyte inflammation and the activation of NF‐κB/NLRP3/pyroptosis pathway in vitro. (A–C) Transcriptome sequencing of myocardial tissue from LPS group and LPS + Kae group. It showed a volcano plot (A), GSEA analysis of NF‐κB pathway (B), and a heat map of genes involved in pyroptosis and NF‐κB signaling pathway (C), N = 3. (D) CCK8 assay evaluating the appropriate concentration of Kaempferitrin Kae). (E) The brief experimental setup of the in vitro experiment. After maltreatment with Kaempferitrin for 2 h, cardiomyocytes were then exposed to LPS (1 μg/L) for 12 h. (F) The relative RNA levels of Il‐1β, Il‐6 , and Tnf‐ α in H9c2 cells. (G) The relative protein levels of NLRP3, p65, p‐p65, Ik‐Bα, p‐Ik‐Bα and GAPDH in H9c2 cell were detected using western blot assay. (H) The densitometric quantification of NLRP3, p‐Ik‐Bα/Ik‐Bα and p‐P65/P65 in H9c2 cells. (I) The relative mRNA levels of Il‐1β , Il‐6 , and Tnf‐α in AC16 cells. (J) The relative protein levels of NLRP3, p65, p‐p65, Ik‐Bα, p‐Ik‐Bα, and GAPDH in AC16 cell were detected using western blot assay. (K) Densitometric quantification of NLRP3, p‐Ik‐Bα/Ik‐Bα, and p‐P65/P65 in AC16 cells. (L, M) The relative protein levels of Cleaved‐GSDMD, Cleaved‐caspase1, and GAPDH in H9c2 cells (L) and AC16 cells (M) were detected using western blot assay. N = 3. (One‐way analysis of variance [ANOVA] followed by Bonferroni post hoc test for comparing multiple groups. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001, ns = not significant).

Article Snippet: To evaluate the impact of pyroptosis in Kae‐mediated cardioprotection against LPS‐induced cardiotoxicity, the caspase‐1 inhibitor VX765 (MCE, #HY‐13205, 25 μM) [ ] was utilized in accordance with the manufacturer's instructions.

Techniques: Activation Assay, In Vitro, Sequencing, CCK-8 Assay, Concentration Assay, Western Blot

Journal: Immunity, Inflammation and Disease

Article Title: Kaempferitrin Attenuates Lipopolysaccharide‐Induced Cardiac Dysfunction Through Suppression of the NF‐κB/NLRP3 Signaling Pathway

doi: 10.1002/iid3.70323

Figure Lengend Snippet: Kaempferitrin may ameliorate LPS‐induced cardiac injury by inhibiting pyroptosis via the NF‐κB/NLRP3 pathway in vivo . (A) The relative protein levels of NLRP3, P65, p‐P65, IkBα, p‐IkBα, and GAPDH in the heart tissue were detected using western blot assay. (B)The relative protein levels of GSDMD, Cleaved‐caspase1, and GAPDH in the heart tissue were detected using western blot assay. C‐E, Densitometric quantification of NLRP3 (C), p‐IkBα/IkBα (D), and PP65/P65 (E). (F, G) Densitometric quantification of Cleaved‐GSDMD and Cleaved‐caspase 1. (H, I) Levels of IL‐1β (H), IL‐18 (I) in heart tissue detected by ELISA assay. N = 6. (One‐way analysis of variance [ANOVA] followed by Bonferroni post hoc test for comparing multiple groups. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001, ns = not significant).

Article Snippet: To evaluate the impact of pyroptosis in Kae‐mediated cardioprotection against LPS‐induced cardiotoxicity, the caspase‐1 inhibitor VX765 (MCE, #HY‐13205, 25 μM) [ ] was utilized in accordance with the manufacturer's instructions.

Techniques: In Vivo, Western Blot, Enzyme-linked Immunosorbent Assay

Journal: Immunity, Inflammation and Disease

Article Title: Kaempferitrin Attenuates Lipopolysaccharide‐Induced Cardiac Dysfunction Through Suppression of the NF‐κB/NLRP3 Signaling Pathway

doi: 10.1002/iid3.70323

Figure Lengend Snippet: Kaempferitrin improves LPS‐induced myocardial inflammatory damage by inhibiting the NF‐κB/NLRP3/pyroptosis pathway. After pretreatment with Kaempferitrin, VX765 or Kaempferitrin+VX765 for 2 h, cardiomyocytes were then exposed to LPS (1 μg/mL) for 12 h. (A) The relative protein levels of GSDMD, Cleaved‐caspase1 and GAPDH in H9c2 cell were detected using western blot assay. The densitometric quantification of Cleaved‐GSDMD (B), and Cleaved‐Caspase1 (C) in H9c2 cells. (D) The relative mRNA levels of Il‐1β , Il‐6 , and Tnf‐α in H9c2 cells. (E) The relative protein levels of GSDMD, Cleaved‐caspase1, and GAPDH in AC16 cell were detected using western blot assay. The densitometric quantification of Cleaved‐GSDMD (F) and Cleaved‐Caspase1 (G) in AC16 cells (H). N = 3. (One‐way analysis of variance [ANOVA] followed by Bonferroni post hoc test for comparing multiple groups. * p < 0.05, ** p < 0.01, ns = not significant).

