Natural killer cells ameliorate liver fibrosis by killing activated stellate cells in NKG2D-dependent and tumor necrosis factor-related apoptosis-inducing ligand-dependent manners

BACKGROUND & AIMS: Viral hepatitis infection, which is a major cause of liver fibrosis, is associated with activation of innate immunity. However, the role of innate immunity in liver fibrosis remains obscure. METHODS: Liver fibrosis was induced either by feeding mice with the 3,5-diethoxycarbonyl-1,4-dihydrocollidine (DDC) diet or by injecting them with carbon tetrachloride. The Toll-like receptor 3 ligand, polyinosinic-polycytidylic acid, was used to activate innate immunity cells and mediators, including natural killer cells and interferon gamma. RESULTS: In the mouse model of DDC-induced liver fibrosis, natural killer cell activation by polyinosinic-polycytidylic acid induced cell death to activated hepatic stellate cells and attenuated the severity of liver fibrosis. Polyinosinic-polycytidylic acid treatment also ameliorated liver fibrosis induced by carbon tetrachloride. The observed protective effect of polyinosinic-polycytidylic acid on liver fibrosis was diminished through either depletion of natural killer cells or by disruption of the interferon gamma gene. Expression of retinoic acid early inducible 1, the NKG2D ligand, was undetectable on quiescent hepatic stellate cells, whereas high levels were found on activated hepatic stellate cells, which correlated with the resistance and susceptibility of quiescent hepatic stellate cells and activated hepatic stellate cells to natural killer cell lysis, respectively. Moreover, treatment with polyinosinic-polycytidylic acid or interferon gamma enhanced the cytotoxicity of natural killer cells against activated hepatic stellate cells and increased the expression of NKG2D and tumor necrosis factor-related apoptosis-inducing ligand on liver natural killer cells. Blocking NKG2D or tumor necrosis factor-related apoptosis-inducing ligand with neutralizing antibodies markedly diminished the cytotoxicity of polyinosinic-polycytidylic acid-activated natural killer cells against activated hepatic stellate cells. CONCLUSIONS: Our findings suggest that natural killer cells kill activated hepatic stellate cells via retinoic acid early inducible 1/NKG2D-dependent and tumor necrosis factor-related apoptosis-inducing ligand-dependent mechanisms, thereby ameliorating liver fibrosis.     

Peritoneoscopic study on rat liver cell necrosis induced by carbon tetrachloride and allyl formate

The surface changes of acute centrolobular and perilobular necrosis were studied in rat liver using peritoneoscopy. Centrolobular necrosis as induced by carbon tetrachloride and perilobular necrosis by allyl formate. Vessels observed on the control rat liver surface were identified as terminal hepatic veins by irrigation of the hepatic vein with a barium solution. Polygonal whitish markings were observed on the liver surface 48 hours after carbon tetrachloride administration. Terminal hepatic veins were regularly distributed throughout the central portions of the whitish areas. Reddish spots about 0.1 mm in diameter were scattered about the terminal hepatic veins. The reddish spots and whitish areas corresponded to the histological findings of central liver cell necrosis and infiltration by Kupffer and white blood cells. Six hours after allyl formate administration, round reddish spots about 0.3 mm in diameter were regularly distributed on the liver surface among the terminal hepatic veins. These spots corresponded to the histological findings of liver cell necrosis around the portal area. In summary, acute centrolobular and perilobular liver cell necrosis were newly observed as reddish spots by peritoneoscopy.     

Death receptor-mediated liver injury

Apoptosis is a cardinal feature of liver injury. Death receptors are major mediators of the apoptotic pathway in the liver and have been implicated in the pathogenesis of diverse human diseases. More importantly, several studies have demonstrated a link between apoptosis and hepatic fibrosis; the latter is the most ominous consequence of chronic liver injury. In this article, we focus on the four death receptors: Fas, tumor necrosis factor receptor 1, tumor necrosis factor-related apoptosis-inducing ligand receptor 1, and tumor necrosis factor-related apoptosis-inducing ligand receptor 2. Although the death receptors have similar structures, they also possess distinct characteristics in their signaling pathways. Fas is abundantly expressed by all cells in the liver and plays a central role in variety of liver diseases. Tumor necrosis factor receptor 1 can induce both proapoptotic and prosurvival pathways. Tumor necrosis factor-related apoptosis-inducing ligand receptors likely trigger selective cell death in malignant and viral infected cells. Understanding the mechanism of liver injury caused by death receptors will enable therapeutic strategies to ameliorate human liver diseases.     

