| Ischemia/reperfusion injury is a phenomenon whereby tissues experience damage as a result of temporarily interrupted blood flow (ischemia) followed by its restoration (reperfusion). Clinically, liver I/R occurs in settings of elective liver surgery, trauma, and transplantation. Liver I/R is a pathophysiologic process whereby hypoxic organ damage is accentuated following return of blood flow and oxygen delivery. In fact,hepatic ischemia-reperfusion injury can also be considered an inflammatory injury.IRI activation of the immune response, caused local and systemic inflammatory response, increased tissue, organ damage, severe cases can lead to death.Thus, understanding the pathophysiology and minimizing the effects of hepatic IRI would probably improve liver surgery and transplant outcomes.P-selectin is a relative molecular mass of about140kU transmembrane glycoprotein, which is thrombin-activated platelets and white blood cells of vascular adhesion receptors on endothelial cells, endothelial cells are usually stored in the W-P bodies and platelets.When subjected to inflammation, ischemia, hypoxia, P-selectin on endothelial cells can be rapidly plasma membrane of endothelial cells,and can be combined with the TNF-a induced by an unknown ligand, involved in the inflammatory response. E-selectin is a relative molecular mass of110kD transmembrane glycoprotein, is a human leukocyte differentiation antigen CD62E, also known as endothelial leukocyte adhesion molecule-1, mainly expressed on endothelial cells.In the absence of any stimulation, E-selectin basic not express, but when subjected to TNF-a, IL-1, IL-6and other stimuli when, E-selectin expression in endothelial cells increased, and plays an important role in inflammation and lymphocyte homing in the whole process, such as the neutrophils and activated T cells. Related literature, P-selectin and E-selectin plays an important role in brain, kidney, pancreas, intestine and other important organs in ischemia-reperfusion, and its biological behavior and relationships a lot of research, but relatively few studies P-selectin and E-selectin role in hepatic ischemia-reperfusion and expression laws.[Objective]In this study,70%by rat liver ischemia-reperfusion injury model, we employed a rat model of partial liver warm IRI in an attempt to analyze the exact mechanismsof liver IRI,application P-selectin and E-selectin antibody pretreatment of experimental animals were observed P-selectin and E-selectin role of hepatic ischemia-reperfusion injury in.[Method]1,We refer to the method of operation, such as Nauta RJ,70%of rat hepatic warm ischemia-reperfusion injury model.2, Specific pathogen-free (SPF) inbred male cleaning closed group of90SD rats, weighing220-250g, all animals are Kunming Medical University Experimental Animal Center.3,Animal grouping and processing method:Were randomly divided into five groups:sham operation group (group1):Preoperative not give injections reagent free only after laparotomy ligament liver and abdomen was closed at60min after;(2) ischemia-reperfusion group (group2):Preoperative not give injections reagent line hepatic ischemia-reperfusion treatment;(3) P-selectin antibody injection+ischemia-reperfusion group treated group (group3):In accordance with the latter into the abdominal cavity2mg/kg intravenous administration, after administration5min, establish warm ischemia model;(4) injection of E-selectin antibody+ischemia-reperfusion group treated group (group4):after laparotomy according12.5ug/kg into the inferior vena cava to drugs, over the same three groups;(5) injection of P/E selectin antibody+ischemia-reperfusion treatment groups (4groups):after laparotomy were given injections P/E-selectin antibody, I was in different parts of the inferior vena cava with3groups;4,Detection of indicators Respectively after th,6h,24h5ml blood through the aorta and get fresh liver tissue, serum ALT and AST liver function evaluation. ELISA was used to detect the relative expression of SOD and MDA. Immunohistochemical staining observed P-selectin, E choose express hormone. Morphological changes observed in liver biopsy, evaluation of liver tissue injury.[Results]1. We refer to such methods Nauta RJ successfully prepared70%of liver ischemia-reperfusion injury model in78cases, hepatic warm ischemia time was60min.2. Affect the expression of P-selectin and E-selectin in the liver:①P-selectin and E-selectin expression. Immunohistochemistry showed that the two groups (I/R group) with a group (sham group) compared to the expression of P-selectin and E-selectin increased (P<0.05);3groups (P selectin monoclonal antibody treatment group) compared with group1, group2, group3P-selectin expression was significantly lower (P<0.05), E-selectin expression with two groups;4group (E-selectin antibody treatment group), E-selectin the expression was significantly lower (P<0.05), P-selectin expression with the two groups (P<0.05);5group (P/E-selectin antibody treatment group) compared with2,3,4group, P-selectin and E selectin expression was significantly lower (P<0.05).