Journal: iScience

Article Title: IL-30 protects against sepsis-induced myocardial dysfunction by inhibiting pro-inflammatory macrophage polarization and pyroptosis

doi: 10.1016/j.isci.2023.107544

Figure Lengend Snippet: IL-30 knockout aggravates sepsis-induced inflammatory responses in the heart by promoting proinflammatory macrophage polarization and pyroptosis (A–E) The cardiac mRNA levels of IL-1β, IL-6, TNF-α, MCP-1 and CXCL2 in each group (n = 8). (F) Representative flow cytometry images and corresponding quantification showing the ratio of CD11B + F4/80 + macrophages in each group (n = 6). (G) Representative flow cytometry images and corresponding quantification showing the ratio of F4/80 + Ly6C high macrophages in each group (n = 6). (H) mRNA levels of CD80, CD86 and INOS in each group (n = 8). (I) mRNA levels of CD163 and CD206 in each group (n = 8). (J) Representative immunofluorescence staining images for F4/80 and caspase1 (n = 4). (K) Representative immunoblots and corresponding quantification showing cardiac tissue levels of NLRP3, IL-1β, IL-18 and GSDMD-nt (n = 4). Values represent the mean ± SEM. ∗ indicates p < 0.05, ∗∗ indicates p < 0.01, ∗∗∗ indicates p < 0.001. IL, interleukin; MCP-1, monocyte chemoattractant protein-1; CXCL2, C-X-C motif chemokine 2; INOS, inducible nitric oxide synthase; NLRP3, NOD-like receptor thermal protein domain associated protein 3; caspase 1, cysteinyl aspartate specific proteinase-1; GSDMD-nt, N-terminal gasdermin domain-containing protein 1.

Article Snippet: Caspase1 , Santa Cruz , Cat#SC-392736.

Techniques: Knock-Out, Flow Cytometry, Immunofluorescence, Staining, Western Blot

Journal: iScience

Article Title: IL-30 protects against sepsis-induced myocardial dysfunction by inhibiting pro-inflammatory macrophage polarization and pyroptosis

doi: 10.1016/j.isci.2023.107544

Figure Lengend Snippet: IL-30 deficiency promoted LPS-induced proinflammatory macrophage polarization and pyroptosis in vitro (A) Representative flow cytometry images and corresponding quantification showing the ratio Ly6C high macrophages in each group (n = 4). (B–D) mRNA level of IL-6, TNF-α, CD80 and CD86 (n = 4). (E) Representative immunoblots and corresponding quantification showing levels of NLRP3, caspase1, IL-1β and GSDMD-nt (n = 4). Values represent the mean ± SEM. ∗ indicates p < 0.05, ∗∗ indicates p < 0.01, ∗∗∗ indicates p < 0.001. IL, interleukin; NLRP3, NOD-like receptor thermal protein domain associated protein 3; caspase 1, cysteinyl aspartate specific proteinase-1; GSDMD-nt, N-terminal gasdermin domain-containing protein 1.

Article Snippet: Caspase1 , Santa Cruz , Cat#SC-392736.

Techniques: In Vitro, Flow Cytometry, Western Blot

Journal: iScience

Article Title: IL-30 protects against sepsis-induced myocardial dysfunction by inhibiting pro-inflammatory macrophage polarization and pyroptosis

doi: 10.1016/j.isci.2023.107544

Figure Lengend Snippet: Inhibiting NLRP3 with MCC950 blocked proinflammatory macrophage polarization and pyroptosis induced by IL-30 deficiency (A and B) LVEF and FS (n = 6). (C) Cardiac CK-MB (n = 6). (D) mRNA level of IL-6 and TNF-α (n = 8). (E) Representative flow cytometry images and corresponding quantification showing the ratio of F4/80 + Ly6C high macrophages in each group (n = 6). (F) Representative immunoblots and corresponding quantification showing cardiac tissue levels of caspase1, IL-1β and GSDMD-nt (n = 4). (G) Representative flow cytometry images and corresponding quantification showing the ratio of Ly6C high macrophages in each group (n = 4). (H) mRNA level of IL-6 and INOS (n = 4). (I) Representative immunoblots and corresponding quantification showing levels of caspase1, IL-1β and GSDMD-nt (n = 4). Values represent the mean ± SEM. ∗ indicates p < 0.05, ∗∗ indicates p < 0.01, ∗∗∗ indicates p < 0.001. IL, interleukin; CK-MB, creatine kinase-myocardial band; caspase 1, cysteinyl aspartate specific proteinase-1; GSDMD-nt, N-terminal gasdermin domain-containing protein 1.

