V. Arroyo

Complications of cirrhosis. II. Renal and circulatory dysfunction. Lights and shadows in an important clinical problem

The pathophysiology of circulatory and renal dysfunction in cirrhosis and the treatment of ascites and related conditions (hepatorenal syndrome and spontaneous bacterial peritonitis) have been research topics of major interest during the last two decades. However, many aspects of these problem remain unclear and will constitute major areas of investigation in the next millennium. The pathogenesis of sodium retention, the most prevalent renal function abnormality of cirrhosis, is only partially known. In approximately one third of patients with ascites, sodium retention occurs despite normal activity of the renin-aldosterone and sympathetic nervous systems and increased circulating plasma levels of natriuretic peptides and activity of the so-called natriuretic hormone. These patients present an impairment in circulatory function which, although less intense, is similar to that of patients with increased activity of the renin-aldosterone and sympathetic nervous systems, suggesting that antinatriuretic factors more sensitive to changes in circulatory function that these systems may be important in the pathogenesis of sodium retention in cirrhosis. The development of drugs that inhibit the tubular effect of antidiuretic hormone and increase renal water excretion without affecting urine solute excretion has opened a field of great interest for the management of water retention and dilutional hyponatremia in cirrhosis. Two families of drugs, the V2 vasopressin receptor antagonists and the kappa-opioid agonists, have been shown to improve free water clearance and correct dilutional hyponatremia in human and experimental cirrhosis with ascites. The first type of drugs blocks the tubular effect of antidiuretic hormone and the second inhibits antidiuretic hormone secretion by the neurohypophysis. On the other hand, two new treatments have also been proved to reverse hepatorenal syndrome in cirrhosis. The most interesting one is that based on the simultaneous administration of plasma volume expansion and vasoconstrictors. The second is transjugular intrahepatic porto-systemic shunt. The long-term administration (1-3 weeks) of analogs of vasopressin (ornipressin or terlipressin) or other vasoconstrictors together with plasma volume expansion with albumin is associated with a dramatic improvement in circulatory function and normalization of serum creatinine concentration in patients with severe hepatorenal syndrome. Of interest is the observation that in many of these patients, hepatorenal syndrome does not recur following discontinuation of the treatment, thus raising important questions about the mechanism by which hepatorenal syndrome follows a progressive course in most untreated cases. The pathogenesis of circulatory dysfunction in cirrhosis and the role of local mechanisms in the development of the splanchnic arteriolar vasodilation associated with portal hypertension will continue as important topics in clinical and basic research in Hepatology. Of special interest is the study of the mechanism by which circulatory function further deteriorates following complications such as severe bacterial infection or therapeutic interventions such as therapeutic paracentesis, and the adverse consequences of the impairment in circulatory function on renal and hepatic hemodynamics. Finally, although major advances have been made concerning the treatment and secondary prophylaxis of spontaneous bacterial peritonitis in cirrhosis, many aspects of the pathogenesis of this infection remain unclear. The mechanism of bacterial translocation and of the colonization of bacteria in the ascitic fluid are particularly important to design adequate measures for primary prophylaxis of this severe bacterial infection.

Increased anandamide induced relaxation in mesenteric arteries of cirrhotic rats: role of cannabinoid and vanilloid receptors

Background and aims: Anandamide is an endocannabinoid that evokes hypotension by interaction with peripheral cannabinoid CB1 receptors and with the perivascular transient receptor potential vanilloid type 1 protein (TRPV1). As anandamide has been implicated in the vasodilated state in advanced cirrhosis, the study investigated whether the mesenteric bed from cirrhotic rats has an altered and selective vasodilator response to anandamide. Methods: We assessed vascular sensitivity to anandamide, mRNA and protein expression of cannabinoid CB1 receptor and TRPV1 receptor, and the topographical distribution of cannabinoid CB1 receptors in resistance mesenteric arteries of cirrhotic and control rats. Results: Mesenteric vessels of cirrhotic animals displayed greater sensitivity to anandamide than control vessels. This vasodilator response was reverted by CB1 or TRPV1 receptor blockade, but not after endothelium denudation or nitric oxide inhibition. Anandamide had no effect on distal femoral arteries. CB1 and TRPV1 receptor protein was higher in cirrhotic than in control vessels. Neither CB1 mRNA nor protein was detected in femoral arteries. Immunochemistry showed that CB1 receptors were mainly in the adventitia and in the endothelial monolayer, with higher expression observed in vessels of cirrhotic rats than in controls. Conclusions: These results indicate that anandamide is a selective splanchnic vasodilator in cirrhosis which predominantly acts via interaction with two different types of receptors, CB1 and TRPV1 receptors, which are mainly located in perivascular sensory nerve terminals of the mesenteric resistance arteries of these animals.

