混合型生物人工肝治疗急性肝衰竭动物的实验研究

目的评估混合型生物人工肝对急性肝衰竭动物的治疗效果。方法混合型生物人工肝由血浆置换、血液滤过和猪肝细胞型生物人工肝构成。6只中国实验小型猪采用D-氨基半乳糖静脉注射建立急性肝衰竭动物模型,在给药48h后给予混合型生物人工肝治疗,先进行血浆置换联合血液滤过,再经过猪肝细胞型生物人工肝处理。观察比较实验动物治疗前后的临床表现及各相关指标的变化;并比较血浆置换联合血液滤过后猪肝细胞型生物人工肝处理后血液中相应指标的变化。结果本组实验动物在混合型生物人工肝治疗过程中生命体征平稳,未发生严重不良反应。与治疗前相比,治疗后实验动物血液中总胆红素、氨、丙氨酸转移酶、内毒素、肌酐明显减少(P0.01),纤维蛋白原、凝血酶原活动度和甲胎蛋白明显增加(P0.01);与血浆置换联合血液滤过比较,猪肝细胞型生物人工肝治疗后血液中纤维蛋白原、凝血酶原活动度有所增加(P0.05),血氨、总胆红素明显降低(P0.01)。结论应用混合型生物人工肝治疗急性肝衰竭安全、有效。混合型生物人工肝较非生物型人工肝(血浆置换+血液滤过)效果好。     

人工肝治疗前后实验性急性肝衰竭动物细胞因子变化的研究

目的探讨人工肝血浆置换联合血液滤过治疗前后对急性肝衰竭动物细胞因子变化的影响。方法28头中国实验小型猪随机分为对照组(n=13)和治疗组(n=15),在给予1.2g/kg的D-氨基半乳糖后48h,治疗组进行血浆置换联合血液滤过治疗,对照组未进行任何治疗。观察比较治疗前后两组动物细胞因子的变化。结果在经过人工肝治疗后,治疗组的生存时间(128.7±11.3)h较对照组(67.9±9.4)h明显延长(t=15.9,P<0.01)。肿瘤坏死因子-α(TNF-α)、白细胞介素-6(IL-6)、白细胞介素-8(IL-8)均出现暂时性下降,同一时间点与对照组相比差异有显著性(t=1.9～4.7,P<0.05)。结论1次人工肝治疗可以使炎性细胞因子暂时下降,缓解急性肝衰竭的进程,但尚不能完全阻断病情的进展。     

Reduced encephalopathy in pigs with ischemia-induced acute hepatic failure treated with a bioartificial liver containing alginate-entrapped hepatocytes

OBJECTIVE: To analyze the effects of an extracorporeal bioartificial liver containing alginate bead-entrapped hepatocytes on pigs with ischemia-induced acute hepatic failure. DESIGN: Prospective animal study. SETTING: University and INSERM laboratory. SUBJECTS: Fifteen Large White/Pietrin female pigs weighing 20-30 kg. INTERVENTIONS: Acute hepatic failure was induced by end-to-side portocaval shunt and ligature of the whole porta hepatitis. The bioartificial liver was in a thermostabilized column, containing a fluidized bed of alginate beads that embedded porcine hepatocytes, connected to a plasmapheresis system. Intracranial pressure; survival; ammonia, total bilirubin, aminotransferases, alkaline phosphatase, and lactate concentrations; and clotting factors were studied. The groups were pigs with acute hepatic failure (group 1, n = 4), pigs with acute hepatic failure treated with bioartificial liver containing empty beads (group 2, n = 4), or porcine hepatocytes (group 3, n = 5). MEASUREMENTS AND MAIN RESULTS: In group 1, survival of pigs averaged 10.9 +/- 1.0 hrs; intracranial pressure reached 32.3 +/- 3.8 mm Hg and was associated with coma and cerebral edema. After connection to the bioartificial liver, the survival of acute hepatic failure pigs was 12.1 +/- 1.4 hrs in group 2 and 14.8 +/- 2.5 hrs in group 3. In group 3, intracranial pressure and bilirubin concentrations were reduced significantly compared with both group 1 and group 2. Neither signs of encephalopathy nor cerebral edema was observed in any animal of group 3. In all animals, plasma ammonium, aminotransferases, alkaline phosphatase, and lactate concentrations increased and clotting factors decreased with no significant differences between the three groups. Autopsy revealed a total necrosis of the liver, which was histologically confirmed. CONCLUSIONS: The ischemia-induced model of acute hepatic failure in pigs is reproducible and provides measurable clinical and biological features. A bioartificial liver containing alginate bead-entrapped hepatocytes improves the signs of encephalopathy in pigs with ischemia-induced acute hepatic failure, suggesting that the bioartificial liver can clear out toxic compounds that are released from necrotic livers.     