Article Snippet: Caspase1 , Santa Cruz , Cat#SC-392736.

Techniques: Flow Cytometry, Western Blot

Journal: iScience

Article Title: IL-30 protects against sepsis-induced myocardial dysfunction by inhibiting pro-inflammatory macrophage polarization and pyroptosis

doi: 10.1016/j.isci.2023.107544

Figure Lengend Snippet: Recombinant IL-30 attenuates sepsis-induced cardiac dysfunction and injury by inhibiting proinflammatory macrophage polarization and pyroptosis (A) Survival rate (n = 10). (B) Cardiac function (n = 6). (C–F) The cardiac and serum levels of CK-MB and LDH were measured in each group (n = 6). (G–J) mRNA level of IL-6, TNF-α, MCP-1 and CXCL2 (n = 8). (K) Representative flow cytometry images and corresponding quantification showing the ratio of F4/80 + Ly6C high macrophages in each group (n = 4). (L) mRNA levels of CD80, CD86 and INOS in each group (n = 8). (M) mRNA levels of CD163 and CD206 in each group (n = 8). (N) Representative immunoblots and corresponding quantification showing levels of NLRP3, caspase1, IL-1β and IL-18 (n = 4). Values represent the mean ± SEM. ∗ indicates p < 0.05, ∗∗ indicates p < 0.01, ∗∗∗ indicates p < 0.001. IL, interleukin; CK-MB, creatine kinase-myocardial band; LDH, lactate dehydrogenase; MCP-1, monocyte chemoattractant protein-1; CXCL2, C-X-C motif chemokine 2; INOS, inducible nitric oxide synthase; NLRP3, NOD-like receptor thermal protein domain associated protein 3; caspase 1, cysteinyl aspartate specific proteinase-1.

Article Snippet: Caspase1 , Santa Cruz , Cat#SC-392736.

Techniques: Recombinant, Flow Cytometry, Western Blot

Journal: iScience

Article Title: IL-30 protects against sepsis-induced myocardial dysfunction by inhibiting pro-inflammatory macrophage polarization and pyroptosis

doi: 10.1016/j.isci.2023.107544

Figure Lengend Snippet: IL-30 inhibited proinflammatory macrophage polarization and pyroptosis in vitro (A) Representative flow cytometry images and corresponding quantification showing the ratio of Ly6C high macrophages in each group (n = 4). (B) mRNA levels of IL-6 and INOS (n = 4). (C) Representative immunoblots and corresponding quantification showing levels of NLRP3, caspase1, IL-1β and IL-18 (n = 4). (D) Representative immunoblots and corresponding quantification showing levels of JAK1, JAK2, STAT1, STAT3, P38, ERK, JNK and P65 (n = 6). Values represent the mean ± SEM. ∗ indicates p < 0.05, ∗∗ indicates p < 0.01, ∗∗∗ indicates p < 0.001, ns indicates no significance.

Article Snippet: Caspase1 , Santa Cruz , Cat#SC-392736.

Techniques: In Vitro, Flow Cytometry, Western Blot

Journal: iScience

Article Title: IL-30 protects against sepsis-induced myocardial dysfunction by inhibiting pro-inflammatory macrophage polarization and pyroptosis

doi: 10.1016/j.isci.2023.107544

Figure Lengend Snippet:

Article Snippet: Caspase1 , Santa Cruz , Cat#SC-392736.

Techniques: Staining, Recombinant, Liposomes, Bicinchoninic Acid Protein Assay, AST Assay, Lactate Dehydrogenase Assay, cDNA Synthesis, Enzyme-linked Immunosorbent Assay, Knock-Out, Software

Journal: Journal of immunology (Baltimore, Md. : 1950)

Article Title: Gasdermin D Drives the Nonexosomal Secretion of Galectin-3, an Insulin Signal Antagonist.