Hepatocyte transplantation activates hepatic stellate cells with beneficial modulation of cell engraftment in the rat

We investigated whether transplanted hepatocytes interact with hepatic stellate cells, as cell-cell interactions could modulate their engraftment in the liver. We transplanted Fischer 344 rat hepatocytes into syngeneic dipeptidyl peptidase IV-deficient rats. Activation of hepatic stellate cells was analyzed by changes in gene expression, including desmin and alpha-smooth muscle actin, matrix proteases and their inhibitors, growth factors, and other stellate cell-associated genes with histological methods or polymerase chain reaction. Furthermore, the potential role of hepatic ischemia, Kupffer cells, and cytokine release in hepatic stellate cell activation was investigated. Hepatocyte transplantation activated desmin-positive hepatic stellate cells, as well as Kupffer cells, including in proximity with transplanted cells. Inhibition of Kupffer cells by gadolinium chloride, blockade of tumor necrosis factor alpha (TNF-alpha) activity with etanercept or attenuation of liver ischemia with nitroglycerin did not decrease this hepatic stellate cell perturbation. After cell transplantation, soluble signals capable of activating hepatic stellate cells were rapidly induced, along with early upregulated expression of matrix metalloproteinases-2, -3, -9, -13, -14, and their inhibitors. Moreover, prior depletion of activated hepatic stellate cells with gliotoxin decreased transplanted cell engraftment. In conclusion, cell transplantation activated hepatic stellate cells, which, in turn, contributed to transplanted cell engraftment in the liver. Manipulation of hepatic stellate cells might provide new strategies to improve liver repopulation after enhanced transplanted cell engraftment.     

Caspase 9-dependent killing of hepatic stellate cells by activated Kupffer cells

BACKGROUND & AIMS: Hepatic stellate cells play an important role in liver fibrogenesis, and hepatic stellate cell death may be involved in the termination of this response. METHODS: Molecular mechanisms of hepatic stellate cell killing were studied in hepatic stellate cell/Kupffer cell cocultures. RESULTS: Lipopolysaccharide stimulation of hepatic stellate cell/Kupffer cell cocultures, but not of hepatic stellate cell monocultures, induced profound alterations of hepatic stellate cell morphology and hepatic stellate cell death. Kupffer cell-induced hepatic stellate cell killing required hepatic stellate cell/Kupffer cell contacts and was prevented by dexamethasone, prostaglandin E(2), tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) receptor 2 antagonists, and down-regulation of receptor-interacting protein, but not by antioxidants, tumor necrosis factor receptor, or CD95 antagonists. Hepatic stellate cell death was characterized by activation of caspases 3, 8, and 9, terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling negativity, lack of gross calcium overload, and TRAIL trafficking to the plasma membrane. Inhibition of caspase 9, but not of caspases 3, 8, or 10, prevented hepatic stellate cell death. Lipopolysaccharide induced a dexamethasone- and prostaglandin E(2)-sensitive expression of TRAIL in Kupffer cells. TRAIL receptors 1 and 2, FLIP (caspase 8-inhibitory protein), and receptor-interacting protein were up-regulated during hepatic stellate cell transformation; however, TRAIL addition did not induce hepatic stellate cell death. Hepatic stellate cell susceptibility toward Kupffer cell-induced death paralleled receptor-interacting protein and TRAIL-receptor expression levels. CONCLUSIONS: Activated Kupffer cell can effectively kill hepatic stellate cell by a caspase 9- and receptor-interacting protein-dependent mechanism, possibly involving TRAIL. The data may suggest a novel form of hepatic stellate cell death.     

Heatstroke: a rare cause of massive hepatic necrosis due to hypoxia

The authors report the case of a 76 year-old woman affected by heatstroke complicated by massive liver cell necrosis and fulminant liver failure, with a favorable outcome. Liver cell necrosis was localized in centro- and medio-lobular areas and was not associated with hepatic vein congestion. These histological features suggest that the main causal factor of liver cell necrosis was relative liver cell hypoxia secondary to increased oxygen tissular requirements induced by hyperthermia.     