Acetylsalicylic Acid Suppresses the Renal Hemodynamic Effect and Reduces the Diuretic Action of Furosemide in Cirrhosis With Ascites

To investigate if lysine acetylsalicylate influences the hemodynamic and diuretic responses to furosemide in cirrhosis, 21 nonazotemic patients with ascites were studied. In 8 patients (group 1), the renal plasma flow and glomerular filtration rate were serially measured before and during three 30-min periods after the i.v. administration of lysine acetylsalicylate (450 mg). In 7 patients (group 2), renal plasma flow, glomerular filtration rate, urine volume, and sodium excretion were measured before and during three 20-min periods after the i.v. administration of furosemide (40 mg). After a 45-min period in which urinary losses were restored, a similar study was performed before and after a second injection of furosemide (40 mg). Six patients (group 3) were studied with an identical protocol as group 2, except that the second injection of furosemide was preceded by the administration of lysine acetylsalicylate (450 mg). In 6 patients of group 1, lysine acetylsalicylate caused a marked and reversible reduction of renal plasma flow and glomerular filtration rate. In groups 2 and 3, the i.v. injections of furosemide alone produced a significant increase in renal plasma flow and glomerular filtration rate, and a marked diuresis and natriuresis. In patients of group 3, pretreatment with lysine acetylsalicylate suppressed the renal hemodynamic effect and markedly reduced the diuretic effect of the second injection of furosemide. Lysine acetylsalicylate did not cause the appearance of renal insufficiency in any of these patients. These results suggest that prostaglandins are involved in the renal response to furosemide in cirrhosis with ascites and that furosemide protects these patients from developing renal insufficiency after acute administration of nonsteroidal antiinflammatory agents.

Oral misoprostol or intravenous prostaglandin E2 do not improve renal function in patients with cirrhosis and ascites with hyponatremia or renal failure

Prostaglandins play an important role in the maintenance of renal hemodynamics and water excretion in cirrhosis. To investigate whether the administration of prostaglandins improves renal function in cirrhotic patients with ascites, 16 patients with functional renal failure and/or dilutional hyponatremia were given oral misoprostol, a prostaglandin E1 analogue (200 micrograms/6 h for 4 days; n = 9) or intravenous prostaglandin E2 (0.5 microgram/min for 1 h followed by 1 microgram/min for another hour; n = 7). The administration of misoprostol did not induce significant changes in the glomerular filtration rate (59 +/- 11 vs. 54 +/- 11 ml/min), sodium excretion (4.0 +/- 1.3 vs. 4.1 +/- 2.1 microEq/min), and free water clearance (2.4 +/- 0.8 vs. 2.1 +/- 0.9 ml/min), nor did it improve the natriuretic response to an intravenous bolus of 40 mg of furosemide (486 +/- 124 vs. 406 +/- 88 microEq/min). Similarly, an infusion of prostaglandin E2 did not induce significant changes in the glomerular filtration rate (baseline: 33 +/- 6; 0.5 microgram/min: 31 +/- 5; 1 microgram/min: 31 +/- 6 ml/min) and sodium excretion (5.7 +/- 2.7; 3.2 +/- 1.4; and 1.5 +/- 0.7 microEq/min, respectively), whereas free water clearance decreased significantly (1.1 +/- 0.7; 0.5 +/- 0.5; and -0.1 +/- 0.2 ml/min, respectively, p < 0.05). These results indicate that oral misoprostol or the intravenous infusion of prostaglandin E2 do not improve renal function in cirrhosis with ascites.