Hepatocellular transplantation in acute hepatic failure and targeting genetic markers to hepatic cells.

Orthotopic liver transplantation (OLT) represents the only therapeutic option for many patients with end-stage liver disease as well as many inborn genetic errors of hepatic metabolism. Despite dramatic progress in methods for OLT, the utilization of this procedure is limited by its considerable morbidity and mortality, by a chronic shortage of organs for transplant, and by difficulty arranging funding for many patients. Many children with fulminant hepatic failure do not receive OLT because this technology is unavailable or unaffordable. Hepatocellular transplantation (HCT), in which isolated, heterologous hepatocytes from a donor liver would be infused into the diseased organ in order to provide essential hepatic functions, could provide a much needed therapeutic alternative to OLT in the treatment of some causes of hepatic insufficiency. Experiments in animals have demonstrated that several genetic deficiencies of hepatic metabolism as well as experimental induced hepatic failure in animals can be reversed by HCT. Despite this experience, HCT has never been attempted in human subjects. This protocol represents the first proposed clinical trial of HCT. We are proposing a clinical trial in which HCT would be attempted as a therapeutic intervention in children with acute hepatic failure who have no other medical or surgical options. This proposal is intended to establish surgical methods for HCT and to evaluate the feasibility of this procedure for treating hepatic disease in humans. It is our expectation that HCT may provide short-term support for patients awaiting organ availability, a "bridge to recovery" allowing patients with fulminant hepatic failure to recover, or a long-term repopulation of the patient's liver with healthy donor cells. One of the major limitations of many animal studies in HCT is that, since the donor hepatocytes are often indistinguishable from those of the host, it has often been difficult to demonstrate a clear correlation between engraftment and the therapeutic effect. In order to verify engraftment independent of the therapeutic response, we propose to "mark" the donor hepatocytes by transducing these cells with a recombinant retroviral vector (LNL6) carrying a marker gene (NEO-R, neomycin phosphoribosyl transferase). The presence of this marker will enhance the ability to identify transplanted cells in the host using assays for the NEO-R gene or transcribed NEO-R mRNA. The LNL6 vector has been approved for human use and has been used as a marker gene for transplanted cells in human subjects without any reported adverse effects. We would like to emphasize that this is a proposal with therapeutic intent.(ABSTRACT TRUNCATED AT 400 WORDS)     

Clinical management of acute hepatic failure

Acute hepatic failure is a rare clinical syndrome associated with high mortality. Hepatic failure leads to a well-recognised pattern of clinical signs and symptoms, sometimes with rapid deterioration and progression to multi-organ failure. Early recognition of this syndrome is essential for appropriate treatment; once identified, patients benefit from early interventional support and treatment in the intensive care unit. Aggressive management may allow stabilisation of patients before their transfer to specialist liver units. At present, orthotopic liver transplantation is the only treatment modality that provides significant improvement in outcome. This review examines the aetiology and clinical presentation of acute hepatic failure, providing guidelines regarding patient management. We present a critical appraisal of specific clinical areas, including the management of cardiovascular, cerebral, renal, coagulopathic and infective complications. Liver transplantation is discussed as well as emerging therapies including non-biological and hybrid liver support systems that may provide a "bridge to transplantation".     

