Fig 1: NK Ameliorated ALD via Enhanced Nur77-Mediated P2X7r Signaling and Lipid Accumulation. Representative western blotting analysis for expression of (A) Nur77, (B) P2X7r, (C) NLRP3, (D) Lipin-1, and (E) SREBP1. GAPDH was used as the loading control. aaa p < 0.001 vs. EtOH group. Representative western blotting analysis for expression of (F) Nur77, (G) P2X7r, caspase-1, and (H) Lipin-1. GAPDH was used as the loading control. bbb p < 0.001 vs. EtOH group. (I) Immunofluorescence staining of Lipin-1 presented at 600× magnification. Representative western blotting analysis for expression of (J) Nur77, (K) P2X7r, NLRP3, (L) caspase-1, and (M) Lipin-1. GAPDH was used as the loading control. c p < 0.05, cc p < 0.01, ccc p < 0.001 siControl vs. siControl-NK, siRNA (Nur77)-NK groups; dd p < 0.01, ddd p < 0.001 siRNA (Nur77) vs. siRNA (Nur77)-NK group; e p < 0.05, eee p < 0.001 siControl-NK vs. siRNA (Nur77)-NK group.
Fig 2: NK Attenuates the Inflammatory Response to Macrophage-to-Hepatocyte Communication. (A) MTT assay measuring cell viability of MPMs and mouse primary hepatocytes with NK treatment. * p < 0.05 vs. negative control; (B) The IL-1β and IL-6 protein released from MPMs into culture medium was measured by an ELISA assay. Representative western blotting analysis for expression of (C) Nur77, (D) P2X7r, (E) caspase-1, (F) IL-1β and IL-23. GAPDH was used as the loading control. ## p < 0.01, ### p < 0.001 vs. normal group; ** p < 0.01, *** p < 0.001 vs. LPS+ATP group. (G) Immunofluorescence staining of NLRP3 and IL-23 presented at 600× magnification. Representative western blotting analysis for expression of (H) P2X7r, IL-23, (I) ASC, and caspase-1 in primary hepatocytes. Densitometric values were normalized to GAPDH. hhh p < 0.001 vs. normal group; zzz p < 0.001 vs. LPS+ATP group; ns, not significant.
Fig 3: NK Regulates Lipid Deposition and Inflammatory Response in EtOH-Stimulated AML-12 Cells. (A) Chemical structure of Nodakenin. (B) MTT assay measuring cell viability of AML-12 with NK treatment. (C) The IL-1β and IL-6 protein levels released from AML-12 cells into culture medium were determined by an ELISA assay. (D) Real-time PCR was used to determine the mRNA expression of fasn, p2x7r, tnf-α, and Il-18. (E) Representative western blotting analysis for expression of PPARα, Lipin-1, and SREBP1. (F) Representative western blotting analysis for expression of Nur77, P2X7r, and IL-23. (G) Representative western blotting analysis for expression of NLRP3, ASC, and caspase-1. GAPDH was used as the loading control. (H) Immunofluorescence staining of Nur77 and P2X7r presented at 600× magnification. # p < 0.05, ## p < 0.01, ### p < 0.001 vs. normal group; * p < 0.05, ** p < 0.01, *** p < 0.001 vs. TGF-β group.
Fig 4: Nur77 May Be Involved in the Regulatory Effects of NK against the P2X7r-Mediated Inflammatory Response in ALD Mouse Livers. (A) Representative western blotting analysis for expression of Nur77 and P2X7r. (B) Representative western blotting analysis for expression of NLRP3 and caspase-1. (C) Representative western blotting analysis for expression of IL-23 and IL1R1. (D) Representative western blotting analysis for expression of MPO. (E) Immunofluorescence staining of P2X7r and NLRP3 presented at 600× magnification. Immunofluorescence staining of MPO presented at 600× magnification. ### p < 0.001 vs. normal group; ** p < 0.01, *** p < 0.001 vs. EtOH group; ns, not significant.
Fig 5: NK ameliorates hepatic fibrosis in a manner potentially involving Nur77-mediated P2X7r inflammatory signaling and lipogenesis.
Supplier Page from Abcam for Recombinant Human NUR77 protein (GST tag N-Terminus)