The Interactions between the Chronic Exposure to Aluminum and Liver Regeneration on Bile Flow and Organic Anion Transport in Rats

The chronic exposure to Aluminum (Al) may compromise different liver functions, mainly during the hepatic regeneration. The aim of this study is to investigate the interactions between the chronic i.p. exposure to Al and hepatic regeneration (HR) on bile flow and organic anion transport in experimental animals. For this purpose, we studied bile flow and fractional transfer rates for the transport of hepatic organic anions (hepatic uptake, sinusoidal efflux, and canalicular excretion), as well as parameters related with the oxidative stress (OS), on rats chronically treated with Al at 0 and 2 days of HR. The Al treatment and time of HR caused a decrease in the biliary flow and in the hepatic uptake and canalicular excretion constants. In addition, Al and HR increased the lipoperoxidation associated with a reduction of the glutathione content and glutathione peroxidase and catalase enzyme’s activities. Since the effects of Al and HR on biliary flow and transport systems were additive, but not on the oxidative status, different mechanisms might be involved on these alterations. Even though the OS may play a key role on the hepatic deleterious effects, there is no unique cause–effect relationship between OS and liver dysfunction in this experimental animal model.     

Partial hepatectomy hemodynamics changes: Experimental data explained by closed-loop lumped modeling

The liver function may be degraded after partial liver ablation surgery. Adverse liver hemodynamics have been shown to be associated to liver failure. The link between these hemodynamics changes and ablation size is however poorly understood. This article proposes to explain with a closed-loop lumped model the hemodynamics changes observed during twelve surgeries in pigs. The portal venous tree is modeled with a pressure-dependent variable resistor. The variables measured, before liver ablation, are used to tune the model parameters. Then, the liver partial ablation is simulated with the model and the simulated pressures and flows are compared with post-operative measurements. Fluid infusion and blood losses occur during the surgery. The closed-loop model presented accounts for these blood volume changes. Moreover, the impact of blood volume changes and the liver lobe mass estimations on the simulated variables is studied. The typical increase of portal pressure, increase of liver pressure loss, slight decrease of portal flow and major decrease in arterial flow are quantitatively captured by the model for a 75% hepatectomy. It appears that the 75% decrease in hepatic arterial flow can be explained by the resistance increase induced by the surgery, and that no hepatic arterial buffer response (HABR) mechanism is needed to account for this change. The different post-operative states, observed in experiments, are reproduced with the proposed model. Thus, an explanation for inter-subjects post-operative variability is proposed. The presented framework can easily be adapted to other species circulations and to different pathologies for clinical hepatic applications.     

Derecruitment in cat liver: extension of undistributed parallel tube model to effects of low hepatic blood flow on ethanol uptake

Previous studies showed two deviations from the predictions of the undistributed parallel tube model for hepatic uptake of substrates: a small deviation at high flows and a large deviation at low flows. We have examined whether these deviations could be described by a single correction factor. In cats anesthetized with pentobarbital, a hepatic venous long-circuit technique with an extracorporeal reservoir was used to vary portal flow and hepatic venous pressure, and allow repeated sampling of arterial, portal, and hepatic venous blood without depletion of the cat's blood volume. Hepatic uptake of ethanol was measured over a wide range of blood flows and when intrahepatic pressure was increased at low flows. This uptake could be described by the parallel tube model with a correction for hepatic blood flow: Uptake = Vmax max.(1 - e-kF).c/(Km + c). In 22 cats, Vmax max = 90 +/- 5 mumols/(min.100 g liver), k = 0.021 +/- 0.0015 when flow (F) was in millilitres per minute per 100 g liver, and Km = 150 +/- 20 microM when c is the log mean sinusoidal concentration. (1 - e-kF) represents the proportion of sinusoids perfused and metabolically active. A dynamic interpretation of this proportion is related to intermittency (derecruitment) of sinusoidal flow. Half the sinusoids were perfused at a flow of 33 mL/(min.100 g liver) and the liver was essentially completely perfused (greater than 95%) at the normal flow of 150 mL/(min.100 g liver). Derecruitment was not changed by raising hepatic venous pressure, and it was not related to hepatic venous resistance.     

