INTRODUCTION.

Gut is a vast pool of bacteria and endotoxins (lipopolysaccharide, LPS) in the body. There are endotoxemia in severe trauma, burn or scald, Intestinal ischemia and liver diseases. They are mainly resulted from large quantities of endotoxin produced by overgrowth of gram-negative bacteria in gut that are increasingly taken into portal vein because of the incereased permeability of intestinal wall. Endotoxemia will be generated if the level of endotoxin surpasses the hepatic capacity for endotoxin scavenging due to decreased phagocytic ability of liver kupffer cells (KCs) and then endotoxin spills over into systemic circulation. For these reasons, it is named IETM because endotoxin comes from gut.

In recent years, the relationship between IETM and liver diseases has been increasingly paid much attention to. Animal studies have demonstrated that various experimental liver diseases were commonly accompanied by IETM. Clinical observations have also showed evidences that the highest incidence(80%-100%) and severe degree of IETM in patients with severe hepatitis was generally recognized, though the incidences of IETM in patients with acute or chronic hepatitis, severe hepatitis, cirrhosis and hepatocarcinoma were not reported equally. Incidences of IETM investigated in our institute in acute or chronic hepatitis, severe hepatitis and cirrhosis were 75%, 79%, 93.3% and 84.3% respectively[1].

LPS activates KCs via two pathways: one is membrane attachment from mCD₁₄ which is the classical CD₁₄ dependent pathway, that requires LPS binding protein (LBP) as a cofactor carring LPS to the membrane of KCs bound to the receptor CD₁₄ ; the soluble compound formed by combining LBP with LPS increases affinity of LPS with CD₁₄. Whereas the other pathway-sCD₁₄ may not require LBP, combines with corresponding receptor on KCs by aid of other proteins (such as HDL, LDL etc). These two pathways finally activate signal transduction system and trigger the synthesis and release of cytokines and inflammatory mediators[2].

KCs stimulated by LPS release chemostatic substances such as TNF-α, interleukins, leukotriene B₄ and complement C₅, which attract circulating neutrophilic leucocytes. Activated neutrophilic leucocytes up-regulate adhesion molecules receptor (CD_11b/CD₁₈) on its membrane surface, then adhere to endothelial cells of hepatic sinusoids. In the meanwhile, adhesion molecules ICAM-1 and ELAM-1 on surface of endothelial cells are also up-regulated and precipitate neutrophilic leucocytes moving to hepatic sinusoids. Oxygen-derived free radicals produced by these activated neutrophilic leucocytes cause lipid peroxidation of cells. Moreover, toxic mediators (PAF, NO, ET-1 etc) released from KCs enhance further liver injury and make platelets aggregated into microthrombi. Hepatic sinusoids being blocked by adhering neutrophilic leucocytes, Platelets and swelling KCs induce slowing of blood flow with following hypoxia. The onset of liver necrosis in this situation is ascribed mainly to abnormal microcirculation. Therefore, liver damage is an overall outcome that is induced by the interactions between inherent KCs, immigranting neutrophilic leucocytes and hepatocytes[3].

Additionally, it shoud be mentioned that LPS not only induces liver necrosis directly or indirectly by chemical mediators released from activated KDs, but also is a promoting factor for other hepatotoxicants (such as D-galactosamine, tetrachloride [CCl₄],ethanol, etc) to induce liver necrosis. The use of anti-LPS antibody can significantly attenuate liver necrosis induced by hepatotoxicants and prevent from the occurrence of acute liver failure[4].

There are still some arguments on whether or not LPS directly injures hepatocytes at present. It is reported that there is high content of lipoid (phospholipid mainly) in the membrane of hepatocellular mitochondria. The affinity of diphosphatidyl glycerol, which is the special component of the lipoid, with is the special component of the lipoid, with LPS is very strong. Its acyl group that binds to LPS induces the structural damage of mitochondria, interferes with biological oxidation, inhibits production of ATP and causes hepatocellular injury. LPS can directly insert into double layers of lipid molecule of membrane or conjugate to membrane receptors and initiate the metabolism of membrane phosphatidyl inositol. Large quantities of phosphatidic acid (PA) and inositol triphosphate (IP₃) are produced. Degeneration and necrosis of hepatocytes are induced by elevated Ca²⁺ in cytoplasm, which is resulted from PA with Ca²⁺ carrier-like action and IP₃ which causes the opening of Ca²⁺ channel in membrane, with great deal of Ca²⁺ flowing the cell. Structural and functional injuries of hepatocytes and mitochondria can also be caused by degradation of membrane phospholipid and lipid peroxidation mediated by PLA₂ activation induced by LPS. LPS phagocytosed by KCs can damage the membrane of lysosome and lead to release of various lysosomal enzymes resulting in cell autolysis.

