Liver Injury In Rat Model Research Articles

Effect of Hepatic Stimulator Substance Administration on Tissue Regeneration Due to Thioacetamide-Induced Liver Injury in Rats

Background: Hepatic stimulator substance (HSS) is a known hepatic growth factor that appears to be organ-specific but species-nonspecific. In the present study we investigated the effect of HSS administration in a rat model of liver injury and regeneration induced by thioacetamide (TAA) injection. Methods: TAA (300 mg/kg body weight) was injected intraperitoneally in male Wistar rats (group I). HSS (50 mg protein/kg body weight) was administered intraperitoneally either at 24 h (group II) or at 36 h (group III) after TAA treatment. The animals were killed at different time points after TAA injection, and the rate of tritiated thymidine incorporation into hepatic DNA, the activity of the enzyme thymidine kinase in liver, and the assessment of the mitotic index in hepatocytes were used to estimate liver regeneration. Results: The administration of TAA caused severe hepatic injury recognized histopathologically and by increased activities of the serum hepatic enzymes aspartate and alanine aminotrasferases. The hepatic injury, which peaked at 24 h and 36 h after TAA injection, was followed by a regenerative process of hepatocytes which presented peaks after 48 h and 60 h (group I). The regenerative process of hepatocytes remained unaffected when HSS was administered 24 h after the injection of TAA (group II). In the case of HSS administration 36 h after the injection of TAA (group III) the examined indices of hepatocyte proliferation were statistically significantly increased at 48 h (P < 0.001), compared with those observed in group I. Conclusions: The administration of HSS enhanced the hepatocyte proliferative capacity, induced by TAA treatment, depending on the time of its administration.

Immunomodulatory action of G-CSF In a rat model of endotoxin-induced liver injury: an intravital microscopic analysis of Kupffer cell and leukocyte response

In contrast to the anticipation that in sepsis granulocyte colony-stimulating factor (G-CSF) would overactivate the nonspecific immune system by recruiting and priming leukocytes with consequent aggravation of inflammatory tissue lesions, recombinant (r) G-CSF pretreatment was protective in various experimental non-neutropenic models of inflammation. The mechanisms of protection, however, are not fully understood. Using intravital fluorescence microscopy, we show that rG-CSF enhances leukocyte endothelial cell interaction within the microvasculature of normal rat livers, whereas rG-CSF pretreatment of animals exposed to lipopolysaccharide (LPS) attenuates the LPS-induced leukocytic response, including stasis in sinusoids as well as rolling and adherence in postsinusoidal venules with subsequent tissue infiltration. Moreover, rG-CSF, which did not affect Kupffer cell activity in normal rat livers, reduced the immediate activation of Kupffer cells on LPS exposure, as indicated in vivo by the delayed adherence/phagocytosis of intra-arterially administered latex particles associated with attenuation of proinflammatory cytokine release (tumor necrosis factor alpha and interleukin-6). Finally, rG-CSF reduced LPS-induced nutritive perfusion failure and hepatocellular excretory dysfunction. This study provides evidence for a distinct, possibly tumor necrosis factor alpha-dependent modulation of LPS-induced cellular response within the liver by rG-CSF, thereby achieving protection against microcirculatory perfusion failure and hepatic dysfunction.

Tissue inhibitor of metalloproteinase-1 messenger RNA expression is enhanced relative to interstitial collagenase messenger RNA in experimental liver injury and fibrosis