Dietary factors alter hepatic innate immune system in mice with nonalcoholic fatty liver disease

Dietary factors promote obesity and obesity-related disorders, such as fatty liver disease. Natural killer T (NKT) cells are components of the innate immune system that regulate proinflammatory (Th-1) and anti-inflammatory (Th-2) immune responses. Previously, we noted that NKT cells are selectively reduced in the fatty livers of obese, leptin-deficient ob/ob mice and demonstrated that this promotes proinflammatory polarization of hepatic cytokine production, exacerbating lipopolysaccharide (LPS) liver injury in these animals. In the current study, we show that hepatic NKT cells are also depleted by diets that induce obesity and fatty livers in wild-type mice, promoting Th-1 polarization of hepatic cytokine production and sensitization to LPS liver injury despite persistent leptin. Adult male C57BL6 mice fed diets containing high amounts of either fat or sucrose, or combined high-fat, high-sucrose, develop increased hepatic NKT cell apoptosis and reduced liver NKT cells. The hepatic lymphocytes are more Th-1 polarized with increased intracellular interferon gamma and tumor necrosis factor alpha. Mice fed high-fat diets also exhibit more liver injury, reflected by 2-fold greater serum alanine aminotransferase (ALT) than control animals after receiving LPS. In conclusion, when otherwise normal mice are fed with high-fat or sucrose diet, they become obese, develop fatty livers, and acquire hepatic innate immune system abnormalities, including increased NKT cell apoptosis. The latter reduces liver NKT cell populations and promotes excessive hepatic production of Th-1 cytokines that promote hepatic inflammation. These diet-induced alterations in the hepatic innate immune system may contribute to obesity-related liver disease.     

Hypoxic hepatitis. Prospective, clinical and hemodynamic study of 45 cases

The authors report 45 episodes of centrilobular liver cell necrosis, called ischemic hepatitis, in 43 cardiac patients. In 75 percent of the episodes, centrilobular liver cell necrosis was preceded by a period of progressive deterioration of myocardiac function. In 100 percent of the episodes, liver cell necrosis occurred after an acute clinical event inducing a transient fall of cardiac output. Shock was observed in only 47 percent of the episodes. The biological hallmarks of this centrilobular liver cell necrosis were a massive increase in serum aminotransferase levels and in 85 percent of the episodes, a decrease in the prothrombine time below 50 percent of control level. The mortality rate, 15 days after admission, was 42 percent. Prognosis was mainly related to cardiac function. The hemodynamic comparison between the 45 episodes of centrilobular liver cell necrosis and 22 cases of cardiogenic shock without liver cell necrosis showed that, besides hepatic ischemia, passive venous congestion of the liver and arterial hypoxemia were also involved in the onset of liver cell necrosis in these cardiac patients. Among these 45 episodes of liver cell necrosis of cardiac origin, a unique case of hepatic necrosis secondary to major hypoxemia and passive venous congestion, despite an high cardiac output was observed and is reported in detail. Accordingly, the appellation "hypoxic hepatitis" seems to be more appropriate than "ischemic hepatitis".     

Attenuation of microcirculatory disturbance after liver ischemia by newly synthesized inflammatory cytokine suppressor,FR167653

BACKGROUND/AIMS: Inflammatory cytokines, such as interleukin-1 beta and tumor necrosis factor-alpha, which activate neutrophils, contribute to hepatic warm ischemia-reperfusion injury. However, the role of the cytokines in hepatic microcirculation immediately after reperfusion is still unclear. This study was carried out to investigate whether FR167653, a dual inhibitor of interleukin-1 beta and tumor necrosis factor-alpha, attenuates hepatic microcirculatory disturbance at the initial phase of reperfusion following liver ischemia. METHODOLOGY: Adult mongrel dogs were subjected to 90 minutes of liver ischemia by a Pringle's maneuver under portosystemic bypass. The animals were divided into two groups: a control group (n = 10), subjected to hepatic warm ischemia only, and a FR167653 administered group (n = 5), which received 1 mg/kg/h FR167653 for 4 hours since 30 minutes before the ischemia to 2 hours after the reperfusion continuously. Seven days animal survival, hepatic tissue blood flow, liver function test, hepatic venous blood concentration of endothelin-1 and plasminogen activator inhibitor-1, liver tissue biochemistry, and histopathology were analyzed. RESULTS: The treatment with FR167653 attenuated microcirculatory disturbance, lessened liver injury, enhanced adenine nucleotides resynthesis, and improved animal survival after liver ischemia. In addition, FR167653 significantly inhibited release of both endothelin-1 and plasminogen activator inhibitor-1 from the liver cells. CONCLUSIONS: These results suggest that the inflammatory cytokines induce microcirculatory disturbance in the initial phase of reperfusion following liver ischemia.     