Temporal Relationship Between the Impairment of Free Water Excretion and Antidiuretic Hormone Hypersecretion in Rats With Experimental Cirrhosis

To investigate the temporal relationship between the impairment of water excretion, sodium retention, and antidiuretic hormone hypersecretion in cirrhosis, free water excretion (estimated by the minimum urinary osmolality) and urinary antidiuretic hormone excretion (which correlates with the plasma levels of this hormone) were measured weekly after an oral water load in 18 rats with carbon tetrachloride-induced cirrhosis and in 20 control animals. The onset of ascites (as an index of sodium retention) in cirrhotic rats was estimated by sequential paracentesis. Thirteen cirrhotic animals developed an impairment of water excretion 2-5 wk after the onset of ascites. The urinary excretion of antidiuretic hormone in these animals, which was normal before the impairment of water excretion, increased markedly within the week in which this abnormality was first detected and remained high thereafter. The remaining 5 cirrhotic rats did not experience an impairment of free water excretion in spite of developing ascites. The urinary excretion of antidiuretic hormone in these animals was similar to that of control rats during the entire study. In all urine samples obtained from cirrhotic rats, there was a highly significant direct linear correlation between the urinary excretion of antidiuretic hormone and the minimum urinary osmolality. Our results show the following: in rats with carbon tetrachloride-induced cirrhosis, sodium retention preceded the impairment of water excretion; and in these animals, the defect in water metabolism correlated chronologically and quantitatively with antidiuretic hormone hypersecretion. These findings are consistent with the concept that antidiuretic hormone is a major determinant of the impaired water metabolism in cirrhosis.

Neutrophil gelatinase-associated lipocalin is a biomarker of acute-on-chronic liver failure and prognosis in cirrhosis

BACKGROUND & AIMS Acute-on-chronic liver failure (ACLF) is a syndrome that occurs in cirrhosis characterized by organ failure(s) and high mortality rate. There are no biomarkers of ACLF. The LCN2 gene and its product, neutrophil gelatinase-associated lipocalin (NGAL), are upregulated in experimental models of liver injury and cultured hepatocytes as a result of injury by toxins or proinflammatory cytokines, particularly Interleukin-6. The aim of this study was to investigate whether NGAL could be a biomarker of ACLF and whether LCN2 gene may be upregulated in the liver in ACLF. METHODS We analyzed urine and plasma NGAL levels in 716 patients hospitalized for complications of cirrhosis, 148 with ACLF. LCN2 expression was assessed in liver biopsies from 29 additional patients with decompensated cirrhosis with and without ACLF. RESULTS Urine NGAL was markedly increased in ACLF vs. no ACLF patients (108(35-400) vs. 29(12-73)μg/g creatinine; p<0.001) and was an independent predictive factor of ACLF; the independent association persisted after adjustment for kidney function or exclusion of variables present in ACLF definition. Urine NGAL was also an independent predictive factor of 28day transplant-free mortality together with MELD score and leukocyte count (AUROC 0.88(0.83-0.92)). Urine NGAL improved significantly the accuracy of MELD in predicting prognosis. The LCN2 gene was markedly upregulated in the liver of patients with ACLF. Gene expression correlated directly with serum bilirubin and INR (r=0.79; p<0.001 and r=0.67; p<0.001), MELD (r=0.68; p<0.001) and Interleukin-6 (r=0.65; p<0.001). CONCLUSIONS NGAL is a biomarker of ACLF and prognosis and correlates with liver failure and systemic inflammation. There is remarkable overexpression of LCN2 gene in the liver in ACLF syndrome. LAY SUMMARY Urine NGAL is a biomarker of acute-on-chronic liver failure (ACLF). NGAL is a protein that may be expressed in several tissues in response to injury. The protein is filtered by the kidneys due to its small size and can be measured in the urine. Ariza, Graupera and colleagues found in a series of 716 patients with cirrhosis that urine NGAL was markedly increased in patients with ACLF and correlated with prognosis. Moreover, gene coding NGAL was markedly overexpressed in the liver tissue in ACLF.

