Korean J Physiol Pharmacol.  2019 Jan;23(1):21-28. 10.4196/kjpp.2019.23.1.21.

Swertiamarin ameliorates carbon tetrachloride-induced hepatic apoptosis via blocking the PI3K/Akt pathway in rats

• Zhang Q
• Chen K
• Wu T
• Song H

• KMID: 2443612
• DOI: http://doi.org/10.4196/kjpp.2019.23.1.21

Abstract

Swertiamarin (STM) is an iridoid compound that is present in the Gentianaceae swertia genus. Here we investigated antiapoptotic effects of STM on carbon tetrachloride (CCl₄)-induced liver injury and its possible mechanisms. Adult male Sprague Dawley rats were randomly divided into a control group, an STM 200 mg/kg group, a CCl₄ group, a CCl₄+STM 100 mg/kg group, and a CCl₄+STM 200 mg/kg group. Rats in experimental groups were subcutaneously injected with 40% CCl₄ twice weekly for 8 weeks. STM (100 and 200 mg/kg per day) was orally given to experimental rats by gavage for 8 consecutive weeks. Hepatocyte apoptosis was determined by TUNEL assay and the expression levels of Bcl-2, Bax, and cleaved caspase-3 proteins were evaluated by western blot analysis. The expression of TGF-β1, collagen I, collagen III, CTGF and fibronectin mRNA were estimated by qRT-PCR. The results showed that STM significantly reduced the number of TUNEL-positive cells compared with the CCl₄ group. The levels of Bax and cleaved caspase-3 proteins, and TGF-β1, collagen I, collagen III, CTGF, and fibronectin mRNA were significantly reduced by STM compared with the CCl₄ group. In addition, STM markedly abrogated the repression of Bcl-2 by CCl₄. STM also attenuated the activation of the PI3K/Akt pathway in the liver. These results suggested that STM ameliorated CCl₄-induced hepatocyte apoptosis in rats.

Keyword

Akt; Carbon tetrachloride; Collagen; Hepatocyte apoptosis; Swertiamarin

MeSH Terms

Adult
Animals
Apoptosis*
Blotting, Western
Carbon Tetrachloride
Carbon*
Caspase 3
Collagen
Fibronectins
Gentianaceae
Hepatocytes
Humans
In Situ Nick-End Labeling
Liver
Male
Rats*
Rats, Sprague-Dawley
Repression, Psychology
RNA, Messenger
Swertia
Carbon
Carbon Tetrachloride
Caspase 3
Collagen
Fibronectins
RNA, Messenger

Figure

• 

  Fig. 1 Effects of STM on hepatic fibrotic genes mRNA expression in CCl4-induced rat liver. The mRNA expression of TGF-β1, collagen I, collagen III, CTGF, and fibronectin was significantly increased following 8 weeks of chronic CCl4 exposure compared with the control group. STM (100 and 200 mg/kg) treatment obviously decreased the mRNA expression of TGF-β1, collagen I, collagen III, CTGF, and fibronectin compared with the CCl4 group (A–E). Data are represented as means±S.D. for 3–4 animals per group. *p<0.05 vs. control, **p<0.01 vs. control; #p<0.05 vs. CCl4, ##p<0.01 vs. CCl4 by one-way ANOVA and LSD post hoc test.

• 

  Fig. 2 Effects of STM on hepatic apoptosis in CCl4-induced rat liver were detected by TUNEL assay. Representative photographs of TUNEL assay performed on liver sections of (A) control, (B) STM 200 mg/kg, (C) CCl4, (D) CCl4+STM 100 mg/kg, (E) CCl4+STM 200 mg/kg. The black arrows point to the apoptotic cells (TUNEL-positive cells) (magnification × 200). (F) The bar graphs show the total number of TUNEL-positive cells/10 fields in liver sections of control, STM 200 mg/kg, CCl4, CCl4+STM-treated rats. Data are represented as means±S.D. for 3–4 animals per group. *p<0.05 vs. control, **p<0.01 vs. control; #p<0.05 vs. CCl4, ##p<0.01 vs. CCl4 by one-way ANOVA and LSD post hoc test.

• 

  Fig. 3 Western blot assay of Bcl-2, Bax, and cleaved caspase-3 expression of livers in CCl4-induced rat liver. CCl4 treatment obviously suppressed the expression of Bcl-2 compared with the control group. In CCl4+STM groups, Bcl-2 protein levels were notably restored when compared with the CCl4 group. The expression of Bax and cleaved caspase-3 were induced following CCl4 treatment. STM markedly reduced the expression of Bax and cleaved caspase-3 compared with the CCl4 group. Data are represented as means±S.D. for 3–4 animals per group. *p<0.05 vs. control, **p<0.01 vs. control; #p<0.05 vs. CCl4, ##p<0.01 vs. CCl4 by one-way ANOVA and LSD post hoc test.

• 

  Fig. 4 STM inhibits hepatocyte apoptosis by blocking the activation of the PI3K/Akt pathway. CCl4 treatment notably elevated the ratio of pAkt/Akt as compared with the control group. In addition, CCl4 treatment increased the ratio of pPI3K/PI3K as compared with the control group. In CCl4+STM groups, STM markedly reduced the expression of pAkt/Akt and pPI3K/PI3K as compared with the CCl4 group. Data are represented as means±S.D. for 3–4 animals per group. *p<0.05 vs. control, **p<0.01 vs. control; #p<0.05 vs. CCl4, ##p<0.01 vs. CCl4 by one-way ANOVA and LSD post hoc test.

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