Alterations in soluble guanylate cyclase content and modulation by nitric oxide in liver disease

Hyperammonemia is the main responsible for the neurological alterations in hepatic encephalopathy in patients with liver failure. We studied the function of the glutamate-nitric oxide (NO)-cGMP pathway in brain in animal models of hyperammonemia and liver failure and in patients died with liver cirrhosis. Activation of glutamate receptors increases intracellular calcium that binds to calmodulin and activates neuronal nitric oxide synthase, increasing nitric oxide, which activates soluble guanylate cyclase (sGC), increasing cGMP. This glutamate-NO-cGMP pathway modulates cerebral processes such as circadian rhythms, the sleep-waking cycle, and some forms of learning and memory. These processes are impaired in patients with hepatic encephalopathy. Activation of sGC by NO is significantly increased in cerebral cortex and significantly reduced in cerebellum from cirrhotic patients died in hepatic coma. Portacaval anastomosis in rats, an animal model of liver failure, reproduces the effects of liver failure on modulation of sGC by NO both in cerebral cortex and cerebellum. In vivo brain microdialisis studies showed that sGC activation by NO is also reduced in vivo in cerebellum in hyperammonemic rats with or without liver failure. The content of alpha but not beta subunits of sGC are increased both in frontal cortex and cerebellum from patients died due to liver disease and from rats with portacaval anastomosis. We assessed whether determination of activation of sGC by NO-generating agent SNAP in lymphocytes could serve as a peripheral marker for the impairment of sGC activation by NO in brain. Chronic hyperammonemia and liver failure also alter sGC activation by NO in lymphocytes from rats or patients. These findings show that the content and modulation by NO of sGC are strongly altered in brain of patients with liver disease. These alterations could be responsible for some of the neurological alterations in hepatic encephalopathy such as sleep disturbances and cognitive impairment.     

MRI-based modeling of spleno-mesenteric confluence flow

Characterization of hepatic blood flow magnitude and distribution can lead to a better understanding of the pathophysiology of liver disease. However, the underlying patterns and dynamics of hepatic flow, such as the helical flow structure that often develops following the spleno-mesenteric confluence (SMC) of the hepatic portal vein, have not yet been comprehensively studied. In this study, we used magnetic resonance image (MRI)-based computational models to study the effects of the helical flow structure and SMC geometry on portal blood flow distribution. Additionally, we examined these flow dynamics with four-dimensional (4D) flow MRI in a group of 12 cirrhotic patients and healthy subjects. A validation model was also created to compare computational data to particle image velocimetry (PIV) data. We found significant correlations between flow structure development, vessel geometry, and blood flow distribution in both virtually modified models and in healthy and cirrhotic subjects. However, the direction of these correlations varied among vessel configuration types. Nonetheless, validation model results displayed good qualitative agreement with computational model data.     

Perfusion-limited effects of plasma drug binding on hepatic drug extraction

The hepatic elimination of phenytoin has been studied in the isolated rat liver perfused at constant flow with Krebs solution alone and in the presence of albumin. At an albumin concentration of 0.5 g/dl, 46.6% of the phenytoin was bound in the perfusate and the comparable value at 5.0 g/dl was 87.4%. The increase in binding resulted in a reduction in the hepatic extraction ratio from 0.67 in Krebs to 0.54 and 0.28 at the two albumin concentrations, respectively. Analysis of this data together with that from the literature on propranolol and warfarin indicated that they were consistent with the perfusion-limited model of hepatic clearance. Accordingly, the general relationship between the extraction ratio and the free fraction of drug in the blood is hyperbolic with the precise shape being determined by the ratio of the clearance of the drug from liver water to the hepatic blood flow rate.     

