Figure 1.
The molecular structure of Esculentoside A (EsA).
Figure 2.
Measurement of EsA cytotoxicity and effects of EsA on CCl4-induced LO2 cell injury in vitro.
The EsA cytotoxicity was measured using CCK-8 assays (A). Levels of TNF-α in LO2 culture mediums challenged by CCl4 increased to approximately 4-fold and which was dramatically prevented by EsA treatment, and the concentration of 2.5 mg/L reduced the level of TNF-α most obviously (B). The cell death rate was observed by flow cytometric analysis (C). The values presented are the means ± standard error of the mean (n = 5). ##P<0.01 versus the Control group. *P<0.05, **P<0.01 versus the Injury group.
Figure 3.
Effects of EsA on CCl4-induced LO2 cell injury and PPAR-γ expression.
The treatment effects of EsA and protein expression of PPAR-γ were measured using western blot (A). Levels of ROS in LO2 cells challenged by CCl4 were shown (B Magnification, 200×). The mRNA expression of PPAR-γ was measured using quantitative real-time PCR (C). The values presented are the means ± standard error of the mean (n = 5). *P<0.05, **P<0.01.
Figure 4.
EsA protected against CCl4-induced histopathological damage and hepatic dysfunction.
Hematoxylin and eosin staining (A Magnification, 200×) showed that livers in Injury group exhibited more ballooned hepatocytes than those in Control group and EsA group, and symptoms of those histopathological damage were significantly alleviated by EsA treatment (n = 6). Photographs of livers were taken 12 hours post-CCl4 injection, livers in Injury group turned white (B). Levels of AST and ALT increased obviously after CCl4 challenge. However, AST and ALT levels did not markedly increase in mice treated with EsA alone, and AST and ALT levels were significantly decreased with EsA treatment. (n = 6 C). The values presented are the means ± standard error of the mean. ##P<0.01 versus the Control group. *P<0.05, **P<0.01 versus the Injury group.
Figure 5.
Effects of EsA on CCl4-induced liver oxidative stress.
EsA treatment significantly decreased levels of MDA (A) and increased the activity of GSH-Px (B) compared with the Injury group. The values presented are the means ± standard error of the mean (n = 6). ##P<0.01 versus the Control group. *P<0.05 versus the Injury group.
Figure 6.
Effects of EsA on CCl4-induced liver inflammation.
mRNA expression of TNF-a, IL-1β and IL-6 (A) and Immunohistochemical staining of F4/80 and CD11b cells (B) accumulating in liver tissues were determined at 12 hours post CCl4-induced acute liver injury (Magnification, 200×). The values presented are the means ± standard error of the mean (n = 6). ##P<0.01 versus the Control group. *P<0.05, **P<0.01 versus the Injury group.
Figure 7.
The underlying mechanism of EsA against CCl4-induced acute liver injury in mice.
The activity of ERK (A) and IκB (B) were determined by western blot. Relative protein levels were quantified by densitometry and expressed as optical density ratio. The values presented are the means ± standard error of the mean (n = 6). ## P<0.01 versus the Control group. **P<0.01 versus the Injury group.
Figure 8.
Effects of EsA on cell apoptosis at 12 hours post-CCl4 injection.
Liver tissues sections were stained with TUNEL method (Magnification, ×200). There were no obvious difference for rates of positive TUNEL stained cells between the Injury and Injury+EsA groups (A). The activity of Bax, Caspase-3 and cleaved Caspase-3 were determined by western blot. Relative protein levels were quantified by densitometry and expressed as optical density ratio (B). The values presented are the means ± standard error of the mean (n = 6). #P<0.05, ##P<0.01 versus the Control group.
Figure 9.
EsA protected against GalN/LPS-induced histopathological damage and hepatic dysfunction.
Hematoxylin and eosin staining (A Magnification, 200×) showed that livers in Injury group exhibited more inflammatory cells than those in Control group and EsA group, which were significantly alleviated by treatment of EsA (n = 6). Liver photographs were taken 12 hours post-GalN/LPS administration, and livers in Injury group turned white (B). Levels of AST and ALT increased obviously after GalN/LPS challenge, and which were significantly decreased with EsA treatment (n = 6 C). The values presented are the means ± standard error of the mean. ##P<0.01 versus the Control group. *P<0.05, **P<0.01 versus the Injury group.