| ObjectiveObjective:to study the role of lycopene on the process of apoptosis and autophagy in the liver ischemia-reperfusion injury of rats.MethodsGrouping:80 SPF SD rats were randomly divided into Sham-operated group, ischemia reperfusion group, treatment group with lycopene, inhibiting group with lycopene and 3-ma,20 rats in each group.Establishing the liver ischemia reperfusion model,the rat was intraperitoneal injection of 1% sodium pentobarbital anesthesia (0.2ml/100g) for anesthesia. In the center of the abdominal longitudinal incision into the abdomen, fully exposed after intra-abdominal viscera, free out of the first porta hepatis. Using nondestructive microscopic vascular clamp clip liver left,middle vascular stem for 40 minutes, making the middle and the left of hepatic lobe to achieve 70% of the liver tissue ischemia.Incision gauze covering physiological saline, achieving loosening vascular clamp after getting the time, then closed abdomen.The determination of sample collection and observation of indicators:after ischemia reperfusion 1 h,6 h,12 h,24 h, to be drawn to detect the biochemical indexes. Using automatic biochemical analyzer test groups of rats serum glutamic acid transaminase (ALT) and aspertate aminotransferase (AST), using double antibody sandwich ELISA method for the quantitative determination of rat serum related inflammatory cell factor of TNF-a, IL-6;Western immunoblot method to detect BCL-1 and bax in the liver tissue; Through the hematoxylin-Yin Gong (HE) staining to observe each group liver tissue pathology change. Compared with T test between two groups of measuring data, categorical data line comparison between chi-square.Statistically significant P values<0.05, all statistical analysis using the original SPSS19.0 statistical analysis software to complete.Results1. The change of ALT and AST levels in the serum:compared with control group of ischemia reperfusion group of serum ALT and AST were significantly increased (P<0.05), the peak after 6 h, then gradually decline,12 h,24 h reperfusion serum ALT and AST level gradually decline. Lycopene treatment group compared with ischemia-reperfusion group, groups of lycopene and 3-ma inhibition blood serum ALT and AST levels relatively higher (P<0.05).2. Serum of TNF-a and IL-6 levels of inflammatory factors change:in the liver ischemia-reperfusion group significantly increased in serum of rats, at 6h after reperfusion reached its peak, and then at 12 h,24 h gradually decline, the treatment group was significantly lower than ischemia-reperfusion group and 6 h after reperfusion there was a significant difference (P<0.01).And the inhibition of TNF alpha, IL-6 in each period were higher than the treatment group, there was a significant difference in 6h after reperfusion (P<0.01)3. The change of liver cell apoptosis protein expression levels:according to the expression of apoptosis related proteins bax liver ischemia-reperfusion group of rats after reperfusion bax expression level rise, and the expression of bax lycopene treatment group liver, lower than only 1 h reperfusion no statistical difference between the two groups.And the inhibiting group and treatment group compared with decreased the expression of bax, only 1 h reperfusion no statistical difference between the two groups.4. The change of the level autophagy protein expression in liver cells, by western blot detection autophagy related liver protein Beclin 1, protein expression showed that Beclin 1 peaked at 6h after reperfusion, in 12h and 24h gradually reduce, lycopene treatment group compared with ischemia-reperfusion group increased the expression of Beclin 1, and join the autophagy inhibitor 3-MA, inhibit the Beclin 1 expression, compared with treatment group showed that lycopene promotes liver ischemia-reperfusion hepatocyte Beclin 1 expression level, promoting the autophagy.5. Liver HE staining observation:control way form normal liver cells, hepatic lobule, central veins, collect abbacy structure is clear, neat liver cell line.Rat liver ischemia reperfusion model control group compared with lycopene treatment group each time point, liver tissue texture, appear all sorts of degree of edema and necrosis, the most significant difference at 6 h after reperfusion, vacuoles degeneration and necrosis of hepatic cells, solid condensed and hyperchromatic nuclei, and lycopene in each treatment group liver tissue pathology morphology obviously improved.ConclusionFound that lycopene pretreatment can improve the level of ischemia reperfusion in rat liver autophagy, reduce serum ALT/AST levels and reduce inflammation reaction and reduce liver cell apoptosis and protect ischemic reperfusion injury. Further by joining autophagy Piao blocker 3-methyl gland (3-MA), found that weakens the protection of the lycopene. In the liver ischemia-reperfusion injury, suggesting the appearance and functions of autophagy and tissue ischemia, extent of damage and duration, ischemia time is short, the blow to the body is weak, autophagy in a someway can reduce the occurrence of apoptosis.This research shows that lycopene can protect ischemia-reperfusion liver injury in rats by promoting autophagy. The results for lycopene protects liver ischemia-reperfusion injury provides a new theoretical basis, which have a important clinical value. |