Activation of c-Jun N-terminal kinase during ischemia and reperfusion in mouse liver

We have generated a mouse model for hepatic ischemia in which surgical subcutaneous transposition of the spleen allows hepatic ischemia to be applied without affecting other tissues. Using this mouse model we investigated the relationship between the length of ischemic periods in the liver and subsequent liver function; furthermore, we assayed the activation of c-Jun N-terminal kinase (JNK) during ischemia and reperfusion. Although prior to this study only the activated form of JNK was known to be translocated to the nucleus, we found that JNK translocates to the nucleus during ischemia without activation and is then activated during reperfusion. These results suggest a novel mechanism of JNK activation.     

Nitric oxide-deficiency regulates hepatic heme oxygenase-1.

Nitric oxide plays a crucial role in the maintenance of liver function and integrity. During stress, the inducible heme oxygenase-1 protein and its reaction products, including carbon monoxide, also exert potent hepatoprotective effects. We investigated a potential relationship between endogenous nitric oxide synthesis and the hepatic regulation of heme oxygenase-1. Inhibition of nitric oxide synthesis in vivo by injection of l-NAME led to a dose-dependent induction of heme oxygenase-1 mRNA, protein and activity in the rat liver, whereas did not affect the expression of other heat shock proteins. The effect of l-NAME was demonstrated by hemodynamic changes within the liver circulation as measured by ultrasonic flow probes. Inhibition of nitric oxide synthase led to a decline in hepatic arterial and portal venous blood flow, and subsequently caused liver cell damage. In contrast, the combined administration of l-NAME and the nitric oxide-independent intestinal vasodilator dihydralazine completely restored portal venous flow, abolished the liver cell damage, and prevented the upregulation of heme oxygenase-1, despite inhibition of nitric oxide production. In conclusion, nitric oxide deficiency upregulates hepatic heme oxygenase-1, which is reversible by maintaining hepatic blood flow. This interdependence has important implications for the development of therapeutic strategies aimed at modulating the activity of these hepatoprotective mediator systems.     

Improvement of rat liver function by energy repletion after the preservation period: implications for hepatic graft management

We very recently showed (using a blood-free perfusion model) that cold preservation sensitized rat hepatocyte functions to rewarming ischemic injury and that the injury can be prevented by repleting high-energy adenylates in the liver by short-term oxygenated warm reperfusion. Here we investigated whether short-term reperfusion after the preservation period can improve hepatic graft function in a blood reperfusion model. Eighteen-hour cold-preserved rat livers either untreated (Group A) or pretreated by 30-min oxygenated warm reperfusion after preservation (Group B) were subjected to 20-min ischemic rewarming and then reperfused with blood. Livers in Group B compared to Group A exhibited approx. three times increased bile production and bromosulfophthalein excretion, nearly 7-fold decreased swelling, and 1.2-fold improved blood flow. These results suggest that repletion of the energy by short-term oxygenated reperfusion after prolonged preservation may improve markedly initial hepatic graft function.     

Establishment of animal model of dual liver transplantation in rat

The animal model of the whole-size and reduced-size liver transplantation in both rat and mouse has been successfully established. Because of the difficulties and complexities in microsurgical technology, the animal model of dual liver transplantation was still not established for twelve years since the first human dual liver transplantation has been made a success. There is an essential need to establish this animal model to lay a basic foundation for clinical practice. To study the physiological and histopathological changes of dual liver transplantation, "Y" type vein from the cross part between vena cava and two iliac of donor and "Y' type prosthesis were employed to recanalize portal vein and the bile duct between dual liver grafts and recipient. The dual right upper lobes about 45-50% of the recipient liver volume were taken as donor, one was orthotopically implanted at its original position, the other was rotated 180° sagitally and heterotopically positioned in the left upper quadrant. Microcirculation parameters, liver function, immunohistochemistry and survival were analyzed to evaluate the function of dual liver grafts. No significant difference in the hepatic microcirculatory flow was found between two grafts in the first 90 minutes after reperfusion. Light and electronic microscope showed the liver architecture was maintained without obvious features of cellular destruction and the continuity of the endothelium was preserved. Only 3 heterotopically positioned graft appeared patchy desquamation of endothelial cell, mitochondrial swelling and hepatocytes cytoplasmic vacuolization. Immunohistochemistry revealed there is no difference in hepatocyte activity and the ability of endothelia to contract and relax after reperfusion between dual grafts. Dual grafts made a rapid amelioration of liver function after reperfusion. 7 rats survived more than 7 days with survival rate of 58.3.%. Using "Y" type vein and bile duct prosthesis, we successfully established a novel rat model of dual right upper liver lobe transplantation.     