Relationship between degree of portal hypertension and liver histologic lesions in patients with alcoholic cirrhosis

The relationship between the degree of portal hypertension and histologic liver lesions was studied in a group of 84 patients with histologically proven alcoholic cirrhosis. The degree of portal hypertension was evaluated by the gradient between wedged and free hepatic venous pressures. Five histologic lesions were quantified: liver cell necrosis, Mallory bodies, neutrophilic infiltrate, fibrosis, and fatty infiltration. The gradient between wedged and free hepatic venous pressures was significantly correlated with the degree of liver cell necrosis and the degree of neutrophilic infiltrate. The stepwise regression analysis showed that only liver cell necrosis has a significant and independent correlation for the degree of portal hypertension. The value for the gradient between wedged and free hepatic venous pressures was significantly higher in patients with (N=48) than in those without (N=36) acute alcoholic hepatitis (19.4±0.8 and 16.5±0.7 mmHg, respectively). Thus, histologic liver lesions observed in acute alcoholic hepatitis may play a role in the risk of complications of portal hypertension in patients with alcoholic cirrhosis.     

Effects of porto-caval end-to-side-anastomosis on the liver of rats (author's transl)

In young rats (n = 117), with a liver weight of 9.9 +/- 1.6 g ("small livers"), the decrease of total liver blood flow was followed by necrosis of liver parenchymal cells in the first two postoperative weeks. These alterations induced by haemodynamic disorders occured since the hepatic artery-although dilated to its maximum--was insufficient to compensate completely for the decrease of the portal hepatic blood flow. In old rats (n = 33) with a liver weight of nearly the double (19.5 +/- 1.9 g; "big livers") the extrem haemodynamic situation after PCA with a more less sufficient arterial hepatic blood supply was followed by complete liver necrosis including necrosis of the v. Kupffer cells. Rats having survived the PCA-operation showed liver cell necrosis in a more less degree as opposed to the early changes. These changes were not thought to be due to persisting haemodynamic disorders but due to endotoxaemia as found in 85% of the animals by means of the Limulus Gelation Test.     

Enhanced tumor necrosis factor and interleukin-1 in an experimental model of massive liver cell necrosis/fatal hepatitis in mice

Studies were conducted to investigate possible roles of tumor necrosis factor (TNF) and interleukin-1 (IL-1) in liver cell necrosis/fatal hepatitis in mice which were injected with Propionibacterium acnes (P.acnes) and subsequently 7 days later with a small dose of lipopolysaccharide-endotoxin (LPS). Higher serum levels of TNF were observed in the model, and enhanced production of both TNF and IL-1 was also found in hepatic as well as splenic adherent cells that were isolated from mice pretreated with P.acnes and were stimulated by LPS in vitro. When TNF substituted for LPS in the model, fatal hepatitis was also induced within 24 hrs, although the replacement of LPS by IL-1 resulted in no lethality. Moreover, when a combination of a near non-lethal doses of TNF and IL-1 substituted for LPS, 100% lethality was observed within 4 hrs. These results strongly suggest that both TNF and IL-1 are crucial soluble factors which are released by infiltrating macrophages in both liver and spleen, and are responsible for the development of liver cell necrosis/fatal hepatitis. In particular, TNF is one of the principal mediators of liver injury and IL-1 may potentiate the lethal effect of TNF in an LPS-related experimental model of massive liver cell necrosis.     

Methylprednisolone injection via the portal vein suppresses inflammation in acute liver failure induced in rats by lipopolysaccharide and d-galactosamine.

