| Objective:There is a high mortality rate and incidence of postoperative complications after emergency operation of liver trauma caused by war or external injury. It is an important factor that surgical procedures to block hepatic blood flow are often used to control bleeding or facilitate debridement (removal) of trauma liver. Hepatic inflow occlusion, no doubt, will bring about the liver warm ischemia injury that can not be underestimated. After the successful operation to stop bleeding or debride, recovery of liver blood supply will bring about the liver injury again, which to some extent can be more serious than damage of the liver warm ischemia caused by pre-blocking blood flow. Moreover, during treatment process of liver trauma, the body is in a state of acute stress, and this time a series of pathophysiological changes in the liver is ischemia reperfusion injury of the liver under acute stress!Now It is widely recognized that energy deficiency of liver cells is the initial factor inducing a series of pathological changes of hepatic ischemia reperfusion injury. It has been proved that if effectively increasing liver glycogen reserves before blocking hepatic blood flow would effectively reduce subsequent ischemic reperfusion injury. However, in order to avoid the shock caused by loss of blood, rescue personnel must stop bleeding as soon as possible in the treatment process. So it is imperative to increase the liver glycogen reserves within a short time. How to increase the liver glycogen reserves within a short time? One is to increase the effective substrate for glycogen synthesis, and the other is to speed up the synthesis of glycogen.It is known that fructose-1, 6-diphosphate (FDP) as a regulator of energy metabolism can reduce ischemia reperfusion injury of a variety of tissues and organs including the liver, meanwhile FDP as the middle product in intracellular glucose metabolism is the common pathway of glycolysis and gluconeogenesis; in addition, glycogen synthase kinase-3 (GSK-3) can phosphorylate serine sites of glycogen synthase to inhibit the synthesis of glycogen, oppositely GSK-3 inhibitor lithium chloride can inhibit GSK-3 activity to activate glycogen synthase and promote glycogen synthesis.Based on the above understanding, this study investigated effects of associated application of GSK-3 inhibitor lithium chloride and FDP on warm ischemia reperfusion injury of liver in rats caused by surgery during the emergency through the establishment of liver trauma model.Methods:1. 42 healthy Sprague-Dawley (SD) rats were randomly divided into 3 groups (mild injury group, moderate injury group and severe injury group, n=14). By improving the biological impact machine-Ⅳdesigned by Trauma Center of PLA, Institute of Surgery Research, Daping Hospital, Third Military Medical University with a T-type second-hit head, 3 groups of liver blunt impact injury model were established respectively with three different-quality steel ball. Then each group was randomly divided into A group and B group according to the difference of measured items. The diversity in vital signs before and after injury and the number of natural survival animals after injury in A group were measured. AST and ALT content in serum before and after injury, TNF-αcontent, the quality of intra-abdominal hemorrhage of animals after injury in B group were measured. The liver injury of all rats was classified and given a score after the death.2. After the establishment of a liver trauma model on 49 SD rats, 42 out of the animals were randomly divided into control group, glucose group and FDP group (n=14), which were respectively injected 2ml of 0.9% sodium chloride, 5% glucose and 15% FDP injection at 10min after injury. Then each group was randomly divided into pre-ischemia group and 4h reperfusion group (n=7) according to time points when animals were executed before and after ischemia. The remaining seven for sham operation group (SH group) were executed in 4h reperfusion time point. The AST and ALT content in serum and glycogen content, SOD vitality, MDA content and ATP content in liver tissue were determined. It was also carried out that HE staining, fluorescence real-time quantitative PCR detection of liver Bcl-2 mRNA expression, cell apoptosis detection (TUNEL method).3. After the establishment of a liver trauma model in 49 SD rats, 42 out of the animals were randomly divided into control group, FDP group, FGI group (FDP and GSK-3 inhibitor in combination group) (n=14). control group was injected 2ml of 0.9% sodium chloride injection. FDP group was injected 2ml of 15% FDP injection. FGI group was injected 2ml of 15% FDP injection, in addition, 0.5% GSK-3 inhibitor lithium chloride (3mmol/kg) by intraperitoneal injection (all at 10min after injury). Then each group was randomly divided into pre-ischemia group and 4h reperfusion group (n=7) according to time point when animals were executed before and after ischemia. The remaining seven for sham operation group (SH group) were executed in 4h reperfusion time point. The AST and ALT content in serum and glycogen content, SOD vitality and MDA content, liver glycogen synthase activity and immunohistochemical determination of GSK-3 expression in liver tissue were determined.Results:1. The average femoral artery pressure in each group after injury was significantly lower than that before injury (P<0.01), and decreased with the increased degree of injury (P<0.01 or P<0.05). AST and ALT content in each group after injury were significantly higher than those before injury (P<0.01 or P<0.05), and increased with the increased degree of injury (P<0.01 or P<0.05). The quality of intra-abdominal hemorrhage in each group significantly increased with the increased degree of injury (P<0.01 or P<0.05). The score for the hepatic injury significantly increased with the increased degree of injury (P<0.01). TNF-αcontents in serum were significantly higher than those before injury (P<0.01), and increased with the increased degree of injury (P<0.01).2. At pre-ischemia time point, liver glycogen content, control groupglucose group>FDP group>SH group (P<0.01 or P<0.05); except that differences between glucose group and FDP group, AST content, control group>glucose group/FDP group>SH group (P<0.01 or P<0.05); SOD vitality, control groupglucose group>FDP group>SH group, (P<0.01 or P<0.05); ATP content, the control groupglucose group>FDP group>SH group (P<0.01 or P<0.05).3. At pre-ischemia time point, liver glycogen content, the control groupFDP group>FGI group>SH group (P<0.01); SOD vitality, control groupFDP group>FGI group>SH group (P<0.01). As compared with pre-ischemia time point, liver glycogen synthase activity of the control group and the FDP group at 4h reperfusion time point were significantly lower (P<0.01); at pre-ischemia time point, liver glycogen synthase activity, FGI group>FDP group (P<0.01); at 4h reperfusion time point, liver glycogen synthase activity, control groupFDP group>FGI Group>SH group (P<0.01).Conclusion:1. The degree of liver damage and stress in each group obviously increases with the increase of quality of the steel ball. As an ideal experimental animal model, this liver trauma model in rat has simple, stable features of liver injury, good repeatability, adjustability and high cost-effective rate. The moderate injury group can be used in this study for further pathological mechanisms and treatment of injury.2. As compared with glucose, the application of FDP has better protective role on rat liver trauma. The mechanism may be related to that it is more effective that the glycogen synthesis of FDP as substrate in the liver in stress conditions, which more increase reserves of liver glycogen before ischemia, thus reducing warm ischemia reperfusion injury caused by surgery during the emergency after rat liver trauma.3. The associated application of FDP and GSK-3 inhibitor lithium chloride can enhance the effect of FDP on the protective role of rat liver trauma. The mechanism may be related to that GSK-3 inhibitor can effectively enhanced the role of glycogen synthesis of FDP as substrate before liver ischemia, which increases the liver glycogen reserves more effectively than the single application of FDP in a short period of time, thus more reducing warm ischemia reperfusion injury caused by surgery during the emergency after rat liver trauma.
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