The role of CYP2A5 in liver injury and fibrosis: chemical-specific difference

Abstract

Liver injuries induced by carbon tetrachloride (CCL4) or thioacetamide (TAA) are dependent on cytochrome P450 2E1 (CYP2E1). CYP2A5 can be induced by TAA but not by CCL4. In this study, liver injury including fibrosis induced by CCL4 or TAA were investigated in wild-type (WT) mice and CYP2A5 knockout (cyp2a5 (-/-) ) mice as well as in CYP2E1 knockout (cyp2e1 (-/-) ) mice as a comparison. Acute and subchronic liver injuries including fibrosis were induced by CCL4 and TAA in WT mice but not in cyp2e1 (-/-) mice, confirming the indispensable role of CYP2E1 in CCL4 and TAA hepatotoxicity. WT mice and cyp2a5 (-/-) mice developed comparable acute liver injury induced by a single injection of CCL4 as well as subchronic liver injury including fibrosis induced by 1 month of repeated administration of CCL4, suggesting that CYP2A5 does not affect CCL4-induced liver injury and fibrosis. However, while 200 mg/kg TAA-induced acute liver injury was comparable in WT mice and cyp2a5 (-/-) mice, 75 and 100 mg/kg TAA-induced liver injury were more severe in cyp2a5 (-/-) mice than those found in WT mice. After multiple injections with 200 mg/kg TAA for 1 month, while subchronic liver injury as indicated by serum aminotransferases was comparable in WT mice and cyp2a5 (-/-) mice, liver fibrosis was more severe in cyp2a5 (-/-) mice than that found in WT mice. These results suggest that while both CCL4- and TAA-induced liver injuries and fibrosis are CYP2E1 dependent, under some conditions, CYP2A5 may protect against TAA-induced liver injury and fibrosis, but it does not affect CCL4 hepatotoxicity.

Keywords: Cytochrome P450; Fibrosis; Hepatic stellate cell; Liver injury; Metabolism; Oxidative stress.

Figure 1

CCL4-induced liver injury and oxidative stress are CYP2E1 dependent but independent of CYP2A5. (A) Serum ALT activity and (B) pathological observation in liver sections with H&E staining (×200) after CCL4 was injected i.p. at 0.5 ml/kg into WT, cyp2a5^−/− mice and cyp2e1^−/− mice. CV, central vein. Arrows show small single necrotic foci. (C) Liver TBARS and GSH and (D) liver microsomal CYP2E1 and CYP2A5 activity after CCL4 was injected i.p. at 0.5 ml/kg into WT and cyp2a5^−/− mice. (E) Serum ALT activity and (F) Serum AST activity after CCL4 was injected i.p. at 15 µl/kg into WT and cyp2a5^−/− mice. * P<0.05, compared with Control; # P<0.05, compared with cyp2e1^−/− mice group.

Figure 1

CCL4-induced liver injury and oxidative stress are CYP2E1 dependent but independent of CYP2A5. (A) Serum ALT activity and (B) pathological observation in liver sections with H&E staining (×200) after CCL4 was injected i.p. at 0.5 ml/kg into WT, cyp2a5^−/− mice and cyp2e1^−/− mice. CV, central vein. Arrows show small single necrotic foci. (C) Liver TBARS and GSH and (D) liver microsomal CYP2E1 and CYP2A5 activity after CCL4 was injected i.p. at 0.5 ml/kg into WT and cyp2a5^−/− mice. (E) Serum ALT activity and (F) Serum AST activity after CCL4 was injected i.p. at 15 µl/kg into WT and cyp2a5^−/− mice. * P<0.05, compared with Control; # P<0.05, compared with cyp2e1^−/− mice group.

Figure 1

CCL4-induced liver injury and oxidative stress are CYP2E1 dependent but independent of CYP2A5. (A) Serum ALT activity and (B) pathological observation in liver sections with H&E staining (×200) after CCL4 was injected i.p. at 0.5 ml/kg into WT, cyp2a5^−/− mice and cyp2e1^−/− mice. CV, central vein. Arrows show small single necrotic foci. (C) Liver TBARS and GSH and (D) liver microsomal CYP2E1 and CYP2A5 activity after CCL4 was injected i.p. at 0.5 ml/kg into WT and cyp2a5^−/− mice. (E) Serum ALT activity and (F) Serum AST activity after CCL4 was injected i.p. at 15 µl/kg into WT and cyp2a5^−/− mice. * P<0.05, compared with Control; # P<0.05, compared with cyp2e1^−/− mice group.

Figure 1

CCL4-induced liver injury and oxidative stress are CYP2E1 dependent but independent of CYP2A5. (A) Serum ALT activity and (B) pathological observation in liver sections with H&E staining (×200) after CCL4 was injected i.p. at 0.5 ml/kg into WT, cyp2a5^−/− mice and cyp2e1^−/− mice. CV, central vein. Arrows show small single necrotic foci. (C) Liver TBARS and GSH and (D) liver microsomal CYP2E1 and CYP2A5 activity after CCL4 was injected i.p. at 0.5 ml/kg into WT and cyp2a5^−/− mice. (E) Serum ALT activity and (F) Serum AST activity after CCL4 was injected i.p. at 15 µl/kg into WT and cyp2a5^−/− mice. * P<0.05, compared with Control; # P<0.05, compared with cyp2e1^−/− mice group.

