Protective Effect Of Necrostatin-1on The Liver Of Rats With Trauma-hemorrhagic Shock

Objective:To investigate the protective effects of Necrostatin-1on the liver of rats with trauma-hemorrhagic shock.Methods:Adult male Sprague-Dawley rats, with weight250-300g, were used. Experimental design is divided into two parts:the first part is the establishment of the trauma-hemorrhagic shock model. A total of40rats were randomly divided into sham group and model group, with20rats in each group. Rats model was produced by adopting hemorrhagic shock/reperfusion in accompany with the left femur, tibia fracture and soft tissue injury. While rats in sham group were only received anesthesia, separating and ligating blood vessels, without trauma-hemorrhagic shock and reperfusion. The24hours mortality was observed, and serum ALT and AST were detected by automatic biochemistry analyzer. The pathology changes in liver tissues were observed by HE staining. The second part is the application of Nec-1, and to investigate the effects of Nec-1on the liver of rats with trauma-hemorrhagic shock. So72rats were randomly divided into sham group, DMSO control group, Nec-1group with24rats in each group. And the rats in Nec-1group were received1mg/kg Nec-1through femoral vein5minutes before reperfusion, while the rats in DMSO control group were received the same amount of solvent. The serum and liver tissues of each group were collected at2,4,8hours after reperfusion. Serum ALT and AST were detected by automatic biochemistry analyzer. The pathology changes in liver tissues were observed by HE staining. Serum TNF-a and IL-1β were detected by ELISA. The mRNA expressions of TNF-α and IL-1β in the liver tissues were determined by RT-PCR. The protein expressions of RIPl and RIP3were also assessed by Western Blot analysis.Results:The first part:To establish the trauma-hemorrhagic shock model of rats. And the24hours mortality of model group is30.00%(P<0.01). Compared with sham group, the levels of serum ALT and AST were significantly increased (P<0.01). Under light microscopy, it was noted that there was no obvious change in sham group. But there were the hepatic sinus expansion, infiltration of inflammatory cells, the disorder of hepatic lobule, as well as liver cells degeneration, necrosis in model group.The second part:(1)Compared with sham group, the levels of serum ALT and AST were significantly increased (P<0.05), and reached the peak at8hours in DMSO control group (P<0.01). But the levels of serum ALT and AST were significantly decreased in Nec-1group at4,8hours (P<0.01).(2)Under light microscopy, it was noted that there were the hepatic sinus expansion, the disorder of hepatic lobule, infiltration of inflammatory cells, as well as liver cells degeneration, necrosis in DMSO control group. After the application of Nec-1, the pathology changes in hepatic tissues were significantly mitigated.(3)Compared with sham group, the expressions of serum TNF-α and IL-1β were significantly increased at2hours, and reached the peak at8hours (P<0.01). While the expressions of serum TNF-α and IL-1β were significantly decreased in Nec-1group compared with those in DMSO control group (P<0.01).(4)Along with the time extension, the mRNA expressions of TNF-α and IL-1βof liver tissues were consistent with the expressions of serum TNF-α and IL-1(3in each group.(5)The protein expression of RIP1and RIP3of liver tissues were markedly up-regulated and the most obvious difference was at8hours by Western Blot (P<0.01). But there was no significantly significant difference in RIP1and RIP3between Nec-1group and DMSO control group.Conelusion:The experiment confirmed that there was acute liver damage after trauma-hemorrhagic shock. After the application of Nec-1, the liver function was improved, the pathology changes were mitigated, the levels of inflammatory factors were significantly reduced. It indicated that Nec-1may be remarkable protect effect on the liver of rats with trauma-hemorrhagic shock. But the protein expressions of RIP1and RIP3were no statistically significant difference, so the intrinsic mechanisms need further investigation.