Mechanism of the Inhibitory Effects of Eucommia ulmoides Oliv. Cortex Extracts (EUCE) in the CCl 4 -Induced Acute Liver Lipid Accumulation in Rats

Abstract

Eucommia ulmoides Oliv. (EU) has been used for treatment of liver diseases. The protective effects of Eucommia Ulmoides Oliv. cortex extracts (EUCE) on the carbon tetrachloride- (CCl4-) induced hepatic lipid accumulation were examined in this study. Rats were orally treated with EUCE in different doses prior to an intraperitoneal injection of 1 mg/kg CCl4. Acute injection of CCl4 decreased plasma triglyceride but increased hepatic triglyceride and cholesterol as compared to control rats. On the other hand, the pretreatment with EUCE diminished these effects at a dose-dependent manner. CCl4 treatment decreased glutathione (GSH) and increased malondialdehyde (MDA) accompanied by activated P450 2E1. The pretreatment with EUCE significantly improved these deleterious effects of CCl4. CCl4 treatment increased P450 2E1 activation and ApoB accumulation. Pretreatment with EUCE reversed these effects. ER stress response was significantly increased by CCl4, which was inhibited by EUCE. One of the possible ER stress regulatory mechanisms, lysosomal activity, was examined. CCl4 reduced lysosomal enzymes that were reversed with the EUCE. The results indicate that oral pretreatment with EUCE may protect liver against CCl4-induced hepatic lipid accumulation. ER stress and its related ROS regulation are suggested as a possible mechanism in the antidyslipidemic effect of EUCE.

Figure 1

Effect of EUCE on CCl₄-induced histological changes in liver. Rats were injected with 1 mg/kg CCl₄, and livers were isolated after 4 hours. Representative photomicrographs (200x) of liver sections from rats (n = 8) stained with hematoxylin and eosin (a) and oil red O (b) are shown.

Figure 2

Effect of EUCE on CCl₄-induced inhibition of hepatic triglyceride and cholesterol secretion. Rats were injected with 1 mg/kg CCl₄, and livers were isolated after 4 hours. Triglyceride and cholesterol levels were measured in the liver (a), (c) and plasma (b), (d), respectively. Values are mean ± SD, n = 8. Asterisks indicate differences from the group treated with CCl₄ only (*P < 0.05; **P < 0.001). ^# P < 0.05 indicates a significant difference compared with the control group.

Figure 3

Effect of EUCE on free radical scavenging activity and CCl₄-induced increases in liver glutathione (GSH) and malondialdehyde (MDA). A difference was found in ALT and AST levels after injection of CCl₄. Values are mean ± SD, n = 8. Asterisks indicate differences from the group treated with CCl₄ only (*P < 0.05). ^# P < 0.05 indicates a significant difference compared with the control group.

Figure 4

Effect of EUCE on CCl₄-induced ER stress response and expression of ApoB and P450 2E1. Rats were injected with 1 mg/kg CCl₄, and livers were isolated after 4 hours. (a) Western blotting was performed on liver protein extracts with anti-GRP78, CHOP, p-eIF-2α, eIF-2α, ATF-6α, IRE-1α, sXBP-1, ApoA1, ApoB, P450 2E1, or GAPDH. (b) Graphs showing quantification of (a). (c) P450 2E1 activity was measured in liver. Asterisks indicate differences from the group treated with CCl₄ only (*P < 0.05). ^# P < 0.05 indicates a significant difference compared with the control group.

Figure 5

Effect of EUCE on CCl₄-induced lysosomal enzymes activity. Rats were injected with 1 mg/kg CCl₄, and livers were isolated after 4 hours. The lysosomal enzymes activity was measured by a kit. Values are mean ± SD, n = 8. Asterisks indicate differences from the group treated with CCl₄ only (*P < 0.05). ^& P < 0.05, ^# P < 0.05 indicate a significant difference compared with the control group.