AIM: To analyze hepatic, mesenteric and mucosal microcirculation and leukocyte-endothelium interaction (LEI) in a rat model with liver cirrhosis.

METHODS: Hepatic cirrhosis was induced in Wistar rats by gavage with carbon tetrachloride, and intravital videomicroscopy was performed in liver, mesentery and small intestine mucosa. Special emphasis is given on microcirculatory and morphometric changes during cirrhotic portal hypertension.

RESULTS: LEI was influenced significantly in the cirrhotic liver but not in the gut. Blood flow measurement showed significant differences among liver, main mesenteric vessels and the mucosa. The results of our study indicate that liver cirrhosis leads to alterations in hepatic and mesenteric blood flow but not in mucosal blood flow.

CONCLUSION: The enhanced inflammatory response in hepatic microvessels may be caused by a decrease of antithrombin III levels and could be responsible for disturbances of organ pathology.

Liver cirrhosis is recognized as an important risk factor for the development of severe septical complications such as spontaneous bacterial peritonitis and bacteriaemia[1]. The impairment of intestinal microcirculation and mucosal barrier may contribute to an increased intestinal permeability and bacterial translocation[2-4]. Hypoperfusion of the gut mucosa has been implicated as an important mechanism contributing to gut-derived endotoxinemia and bacterieamia in liver cirrhosis. Due to splanchnic arterial vasodilatation and portal hypertension, intestinal capillary pressure is altered and effective arterial blood volume decreases[5].

It is generally recognized that there is a close relationship between the hepatic microcirculation and liver function and structure[6,7]. Recent studies established the important correlation between hepatic microcirculation and the development of liver pathology[8]. Furthermore, it has been described that leukocyte adherence in liver sinusoids is amplified after gut ischemia and reperfusion due to an up-regulation of adhesion molecules[9].

We still lack quantitative data describing hepatic and mesenteric microvascular parameters such as blood flow velocity, volumetric blood flow, leukocyte-endothelial interaction and vessel size within one experimental setting. Currently, intravital videomicroscopy is the most pretentious technique to measure variables of microvascular perfusion in vivo. In addition to the measurement of mesenteric blood flow and leukocyte kinetics, we quantified mucosal blood flow in terminal arterioles directly, since the assessment of mucosal blood flow has been simplified[10].

The present study was undertaken to quantitate basic microvascular parameters in normal rat liver and intestine and to investigate changes in these parameters associated with the development of cirrhosis.

Male Wistar rats weighing 450±47 g at the time of surgery (initial weight 250±5 g) were used in all experiments. They were housed in a controlled environment with a 12-h light: dark cycle and were fed standard rat diet with water ad libitum.

Chronic, progressive hepatic cirrhosis was induced according to the method of Proctor and Chatamra by gavage with carbon tetrachloride (CCl₄)[11]. Animals were given phenobarbital (35 mg/100 mL) in their drinking water to induce the enzyme cytochrome P450, which has been shown to increase rat liver sensitivity to CCl₄[12]. Two weeks later an initial dose of 0.04 mL CCl₄ was given after orogastric intubation and enhanced weekly in steps of 0.04 mL to a maximum dose of 0.4 mL CCl₄. Animals were weighed weekly.

Six control animals without liver cirrhosis were observed for statistical analysis.

Cardiorespiratory parameters did not differ between groups throughout the observation period (Table 1).

Liver cirrhosis has been detected to be an important risk factor for mortality in critically ill patients[16]. Its incidence in western countries is rising and represents the most common cause of mortality among the non-malignant digestive diseases[17]. The mechanisms for the increased morbidity and mortality may be immunological, mechanical or pharmacological. Special emphasize is given to hepatic and intestinal microcirculatory disorders. However, it is still not clear how alterations in hepatic microcirculation contribute to the progression of the disease and its sequelae.

In the present study, we have applied the technique of intravital videomicroscopy to quantitate intestinal and hepatic microcirculation and LEI in cirrhotic rats. We used a model of CCl₄-induced cirrhosis. Liver damage is mainly caused by hepatotoxic CCl₄ metabolites produced within the liver. The intensity of the hepatic damage is exaggerated when the microsomal enzyme oxidizing system is previously induced, for instance by the administration of phenobarbital[12]. Induced cirrhosis is associated with marked hemodynamic disturbances. These include changes in both the intrahepatic and splanchnic circulation resulting in portal hypertension[18,19]. Our quantitative studies of hepatic and mesenteric microvascular parameters indicate that cirrhosis leads to a marked arterial vasodilatation. This observation is in accordance with other experimental studies. The pathogenesis of vascular tone is explained by biochemical investigations and disorders of vascular heme oxygenase-1 expression and nitric oxide synthase expression in experimental cirrhosis[20,21 ].

In this study, presence of liver cirrhosis had no effect on villus blood flow measured in the terminal, central villus arteriole. To our knowledge, there is no other study where the effects of cirrhosis and portal hypertension on villus blood flow were derived directly from data obtained on the level of single villus arterioles. The disturbance of mucosal blood flow may contribute to an impairment of the intestinal barrier function. It has been described that alterations of the mucosal permeability in cirrhosis are related to the degree of liver failure and to the progression of the liver disease[4].

However, the impact of liver cirrhosis on hepatic microcirculation is more evident. LEI is enhanced significantly and may explain organ disturbances. LEI itself is modulated by a variety of adhesion molecules expressed on the surface of leukocytes and endothelial cells. These adhesion molecules mediate the decrease in leukocyte rolling and the increase in leukocyte adherence and migration.

In summary, this study demonstrates that in the gut mucosa, cirrhosis may not induce disturbances in the villus microcirculation, despite altered levels of mesenteric blood flow.

Experimental liver cirrhosis is associated with a marked increase of liver enzymes and a decrease of antithrombin III levels and prothrombin time. Antithrombin III is one of the most important physiological inhibitors of coagulation. It is synthesized in liver parenchymal cells. Moreover, it has been described that AT III has anti-inflammatory actions in experimental sepsis and hepatic ischemia-reperfusion[22,23]. It could be presumed that in cirrhosis a drop of the antithrombin III level leads to hemostatic disorders similar to disseminated intravasal coagulation (DIC) and inflammatory response in the microcirculatory units, even if clinical observations did not emphasize those findings[24,25 ]. In our study, hemostatic dysfunction could be measured and LEI in liver parenchyma and mesenteric vessels was enhanced significantly.

We conclude that experimental liver cirrhosis leads to disturbances in hepatic and mesenteric but not in mucosal microcirculation. Although not proven by our experiments, lower antithrombin III levels as found in our study could potentially maintain or aggravate hemostatic disorders and alterations in hepatic inflammatory response to portal hypertension associated with a deterioration of liver function.