Cholestatic liver disease is associated with widespread derangements in the cardiovascular system, such as bradycardia, hypotension, QT prolongation and peripheral vasodilation; it is also associated with increased susceptibility to postoperative renal failure and haemorrhagic shock. A number of cellular signalling pathways have been shown to contribute to these abnormalities. In this article, we briefly review recent in vivo and in vitro findings in the field in an attempt to highlight the areas of agreement and areas of controversy. In this review, we will summarize pathogenic mechanisms underlying cardiac and vascular abnormalities in obstructive cholestasis. It seems that cardiovascular dysfunction is likely because of bile acids as one of the predominant factors. Other important factors which might play roles in these abnormalities are increased nitric oxide, endogenous opioids and endocannabinoids. These three factors interact with each other to exert vasodilation and impaired cardiovascular responses to sympathetic stimulation.
For many years, surgeons were puzzled and distressed by the frequent complications of hypotension and kidney failure after surgery on patients with obstructive jaundice [1-3]. Increased recognition and awareness of this clinical problem have led to extensive clinical and laboratory investigations, resulting in a better appreciation of the relationship between the liver, the kidney and the cardiovascular system. The first to make a systematic study of the bradycardia of jaundice was RÖhrig, in 1863, who showed that the bile salts are responsible for the bradycardia and low blood pressure of jaundice . In 1911, Clairmont and von Haberer first described the occurrence of renal failure developing after surgery for obstructive jaundice. Following these original observations, numerous clinical series have been reported in the literature, all of which point to a strong association between post-surgical renal failure and obstructive jaundice. A review of the different series indicates that the overall mortality rate for patients undergoing surgery for obstructive jaundice is 16% to 18%. Acute renal failure occurs in approximately 8% to 10% of patients requiring surgery for relief of obstructive jaundice and contributes to eventual mortality in 70% to 80% of those who develop it .
In this review, we attempt to summarize the pathogenic mechanisms underlying cardiac and vascular abnormalities in cholestasis. Human data in this field is relatively scarce; majority of the findings discussed in this article are based on animal studies. The Figure 1 summarizes the major pathologic mechanisms in the blood vessels and/or the heart.
Cardiac abnormalities in cholestasis
The association of obstructive jaundice with bradycardia has been known for over a century . It is also well known that cholestatic liver disease is associated with hypotension [6, 7], QT prolongation  and hyporesponsiveness of the heart to adrenergic stimulation [9, 10]. Literature reports also have described a cardiomyopathy caused by obstructive jaundice . Our laboratory performed several studies on the isolated hearts from cholestatic animals. These studies showed structural abnormalities and increased apoptosis in the hearts of cholestatic animals compared with control hearts. We also showed that the hearts taken from cholestatic animals had increased level of malondialdehyde, a byproduct of lipid peroxidation and decreased activities of catalase and glutathione peroxidase . Cruz et al., reported that obstructive cholestasis was associated with increased myocardial concentration of malondialdehyde. It was also associated with decreased levels of several antioxidant enzymes including reduced glutathione, catalase, superoxide dismutase and glutathione peroxydase. The authors demonstrated that melatonin administration significantly decreased malondialdehyde levels, and increased activity of all measured antioxidant enzymes in the cholestatic heart. . Kemp et al., showed that cellular respiration of the “jaundiced heart” is depressed. This depression was demonstrated by the reduced capacity of cardiac mitochondria to consume oxygen and synthesize ATP, suggesting a role for latent cardiac impairment in development of haemodynamic decompensation in cholestasis . The precise mechanism(s) of cholestasis-induced cardiac complications are not completely understood. The suggested mechanisms and potential causative molecules of cardiac abnormalities in obstructive cholestasis are discussed below.
Bile acids are amphiphilic steroids synthesized from free cholesterol in the hepatocytes and are essential for the solubilization of lipids in bile and the gastrointestinal tract, the induction and maintenance of bile flow, and the absorption of fat from the gastrointestinal tract .
