The Role Of Hydrogen Peroxide In Hyperdynamic Circulation In Cirrhotic Portal Hypertension

Portal hypertension is one of the severe complications in patients with chronic liver diseases. It is triggered by the increase of intrahepatic resistance and maintained by the overloaded portal venous inflow, which further promotes the formation of portal-systemic collateral vessels. Even though these collaterals aim to lower portal pressure, portal hypertension is maintained by an increased blood flow into the portal system, which thus becomes hyperdynamic.Hyperdynamic splanchnic circulation in portal hypertension is, at least in part, a vascular endothelial growth factor(VEGF)-dependent angiogenic process. Evidence suggests that VEGR-2 blockade decrease the portal venous inflow, the shunting vessels and alleviate the hyperdynamic splanchnic circulation in partial portal vein ligation rats. However,the mechanism under how VEGF been regulated in portal hypertension is still unknow. The underlying mechanisms by which the angiogenesis-associated response are modulated in portal hypertension remain to be explored. Experimental evidence indicates that hydrogen peroxide(H2O2), a form of reactive oxygen species(ROS) in the vasculature, is required for VEGF expression. And interestingly, we have recently found that H2O2 level is increased in mesenteric artery from portal hypertension rats,as compared with normal rats. Moreover, previous studies have also suggested that the portal hypertension syndrome is associated with increased oxidative stress. Therefore, it could be possible that increased level of H2O2 plays a role in angiogenesis-associated responses in portal hypertension. In the present research, we determined the effects of H2O2 blockade on the extent of portosystemic collateral vessels, the development of hyperdynamic splanchnic circulation and the splanchnic neovascularisation, in CCL4-induced portal hypertensive rats and in normal animals.Part 1 The preparation of CCL4 induced rat model of liver cirrhosis and portal hypertensionObjective: To explore the feasibility of preparing liver cirrhosis and portal hypertension rat model with the method of subcutaneous injection of CCL4. Method: Male Sprague-Dawley rats were divided into two groups: PHT group injected with 50% CCl4 oil solution on the back subcutaneously, twice a week for 16 weeks, supplied with 5% ethanol for drinking discontinuously, and normal group injected with physiological saline in the same manner and same dose. After the building up of the model, portal vein pressure, Mean Arterial Pressure, Heart Rate were determined by multifunctional monitor. And liver biopsy HE staining was made. Results:(1)The PHT group rats had the appearance of weight loss, irritability, listlessness, and slow responsiveness, and cirrhotic nodules could be seen clearly in their liver, with intra-abdominal ascites,the model was successful;(2) The HE section of portal hypertensive rats was characterized by the damaged original structure,fibrosis,abnormal nodules and pseudolobule formation.(3) The Mean Arterial Pressure of portal hypertensive rats was lower than normal rats(130.17±5.0 vs 107.86±10.17,p<0.001);the portal vein pressure of portal hypertensive rats was higher than normal rats(15.00±1.64 vs 6.04±0.5,p<0.001).Heart Rate of two group did not be changed. Conclusion: Rat model of liver cirrhosis and portal hypertension can be successfully prepared with the method of subcutaneous injection of CCL4 and supplying with ethanol discontinuously.Part 2 Measurement of the hemodynamic parameters in portal hypertensive rats and the role of H2O2 on hemodynamic parameters of portal hypertensive rats.Objective: To access the DYE-TRAK colored microspheres blood flow system and hydrogen peroxide blockade on hemodynamic parameters in normal and portal hypertensive rats. Method:PE-50 tube or remaining needle were inserted into arteria femoralis, portal venous and left ventricle to applying the colored microspheres blood flow system.Then the hemodynamic parameters were calculated using special formula. Result:(1)The cardiac index, portosystemic collateral vessels, portal venous inflow and intrahepatic resistance of portal hypertensive rats were significantly higher than normal rats;The total peripheric resistance, splanchnic arteriolar resistance and portal-collateral resistance were significantly lower than normal rats;PEG-catalase treatment increased the splanchnic arteriolar resistance, portal-collateral resistance and portal venous resistance,and decreased portosystemic collateral vessels and portal venous inflow.The reduction of portal venous inflow mainly located in the decrease of mesentery and intestine. Conclusion: The DYE-TRAK colored microspheres blood flow system was reliable.PEG-catalase improve the splanchnic hyperdynamic circulation of portal hypertensive rats.Part 3 The mechanism of hydrogen peroxide affecting the expression of angiogenesis markers in four groups.Objective: To explore the effect of hydrogen peroxide on the process of hyperdynamic circulation in cirrhosis and portal hypertension and the possible mechanism. Method: The levels of hydrogen peroxide was measured by the hydrogen peroxide kit; the mesenteric and intestinal angiogenesis markers were measured by western blot. Result:(1)Hydrogen peroxide levels of mesentery and intestine in portal hypertensive rats were higher than that in normal animals, which were decreased by hydrogen peroxide blockade;(2) levels of VEGF,VEGF-2 and CD31 in portal hypertensive rats were higher than that in normal animals, which were decreased by hydrogen peroxide blockade. Conclusion: hydrogen peroxide take part in the formation of hemodynamic circulation in portal hypertension which is associated with VEGF-induced angiogenesis.