Human serum albumin: Not just a plasma volume expander

Human serum albumin (HSA), used as an intravascular volume expander, was introduced during World War II as a substitute for blood or plasma. HSA is responsible for 80% of the colloid osmotic pressure of plasma; therefore, the intravenous administration of albumin is associated with a rapid increase in the circulating blood volume. However, albumin is more than a simple plasma volume expander. HSA has many other physiological functions, including the binding and transport of a wide variety of water-insoluble endogenous and exogenous substances, metals, and drugs.1,2 HSA is also quantitatively the most important circulating antioxidant.3 Some of these properties are the basis for new potential therapeutic indications for HSA, such as in ischemic stroke and Alzheimer’s disease.4,5 It is well known that the oxidation or binding of HSA to endogenous ligands produced or accumulated under pathological conditions such as sepsis, diabetes, chronic renal failure, and cancer6-8 is associated with significant structural and functional modifications of the molecule of albumin that markedly affect its biological activity.9 The article by Jalan et al.10 in this issue of HEPATOLOGY adds liver cirrhosis to this list of diseases with profound structural and functional modifications of HSA. They investigated 80 healthy subjects, 12 patients with decompensated cirrhosis, and 22 patients with acute-on-chronic liver failure as defined by the development of acute deterioration of liver function with hepatic encephalopathy and/or hepatorenal syndrome in a close temporal relationship with a precipitating event. Albumin function was investigated with the spin-trapping technique combined with electron paramagnetic resonance spectroscopy using 16-doxyl stearic acid as the spin label and ethanol as the polar reagent. The electron paramagnetic resonance spectrum generated from the stearic acid spin label bound to albumin allowed the assessment of structural and functional characteristics of the protein. The binding constants and the ability of albumin to bind fatty acids were estimated by measurements of the concentration of the fatty spin label bound to albumin as well as the amount of unbound fatty acid spin label. From these parameters, the transport (substrate sorption, binding, and delivery to target organs) and detoxification (ability of albumin to bind toxic substances produced via metabolism) efficiency of albumin could be estimated. Additionally, certain parameters of the spectrum indicated the mobility of the fatty acid spin label at its binding site on albumin, which depends mainly on the protein conformation at the albumin binding site for fatty acids. The ischemia-modified albumin, measured as the capacity of albumin to chelate cobalt, was also assessed to determine if it could be used as a simple test of albumin function. During the exposure of albumin to ischemic conditions such as acute coronary syndrome, the N-terminal of the albumin molecule is modified by oxidative free radicals and reactive oxygen species, and this results in the generation of a molecule with a low binding affinity to heavy metals. Finally, the plasma malondialdehyde and 8-isoprotane F2 levels were determined as markers of oxidative stress. The study revealed several important findings. First, all parameters estimating albumin function (functional capacity of the binding sites, transport efficiency, and detoxification efficiency) were severely compromised in patients with cirrhosis. Second, impairment in albumin function correlated closely with the degree of hepatic insufficiency; patients with acute-on-chronic liver failure were those with a more severe impairment of albumin function. Third, the plasma levels of ischemia-modified albumin with respect to the total serum albumin concentration were significantly increased in patients with cirrhosis, particularly in those with acute-on-chronic liver failure. Lastly, markers of oxidative stress were significantly increased in cirrhosis and also correlated with the degree of liver impairment. These findings are not surprising. Liver failure results in the accumulation of many water-insoluble endogenous substances that bind albumin sites and may alter albumin structure and function. On the other hand, acute-on-chronic liver failure is generally caused by bacterial infections or other conditions associated with an increased release of inflammatory mediators, which also cause the production of metabolic waste products that bind to albumin for transport and Abbreviations: HSA, human serum albumin; MARS, Molecular Absorbents Recirculating System. Address reprint requests to: Vincente Arroyo, Liver Unit, Hospital Clinic, Villarroel 170, Barcelona 08036, Spain. E-mail: varroyo@clinic.ub.es; fax: (34) 93 451 55 22. Copyright