Liver fibrosis results from a relative imbalance between synthesis and degradation of matrix proteins. We have previously described release of the protein collagenase inhibitor, tissue inhibitor of metalloproteinase-1 (TIMP-1), by culture-activated human hepatic stellate cells (HSCs). In this study, we have investigated the relative expression of TIMP-1 and interstitial collagenase in culture-activated rat HSCs and rat models of liver injury and fibrosis. The complementary DNA (cDNA) for rat TIMP-1 was obtained by homology polymerase chain reaction (PCR) and sequenced. By Northern analysis using this probe, TIMP-1 messenger RNA (mRNA) expression was up-regulated with HSC activation by culture on plastic as defined by cellular expression of procollagen-1. Interstitial collagenase mRNA was expressed in early 1. Interstitial collagenase mRNA was expressed in early culture (<4 days) but became undetectable in more activated cells (7-21 days). By activity assay of serum-free cell-conditioned media, TIMP-1 was found to be released in increasingly concentrations with duration of culture on plastic. Expression of TIMP-1 interstitial collagenase, and procollagen-1 mRNAs were studied in rat models of biliary and parenchymal injury (bile duct ligation and CC14 administration) by ribonuclease protein assay. TIMP-1 mRNA expression was increased at 6, 24 hours, and 3 days after bile duct ligation and was also shown to rise in acute CC14 liver injury and remain elevated as the liver became fibrotic. TIMP-1 expression preceded procollagen-1 expression in both models. In contrasts, interstitial collagenase mRNA levels remained similar to control values throughout both models of liver injury. Total cellular RNA from hepatocytes, HSCs, and kupffer cells freshly isolated from livers after acute CC14 injury was subjected to Northern analysis. TIMP-1 transcripts were observed in nonparenchymal cells only. We suggest that increased expression of TIMP-1 relative to interstitial collagenase by HSCs may promote progression of liver fibrosis in these rat models by preventing degradation of secreted collagens. (Hepatology 1996 Jul;24(1):176-84)

Oral arginine supplementation in acute liver injury

Acute liver failure is accompanied by a high rate of bacterial and septic complications. Arginine has a potent effect on the immune system and modulates bacterial clearance in septic models. We studied the effect of oral arginine supplementation on the extent of liver injury and the associated bacterial translocation in an acute liver injury model in rats. Sprague-Dawley rats were divided into normal, liver injury, and arginine-supplemented groups. In the arginine group, 2% arginine was supplemented daily through a nasogastric tube for 8 d. Acute liver injury was induced on the eighth day by intraperitoneal injection of d-galactosamine (1.1 g/kg body wt). Samples were collected 24 h after the liver injury. In the arginine-supplemented group, alkaline phosphatase, bilirubin, and aspartate aminotransferase were reduced significantly compared with the acute liver injury control group. The results of bacterial translocation in the arginine-supplemented group showed a significantly reduced number of translocated bacteria to the liver and mesenteric lymph nodes than occurred in the acute liver injury group. The histological study of the liver in the arginine-supplemented group showed scattered areas of hepatocellular necrosis and inflammatory cell infiltration, and in the acute liver injury group there were more and widespread hepatocellular necrosis and inflammatory cell infiltration. Oral supplementation of arginine in an acute liver injury model improves significantly the state of the liver injury and reduces bacterial translocation to the liver and mesenteric lymph nodes.

Regulation of monocyte chemoattractant protein 1 by cytokines and oxygen free radicals in rat hepatic fat-storing cells

BACKGROUND & AIMS: Monocyte chemoattractant protein 1 (MCP-1) is a potent monocyte/macrophage chemoattractant expressed by fat-storing cells (FSCs) in rat models of liver injury. This study investigated the mechanism of this activation of hepatic MCP-1 expression. METHODS: The regulation of MCP-1 messenger RNA (mRNA) expression and protein synthesis was examined in FSC lines derived from CCl4-induced cirrhotic rat liver (cirrhotic FSCs) and normal rat liver (normal FSCs). RESULTS: Northern blot hybridization analysis revealed low levels of MCP-1 mRNA in cultured cirrhotic FSCs that increased markedly after treatment with tumor necrosis factor alpha, interleukin 1 alpha, or transforming growth factor beta 1. All three cytokines increased the synthesis and secretion of MCP-1 protein. Oxygen free radical production also increased MCP-1 mRNA levels. These increases in MCP-1 mRNA were blocked by dexamethasone. In normal FSCs, levels of MCP-1 mRNA and secreted protein were increased in response to cytokines or oxygen free radical production, but the magnitude and duration of this increase was less than in cirrhotic FSCs. CONCLUSIONS: In liver injury, monocyte/macrophage recruitment and activation from FSC production of MCP-1 may be stimulated by cytokines and oxygen free radicals. During chronic liver injury leading to cirrhosis, FSCs may become hypersensitive to these stimuli, further fueling the inflammatory response. (Gastroenterology 1996 Jun;110(6):1870-7)

Ethanol-related liver injury in the rat: a model of steatosis, inflammation and pericentral fibrosis

