pbs  (MedChemExpress)

Journal: Cell Discovery

Article Title: Breast cancer induces CD62L + Kupffer cells via DMBT1 to promote neutrophil extracellular trap formation and liver metastasis

doi: 10.1038/s41421-025-00819-8

Figure Lengend Snippet: a Co-IP assays for DMBT1 and MUC1-C in 293 T and HL60 cells. Ctrl, Ni-NTA bead control sample. b p65 phosphorylation and nuclear distribution of MUC1-C in KCs after treatment of CM of cancer cells with or without DMBT1 knockdown/overexpression. c ChIP-qPCR analysis of p65 binding to Ccl8 or Cd62l promoters in monocytes. d p65 phosphorylation and CCL8 expression of KCs cultured with DMBT1 -overexpressing MCF7 CM and/or JSH-23 (30 μM). e – g CD62L induction ( e ), Ccl8 and Cd62l expression ( f ) of KCs after treatment of LvM16 CM, with or without GO203, and the NET-inducing effects of the CM of such KCs ( g ). h – m GO203 treatment (15 mg/kg) of NOD-SCID mice with intrasplenic injection of MCF7 cells with DMBT1 overexpression. Shown are flow cytometry analysis of CD62L + KCs in livers ( h ), in vivo BLI ( i ), ex vivo BLI of livers ( j ), representative of BLI and metastases ( k ), NETosis in livers ( l ) and body weight changes of mice ( m ) at week 4 after cancer inoculation. n = 3 biological repeats ( c , e – g ), 7 ( h , i , m ), 6 ( l ) or 3 ( j ) mice. P values were obtained by two-tailed unpaired t -test (others). Scale bars, 50 μm. Data are shown as mean ± SD (others).

Article Snippet: The recombinant proteins, neutralizing antibody and reagents used in vitro and in vivo assays are human DMBT1 (11678-H08H, SinoBiological), murine CCL8 (AF790-SP, R&D), SAA1 (CSB-EP020656M0a0, CUSABIO), SAA3 (CSB-YP361411M, CUSABIO), GO203 (1222186-26-6, MCE), DNase I (4536282001, ROCHE), anti-mouse Ly6G (BE0075-1, BioXcell), GSK484 (HY-100514, MCE), estradiol valerate (S3149, Selleck), JSH-23 (749886-87-1, MCE) and DMBT1 ELISA kits (EH1756, FineTest).

Techniques: Co-Immunoprecipitation Assay, Control, Phospho-proteomics, Knockdown, Over Expression, ChIP-qPCR, Binding Assay, Expressing, Cell Culture, Injection, Flow Cytometry, In Vivo, Ex Vivo, Two Tailed Test

Journal: Respiratory research

Article Title: MUC1 attenuates neutrophilic airway inflammation in asthma by reducing NLRP3 inflammasome-mediated pyroptosis through the inhibition of the TLR4/MyD88/NF-κB pathway.

doi: 10.1186/s12931-023-02550-y

Figure Lengend Snippet: Fig. 6 TAK-242 partially ameliorates GO-203-exacerbated neutrophilic airway inflammation in OVA/LPS-induced asthmatic mouse. (A, B) HE staining and PAS staining were performed on lung sections in the indicated groups. Images were taken under a microscope at 200× magnification. Scale bar: 50 μm. (C, D) Inflammatory infiltration and hyperplasia of goblet cells were quantified using inflammation and PAS scores. (E) Statistical analysis of the total in flammatory cells, macrophages, neutrophils, eosinophils, and lymphocytes in BALF. (F) The mRNA expression levels of IL-6, IL-8, IL-18, IL-1β, and TNF-α in lung tissues were detected using RT-qPCR. LDH = lactate dehydrogenase. Data are expressed as mean ± SD (n = 6 animals per group) and were analyzed using one-way analysis of variance,*p < 0.05; **p < 0.01; ***p < 0.001; and ****p < 0.0001

Article Snippet: Mice in the OVA/LPS + GO-203 and OVA/ LPS + GO-203 + TAK-242 groups were intraperitoneally injected with the MUC1 inhibitor GO-203 (MedChemExpress, NJ, USA) (20 mg/kg) 2 h before each challenge.