Article Snippet: To evaluate the impact of pyroptosis in Kae‐mediated cardioprotection against LPS‐induced cardiotoxicity, the caspase‐1 inhibitor VX765 (MCE, #HY‐13205, 25 μM) [ ] was utilized in accordance with the manufacturer's instructions.

Techniques: Western Blot

Journal: Immunity, Inflammation and Disease

Article Title: Kaempferitrin Attenuates Lipopolysaccharide‐Induced Cardiac Dysfunction Through Suppression of the NF‐κB/NLRP3 Signaling Pathway

doi: 10.1002/iid3.70323

Figure Lengend Snippet: NLRP3 is required for Kae to defend against cardiotoxicity induced by LPS in vivo. The WT mice and NLRP3 knockout mice were exposed to LPS (10 mg/kg) for 12 h. (A) Representative transthoracic echocardiography M‐model images from each group. (B) EF% and (C) FS% of the mice in each group. (D, E) Serum levels of CK‐MB and LDH in mice from each group. (F) Representative images of HE staining and TUNEL staining for heart tissue. Scale bar = 50 μm. N = 6. (G) The relative protein levels of Cleaved‐GSDMD, Cleaved‐caspase1, and GAPDH in the heart tissue were detected using western blot assay. Densitometric quantification of C‐GSDMD (H) and C‐Caspase 1 (I) were shown. N = 6. (One‐way analysis of variance [ANOVA] followed by Bonferroni post hoc test for comparing multiple groups. * p < 0.05, ** p < 0.01, **** p < 0.0001, ns = not significant).

Article Snippet: To evaluate the impact of pyroptosis in Kae‐mediated cardioprotection against LPS‐induced cardiotoxicity, the caspase‐1 inhibitor VX765 (MCE, #HY‐13205, 25 μM) [ ] was utilized in accordance with the manufacturer's instructions.

Techniques: In Vivo, Knock-Out, Staining, TUNEL Assay, Western Blot

Journal: The EMBO Journal

Article Title: Cardiolipin inhibits the non-canonical inflammasome by preventing LPS binding to caspase-4/11

doi: 10.1038/s44318-025-00507-z

Figure Lengend Snippet: ( A ) Human monocyte-derived macrophages (HMDM) from healthy donors or ( B ) Bone marrow-derived macrophages (BMDM) from wild-type (WT) mice were incubated for 4 h with 1 µg/mL Pam 3 CSK 4 . Cell culture medium was replaced with OptiMEM plus HEPES (−), 10 µM CL, or 5 µg/mL of RS-LPS (complexed with LTX) in the absence or presence of 2 µg/mL of LPS complexed with LTX (A) or CTB ( B ). Cells were incubated for 4 h (A) or 18 h ( B ). Cell supernatants were analysed for cleaved IL-1β (ELISA) and LDH (cytotoxicity assay). ( C , D ) BMDM from Nlrp3 −/− mice were incubated for 4 h with 1 µg/mL Pam 3 CSK 4 before cell culture medium was replaced with OptiMEM plus HEPES (Ctrl), or 2 µg/mL of LPS complexed with CTB in the presence of HEPES (−), or 10 µM CL (CL). Cells were incubated for 18 h. GSDMD cleavage and tubulin expression were assessed in mixed supernatants and lysates by western blot ( C ). LDH release was quantified by cytotoxicity assay ( D ). ( E ) HMDM from healthy donors or ( F – H) Human bronchial epithelial cells (HBEC) were incubated for 4 h ( E ) or 18 h ( F – H ) with 1 µg/mL Pam 3 CSK 4 . Cell culture medium was replaced with OptiMEM with vehicle (DMSO) or 10 µM MCC950 (NLRP3 inhibitor), or 10 µM VX765 (CASP1/4 inhibitor). Cells were incubated for 1 h, and then 2 µg/mL of LPS complexed with LTX was added in the presence of HEPES (−) or 10 µM CL (CL). Cells were incubated for 4 h. Cell supernatants were analysed for LDH ( E , F ) (cytotoxicity assay), cleaved IL-1β ( G ) and IL-18 ( H ) (ELISA). Data information: Each symbol is the mean of technical triplicates from an independent biological replicate. Bars are the mean of three or more independent biological replicates ( n = 3–5) ± SEM. Statistical analysis: Data were verified for normality using a Shapiro–Wilk test and analysed by ( A , B , E – H ) one-way ANOVA Šídák’s multiple comparisons test, ( D ) paired t test. P values are reported above bars. Statistical significance was defined as follows: significant difference for P < 0.05 (*), not significant for P ≥ 0.05 (ns). .

Article Snippet: After 4 h, the cell culture medium was replaced with OptiMEM alone or containing 10 μM of the NLRP3 inhibitor MCC950 or the CASP1/4 inhibitor VX765 (MedChem Express).

Techniques: Derivative Assay, Incubation, Cell Culture, Enzyme-linked Immunosorbent Assay, LDH Cytotoxicity Assay, Expressing, Western Blot, Cytotoxicity Assay