BACKGROUND: We have reported that hepatic arterial steroid injection is an effective therapy to rescue patients from fulminant or severe acute hepatic failure. We speculate that a high concentration of steroid suppresses inflammatory processes in the liver directly by restraining activated inflammatory cells, including macrophages. To analyse the detailed mechanism, steroid injection via the portal vein was performed in an experimental model of liver damage. METHODS: Rats subjected to lipopolysaccharide and d-galactosamine injection were treated with a methylprednisolone injection via the tail vein or the portal vein. The survival rate, serum levels of inflammatory cytokines and apoptotic cell counts in the liver were analysed. RESULTS: The survival rate was significantly improved by steroid injection, especially via the portal vein. Serum values of alanine aminotransferase, tumor necrosis factor-alpha and interferon-gamma were reduced in the treated groups, especially the group given portal venous injections. Apoptotic cell counts in the liver were significantly lower in the group injected with steroid via the portal vein. CONCLUSION: In the model rats, high concentrations of steroid in the liver acted on inflammatory cells and suppressed inflammatory cytokines and liver cell death. The mechanism is suggested to be the same for arterial steroid injection therapy in patients with acute hepatic failure.     

Regulation of mucosal addressin cell adhesion molecule 1 expression in human and mice by vascular adhesion protein 1 amine oxidase activity

Primary sclerosing cholangitis (PSC) and autoimmune hepatitis are hepatic complications associated with inflammatory bowel disease (IBD). The expression of mucosal addressin cell adhesion molecule 1 (MAdCAM-1) on mucosal endothelium is a prerequisite for the development of IBD, and it is also detected on the hepatic vessels of patients with liver diseases associated with IBD. This aberrant hepatic expression of MAdCAM-1 results in the recruitment of effector cells initially activated in the gut to the liver, in which they drive liver injury. However, the factors responsible for the aberrant hepatic expression of MAdCAM-1 are not known. In this study, we show that deamination of methylamine (MA) by vascular adhesion protein 1 (VAP-1) [a semicarbazide-sensitive amine oxidase (SSAO) expressed in the human liver] in the presence of tumor necrosis factor α induces the expression of functional MAdCAM-1 in hepatic endothelial cells and in intact human liver tissue ex vivo. This is associated with increased adhesion of lymphocytes from patients with PSC to hepatic vessels. Feeding mice MA, a constituent of food and cigarette smoke found in portal blood, led to VAP-1/SSAO-dependent MAdCAM-1 expression in mucosal vessels in vivo. CONCLUSION: Activation of VAP-1/SSAO enzymatic activity by MA, a constituent of food and cigarette smoke, induces the expression of MAdCAM-1 in hepatic vessels and results in the enhanced recruitment of mucosal effector lymphocytes to the liver. This could be an important mechanism underlying the hepatic complications of IBD.     

Kupffer cell depletion attenuates superoxide anion release into the hepatic sinusoids after lipopolysaccharide treatment

BACKGROUND AND AIM: The mechanisms involved in the beneficial effect of gadolinium chloride against endotoxin-induced liver damage were studied. METHODS: Superoxide anions released into the hepatic sinusoids were examined in a liver perfusion model using the cytochrome C method. RESULTS: Gadolinium chloride treatment fully depleted ED2-positive cells from the liver and significantly attenuated superoxide anion release after a lipopolysaccharide or tumor necrosis factor-alpha (TNF-alpha) challenge. Moreover, gadolinium chloride treatment resulted in a significant decline in endothelial cell damage in the hepatic sinusoids as assessed by the purine nucleoside phosphorylase/glutamic-pyruvic transaminase ratio in the liver perfusate. Although gadolinium chloride treatment did not affect the level of serum TNF-alpha, it significantly reduced that of interleukin (IL)-8 and neutrophil migration in the hepatic sinusoids after the lipopolysaccharide challenge. CONCLUSION: These data suggest that a reduction of the superoxide anion level in the hepatic sinusoids in acute endotoxemia and subsequent reduction of neutrophil migration into the liver may indicate that gadolinium chloride treatment suppresses the progression of liver damage in acute endotoxemia.     