To explore LPS directly acting on hepatocytes in vitro, we found that hepatocytes showed typical ladder pattern of apoptotic cells morphologically and this process could be inhibited by apoptotic inhibitor ATA. The number of apoptotic hepatocytes was proportional with the dosage of LPS; the number of apoptotic hepatocytes treated with LPS within 24 hours was also in positive ratio with time. It implicated that LPS could directly induce apoptosis of hepatocytes and this action was more prominent at the early phase of treatment[5].

The interaction of LPS with KCs plays a determinant role during the occurance of liver injury and acute liver failure. It was confirmed in a large number of animal experiments that liver injury induced by CCl₄, D-galactosamine, thioacetamide (TAA), ethanol, etc. could be reduced or prevented by performing colectomy or administering antibiotics (such as polymyxin B, mycifradin) to decrease the level of intestinal LPS, or by administering gadolinium chloride (GdCl₃), silica (SiO₂)to block KCs.

The research performed by Ahmad et al[6] indicated that there was high expression of ICAM-1 on hepatocytes, KCs, endothelial cells during acute endotoxemia; there was no expression of ICAM-1 on hepatocytes and KCs when KCs were blocked by GdCl₃; there is no occurrence of TNF-α and LTB₄, which is released from KCs activated by LPS, attracting neutrophilic leucocytes to infiltrtate into liver. These results implicated that KCs played a vital role in hepatitis and liver injury induced by LPS.

Many researches suggested that ethanol can increase permeability of intestinal mucosa, macromolecules such as LPS were intaken into blood and the plasma level of LPS thus rose up. While phagocytosing LPS, KCs released chemical mediators such as TNF-α, PGE₂ to promot oxidation of alcohol in the liver. Owing to increased oxygen consumpsion, hepatocytes was injured by free radicals formed in the state of hypoxia. Experiments verified that the level of plasma LPS in alcoholism animals was significantly correlated with hepatic pathological change (cellular steatosis and necrosis, inflammatory infiltration); the steatosis and necrosis of liver induced by alcohol could be prevented by administering GdCl₃ to block KCs[7].

D-galactosamine can induce focal necrosis of whole hepatic lobule with severe infiltration of neutrophlic leucocytes, but liver injury can be thoroughly prevented by GdCl₃ injection in caudal vein before D-galactosamine injection intraperitoneally in rats. Liver injury and apoptosis induced by D-galactosamine and small dose of LPS can be markedly alleviated via treatment with TNF-α antiserum, so did alcoholic liver injury. Because TNF-α was mainly stemmed from activated KCs, the key role of KCs played in the liver injury was thus certified[8,9].

In summary, previous researches on mechanism of liver injury were mostly confined to parenchymal liver cells, whereas large amount of current experiments have demonstrated that activation of KC by LPS plays a more important role in the occurance and development of liver injury.

Based upon the above-mentioned understanding we have conceived the following hypothesis on liver injury induced by various pathogenic factors and the development and prognosis of hepatitis (Figure ​(Figure11)

Figure 1

Diagram of the hypothesis of IETM in liver failure

The "primary liver injury (PLI)" is the hepatic damage that various pathogenic factors (such as hepatitis virus, alcohol, drugs, hepatotoxicants, etc.)induced by specific mechanism separately. In contrast, IETM is formed in the course of the development of hepatitis. Liver injury, which is resulted from LPS, activation of KCs by LPS and others, is refered to as "secondary liver injury (SLI)" The SLI which has lost their own specificities of primary pathogenic factors is ascribed to IETM.

The SLI has an important effect on the development and prognosis of various hepatitis. The severe IETM, which commonly causes over-inflammatory reaction and serious hepatic necrosis, will lead to severe hepatitis and even acute hepatic failure[10]. The mild IETM will successively precipitate a cascade, including the repeated and persistent hepatocellular injury accompanied by infiltration of in- flammatory cells, hepatic fibrosis, cirrhosis and even hepatocarcinoma. Generally, it ends with chronic hepatic failure[11].

Certainly, the mechanism of occurrence and development of various viral hepatitis is not so simple but a considerably complicated issue related to virus itself, host immune state, etc. It is difficult to cover it by only one hypothesis.