N-acetylcysteine increases cerebral perfusion pressure in pigs with fulminant hepatic failure

OBJECTIVE: Intravenous administration of N-acetylcysteine beyond 15 hrs reduces mortality rates in patients suffering from paracetamol-induced fulminant hepatic failure, although the mechanism of the therapeutic benefit remains unclear. We hypothesized increased survival to be caused by improved hemodynamic performance. The main objective for the study was to explore the effect of N-acetylcysteine on hemodynamics, oxygen transport, and regional blood flow in pigs with fulminant hepatic failure. DESIGN: Prospective, randomized, controlled trial. SETTING: Surgical research laboratory in a university hospital. SUBJECTS: Female Norwegian Landrace pigs. INTERVENTIONS: Fulminant hepatic failure was induced by a total liver devascularization procedure. Five hours later, the pigs were allocated to N-acetylcysteine treatment (150 mg.kg-1 in 100 mL of 0.9% saline over 15 mins, followed by 50 mg.kg-1 in 500 mL of 0.9% saline over a period of 4 hrs) or placebo. MEASUREMENTS AND MAIN RESULTS: Mean arterial pressure stabilized in the N-acetylcysteine group and increased slightly during the last 2 hrs (pGT =.009). Thus, mean arterial pressure was significantly higher compared with placebo after 3 hrs (p =.01). Cerebral perfusion pressure was significantly higher during the last 2 hrs in the N-acetylcysteine group (pGT =.033). Common carotid artery flow also increased and was maintained at a higher level compared with placebo (pG =.027). Systemic vascular resistance index initially decreased but then gradually increased (pGT <.001). Cardiac index increased after 15 mins of N-acetylcysteine infusion, causing a significant interaction (pGT =.038), but did not differ after 3 hrs. No significant differences in hindleg and mesentery hemodynamics were found. A short-lived increase in oxygen delivery caused by a temporary increase in cardiac index was observed but without any corresponding increase in oxygen consumption. CONCLUSIONS: Intravenous N-acetylcysteine infusion increases cerebral perfusion pressure in pigs with fulminant hepatic failure. Earlier reported effects on oxygen transport and uptake could not be confirmed.     

Acetaminophen-induced acute pancreatitis: A case report and literature review

Acute pancreatitis is rarely associated with drugs. Acetaminophen overdose is a well-known cause of hepatic toxicity, but drug-induced pancreatitis is rarely reported, especially after mild overdose. A 32-year-old woman presented with nausea and vomiting for 12 h, but no abdominal pain following an overdose of eight Tylenol tablets containing acetaminophen (325 mg acetaminophen per tablet). Laboratory results on admission showed abnormal amylase and lipase levels but completely normal liver function. Magnetic resonance cholangiopancreatography revealed mild swelling of the pancreas without fluid collection around the pancreas. The patient complained of severe abdominal pain five days after admission when attempting to drink water and liquids. Eight days after admission, fluid around the pancreas was observed by computed tomography. The patient was subsequently diagnosed with acetaminophen-induced acute pancreatitis after exclusion of common causes. Routine treatment for pancreatitis and N-acetylcysteine were administered to prevent disease progression. The patient was discharged in good condition.     

Spotlight on animal models of acute traumatic coagulopathy: an update

Critically injured persons suffer trauma, hemorrhage, and high mortality. A subset of such patients develops early coagulation dysfunction characterized as acute traumatic coagulopathy (ATC), with a poor prognosis. The mechanisms contributing to ATC remain incompletely understood. Notwithstanding some successes in conducting clinical trials in early traumatic coagulopathy, conducting clinical research in ATC is ethically and logistically challenging. In vitro studies cannot capture the complex pathophysiological interplay between blood, vasculature, and organ systems relevant to ATC. Animal models are therefore vital for understanding ATC and to test interventions. Previous systematic reviews of animal models of ATC covered progress up to 2014. The current review aimed to extend that coverage to the end of 2021. A structured systematic search of MEDLINE/PubMed was carried out and identified 56 relevant publications. Unlike in previous reviews, where pig models predominated, rat and pig models contributed equally (19 studies each), and non-human primate models entered the field. Most studies now featured defined trauma (39 of 56), and hemorrhage controlled by pressure or volume (42 studies), with some documenting that both were necessary to induce ATC. Most studies documented coagulopathy using clotting or viscoelastometric assays and created an endogenous coagulopathy not dependent on iatrogenic dilution. As before, the diversity of species and experimental protocols may limit the translatability of the identified studies. Thus, while animal research has become more aligned to clinical realities since 2014, further efforts are required to unravel ATC mechanisms and enable the prediction and evaluation of optimal clinical interventions.     

