Journal: Cell Death Discovery

Article Title: Hepatic TNFRSF12A promotes bile acid-induced hepatocyte pyroptosis through NFκB/Caspase-1/GSDMD signaling in cholestasis

doi: 10.1038/s41420-023-01326-z

Figure Lengend Snippet: A TNFRSF12A mRNA transcripts, and ( B ) protein levels in PLC/RPF/5- ASBT cells treated with 100 μM TCA, TCDCA, GCA, GCDCA, or TCDCA. * p < 0.05 compared to control (DMSO); Time-dependent effects of TCA on TNFRSF12A ( C ) mRNA and ( D ) protein expression in PLC/RPF/5- ASBT cells; ( E ) Western blot analysis of effects of conjugated BAs (TCA, GCA, GCDCA, TDCAC, or TCDCA, 100 μM) on protein expression of pro-Caspase-1, cleaved-Caspase-1, GSDMD, cleaved-GSDMD, caspase-3, Bcl2, and Bax in PLC/RPF/5- ASBT cells; F Representative images of TEM showing features of pyroptosis, including membrane pores, cell swelling, and large bubbles blowing from the plasma membrane in PLC/RPF/5- ASBT cells treated with 100 μM TCA (scale bar: 5 µm).

Article Snippet: Before stimulation with LPS (100 ng/ml), TCA (100 μM), or DMSO, PLC/RPF/5- ASBT cells were pretreated with 10 μM BAY 11-7082, a selective inhibitor of NFκB signaling, for 30 min and treated with 100 μM glycocholate acid (GCA), glycocholic acid (GCDCA), taurocholic acid (TCA), taurochenodeoxycholic acid (TCDCA) or taurohyodeoxycholic acid (TDCA) for 24 h. THP-1 cells or THP-1-derived macrophages induced by phorbol 12-myristate 13-acetate (PMA) (MedChemExpress, USA; Cat# 16561-29-8) [ ] were treated with conjugated BAs or used for transwell assays.

Techniques: Control, Expressing, Western Blot, Membrane, Clinical Proteomics

Journal: Cell Death Discovery

Article Title: Hepatic TNFRSF12A promotes bile acid-induced hepatocyte pyroptosis through NFκB/Caspase-1/GSDMD signaling in cholestasis

doi: 10.1038/s41420-023-01326-z

Figure Lengend Snippet: A Serum TWEAK levels in control patients ( n = 20) and OC patients ( n = 34). * p < 0.05 vs. control patients; B The levels of TWEAK mRNA transcripts in PLC/RPF/5- ASBT cells or primary mouse hepatocytes treated with 100 μM TCA, GCA, GCDCA, TDCAC, and TCDCA; ( C ) Immunofluorescence labeling of F4/80 (a specific marker for macrophages) in a human control liver and an OC liver; D The levels of TWEAK mRNA transcripts in THP1 cells treated with 100 μM GCA. * p < 0.05 vs. DMSO control group; E A diagram for transwell assays (left) and migration ability of co-cultured THP1-derived macrophages (right) under conjugated BA stimulation; F The TWEAK levels in cell supernatant of co-cultured THP-derived macrophages and primary mouse hepatocytes following treatment with DMSO as control, GCA or TCDCA; G The levels of Cxcl2, Ccl2, IL-18, IL-1β, and Tweak mRNA transcripts in co-cultured primary mouse hepatocytes following treatment with DMSO as control, GCA or TCDCA. * p < 0.05 vs. DMSO control group; H The levels of Tnfrsf12a, Nlrp3, cleaved-Caspase-1, cleaved-GSDMD, and cleaved- IL-1β in primary mouse hepatocytes treated with DMSO as control, GCA, and/or TWEAK. OC, obstructive cholestasis.

Article Snippet: Before stimulation with LPS (100 ng/ml), TCA (100 μM), or DMSO, PLC/RPF/5- ASBT cells were pretreated with 10 μM BAY 11-7082, a selective inhibitor of NFκB signaling, for 30 min and treated with 100 μM glycocholate acid (GCA), glycocholic acid (GCDCA), taurocholic acid (TCA), taurochenodeoxycholic acid (TCDCA) or taurohyodeoxycholic acid (TDCA) for 24 h. THP-1 cells or THP-1-derived macrophages induced by phorbol 12-myristate 13-acetate (PMA) (MedChemExpress, USA; Cat# 16561-29-8) [ ] were treated with conjugated BAs or used for transwell assays.

Techniques: Control, Immunofluorescence, Labeling, Marker, Migration, Cell Culture, Derivative Assay

Journal: Cell Death Discovery

Article Title: Hepatic TNFRSF12A promotes bile acid-induced hepatocyte pyroptosis through NFκB/Caspase-1/GSDMD signaling in cholestasis

doi: 10.1038/s41420-023-01326-z

Figure Lengend Snippet: A Western blot analysis of protein levels of transcription factors c-JUN, JUN-D, and SMAD2/3 in nuclear extracts from human control and OC livers. * p < 0.05 vs. control patients; B Western blot analysis of nuclear expression of transcription factor c-JUN in the PLC/RPF/5- ASBT cells treated with 100 μM TCA, TCDCA, GCA, GCDCA, or TDCA. C Western blot analysis of nuclear expression of transcription factor c-JUN in PLC/RPF/5- ASBT cells treated with 0, 10, 25, 50, 100 μM TCA; ( D ) ChIP assay data showed increased activities of c-JUN binding to the TNFRSF12A promoter (ChIP site 1) in a dose-dependent manner in PLC/RPF/5- ASBT cells and ( E ) in liver tissues of patients with obstructive cholestasis, compared with control patients. * p < 0.05 vs. their corresponding controls; F A schematic diagram illustrating the potential mechanism by which BAs induced TNFRSF12A expression to trigger hepatocyte pyroptosis in cholestatic liver injury.

Article Snippet: Before stimulation with LPS (100 ng/ml), TCA (100 μM), or DMSO, PLC/RPF/5- ASBT cells were pretreated with 10 μM BAY 11-7082, a selective inhibitor of NFκB signaling, for 30 min and treated with 100 μM glycocholate acid (GCA), glycocholic acid (GCDCA), taurocholic acid (TCA), taurochenodeoxycholic acid (TCDCA) or taurohyodeoxycholic acid (TDCA) for 24 h. THP-1 cells or THP-1-derived macrophages induced by phorbol 12-myristate 13-acetate (PMA) (MedChemExpress, USA; Cat# 16561-29-8) [ ] were treated with conjugated BAs or used for transwell assays.

Techniques: Western Blot, Control, Expressing, Binding Assay