The Role Of Heme Oxygenase In The Bile Duct Around Vascular Regeneration

BackgroundJennings in1960first proposed ischemia and reperfusion injury (ischemia reperfusion injury IRI) the concept of ischemic, organsorganizations regain blood supply, not only can not the organization, organ recovery, actually worsened the function of metabolism and structural damage, This phenomenon is called the IRI. Hepatic ischemia-reperfusion injury (of hepatic ischemia reperfusion injury, HIRI) is a liver transplant, liver tumor and hemorrhagic shock and other common diseases and surgery experienced more than one factor involved in the pathological process involving many factors, the IRI to produce free radicals is an important factor in causing the injury occurred.。The damage is the result of various factors, prevention can be divided into ischemic preconditioning, ischemic postconditioning. Hepatic ischemia-reperfusion injury led directly to the lesions of the biliary system, which biliary ischemia reperfusion injury, biliary ischemia, caused by bile duct epithelial cell necrosis, bile duct peripheral vascular lesions., Biliary system and liver are not same, the liver is to accept the dual hepatic arterial and venous blood vessels that supply blood and nutrition, the main branches of the biliary tract is supplied by the hepatic artery, hepatic artery hepatic bile duct peripheral vascular (peribiliary vascular plexus pbvp) to provide blood and nutrients, so when the liver was injury by ischemia and reperfusion, bile duct blood system bear the brunt of the biliary tract is an important target of the warm ischemia, cold ischemia and reperfusion injury induced biliary Target organ, the complex mechanisms related to many factors, the mechanism may include free radical mediated injury, consumption of three adenosine monophosphate, calcium overload, cytokines, and the damaging effects of microcirculation, hydrophobic bile salts, as well as damage of the cytoskeleton, which warm ischemic vascular plexus around the bile duct, the aggregation of inflammatory cells in the blood vessels cause vascular endothelial cell necrosis, intimal injury, vasoconstriction, and other factors. Bile duct peripheral vascular regeneration after injury have a positive effect on the rehabilitation of patients after liver surgery.Heme oxygenase-1(the heme oxygenase-1HO-1) is a catalytic enzyme of heme degradation, which can be decomposed into carbon monoxide, biliverdin and iron in the body. The study found that the heme oxygenase-1in ischemia and reperfusion injury play the role of protecting the gene. HO-1overexpression can induce vascular endothelial growth factor (vascular of endothelial growth factor VEGF), and enhance the ability to repair after vascular injury.In view of the hemoglobin oxygen and enzyme-1many merits,Ho-lean promote the regeneration of the blood vessels in the heart after the occurrence of the increase in the number of myocardial infarction capillary, all function were obviously improved, so its in the liver tissue, in bile duct peripheral vascular whether it can have a positive effect, still is the research direction of the liver and gallbladder surgery.This experiment used rat liver heat ischemia model, to investigate the effect and mechanism of heme oxygenase-1in biliary ischemia and reperfusion injury especially whether to promote the bile duct peripheral vascular plexus regeneration, reduce biliary ischemia-reperfusion injury, restore biliary blood supply. Provide a theoretical basis and experimental evidence for the diagnosis and treatment of liver transplantation or biliary complications in liver surgery.Purpose:Establish rat liver heat ischemia in perfusion damage model, this paper discusses the heme oxygen and enzyme-1in the liver and biliary hot ischemia-reperfusion injury of the role of endothelial progenitor cells in the blood vessels observation of the repair role, especially in the bile duct peripheral vascular plexus regeneration in the role. After surgery for clinical liver biliary ischemia-reperfusion injury treatment of providing theoretical basis and experimental basis Materials and methods:1. The animal group:128only adult rats randomly divided into four groups:(1) physiological saline groups, will rats after anesthesia, pussy in dorsal vein into0.5ml saline.(2) the blank of adenovirus, namely, in rats back into the blank genitals vein viral0.5ml.(3) HO-1group, that is, in rats back into the genitals vein dilution good HO-1-0.5ml.(4) siRNA group, that is, in rats back into the genitals vein dilution good0.5ml siRNA. According to the time of every group in point, take out the corresponding rat liver tissue, and liver door of bile duct organization.2. Hot liver ischemia-reperfusion injury pathological model:choose clean level health adult male SD (Sprague-Dawley) rats, weight200-250g, blocking the rat liver leaf and left in the middle of the hepatic artery and portal vein,45minutes loosen vascular clip, close the abdominal cavity3Specimen collection:after reperfusion in1hour, respectively1day,7days,14days time point8only randomly selected rats, collect peripheral blood circulation5ml, along with saline water portal into flush the liver, wait for a powder after liver gray, quickly take reperfusion of left lobe and middle of the liver.4. Index test:take the rats after reperfusion injury of liver tissue1) TUNEL liver cell apoptosis detection, observation hot ischemia-reperfusion injury of liver cell apoptosis index;2) the Western blot test vascular endothelial growth factor A (VEGF-A), observe related proteins in the liver vascular regeneration the expression;3) take the liver and liver door of bile duct organization (7days and14days two time points) dual von Willebrand factor immune fluorescent marker (vWF) and a-smooth muscle actin (a-SMA), more around the blood vessels of the small bile duct regeneration;4) flow cytometry test cycle and liver tissue was in the distribution of endothelial progenitor cells. Results:1. TUNEL liver cell apoptosis index test showed that:adenovirus group and the other three groups in the same time point comparison, apoptosis index is low (P<0.05); SiRNA group cell apoptosis index is at the same time points higher to the other three groups (P<0.05); Physiological saline groups and blank adenovirus group of liver cell apoptosis several similar, the difference was not statistically significant (P>0.05)2. Western Blotting test results show that compared with time points, adenovirus HO-1the highest expression of VEGF, siRNA express the lowest, and the rest no significant differences between the two groups;3. Immunofluorescence double dye shows:HO-1group see liver and bile duct small blood vessel is around the number to the other three groups increased significantly (P<0.05), blank adenovirus group and physiological saline groups was statistically significant differences, the number of vessels siRNA than the other three groups (P<0.05) less4. Flow cytometric tests showed that:physiological saline groups and blank adenovirus is not statistically significant difference (P>0.05); Heme oxygen and enzyme-1group at the same time point, liver and blood in the content of EPCs significantly higher than the other three group<0.05); SiRNA group in the liver and blood EPCs content was significantly lower than the saline groups, and blank adenovirus group (P<0.05).conclusion:1. In Fei Xsuch as the establishment of the rat liver heat ischemia models based on improved model is a stable and reliable, simple and feasible heat ischemia-reperfusion injury model,. 2. HO-1can reduce rat liver cell apoptosis, promote the vascular endothelial growth factor and the expression of endothelial cells, around the blood vessels of the small bile duct regeneration. So HO-1in the liver ischemia-reperfusion injury play a protecting effection.