Endotoxin clearance and its relation to hepatic and renal disturbances in rats with liver cirrhosis

BACKGROUND/AIMS: Little is known about endotoxin clearance and secretion of cytokines from macrophages in liver cirrhosis. The aims of this study were to investigate the relationship of endotoxin clearance and release of tumor necrosis factor alpha by various macrophages to hepatic and renal disturbances in liver cirrhosis. Methods: Male Sprague-Dawley rats were given 0.04% thioacetamide orally for 6 or 12 months. The organ distribution of infused [3H]-endotoxin (10 microg/kg b.w.) was analyzed at 30 min or at 24 h. Uptake of [3H]-endotoxin and secretion of tumor necrosis factor alpha by Kupffer cells, splenic macrophages and peripheral blood monocytes (1 x 10(4) cells/ml) from cirrhotic and control rats were determined. RESULTS: In cirrhotic rats, more endotoxin was left in the body and more endotoxin accumulated in the spleen and kidney, and thus was related to elevation of serum total bilirubin, aspartate aminotransferase, alanine aminotransferase, blood urea nitrogen, creatinine and tumor necrosis factor alpha. Endotoxin uptake and tumor necrosis factor alpha release by the Kupffer cells were decreased and those by the splenic macrophages and peripheral blood monocytes were increased in cirrhotic rats. CONCLUSIONS: In liver cirrhosis, impaired clearance of endotoxin together with increased secretion of tumor necrosis factor alpha by extrahepatic macrophages may play an important role in the progression of hepatic and renal disturbances.     

Oxidative stress in fulminant hepatic failure: comparison of two pig models with and without liver necrosis.

BACKGROUND/AIMS: No experimental study has clearly demonstrated how liver necrosis worsens the evolution of fulminant hepatic failure. Considering that several types of liver injury are associated with oxidative stress, we decided to measure plasma oxidative markers in two pig models of fulminant hepatic failure without and with liver necrosis. METHODOLOGY: Fulminant hepatic failure was produced in two groups of six pigs each by either total hepatectomy or complete hepatic devascularization. The following parameters were recorded before and during the course of hepatic failure: electrocerebral activity, plasma vitamin E, malondialdehyde and fluorescent protein-aldehyde adducts, total cholesterol, lactate-dehydrogenase, creatine phosphokinase, and ammonium. RESULTS: Despite comparable survival periods, hepatic necrosis was associated with earlier electrocerebral deterioration. Plasma concentration of malondialdehyde and fluorescent protein-aldehyde adducts rose and vitamin E content decreased in both groups. However, while in the group without liver necrosis the rates of cholesterol and vitamin E decay were identical, in the group with liver necrosis cholesterol concentration decreased less than vitamin E concentration, strongly indicating a true intravascular oxidation of vitamin E. Interestingly, in both models the rise of oxidative parameters preceded the development of cell injury. CONCLUSIONS: Oxidative stress, although present in both models, was significantly higher in the group with liver necrosis.     

短期二氧化硫吸入的肝脏毒性研究

目的　探讨短期二氧化硫吸入染毒对小鼠肝脏的损伤作用。方法　通过测定血清和肝脏中乳酸脱氢酶(LDH)活性、苏木精 伊红染色观察肝脏的病理组织学改变和DNA凝胶电泳分析来观察二氧化硫对肝脏的损伤作用。结果　 (1)肝脏中LDH活性随剂量增加而逐渐下降 ,并且有一定的剂量反应关系 (r =-0 .872 ,P <0 .0 1) ;(2 )苏木精 伊红染色发现二氧化硫染毒可引起肝实质细胞坏死 ,汇管区血管壁增厚并伴有淋巴细胞和单核细胞浸润 ,中央静脉以及肝窦充血 ,库普弗细胞增生 ;(3 )DNA凝胶电泳分析发现二氧化硫染毒可引起肝脏DNA随机降解 ,出现涂片状DNA涂片带。结论　短期二氧化硫吸入染毒对小鼠具有明显的肝脏损伤作用。     

肿瘤坏死因子-α诱导肝细胞凋亡在暴发性肝衰竭中的作用

目的 研究肿瘤坏死因子－α（ｔｕｍｏｒ ｎｅｃｒｏｓｉｓ ｆａｃｔｏｒ－α,ＴＮＦ－α）诱导肝细胞凋亡细胞凋亡在暴发性肝衰竭中的作用机制。方法 分别注射脂多糖（ｌｉｐｏｐ ｏｌｙｓａｃｃｈａｒｉｄｅ,ＬＰＳ）和ＴＮＦ－α于Ｄ－氨基半乳糖（Ｄ－ｇａｌａｃｔｏｓａｍｉｎｅ,ＧａｌＮ）致敏的ＢＡＬＢ／ｃ小鼠造成暴发性肝衰竭模型,用脱氧核糖核酸转移酶介导的缺口原位末端标记（ｉｎ ｓｉｔｅ ｅｎｄ ｌａｂｅ