Many in vitro and in vivo studies have subsequently established the negative chronotropic and inotropic effects of bile acids [15-17]. By exposing isolated atria of Wistar rats to cholic acid, Joubert demonstrated a dose-dependent negative chronotropic effect . This negative chronotropic effect of bile salts was also described by Bogin et al. . Some studies have suggested that the negative chronotropic effect induced by bile acids is mediated through vagal stimulation and can be antagonized by atropine [17, 19]. Dave et al.  observed that in a rat model, the effect of sodium tauroglycocholate itself was variable; it produced an increase in acetylcholine response in low doses and a blockade with higher doses. They also demonstrated that the depressor response to sodium tauroglycocholate was potentiated by physostigmine and blocked by atropine. Binah et al. have shown that, in addition to negative chronotropism, bile acids also exert a concentration-dependent negative inotropic effect on rat papillary muscle and isolated ventricular myocytes. This effect is attributed to a reduction in the duration of the action potential because of the suppression of the slow inward current of calcium . Clinical observations in patients with obstructive jaundice have also established the negative chronotropic effect of bile acids .
Bile acids in the concentration of 10−4M reduced β-adrenoceptor density in rat cardiac membranes. Based on this finding, Gazawi et al. suggested that bile acids, at plasma/serum concentrations, may have an etiologic role in cardiomyopathy of cholestatic liver disease .
Taken together, these studies suggest that bile acids are one of the main factors involved in cardiac complications in cholestasis.
The mRNA of endothelial type nitric oxide synthase has been shown to be expressed abundantly in atria as well as ventricular cardiomyocytes , supporting the role of nitric oxide in the regulation of cardiac function. Nitric oxide is also known to negatively regulate cardiac contractile function. It has been also shown that nitric oxide is involved in some types of cardiac dysfunction including ischaemic heart disease .
Many studies have suggested overproduction of nitric oxide in cholestasis as well as in animal models of cirrhosis [25-28]. Our laboratory performed several studies on the possible involvement of nitric oxide in cardiac dysfunction in cholestatic rats. We showed that heart rate of cholestatic animals was significantly less than that of sham-operated control rats in vivo and this bradycardia was corrected with daily administration of l-NNA, a non-selective nitric oxide synthase inhibitor . In another study from our laboratory by Gaskari et al., we showed that bradycardia of cholestatic rats was also corrected with daily administration of l-NAME, another non-selective nitric oxide synthase inhibitor. In this study, to differentiate which nitric oxide synthase isoenzyme is involved in cardiac abnormalities of cholestasis, both l-NAME and aminoguanidine, a selective inhibitor of inducible nitric oxide synthase, were used. We showed that the decreased chronotropic effect of epinephrine in cholestasis was corrected by daily injections of l-NAME, while it was not corrected by aminoguanidine. This demonstrated a possible involvement of endothelial-, but not inducible- nitric oxide synthase in these abnormalities . In another study, we demonstrated that cholestasis-induced decrease in chronotropic effect of epinephrine was corrected by daily administration of l-NNA in cholestatic rats which is in agreement with the previous study performed by Gaskari et al. . Surprisingly l-arginine, a precursor of nitric oxide, was as effective as l-NNA and increased the chronotropic effect of epinephrine in cholestatic rats but not in control animals. The opposite effect of chronic l-arginine administration in cholestasis and in control rats might be explained by an amelioration of cholestasis-induced liver damage by chronic l-arginine administration in bile duct-ligated rats . In another study, we demonstrated that the basal contractile force of papillary muscle was significantly decreased in bile duct-ligated rats compared with controls. The concentration–response curve for phenylephrine and isoproterenol demonstrated a reduced maximum effect in bile duct-ligated rats. Basal contractile abnormalities of bile duct-ligated rats were corrected by either acute or chronic administration of l-NAME . We also found that cholestatic animals are resistant to epinephrine-induced arrhythmias and that this process depends, at least in part, on long-term nitric oxide overproduction in cholestasis . In a recent study, we showed that treatment with l-NAME, but not d-NAME, improved both structural abnormalities and enhanced apoptosis of cholestatic hearts. Increased level of malondialdehyde, a byproduct of lipid peroxidation, and decreased activities of catalase and glutathione peroxidase in cholestatic hearts were not modified by d-NAME treatment. l-NAME treatment resulted in decreased levels of malondialdehyde and increased activities of catalase, glutathione peroxidase and superoxide dismutase in bile duct ligated mice. The content of nitric oxide was also higher in cholestatic cardiac tissue which was decreased by l-NAME treatment .