②affect liver function.2groups (I/R group) compared with group1(sham group), group2transaminase (ALT, AST) was significantly higher (P<0.05);3group compared with4groups with2, Group3,4group transaminase (ALT, AST) lower (P<0.05),5group (P/E-selectin antibody treatment group) compared with2,3,4group, transaminase (ALT, AST) was significantly lower (P<0.05).③change in liver morphology. Group1(sham group), showing liver size, color, normal under the microscope, no degeneration or necrosis of the liver tissue structure clear, clear hepatic cord structure, arranged in neat rows, hepatic sinusoid does not expand, only to see a little inflammatory cell infiltration;2groups (ischemia-reperfusion group), liver cell edema, liver basic organizational structure is not clear, hepatic cord, liver sinusoidal structural changes, hepatocyte ballooning degeneration, periportal inflammatory cell infiltration, optical microscope, multiple necrosis.3groups (P selectin antibody group), compared two groups, improving the overall organizational structure of the liver, liver volume seen any larger, swelling of the liver cells, liver cells were ballooning and ballooning degeneration of both mixed periportal inflammatory cell infiltration, light microscope spotty necrosis and focal necrosis;4groups (E-selectin antibody group), compared two groups, improving the overall organizational structure of the liver, liver volume was slightly larger than normal liver, swelling of the liver cells is not obvious, liver vacuolar degeneration reduced hepatic sinusoid small amount of mild expansion periportal inflammatory cell infiltration, showing spotty necrosis and focal necrosis;5groups (P/E-selectin antibody group), compared with2,3,4, mild hepatic cell damage, hepatic cords neat and hepatic sinusoid expansion is not obvious, and periportal hepatocytes normal, normal lobular architecture, only a minority of hepatocellular edema, inflammatory cell infiltration decreased.3. Changes in liver tissue MDA and SOD:The ELISA test methods, sample test rats MDA content and SOD. Compared with group1, group2, group3and group4and5in liver tissue MDA levels were increased (P<0.05), the difference was statistically significant. Compared with group2, group3to reduce liver tissue MDA levels (P<0.05), tissue MDA levels four groups to reduce the liver (P<0.05),5of liver tissue MDA levels below3,4group, significantly lower than group2(P<0.05); SOD in ischemia-reperfusion lh began to decline, showing significant changes24h. Compared with group1, group2, group3and group4and5in liver tissue SOD levels were elevated (P<0.05), the difference was statistically significant. Compared with group2, group3liver tissue SOD levels increased (P<0.05),4of liver tissue SOD levels increased (P<0.05),5groups of SOD in liver tissue was significantly higher than3,4group (P<0.05), were also significantly higher than those not suffering from any liver ischemic injury in a group;4. Changes in liver apoptosis:apoptotic cells have the following characteristics: single cells, without inflammation, cell curl brown particles or broken nuclei. TUNEL assay was found, a group (sham group) rarely occurs in liver tissue apoptotic bodies.2groups (I/R group)1h reperfusion lh, only a few apoptotic cells; liver tissue reperfusion6h increased apoptotic bodies;24h reperfusion apoptotic bodies seen a lot of aggregation, and the emergence of necrosis. Each time period, the number of apoptotic bodies in liver tissue are three groups of less than non-treated group2;4hepatic tissue each time period, the number of apoptotic bodies are less than the non-treatment group2; five groups of apoptotic bodies fewer than3,4group, significantly less than the2groups.[Conclusion]1, The modified Nauta’s "local70%liver ischemia reperfusion model" is a stable, reliable, simple,convenient and practicable model.2, Rat hepatic ischemia-reperfusion injury, P-selectin and E-selectin expression are increased, both to participate in the hepatic ischemia-reperfusion injury, injury-mediated activation of leukocytes and endothelial cells in tissue; different that P-selectin mediates the initial stage of inflammation, then slowly weakened after2h, E-selectin role greater than P-selectin plays an important role in the later stages of the process of inflammation.3,P-selectin and E-selectin monoclonal antibody on hepatic ischemia-reperfusioninjury has a protective effect, using both antibody therapy is better than an antibody. |
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