Background/Aims: While several animal models exist for the study of ethanol heptotoxicity, they are limited in their applicability. This paper describes a relatively simple rat model of alcohol-related liver injury. Methods: Ethanol was supplied in the drinking water in a concentration of 40% v/v for up to 29 weeks. Animals are concurrently supplied a chow diet which provides adequate protein and choline for normal growth. Total fat intake is low (7% of consumed calories). Results: Histological changes of steatosis, inflammation, hepatocyte necrosis and pericentral sclerosis were evident in ethanol-treated rat livers. Littermate controls with and without pair-feeding had normal livers. Electron microscopy revealed abnormal mitochondria and a marked proliferation of smooth endoplasmic reticulum in livers of animals fed ethanol. Biochemical analysis revealed that levels of hepatic-free choline were similar in treated and pair-fed control rats. There was an expected increase in the activity of the microsomal enzyme cytochrome P450 2E1 in ethanol-fed rats. Conclusions: The model provides a convenient method for the production of alcoholic liver inquiry, and it may be useful for the study of the pathogenesis of ethanol-related liver disease.

Protective action of putrescine against rat liver injury.

The hepatoprotective action of orally dosed putrescine was investigated using rat models of liver injury. When rats received putrescine orally soon after a dose of carbon tetrachloride or D-galactosamine, deranged serum alanine aminotransferase values and prothrombin times were significantly attenuated compared with control levels, with improved histologic extent of liver injury. Putrescine addition to the medium of rat hepatocytes in primary culture reduced cell killing induced by D-galactosamine or the membrane detergents chenodeoxycholic acid and Triton X-100. Similar reduction was seen in cells exposed to tert-butyl hydroperoxide (TBHP), an agent producing cell death through lipid peroxidation, with attenuation of cellular malondialdehyde content. Putrescine also significantly attenuated the extent of increased plasma membrane microviscosity as assessed with 1-[4-(trimethylammonio)phenyl]-6-phenyl-1,3,5-hexatriene in TBHP-treated cells. These results suggest that orally given putrescine protects against liver injury. Plasma membrane stabilization and reduction of lipid peroxidation may contribute to this hepatoprotection.

The Role of Calcium Ions and Calcium Channel Entry Blockers in Experimental Ischemia–Reperfusion-induced Liver Injury

Verapamil administered before treatment, but not after treatment, had a beneficial effect on a 90-minute warm ischemia-reperfusion rat liver injury model. The possible activation of proteases converting the xanthine dehydrogenase to xanthine oxidase, the significant mitochondrial calcium loading during the ischemic period, and the potentiation of calcium and oxygen-derived free radicals to promote injury to mitochondria are mechanisms supported by this study, based on both histologic observations and on the pattern of enzyme leak after the acute ischemic event.

Effect of Endotoxin on Release of Reactive Oxygen Intermediates by Rat Hepatic Macrophages

Reactive oxygen intermediates released by activated hepatic macrophages have been implicated in the pathogenesis of a rat model of liver injury induced by sequential administration of Corynebacterium parvum and endotoxin. In this model, C. parvum causes extensive infiltration of the liver with activated macrophages, but severe liver injury occurs only after subsequent exposure to endotoxin. We have therefore investigated the effects of endotoxin on the release of reactive oxygen intermediates by C. parvum-activated hepatic macrophages. After in vitro exposure to zymosan, opsonized zymosan, or phorbol myristate acetate, hepatic macrophages isolated from C. parvum- and endotoxin-treated rats demonstrated significantly (1.5-2-fold) increased release of superoxide and oxidation of [1-14C]glucose via the hexose monophosphate shunt compared with hepatic macrophages isolated from C. parvum- and saline-treated control rats. These results indicate that endotoxin enhances the state of activation of hepatic macrophages already partially activated by C. parvum. We suggest that the increased release of reactive oxygen intermediates by these cells promotes liver injury in this model.