A biphasic model for sinusoidal liver perfusion remodeling after outflow obstruction

Liver resection can lead to focal outflow obstruction due to transection of hepatic veins. Outflow obstruction may cause additional damage to the small remnant liver. Drainage of the obstructed territories is reestablished via dilatation of sinusoids. Subsequently, sinusoidal canals are formed draining the blood from the obstructed territory to the neighboring unobstructed territories. We raised the phenomenological hypothesis that the blood pressure gradient is the main driving force for the formation of sinusoidal vascular canals. We generated a biphasic mechanical model to describe this vascular remodeling process in relation to the variable pressure gradient. Therefore, we introduced a transverse isotropic permeability relation as well as an evolutional optimization rule to describe the relationship between pressure gradient and the direction of the sinusoidal blood flow in the fluid phase. As a next step, we developed a framework for the calculation concept including the representation of the governing weak formulations. Then, we examined a representative numerical example with simulation of the blood flow under both conditions, the physiological situation as well as after outflow obstruction. Doing so, we were able to reproduce numerically the experimentally observed process of reestablishing hepatic venous drainage via redirection of blood flow and formation of new vascular structures in respect to the fluid flow. The calculated results support the hypothesis that the reorientation of blood flow mainly depends on the pressure gradient. Further investigations are needed to determine the micromechanical influences on the reorientation of the sinusoids.     

SD大鼠酒精性脂肪肝模型的监测分析

目的：通过血清生化指标和病理学的监测分析来建立标准的SD大鼠酒精性脂肪肝动物模型。方法：选取40只SD大鼠,随机分为两组,模型组采用直接饮酒法,于第8、12和20周时检测大鼠血清生化指标：丙氨酸氨基转移酶（ALT）、天门冬氨酸氨基转移酶（AST）、甘油三酯（TG）,并于第8、12周时随机采集5只大鼠肝组织,20周时采集剩余所有大鼠肝组织并进行病理学分析。结果：模型组于第8、12和20周时体重增长量均低于对照组（P〈0．01）,血清ALT、AST均高于对照组（P〈0．01）,第8周和12周时TG高于对照组（P〈0．01）。病理学结果显示肝组织从8周至20周呈现出酒精性脂肪肝、重度酒精性脂肪肝伴肝炎和酒精性肝纤维化等演变过程。结论：直接饮酒法可成功地复制出酒精性脂肪肝动物模型,通过监测分析可了解酒精性脂肪肝病变的整个过程,为今后建立酒精性脂肪肝和肝纤维化动物模型提供了理论参考。     

Effects of positive end-expiratory pressure on hepatic blood flow and performance