Figure 2

CCL4-induced chronic liver injury and fibrosis are dependent on CYP2E1 but independent of CYP2A5. CCL4 was injected i.p. at 0.5 ml/kg into WT, cyp2a5^−/− mice and cyp2e1^−/− mice, twice a week, for one month. The mice were sacrificed 48 h after the last CCL4 injection. (A) Serum ALT activity. (B) H&E staining (×100). (C) Sirius Red/Fast Green staining(×100). Lower panels show the coarse surfaces of liver. Arrows show red stained fibers. (D) Quantification for Sirius Red staining. (E) IHC for collagen I (×100). Arrows show red stained collagen I. (F) Quantification for Collagen I IHC staining. * P<0.05, compared with Control; # P<0.05, compared with WT group; & P<0.05, compared with cyp2a5^−/− mice group.

Figure 2

CCL4-induced chronic liver injury and fibrosis are dependent on CYP2E1 but independent of CYP2A5. CCL4 was injected i.p. at 0.5 ml/kg into WT, cyp2a5^−/− mice and cyp2e1^−/− mice, twice a week, for one month. The mice were sacrificed 48 h after the last CCL4 injection. (A) Serum ALT activity. (B) H&E staining (×100). (C) Sirius Red/Fast Green staining(×100). Lower panels show the coarse surfaces of liver. Arrows show red stained fibers. (D) Quantification for Sirius Red staining. (E) IHC for collagen I (×100). Arrows show red stained collagen I. (F) Quantification for Collagen I IHC staining. * P<0.05, compared with Control; # P<0.05, compared with WT group; & P<0.05, compared with cyp2a5^−/− mice group.

Figure 3

Relationship of CYP2A5 and TAA-induced liver injury and oxidative stress. (A) Liver microsomal activities of CYP2E1 and CYP2A5 and (B) serum levels of ALT and AST after the WT mice were injected with different doses of TAA. (C) Serum levels of ALT, (D) serum AST and (E) hepatic TBARS and GSH after different doses of TAA were injected into WT mice and cyp2a5^−/− mice. * P<0.05, compared with Control group (0 mg/kg); # P<0.05, compared with WT group.

Figure 3

Relationship of CYP2A5 and TAA-induced liver injury and oxidative stress. (A) Liver microsomal activities of CYP2E1 and CYP2A5 and (B) serum levels of ALT and AST after the WT mice were injected with different doses of TAA. (C) Serum levels of ALT, (D) serum AST and (E) hepatic TBARS and GSH after different doses of TAA were injected into WT mice and cyp2a5^−/− mice. * P<0.05, compared with Control group (0 mg/kg); # P<0.05, compared with WT group.

Figure 3

Relationship of CYP2A5 and TAA-induced liver injury and oxidative stress. (A) Liver microsomal activities of CYP2E1 and CYP2A5 and (B) serum levels of ALT and AST after the WT mice were injected with different doses of TAA. (C) Serum levels of ALT, (D) serum AST and (E) hepatic TBARS and GSH after different doses of TAA were injected into WT mice and cyp2a5^−/− mice. * P<0.05, compared with Control group (0 mg/kg); # P<0.05, compared with WT group.

Figure 4

TAA-induced liver fibrosis but not chronic liver injury is enhanced in the cyp2a5^−/− mice compared to WT mice. TAA was injected at 200 mg/kg into WT mice, cyp2a5^−/− mice and cyp2e1^−/− mice, twice a week, for one month. The mice were sacrificed 48 h after the last TAA injection. (A) Serum ALT activity. (B) H&E staining (×100). (C) Sirius Red/Fast Green staining(×100). Lower panels show the coarse surfaces of liver. Arrows show red stained fibers. (D) Quantification for Sirius Red staining. (E) IHC for collagen I (×100 upper panels; ×200 lower panels). Arrows show red stained collagen I. (F) Quantification for Collagen I IHC staining. * P<0.05, compared with Control; # P<0.05, compared with WT group; & P<0.05, compared with cyp2a5^−/− mice group.

Figure 4

TAA-induced liver fibrosis but not chronic liver injury is enhanced in the cyp2a5^−/− mice compared to WT mice. TAA was injected at 200 mg/kg into WT mice, cyp2a5^−/− mice and cyp2e1^−/− mice, twice a week, for one month. The mice were sacrificed 48 h after the last TAA injection. (A) Serum ALT activity. (B) H&E staining (×100). (C) Sirius Red/Fast Green staining(×100). Lower panels show the coarse surfaces of liver. Arrows show red stained fibers. (D) Quantification for Sirius Red staining. (E) IHC for collagen I (×100 upper panels; ×200 lower panels). Arrows show red stained collagen I. (F) Quantification for Collagen I IHC staining. * P<0.05, compared with Control; # P<0.05, compared with WT group; & P<0.05, compared with cyp2a5^−/− mice group.