Figures & data
Table 1. Primer sequence of target gene.
Figure 1. AS-IV improved liver function and alleviated liver injury in mice with acute alcohol-induced liver injury (n = 5–10). (A) Body weight gain. (B) Liver index. (C) Serum levels of ALT. (D) Serum levels of AST. (E) H&E staining of liver tissue samples (magnification, 200×, the arrows indicate cell degeneration, hepatocyte necrosis, and nuclear necrosis). The data are represented as means ± SEM. #p < 0.05 vs. control group, *p < 0.05 and **p < 0.01 vs. model group. ns - no significance.
Figure 2. AS-IV alleviates oxidative stress in the liver tissue of ALD mice (n = 9). (A–D) The markers of oxidative stress are SOD, GSH-Px, 4-HNE, and MDA. The data are represented as means ± SEM. #p < 0.05, ##p < 0.01, and ###p < 0.001 vs. control group, *p < 0.05, **p < 0.01, and ***p < 0.001 vs. model group.
Figure 3. AS-IV reduced the Inflammation in serum and liver tissue of ALD mice (n = 5–10). (A) The expression of F4/80 was detected by immunohistochemical staining (Brown color means positive staining) and western blotting, and presented as fold change relative to control. (B–E) concentration of serum TNF-α, IL-1β, IL-6, and MPO. (F–I) concentration of liver tissue TNF-α, IL-1β, IL-6, and MPO. The data are represented as means ± SEM. #p < 0.05, ##p < 0.01, and ###p < 0.001 vs. control group, *p < 0.05, **p < 0.01, and ***p < 0.001 vs. model group.
Figure 4. AS-IV reduced the serum levels of DAO, LPS, and LBP and improved intestinal barrier functions. (n = 6–8). (A–C) serum DAO, LPS, and LBP levels. (D–F) protein expression of Occludin and Claudin 4 in the small intestine was detected by western blotting, and presented as fold change relative to control. The data are represented as means ± SEM. ##p < 0.01 and ###p < 0.001 vs. control group, *p < 0.05, **p < 0.01, and ***p < 0.001 vs. model group.
Figure 5. AS-IV affects the composition of gut microbiota in mice with acute alcohol-induced liver injury (n = 5). (A) Venn analysis of bacterial OTUs composition between Control, Model, AS-IV-L, AS-IV-M, and AS-IV-H groups. (B,C) Principal component analysis based on weighted and unweighted UniFrac. (D) Distribution of intestinal flora at phylum and genus level, respectively.
Figure 6. The Lefse analysis among five groups (LDA score > 3, p < 0.05, FDR < 0.1). (A) cladogram in control and model groups at each classification level (Phylum, Class, Order, Family, Genus). (B) LDA results between control and model groups at each classification level (Phylum, Class, Order, Family, Genus). (C) LDA results in all groups at genus level. (D-H) The significantly changed bacteria at the genus level based on LDA results in all groups.
Figure 7. Correlation between inflammatory factors, endotoxemia, and gut microbiota. Phylum (A) and Genus (B) levels. *p < 0.05, **p < 0.01.
Figure 8. AS-IV inhibited hepatic inflammation in mice with acute alcohol-induced liver injury through NLRP3/Caspase-1 signaling pathway (n = 6–9). (A–D) the mRNA expression of NLRP3, caspase-1, IL-1β, and IL-18 in liver. (E–J) protein expression of NLRP3, Pro-Caspase-1, IL-1β, IL-18, and Cleaved-Caspase-1 in liver assayed by western blotting, and presented as fold change relative to control. The same internal reference GAPDH bands were used in and . The data are represented as means ± SEM. #p < 0.05, ##p < 0.01, and ###p < 0.001 vs. control group, *p < 0.05, **p < 0.01, and ***p < 0.001 vs. model group.
Figure 9. Schematic diagram of the protective effect of Astragaloside IV in alcohol-induced liver injury via modulating gut microbiota and regulating NLRP3/Caspase-1 signaling pathway, resulting in IL-1β and IL-18 reduction, and eventually alleviating liver inflammation. PAMP: pathogen-associated molecular patterns; NLRP3: nucleotide-binding domain-like receptor protein 3; ASC: apoptosis associated speck-like protein containing a caspase recruitment domain.
Data availability statement
The raw data supporting the conclusions of this article will be made available by the authors, without undue reservation.