Positive end-expiratory pressure (PEEP) may impair extrapulmonary organ function. However, the effects of PEEP on the liver are unclear. We tested the hypothesis that at a constant cardiac output (CO), PEEP does not induce changes in hepatic blood flow (QL) and parenchymal performance. In splenectomized, close-chested canine preparations (group I, n = 6), QL was derived as hepatic outflow using electromagnetic flow probes (QLemf), and hepatic performance was defined by extraction and clearance of indocyanine green (ICG). In a noninvasive model (group II, n = 7), the effects of PEEP on hepatic performance alone were similarly analyzed. Measurements were taken during intermittent positive-pressure ventilation (IPPV1), after addition of 10 cmH2O PEEP to IPPV (PEEP1), during continued PEEP but after return of CO to IPPV1 levels by intravascular volume infusions (PEEP2), and after removal of both PEEP and excess blood volume (IPPV2). Phasic inspiratory decreases in QLemf present during positive-pressure ventilation were not increased during either PEEP1 or PEEP2. Mean QLemf decreased proportionately with CO during PEEP1 (P less than 0.05), but was restored to IPPV1 levels in a parallel fashion with CO during PEEP2. The ICG pharmacokinetic responses to PEEP were complex, with differential effects on extraction and clearance. Despite this, hepatic performance was not imparied in either group. we conclude that global QL reductions during PEEP are proportional to PEEP-induced decreases in CO and are preventable by returning CO to pre-PEEP levels by intravascular volume infusions. However, covarying changes in blood volume and hepatic outflow resistance may independently modulate hepatic function.     

The Relationship Between Plasma and Brain Quinolinic Acid Levels and the Severity of Hepatic Encephalopathy in Animal Models of Fulminant Hepatic Failure

Abstract: Quinolinic acid is an excitatory, neurotoxic tryptophan metabolite proposed to play a role in the pathogenesis of hepatic encephalopathy. This involvement was investigated in rat and rabbit models of fulminant hepatic failure at different stages of hepatic encephalopathy. Although plasma and brain tryptophan levels were significantly increased in all stages of hepatic encephalopathy, quinolinic acid levels increased three- to sevenfold only in the plasma, CSF, and brain regions of animals in stage IV hepatic encephalopathy. Plasma-CSF and plasma-brain quinolinic acid levels in rats and rabbits with fulminant hepatic failure were strongly correlated, with CSF and brain concentrations ∼10% those of plasma levels. Moreover, there was no significant regional difference in brain quinolinic acid concentrations in either model. Extrahepatic indoleamine-2,3-dioxygenase activity was not altered in rats in stage IV hepatic encephalopathy, but hepatic l -tryptophan-2,3-dioxygenase activity was increased. These results suggest that quinolinic acid synthesized in the liver enters the plasma and then accumulates in the CNS after crossing a permeabilized blood-brain barrier in the end stages of liver failure. Furthermore, the observation of low brain concentrations of quinolinic acid only in stage IV encephalopathy suggests that the contribution of quinolinic acid to the pathogenesis of hepatic encephalopathy in these animal models is minor.     

肝硬化动物模型造模方法的研究进展

肝硬化(hepatic cirrhosis,HC)是各种慢性肝病发展的晚期阶段,发病初期表现为轻度乏力、腹胀等症状,后期以肝功能损害和门脉高压为主,随病情进一步发展可出现消化道出血、肝性脑病等严重并发症,死亡率极高。然而,目前研究尚未完全明确肝硬化的发病机制。因此,建立肝硬化动物模型,并通过动物实验研究肝硬化的发病机制,探索肝硬化的防治手段是我们需要重视的问题。由于动物与人类种属之间存在巨大差异,若要完全模拟肝硬化的病理特点制备实验动物模型是非常困难的。为深入了解肝硬化的发病机制和特点,本文将对肝硬化动物模型的制备方法进行综述。     

阻断子宫动脉建立FGR大鼠模型的研究

目的通过暂时阻断妊娠期大鼠子宫血供的方法建立子宫缺血引起胎儿生长受限的动物模型。方法根据大鼠子宫动脉是卵巢动脉的一个分支的解剖特点,于孕鼠妊娠第15天时施行手术暂时阻断卵巢动脉并于第21天行剖宫产术,术后称量新生胎仔体重及胎盘、脑、心、肝、肺、肾等重要脏器重量,对比各组间新生胎仔的预后的不同,并对照研究阻断血供10、20、30及40 min对胎仔的不同影响。结果妊娠晚期阻断孕鼠卵巢动脉20min可成功构建胎儿生长受限模型,这种方法与阻断动脉血流30或40 min相比,手术时间短,技术要求不高,胎仔死亡率与对照组差异无显著性(P>0.05)。各实验组较对照组新生胎仔体重及胎盘、各重要脏器重量均明显降低(P<0.05)。结论通过阻断卵巢动脉从而阻断子宫动脉血流,成功建立缺血缺氧性FGR孕鼠模型。该模型重复性好,操作简便,并可成功设立同体对照,为进行FGR相关的产科理论研究提供了一个有利的技术平台。     

