| Objective To investigate the effect of Hyperhomocysteinema over the liver in ovariectomized rats and the positive action of estrogen, folic acid and N-acetylcysteine toward HHcy's effect and to explore the mechanism.Methods 48 SD rats of 4 months were randomly divided into 6 groups, namely group of sham-operated (Sham group), ovariectomy (OVX group), methionine (OVX+L-methionine 1 g. kg-1, d-1, Met group), estrogen replacement therapy (OVX+L-methionine+17-βestradiol 0.1 mg.kg-1.d-1, ERT group), folic acid (OVX+L-methionine+folic 20 mg. kg-1, d-1, FA group) and N-acetylcysteine(OVX+L-methionine+ N-acetylcysteine 100 mg. kg-1, d-1, NAC group).1 week after been ovariectomized,all rats were intervened with drugs and gastric perfusion. Then,9 weeks later, all the rats were sacrificed for checking the level of total homocysteine(tHcy), alanine aminotransferase (ALT), aspartate aminotransferase (AST). The pathological changes were observed after haematoxylin-eosin (HE) staining in liver. Total antioxidative capacity (TAC), malondialdehyde (MDA), superoxide dismutase (SOD) and glutathion peroxidase (GSH-PX) and concentration of glutathione reductase (GR) and reduced glutathione hormone (GSH) in their liver homogenates. Besides, the nitrotyrosine of liver were tested through method of immunohistochemical staining and western blot.Results 1. Compared with Sham group, serum tHcy concentrations were increased significantly in OVX and Met groups(p< 0.01). Compared with OVX group, serum tHcy concentrations were increased distinctly in Met group (p< 0.01). ERT, FA and NAC groups presented a decrease of serum tHcy compared to Met group (p< 0.01), which were still significantly higher than Sham group (p< 0.01).2. Serum ALT and AST levels had significant rise in OVX and Met group (p< 0.01, p < 0.05) compared to Sham group. Compared with OVX group, serum ALT and AST levels were increased distinctly in Met group (p< 0.01). ERT, FA and NAC groups presented a significant decrease of serum ALT and AST levels compared to Met group (p< 0.01), which were still higher than Sham group (p< 0.01, p< 0.05). Moreover, the ALT and AST levels in NAC group were lower than ERT group(p< 0.01).3. Morphological observation in liver tissue:The structure of hepatic lobules is clear in Sham group. In OVX group, the liver cells and hepatic lobules have changed, and there is mild steatosis. The normal liver structure was destroyed and extensive fatty degeneration of liver cells has occurred in Met group. Compared with Met group, hepatic steatosis and structure of hepatic lobules have improved to some degree in ERT, FA and NAC groups.4. OVX and Met groups demonstrated a increase of MDA in liver homogenate compared to Sham group (p< 0.01), while the TAC, SOD, GSH, GR and GSH-PX levels were significantly lower (p<0.01). Compared with OVX group, liver MDA was distinctly increased (p< 0.01) and TAC, SOD, GSH, GR and GSH-PX levels were significantly lower (p< 0.01, p< 0.05) in Met group. ERT, FA and NAC groups improved oxidative stress state versus Met group, MDA levels were significantly decreased (p< 0.01), TAC, SOD, GSH, GR and GSH-PX levels increased distinctly(p< 0.01, p< 0.05).5. Liver nitrotyrosine (NT) expression:The OVX and Met groups were significantly increased (p< 0.01) compared to Sham group, and Met group further increased the expression of NT compared to OVX group(p< 0.01). ERT, FA and NAC groups have decreased in NT expression compared to Met group(p< 0.01), which were still significantly higher than Sham group (p< 0.01).Conclusion 1. Methionine overload caused Hyperhomocysteinema and further induced liver injury in ovariectomized rats, the mechanism might be related to Hyperhomocysteinema induced liver oxidative damage. 2. The treatment of 17-P estrogen, folic acid or N-acetylcysteine can alleviate the liver injury in ovariectomized rats caused by HHcy to some degree, which might be owned to the improving of homocysteine metabolism and state of oxidative stress in rats and other factors.
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