Beneficial effects of superoxide dismutase and ascorbic acid in a rat model of postischemic liver injury

Beneficial effects of superoxide dismutase and ascorbic acid in a rat model of postischemic liver injury

Corynebacterium parvum-elicited hepatic macrophages demonstrate enhanced respiratory burst activity compared with resident kupffer cells in the rat

We have recently demonstrated that release of oxygen-derived free radicals by activated hepatic macrophages may be involved in the pathogenesis of a rat model of liver injury induced by Corynebacterium parvum and endotoxin. In the present study we have compared the respiratory burst activity of isolated normal rat Kupffer cells with that of hepatic macrophages elicited by C. parvum. Superoxide production (O2·−) and glucose oxidation via the hexose monophosphate shunt (HMPS) were low in normal Kupffer cells, but were significantly increased (O2·− 2.1-fold, HMPS 1.7-fold) by phorbol myristate acetate, a stimulant of the respiratory burst. Corynebacterium parvum-elicited hepatic macrophages demonstrated significantly enhanced Superoxide production and HMPS activity compared with normal Kupffer cells, both in the absence of specific stimuli (O2·− 3.3-fold, HMPS 5.3-fold) and after exposure to phorbol myristate acetate (O2·− 4.5-fold, HMPS 5.3-fold). These results demonstrate that normal Kupffer cells are capable of exhibiting respiratory burst activity, but this is markedly increased for hepatic macrophages elicited by an inflammatory stimulus.

Change of hepatic histamine content during hepatic fibrosis

Hepatic function was studied by measuring the time courses of several variables in blood and liver using a chronic liver-injury model produced by administering CCl 4 consecutively for 12 weeks in rats. A marked increase in liver histamine content occurred after 10 weeks of treatment with CCl 4. At weeks 10 and 12, liver histamine levels in the CCl 4-treated group were 1.95 and 4.61 times higher, respectively, than in the control group. This change in liver histamine content appeared after that in other variables such as glutamic pyruvic transaminase, alkaline phosphatase, and white blood cells, but it corresponded to a change in liver hydroxyproline. Increased mast cells were seen in fibrotic foci around Glisson's sheath by microscopic morphological observation of the liver 12 weeks after treatment with CCl 4. The histamine concentration in plasma tended to decrease after CCl 4 treatment, and at week 12 the decrease was statistically significant compared with control. The liver activities of histamine-metabolizing enzymes, histamine- N-methyltransferase and histaminase, decreased to 1 3.4 and 1 6.0 times those of the nontreated group, respectively, 12 weeks after treatment with CCl 4, whereas blood histaminase increased about 9.2 times. The increase in histamine content in injured liver was presumedly derived from the increase in mast cells in the inflamed area of the liver; also, the deficiency of histaminemetabolizing enzymes in liver might have caused the high histamine content in the liver. On the other hand, the decrease in plasma histamine concentration might have occurred as a consequence of the enzyme leakage from hepatocytes that accompanied the breakdown of hepatocytes by CCl 4 and thus, of the histamine metabolism in blood by the leaked enzymes. The same kind of experiment was performed using a dimethylnitrosamine-induced liver injury model in rats. The increase of hydroxyproline in the liver occurred 11 days after that of histamine content in liver. These results suggest the possibility that increased histamine in the liver may participate in the biosynthesis of collagen.

Bile flow and biliary sulfobromophthalein sodium excretion in rats with liver cirrhosis and portacaval shunt.

In order to study the combined effect of end-to-side portacaval shunt and chronic liver damage on biliary organic anion transport, the rat model of thioacetamide-induced chronic liver injury was utilized. Compared with sham-operated animals, bile flow and maximal biliary excretion (Tm) of sulfobromophthalein sodium (BSP) was decreased in rats 9 weeks after shunt operation. If rats with shunts were treated for 8 weeks with thioacetamide, an agent causing hepatic fibrosis and pseudolobule formation, BSP transport from the hepatocyte into bile was further diminished. Compared with the shunted controls, the thioacetamide-treated rats with shunts had elevated serum bilirubin and alkaline phosphatase concentrations, and on electron microscopy their livers had dilated bile canaliculi with a decreased number of microvilli. Non-shunted rats treated with thioacetamide for 8 weeks had similar but less severe changes in the canalicular ultrastructure. Bile flow and BSP Tm were not influenced by thioacetamide treatment alone, perhaps due to the marked liver hypertrophy in these animals. These results indicate that canalicular active transport of organic anions is more sensitive to the effect of thioacetamide in animals with portacaval shunts than in those with sham operations. A similar impairment of hepatic organic anion handling by hepatotoxic compounds might be the consequence of portasystemic shunts in patients with liver cirrhosis.