Role of extracellular cGMP and of hyperammonemia in the impairment of learning in rats with chronic hepatic failure. Therapeutic implications

Hepatic encephalopathy is a complex neuropsychiatric syndrome present in patients with chronic or acute liver disease. We review here some recent advances in the study, in animal models, of the mechanisms involved in the impairment in intellectual function in hepatic encephalopathy. These studies show that the function of the glutamate-nitric oxide-cGMP pathway is impaired in brain in vivo in rats with chronic hyperammonemia or liver failure and from patients died in hepatic encephalopathy. This impairment leads to a reduced extracellular concentration of cGMP in the cerebellum and is associated with reduced learning ability in these animal models. Moreover, learning ability of hyperammonemic rats was restored by increasing cGMP by: (1) continuous intracerebral administration of zaprinast, an inhibitor of the cGMP-degrading phosphodiesterase, (2) chronic oral administration of sildenafil, an inhibitor of the phosphodiesterase that crosses the blood-brain barrier and (3) continuous intracerebral administration of cGMP. The data summarized indicate that impairment of learning ability in rats with chronic liver failure or hyperammonemia is due to impairment of the glutamate-nitric oxide-cGMP pathway. Moreover, increasing extracellular cGMP by pharmacological means may be a new therapeutic approach to improve cognitive function in patients with hepatic encephalopathy.     

四氯化碳诱导大鼠肝性脑病模型的制备

目的建立肝性脑病（HE）动物模型。方法应用四氯化碳联合酒精自饮法制作慢性肝功能衰竭引起HE发生的模型,进行迷宫试验、脑电图、血液生化、形态学、行为活动观察等检测。结果 1.肉眼下可见模型组肝脏普遍肿大,表面较粗糙,有较粗的颗粒,暗红色;正常组肝脏未见异常。模型组肝内有大量纤维增生,肝细胞溶解,弥漫性大片坏死,残留肝细胞成片气球样变。2.与对照组大鼠比较,造模组第九周水迷宫测试逃避潜伏期明显延长,以潜伏期延长超过正常x＋2.5 s为标准,本实验有14/20（55%）只大鼠水迷宫潜伏期明显延长;3.HE模型组血氨浓度明显高于正常对照组,组间比较差异有显著性（P〈0.01）;4.正常组脑电图以α波为主,无明显异常;模型组出现肝性脑病变化。结论 1.建立了肝硬化合并肝性脑病较为理想的动物模型;2.水迷宫测试联合检测更易检出HE。     

自噬在肝再生中的作用

自噬是存在于真核细胞内的一种溶酶体依赖性的降解途径,在肝脏生理和病理过程中发挥着重要作用。肝脏具有强大的再生能力,在受到急、慢性损伤时,残肝细胞将会被激活进入细胞周期进行细胞增殖,以补偿丢失的肝组织和恢复肝功能。文章阐述了各种类型损伤之后的肝再生与自噬的关系。在物理性、酒精、食源性等因素引起的肝损伤中,肝脏通过启动自噬来促进肝再生;在化学性损伤的肝再生模型中,自噬在其中的作用仍然有争议;在病毒感染之后的肝再生中,一些嗜肝病毒（如丙肝病毒和乙肝病毒等）反而利用自噬来促进病毒颗粒复制,抑制肝再生。对自噬和肝再生机制的研究,将有助于进一步阐明再生过程,为治疗肝脏疾病提供新方法。