doi: 10.4049/jimmunol.1900212

Figure Lengend Snippet: FIGURE 1. Identification of galectin-3 as an inflammasome-regulated secretory protein. (A) Diagram of secreted proteins in CM from LPS/ATP–treated WT and Nlrp32/2 BMDMs. (B) Two-dimensional PAGE analysis of supernatant from WT and Nlrp3+/R258W BMDMs treated with 500 ng/ml LPS for 4 h. The labeled dots were dug out for MS analysis. (C) Protein list of identified proteins from (B) by MS. The blue-labeled proteins are at least 2-fold lower in Nlrp32/2 CM than that in WT CM of (A). p # 0.05. (D) Volcano plot of secreted proteins in (A). The pink circles represent individual protein at least 2-fold higher or lower in WT CM than that in Nlrp32/2 CM, with p # 0.05. (E) ELISA detection of serum galectin-3 level in WT mice (n = 5), which received i.p. injection of Ac-YVAD (20 mg/kg) or vehicle daily for 3 d. Data are shown as mean 6 SEM. ***p # 0.001. (F) ELISA detection of serum galectin-3 level in WT and Casp1/112/2 mice (n = 4). Data are shown as mean 6 SEM. ***p # 0.001.

Article Snippet: Casp1 inhibitor Ac-YVAD-CMK (catalog no. sc-300323; Santa Cruz Biotechnology) or galectin-3 inhibitor TD139 (catalog no. HY-19940; MedChemExpress) was administrated daily by i.p. injection.

Techniques: Labeling, Enzyme-linked Immunosorbent Assay, Injection

Journal: Journal of immunology (Baltimore, Md. : 1950)

Article Title: Gasdermin D Drives the Nonexosomal Secretion of Galectin-3, an Insulin Signal Antagonist.

doi: 10.4049/jimmunol.1900212

Figure Lengend Snippet: FIGURE 2. The inflammasome activation drives cellular and systemic secretion of galectin-3. (A) BMDMs from WT or Nlrp3+/R258W mice were stimulated with 500 ng/ml LPS for the indicated times; supernatant (SUP) and whole cell lysate (WCL) were collected for immunoblotting of the indi- cated proteins. Data are representative of at least three independent experiments. (B) LPS-primed WT BMDMs were treated with ATP (5 mM) or nigericin (5 mg/ml) for 30 min, and immunoblotting detection of indicated proteins is shown. Data are representative of at least three independent experiments. (C) LPS-primed peritoneal macrophages were stimulated with ATP (5 mM) for 30 min. Indicated proteins were detected via immunoblotting assay. Data are representative of at least two independent experiments. (D) LPS-primed BMDC were stimulated with ATP (5 mM) for 30 min. Indicated proteins were detected via immunoblotting assay. Data are representative of at least three independent experiments. (E) BMDM were primed with LPS for 10 min, followed by ATP (5 mM) challenge for 30 min. Indicated proteins were detected via immunoblotting assay. Data are representative of at least three in- dependent experiments. (F) Western blotting analysis of SUP and WCL from WT, Nlrp32/2, Asc2/2, or Casp1/112/2 BMDMs stimulated with LPS/ATP. Data are representative of at least three independent experiments. (G) Western blotting analysis of SUP and WCL from LPS-primed THP-1 macrophages stimulated with ATP. Data are representative of at least three independent experiments. (H) Western blotting analysis of SUP and WCL from unprimed WT or Aim22/2 BMDMs transfected with poly(dA:dT) for 2 h. Data are representative of at least two independent experiments. (I) Western blotting analysis of SUP and WCL from unprimed WT or Nlrc42/2 BMDMs infected with S. typhimurium (multiplicity of infection = 10) for 3 h. Data are representative of at least two independent experiments. (J) Western blotting analysis of SUP and WCL from unprimed THP-1 macrophages transfected with poly(dA:dT) for 4 h. Data are representative of at least three independent experiments. (K and L) ELISA analysis of serum galectin-3 (K) and IL-18 (L) level in WT and Nlrp32/2, Aim22/2, Nlrc42/2, and Asc2/2 mice (n = 4); the WT group is the same one used in Fig. 1F. Data are shown as mean 6 SEM. ns: p . 0.05, **p # 0.01, ****p , 0.0001.

Article Snippet: Casp1 inhibitor Ac-YVAD-CMK (catalog no. sc-300323; Santa Cruz Biotechnology) or galectin-3 inhibitor TD139 (catalog no. HY-19940; MedChemExpress) was administrated daily by i.p. injection.

Techniques: Activation Assay, Western Blot, Transfection, Infection, Enzyme-linked Immunosorbent Assay