Antiarrhythmic properties of MJ 9067 in acute animal models.

A novel benzanilide derivative, MJ 9067, has been shown to abolish experimental atrial and ventricular arrhythmiase effectively and promote the return of normal sinus rhythm in a variety of animal models. At intravenous dose levels ranging from 0.5 to 3.2 mg/kg, MJ 9067 successfully converted atrial fibrillation induced by either local application of aconitine or electrical stimulation, and ventricular tachycardia elicited by intravenous injection of ouabain or digoxin. The compound was equally effective in vagotomized or nonvagotomized dogs, and in intact cats and monkeys. The ventricular ectopic rate in conscious dogs 18 to 20 hours after two-stage ligation of a coronary artery was also markedly reduced by the drug at 2 mg/kg i.v. At antiarrhythmic dose levels, there were no undesirable effects noted on peripheral blood pressure, heart rate or the configuration of the electrocardiogram.     

Postdilution hemofiltration in the management of acute hepatic failure: a pilot study

We conducted a pilot study to assess the feasibility and efficacy of postdilution hemofiltration (PDHF) in the management of acute hepatic failure. From January 1984 through May 1986, we encountered seven patients with acute hepatic failure and entered these consecutive patients in the study; three had non-A, non-B hepatitis and one each had type B hepatitis, fulminant Wilson's disease (hepatolenticular degeneration), acute allograft (liver) failure, and acute fatty liver of pregnancy. Two of these seven patients were unable to undergo PDHF because of a precarious hemodynamic status. Of the five patients treated with PDHF, four had amelioration of hepatic encephalopathy; in two of these patients, a close temporal relationship was noted between the improvement and the procedure. Four patients had appreciable thrombocytopenia related to PDHF and bleeding complications. Our preliminary results support a possible role for PDHF as a temporary artificial liver support system for patients with acute hepatic failure.     

Naratriptan: biological profile in animal models relevant to migraine

The biological profile of naratriptan (N-methyl-3-(1-methyl-4-piperidinyl)-1H-indole-5-ethane-sulphona-mide), a novel 5HT_1B/1D receptor agonist, was investigated in a variety of experimental models of relevance to migraine. Naratriptan has high affinity for human recombinant 5HT_1B and 5HT_1D receptors (pKi = 8.70.03 and 8.30.1, respectively) and causes contractions of dog isolated basilar and middle cerebral artery (EC₅₀ values of 0.11 and 0.07 M, respectively). Naratriptan causes small contractions of human isolated coronary arteries (EC₅₀ value of 0.17 M; maximum contraction equivalent to 33% of 5HT maximum). In anaesthetized dogs, naratriptan causes selective vasoconstriction of the carotid arterial bed (CD₅₀ dose = 193 g kg⁻¹) and, in anaesthetized rats, naratriptan selectively inhibits neurogenic plasma protein extravasation in the dura (ID₅₀ = 4.1 g kg⁻¹). In a variety of antinociceptive tests, naratriptan has no effect even at high doses. In conscious rats and dogs, naratriptan has high oral bioavailability (71% and 95%, respectively). The data show that naratriptan is a selective agonist at 5HT_1B/1D receptors, with a pharmacological profile very similar to that of sumatriptan, albeit 2-3 fold more potent. These observations, coupled with high oral bioavailability in animals, suggest that naratriptan has the profile of an orally effective anti-migraine drug.     