Despite the well-known involvement of the peripheral sympathetic abnormalities in the development of cardiovascular complications of cholestasis, the role of the central sympathetic system is still elusive. We demonstrated that both central and peripheral haemodynamic responses to clonidine, an α2 receptor agonist, are altered in cholestasis. We also provided evidence that nitric oxide contributes to the development of both central and peripheral abnormalities . The goal of that study was to evaluate the effects of central sympathetic tone reduction, through clonidine administration, on haemodynamic parameters of 7-day bile duct-ligated rats. Seven days after bile duct ligation, clonidine elicited an initial hypertension (the peripheral effect) followed by persistent hypotension and bradycardia (the central effects). Cholestatic rats demonstrated significant basal bradycardia and hypotension, which were corrected by chronic l-NAME treatment. While the peripheral effect of clonidine was blunted, the central effects were exaggerated in cholestatic rats. Acute l-NAME treatment accentuated the hypertensive phase in sham-operated and cholestatic rats. However, the difference between the two groups was preserved. This treatment attenuated the central effects in both sham-operated and cholestatic rats to the same level. Chronic l-NAME treatment resulted in exaggeration of the peripheral response in cholestatic and central responses in sham-operated rats, and abolished the difference between the groups. This study showed that both central and peripheral haemodynamic responses to clonidine are altered in cholestasis. It also provides evidence that nitric oxide contributes to the development of these abnormalities .
Collectively, these findings suggest an important role for nitric oxide in the pathophysiology of cardiac complications in cholestatic subjects. The exact isoenzymes responsible for these complications are yet to be determined. From one study by Gaskari et al.  we may suggest that inducible nitric oxide synthase does not play a role in the decreased chronotropic effect of epinephrine in cholestatic rats. Further investigation is needed to identify the role of different nitric oxide synthase isoenzymes in cardiac abnormalities in cholestasis.
Endogenous opioid peptides
Studies from the 1980s in patients with primary biliary cirrhosis represent the earliest evidence of a correlation between the plasma levels of endogenous opioids and liver disease . This concept led to the observation that opioid receptor antagonists ameliorate pruritus in patients with cholestasis [33, 34]. Although opioid antagonist therapy has since been used therapeutically for the treatment of pruritus in cholestatic liver disease, the physiological basis of its actions has not completely been understood. Endogenous opioid peptides have been reported to accumulate in plasma of cholestatic subjects [35, 36] and several reports suggest an important role for endogenous opioid accumulation in the pathophysiology of cholestasis [37-40].
Previous experiments have shown that the endogenous opioid peptides are produced and secreted by the cardiac myocytes as well as the sympathetic nerves and adrenal glands [41, 42]. It is well known that the endogenous opioid peptides are involved in the regulation of the cardiovascular system through both peripheral and central receptors. Besides modulating the autonomic nervous system , they have been demonstrated to have effects on the cardiac rhythm  and contractility . Abnormalities of the endogenous opioid system have been reported in several pathophysiological conditions in both human and animal models of cardiovascular diseases such as acute or chronic heart ischaemia and genetic hypertension. [46-48].
We have done several studies to explore the role of endogenous opioid peptides in the cardiac abnormalities of cholestasis. In these studies, we illustrated that bradycardia of cholestatic rats was corrected with daily administration of naltrexone, a non-selective opioid receptor antagonist. The decreased chronotropic effect of epinephrine in cholestasis was also corrected by daily injection of naltrexone . Basal cardiac muscle contractile abnormalities of bile duct-ligated rats were improved by either acute or chronic naltrexone treatment . We also demonstrated that naltrexone restored normal heart rate, blood pressure and susceptibility to arrhythmia in cholestatic animals, with no significant effect on QT interval. These findings also suggest that resistance against ischaemia/reperfusion-induced arrhythmia in short-term cholestasis might be because of increased availability of endogenous opioid peptides . These investigations provided some evidence for the involvement of increased opioidergic tone in cholestasis-induced cardiac impairment. In these experiments, the blockade of endogenous opioidergic system by naltrexone could correct the impaired cardiac function of cholestatic animals. Further studies are needed to identify which opioid receptor subtypes or endogenous opioid peptides play a role in development of these cardiac abnormalities.
Other possible mechanisms
Pereira et al., suggested the involvement of atrial natriuretic peptide (ANP) in cardiovascular abnormalities of cholestasis. ANP levels increase shortly after common bile duct ligation in the rabbit and this increase is paralleled by a simultaneous increase in the percentage of atrial cells staining for ANP. According to these data, the authors suggested that ANP may be involved in the pathogenesis of impaired water and sodium metabolism and renal dysfunction present in obstructive jaundice .
There is also a report about the role of TNF-α in the cardiac dysfunction of cholestatic bile duct-ligated mice. It has been reported that increased TNFα, acting via NFκB–inducible nitric oxide synthase and p38MAPK signaling pathways, plays an important role in the pathogenesis of cholestasis-induced cardiac dysfunction. Additionally, TNFα-accentuated endocannabinoid action and oxidative stress might also be involved in its negative inotropic effects in fibro-cholestatic hearts .
