Fig. 4: PirB mediates ANGPTL8-induced MDM migration and activation.

a Nuclear translocation of P65 in MDMs. Scale bar, 50 µm. b Western blot analysis of the phosphorylation of signaling molecules in MDMs. c Assessment of the tyrosine phosphorylation of the PirB protein and interactions between PirB and SHP-1/2 by Co-IP assays. d Western blot analysis of the phosphorylation of signaling molecules in MDMs pretreated with indicated antibodies. e Effect of anti-PirB antibody on cytokine expression (n = 12 cells examined over four independent experiments). f Effect of neutralizing antibodies against cytokines and PirB on lipogenesis and apoptosis of hepatocytes cocultured with MDMs in the presence of rA8 and PA (n = 4 independent experiments). g Cytokine expression of MDMs from WT or PirB^−/− mice after rA8 or scrambled protein treatment (n = 6 cells examined over three independent experiments). h Effect of anti-PirB antibody and anti-integrin-α5β1 antibody on ANGPTL8-induced MDM migration (n = 6 cells examined over three independent experiments). Scale bar, 200 µm. i PirB^−/− MDMs were resistant to ANGPTL8-induced migration (n = 6 independent experiments). j Migration of MDMs pretreated with phosphorylation inhibitors before rA8 or scrambled protein stimulation (n = 6 cells examined over three independent experiments). U0126, p-ERK1/2 inhibitor; SB203580, p-P38 inhibitor; MK-2206, p-AKT inhibitor; BAY11-7082, p-P65 inhibitor. k Experimental scheme to develop BM chimeras. l Flow cytometry analysis and quantification of KCs (CLEC2^hiF4/80^hiCD11b^lo) and MDMs (CLEC2^loCD11b^hiF4/80^lo) in the livers of the indicated BM chimeras (n = 5 or 6). The data are shown as the mean ± s.e.m. and were statistically analyzed by one-way ANOVA with Tukey’s multiple-comparison test (e, l) or two-tailed Student’s t-test (g, h, i, j). Data shown are representative of three independent experiments with similar results (a–d). Uncropped blots are provided in Source Data. All the p values were two-sided and adjustments were made for multiple comparisons. kD relative molecular weight in kilodalton, rA8 (or A8) recombinant ANGPTL8 protein, KCs Kupffer cells, MDMs monocyte-derived macrophages, PA palmitate, BM bone marrow, NS nonsignificant. Source data are available as a Source Data file.