Role of hemostatic factors in hepatic injury and disease: animal models de‐liver

Chronic liver damage is associated with unique changes in the hemostatic system. Patients with liver disease often show a precariously rebalanced hemostatic system, which is easily tipped towards bleeding or thrombotic complications by otherwise benign stimuli. In addition, some clinical studies have shown that hemostatic system components contribute to the progression of liver disease. There is a strong basic science foundation for clinical studies with this particular focus. Chronic and acute liver disease can be modeled in rodents and large animals with a variety of approaches, which span chronic exposure to toxic xenobiotics, diet‐induced obesity, and surgical intervention. These experimental approaches have now provided strong evidence that, in addition to perturbations in hemostasis caused by liver disease, elements of the hemostatic system have powerful effects on the progression of experimental liver toxicity and disease. In this review, we cover the basis of the animal models that are most often utilized to assess the impact of the hemostatic system on liver disease, and highlight the role that coagulation proteases and their targets play in experimental liver toxicity and disease, emphasizing key similarities and differences between models. The need to characterize hemostatic changes in existing animal models and to develop novel animal models recapitulating the coagulopathy of chronic liver disease is highlighted. Finally, we emphasize the continued need to translate knowledge derived from highly applicable animal models to improve our understanding of the reciprocal interaction between liver disease and the hemostatic system in patients.     

Pathogenic molecular mechanisms in an animal model of fulminant hepatic failure: rabbit hemorrhagic viral disease

In this study we sought to determine whether molecular mechanisms involved in the pathogenesis of fulminant hepatic failure are present in rabbits experimentally infected with rabbit hemorrhagic disease virus (RHDV). The activities of aspartate aminotransferase, alanine aminotransferase, and lactate dehydrogenase, as well as bilirubin concentration, were found to be significantly increased 36 hours after infection. Infected animals also demonstrated significant decreases in factor VII activity, in the Fischer index, and in the deterioration of prothrombin time. The concentration of reduced glutathione was significantly decreased 36 hours after infection, and we noted a marked increase in the ratio of oxidized to reduced glutathione. Infected animals showed progressive decreases in liver activity of the antioxidant enzyme superoxide dismutase. Expression of hepatocyte growth factor and c-met was found to be progressively reduced from 24 hours after infection, during which time we detected no modification in messenger RNA (mRNA) levels of transforming growth factor (TGF)-alpha. TFG-beta 1 was overexpressed 24 and 36 hours after infection, and 36 hours after infection we detected a significant increase in TNF-alpha mRNA levels. Experimental RHDV infection also induced marked activation of nuclear factor-kappaB and a significant increase in inducible nitric oxide synthase mRNA levels from 24 hours after infection. Data obtained from this animal model support its usefulness in the investigation of potential novel therapeutical modalities aimed at neutralizing reactive oxygen species and hepatocyte growth inhibitors or enhancing hepatocyte responsiveness to mitogens.     

Cardiac magnetic resonance imaging in small animal models of human heart failure

The aim of this study was to test the feasibility of cine magnetic resonance imaging (MRI) for assessment of the infarcted rat and mouse heart and to compare the results with established methods. These models have been proven to predict genesis and prevention of heart failure in patients. The value of cine MRI was tested in studies investigating interventions to change the course of the remodeling process. MRI was performed for determination of left ventricular (LV) volumes and mass, myocardial infarct (MI) size and cardiac output. LV wet weight was determined after MRI. Rats underwent conventional hemodynamic measurements for determination of cardiac output and LV volumes by electromagnetic flowmeter and pressure-volume curves. Infarct size was determined by histology. MRI-acquired MI-size (18.5+/-2%) was smaller than that found by histology (22.8+/-2.5%, p<0.05) with close correlation (r=0.97). There was agreement in LV mass between MRI and wet weight (r=0.97, p<0.05) and in the MRI- and flowmeter measurements of cardiac output (r=0.80, p<0.05). Volume by MRI differed from pressure-volume curves with good correlation (r=0.96, p<0.05). In a serial study of mice after coronary ligation, LV hypertrophy at 8 weeks was detected (Sham 105.1+/-7.9 mg, MI 144.4+/-11.7 mg, p<0.05). Left ventricles were enlarged in infarcted mice (end-diastolic volume, week 8: Sham 63.5+/-4 microl, MI 94.2 microl, p<0.05). In conclusion, cine MRI is a valuable diagnostic tool applicable to the rat and mouse model of MI. Being non-invasive and exact it offers new insights into the remodeling process after MI because serial measurements are possible. The technique was applied to study several interventions and proved its usefulness.