Origins of cardiac dysfunction in cirrhosis

Is cirrhotic cardiomyopathy a specific cardiac dysfunction of cirrhotic patients or is it induced by the hyperdynamic circulation in these patients? The clinical course of patients with advanced liver disease is complicated by progressive impairment in circulatory function characterised by low arterial pressure, high cardiac output, and decreased systemic vascular resistance.1 Clinical and experimental investigations performed during the past two decades have shed light on the multiple mechanisms accounting for these disturbances. These studies have also established the pathogenic role of circulatory dysfunction in organ specific syndromes that commonly develop in cirrhotic patients, such as the hepatorenal and the hepatopulmonary syndromes.2,3 The heart is another functionally compromised organ in cirrhotic patients. However, whether the hyperdynamic circulation, by overloading the heart, induces cirrhotic cardiomyopathy or whether this is a specific cardiac dysfunction of cirrhotic patients has been subject of extensive discussions.4 Cardiac function abnormalities in cirrhosis are clinically not apparent. However, when cardiac function is explored, a reduction in right ventricular volume, probably secondary to reduced venous return, and left ventricular dysfunction, characterised by left ventricular preload and volume, are observed.5,6 Moreover, cardiac structural abnormalities, including hypertrophy of the myocardium and increased left ventricle thickness and hence diastolic dysfunction, have also been described.7 Cirrhotic cardiomyopathy is latent, probably because of the low peripheral vascular resistance presented by these patients, which reduces cardiac afterload. The existence of an abnormal ventricular behaviour can however be unveiled during exercise or following pharmacological stress. It has been demonstrated that left ventricular end diastolic pressure increases and stroke index and left ventricular ejection fraction decrease more …

Gene transduction of an active mutant of akt exerts cytoprotection and reduces graft injury after liver transplantation.

Akt is expected to be an effective target for the treatment of ischemia‐reperfusion injury (I/R) due to its anti‐apoptotic properties and its ability to activate the endothelial nitric oxide synthase (eNOS) enzyme. Therefore, this study was aimed to determine the efficacy of an active mutant of Akt (myr‐Akt) to decrease I/R injury in a model of orthotopic liver transplantation in pigs. In addition, we analyzed the contribution of nitric oxide in the Akt‐mediated effects by using an eNOS mutant (S1179DeNOS) that mimics the phosphorylation promoted by Akt in the eNOS sequence. Donors were treated with adenoviruses codifying for myr‐Akt, S1179DeNOS or β‐galactosidase 24 h before liver harvesting. Then, liver grafts were orthotopically transplanted into their corresponding recipients. Levels of transaminases and lactate dehydrogenase (LDH) increased in all recipients after 24 h of transplant. However, transaminases and LDH levels were significantly lower in the myr‐Akt group compared with vehicle. The percentage of apoptotic cells and the amount of activated‐caspase 3 protein were also markedly reduced in myr‐Akt‐treated grafts after 4 days of liver transplant compared with vehicle and S1179DeNOS groups. In conclusion, myr‐Akt gene therapy effectively exerts cytoprotection against hepatic I/R injury regardless of the Akt‐dependent eNOS activation.

Use of piretanide, a new loop diuretic, in cirrhosis with ascites: relationship between the diuretic response and the plasma aldosterone level.

Twenty patients with cirrhosis and ascites but no renal failure were given piretanide, a new loop diuretic, in order to investigate its efficacy and to relate the diuretic response with the pretreatment plasma aldosterone concentration. Eleven patients responded to piretanide 12 mg/day (equivalent in potency to 80 mg furosemide); there was no response in nine patients. Both groups were similar with regard to liver function, plasma urea, serum creatinine, plasma electrolytes, urine volume, and urine potassium concentration. The basal urinary sodium excretion was significantly higher in those patients who responded (23.6 +/- 5.7 mmol/day vs. 4.3 +/- 1.42 mmol/day; P < 0.01) (M +/- SE). Plasma renin activity (PRA) and plasma aldosterone concentration (PAC) were normal or only slightly increased in patients who responded to piretanide (PRA = 1.22 +/- 0.20 ng/ml/h; PAC = 12.25 +/- 2.20 ng/100 ml) and very high in patients who did not respond (PRA = 8.71 +/- 1.18 ng/ml/h; PAC = 84.6 +/- 16.2 ng/100 ml) (P < 0.001). Patients unresponsive to piretanide 12 mg/day also failed to respond when the dose was increased to 24 mg/day. However, the addition of spironolactone, 150 mg/day, to piretanide was followed in these patients by a marked increase in diuresis and natriuresis. These results strongly suggest that the pre-treatment level of aldosterone is an important factor influencing the response to loop diuretics in patients with non-azotaemic cirrhosis and ascites.