Techniques: Staining, Microscopy, Expressing, Quantitative RT-PCR

Journal: Respiratory research

Article Title: MUC1 attenuates neutrophilic airway inflammation in asthma by reducing NLRP3 inflammasome-mediated pyroptosis through the inhibition of the TLR4/MyD88/NF-κB pathway.

doi: 10.1186/s12931-023-02550-y

Figure Lengend Snippet: Fig. 7 TAK-242 downregulates GO-203-caused TLR4/MyD88/NF-κB pathway activation and NLRP3 inflammasome-mediated pyroptosis in OVA/LPS- induced asthmatic mouse. (A) The protein expression levels of MUC1, TLR4, MyD88, p-p65, and p65 in lung tissue were determined via immunoblotting. (B) The expression of MUC1, TLR4, and p-p65 in the bronchi were determined via immunofluorescence staining. Images were taken using a fluorescent microscope at 400× magnification. Red: MUC1; green: TLR4 and p-p65; blue: DAPI; scale bar: 50 μm. (C) The p65 protein levels in the nuclear and cyto plasmic fractions in lung tissue were analyzed via immunoblotting. (D) The protein expression levels of NLRP3, caspase-1, cleaved caspase-1, GSDMD, GSDMD-N, IL-18, and IL-1β in lung tissue were determined via immunoblotting. (E) LDH release into BALF was detected via LDH release assay. (F) The expression of NLRP3 and caspase-1 in the bronchi were determined via immunofluorescence staining. Images were taken using a fluorescent microscope at 400× magnification. Red: NLRP3; green: caspase-1; blue: DAPI; scale bar: 50 μm. LDH = lactate dehydrogenase. Data are expressed as mean ± SD (n = 6 animals per group) and were analyzed using one-way analysis of variance, *p < 0.05; **p < 0.01; ***p < 0.001; and ****p < 0.0001

Article Snippet: Mice in the OVA/LPS + GO-203 and OVA/ LPS + GO-203 + TAK-242 groups were intraperitoneally injected with the MUC1 inhibitor GO-203 (MedChemExpress, NJ, USA) (20 mg/kg) 2 h before each challenge.

Techniques: Activation Assay, Expressing, Western Blot, Immunofluorescence, Staining, Microscopy, Lactate Dehydrogenase Assay

Journal: Respiratory research

Article Title: MUC1 attenuates neutrophilic airway inflammation in asthma by reducing NLRP3 inflammasome-mediated pyroptosis through the inhibition of the TLR4/MyD88/NF-κB pathway.

doi: 10.1186/s12931-023-02550-y

Figure Lengend Snippet: Fig. 5 TAK-242 ameliorates TLR4/MyD88/NF-κB pathway activation and NLRP3 inflammasome-mediated pyroptosis and inflammation exacerbated by MUC1 downregulation in vitro. BEAS-2B cells were treated with TAK-242 (100 nM) for 2 h, followed by LPS (5 µg/mL) stimulating for 24 h, at 24 h after MUC1-siRNA or negative control-siRNA transfection. (A) The mRNA expression level of MUC1 was detected using RT-qPCR. (B, C) The protein expression levels of MUC1, TLR4, MyD88, p-p65, and p65 were determined via immunoblotting. (D) The p65 protein levels in the nuclear and cytoplasmic fractions were analyzed via immunoblotting. (E) The expression and localization of p-p65 were observed via immunofluorescence staining. Images were taken using a fluorescent microscope at 400x magnification. Red: p-p65; blue: DAPI; scale bar: 50 μm. (F) The protein expression levels of NLRP3, caspase-1, cleaved caspase-1, GSDMD, GSDMD-N, IL-18, and IL-1β were determined via immunoblotting. (G) LDH release into the culture medium was detected via LDH release assay. (H) The mRNA expression levels of IL-6, IL-8, IL-18, IL-1β, and TNF-α were detected using RT-qPCR. Data are expressed as mean ± SD (n = 3) and were analyzed using one-way analysis of variance, *p < 0.05; **p < 0.01; ***p < 0.001; and ****p < 0.0001. LPS = lipopolysaccharide; LDH = lactate dehydrogenase

Article Snippet: Mice in the OVA/LPS + GO-203 and OVA/ LPS + GO-203 + TAK-242 groups were intraperitoneally injected with the MUC1 inhibitor GO-203 (MedChemExpress, NJ, USA) (20 mg/kg) 2 h before each challenge.

