Fig. 8.

Macrophage TXNIP deficiency-mediated OASL1 inhibits stress-induced hepatocyte death via modulating Apaf1/Cyt c/caspase-9 activation.

BMMs were isolated from TXNIP^M-KO mice and transfected with the p-CRISPR-OASL1 KO or control vector followed by LPS stimulation. (A) ELISA analysis of supernatant TNF-α levels in LPS-stimulated BMMs (n = 4 samples/group). (B) The schematic figure for the macrophage/hepatocyte coculture system. (C) BMMs transfected with the p-CRISPR-OASL1 KO were stimulated with LPS and then cocultured with primary hepatocytes supplemented with or without H2O2 for 24 h. Western-assisted analysis and relative density ratio of Apaf-1, Cyt c, cleaved caspase-9, cleaved caspase-3, and RIPK3 in hepatocytes after coculture. Representative of 4 experiments. (D) LDH release from hepatocytes in cocultures (n = 4 samples/group). (E) Immunofluorescence staining of TUNEL⁺ hepatocytes after coculture (n = 4 samples/group). Scale bars, 100 μm. All data represent the mean ± SD. Statistical analysis was performed using Permutation t-test. ∗p <0.05, ∗∗p <0.01, ∗∗∗p <0.001. Apaf1, apoptotic peptidase activating factor 1; BMM, bone marrow-derived macrophage; CRISPR, clustered regularly interspaced short palindromic repeats; Cyt c, cytochrome c; KO, knockout; LPS, lipopolysaccharide; OASL1, 2′,5′ oligoadenylate synthetase-like 1; TNF-α, tumour necrosis factor-alpha; TXNIP, thioredoxin-interacting protein; TXNIP^M-KO, myeloid-specific TXNIP KO; RIPK3, receptor-interacting serine/threonine-protein kinase 3.