Techniques: Activation Assay, In Vitro, Negative Control, Transfection, Expressing, Quantitative RT-PCR, Western Blot, Immunofluorescence, Staining, Microscopy, Lactate Dehydrogenase Assay

Journal: Respiratory research

Article Title: MUC1 attenuates neutrophilic airway inflammation in asthma by reducing NLRP3 inflammasome-mediated pyroptosis through the inhibition of the TLR4/MyD88/NF-κB pathway.

doi: 10.1186/s12931-023-02550-y

Figure Lengend Snippet: Fig. 6 TAK-242 partially ameliorates GO-203-exacerbated neutrophilic airway inflammation in OVA/LPS-induced asthmatic mouse. (A, B) HE staining and PAS staining were performed on lung sections in the indicated groups. Images were taken under a microscope at 200× magnification. Scale bar: 50 μm. (C, D) Inflammatory infiltration and hyperplasia of goblet cells were quantified using inflammation and PAS scores. (E) Statistical analysis of the total in flammatory cells, macrophages, neutrophils, eosinophils, and lymphocytes in BALF. (F) The mRNA expression levels of IL-6, IL-8, IL-18, IL-1β, and TNF-α in lung tissues were detected using RT-qPCR. LDH = lactate dehydrogenase. Data are expressed as mean ± SD (n = 6 animals per group) and were analyzed using one-way analysis of variance,*p < 0.05; **p < 0.01; ***p < 0.001; and ****p < 0.0001

Article Snippet: Mice in the OVA/LPS + GO-203 and OVA/ LPS + GO-203 + TAK-242 groups were intraperitoneally injected with the MUC1 inhibitor GO-203 (MedChemExpress, NJ, USA) (20 mg/kg) 2 h before each challenge.

Techniques: Staining, Microscopy, Expressing, Quantitative RT-PCR

Journal: Respiratory research

Article Title: MUC1 attenuates neutrophilic airway inflammation in asthma by reducing NLRP3 inflammasome-mediated pyroptosis through the inhibition of the TLR4/MyD88/NF-κB pathway.

doi: 10.1186/s12931-023-02550-y

Figure Lengend Snippet: Fig. 7 TAK-242 downregulates GO-203-caused TLR4/MyD88/NF-κB pathway activation and NLRP3 inflammasome-mediated pyroptosis in OVA/LPS- induced asthmatic mouse. (A) The protein expression levels of MUC1, TLR4, MyD88, p-p65, and p65 in lung tissue were determined via immunoblotting. (B) The expression of MUC1, TLR4, and p-p65 in the bronchi were determined via immunofluorescence staining. Images were taken using a fluorescent microscope at 400× magnification. Red: MUC1; green: TLR4 and p-p65; blue: DAPI; scale bar: 50 μm. (C) The p65 protein levels in the nuclear and cyto plasmic fractions in lung tissue were analyzed via immunoblotting. (D) The protein expression levels of NLRP3, caspase-1, cleaved caspase-1, GSDMD, GSDMD-N, IL-18, and IL-1β in lung tissue were determined via immunoblotting. (E) LDH release into BALF was detected via LDH release assay. (F) The expression of NLRP3 and caspase-1 in the bronchi were determined via immunofluorescence staining. Images were taken using a fluorescent microscope at 400× magnification. Red: NLRP3; green: caspase-1; blue: DAPI; scale bar: 50 μm. LDH = lactate dehydrogenase. Data are expressed as mean ± SD (n = 6 animals per group) and were analyzed using one-way analysis of variance, *p < 0.05; **p < 0.01; ***p < 0.001; and ****p < 0.0001

Article Snippet: Mice in the OVA/LPS + GO-203 and OVA/ LPS + GO-203 + TAK-242 groups were intraperitoneally injected with the MUC1 inhibitor GO-203 (MedChemExpress, NJ, USA) (20 mg/kg) 2 h before each challenge.

Techniques: Activation Assay, Expressing, Western Blot, Immunofluorescence, Staining, Microscopy, Lactate Dehydrogenase Assay