Hypotension and impaired vascular reactivity in cholestasis
In 1932, Meakin  noted that in a patient who had suffered from essential hypertension, blood pressure became normal when he developed “catarrhal jaundice” because of complete biliary obstruction. Blood pressure returned to elevated values once the jaundice resolved. In a retrospective review of 100 consecutive cases of obstructive jaundice, Meakin found that systolic, diastolic, and pulse pressure tended to be lower than those observed in the normal population. Because most of these patients also had bradycardia, it was presumed that the hypotension is related to the bradycardia.
Independent observations in 1956 by Zollinger and Williams established that jaundiced patients undergoing biliary surgery were more susceptible to a hypotensive crisis and renal failure after haemorrhage during surgery .
Both in vivo and in vitro studies in experimental models have established the vasodilatory properties of jaundice with or without concomitant liver disease.
Common bile duct-ligated dogs manifest systemic hypotension and diminished peripheral vascular resistance . Interestingly, hypertension induced in dogs by unilateral obstruction of the renal artery (a model mimicking renal artery stenosis) could be reversed after bile duct ligation of the same dog . The reduction in systemic blood pressure and vascular resistance in cholestatic dogs is associated with blunted response to vasoactive agents [6, 9]. The susceptibility of the common bile duct-ligated rats to haemorrhagic shock appears because of the pooling of blood in the splanchnic circulation. This pool is unavailable for defence of the circulation during haemorrhage in cholestatic rats. In baboons, in spite of normal systemic blood pressure at rest, there is blunted responsiveness of the skeletal muscle vasculature to norepinephrine . The vascular response to vasoactive agents varies among different vascular beds. In the bile duct ligated baboon model, the blunted vasopressor response of the peripheral vasculature is associated with enhanced pressor responsiveness of the renal and cerebral vessels . Utkan et al.  indicated that changes in the reactivity of arteries can be demonstrated at various times after the onset of bile duct ligation. The nature of these changes in vascular reactivity appears to be dependent on the duration of bile duct ligation. The exact mechanisms of these cholestasis-induced vascular changes are not completely understood. The suggested mechanisms and potential causative molecules of these vascular abnormalities are discussed below.
Bile acids are vasorelaxants . The evidence supporting this action was derived from two separate studies. In 1984 Bomzon et al.  demonstrated that the bile acids, deoxycholic acid, cholic acid and taurocholic acid could attenuate the contractile response to norepinephrine. Furthermore, they noted that deoxycholic acid was more potent than either cholic acid or taurocholic acid. Ten years later, Lee and his associates  reported that incremental doses of the bile acids, tauroursodeoxycholic acid, taurochenodeoxycholic acid and taurodeoxycholic acid caused dose-dependent vasorelaxation in the isolated perfused rat mesentery precontracted with the α1-adrenoceptor agonist, cirazoline. Bile acids are naturally occurring amphiphilic steroids derived from cholesterol whose relative solubility in aqueous and lipid media is commonly referred to as the ‘hydrophobic-hydrophilic balance’. This balance is determined by the state of ionization, the orientation, position and number of hydroxyl groups, and by the presence of the side chain ester. Conjugation and the presence of hydroxyl groups increase hydrophilicity . The vasorelaxant action of bile acids is linked to their lipophilicity. Lipophilic bile acids exhibit greater vasoactivity than the modest to negligible vasoactivity exhibited by hydrophilic bile acids .
One of the mechanisms whereby bile acids exert their vasorelaxant effects has been attributed to their ability to restrict calcium entry through voltage-dependent calcium channels . However, a direct action on vascular contractile receptor systems with consequent restriction of calcium entry through receptor-operated calcium channels has not yet been excluded. As bile acids attenuated the contractile vascular response to α1-adrenoceptor agonists [59, 60], it is possible that calcium entry through receptor-operated calcium channels was impeded because of bile acids behaving as a receptor antagonist. In 1994, Lee et al.  reported that bile acid-induced vasorelaxation was endothelium- and nitric oxide-independent. Ljubuncic et al.,  reported that lipophilic bile acids are non-selective vasorelaxants whose mechanism of action is a multifaceted process involving antagonism of contractile surface membrane receptors possibly affected by an increased extent of lipid peroxidation and/or membrane fluidity. They concluded that these actions were independent of the release of endothelial-derived relaxant factors or stimulation of a surface membrane bile acid binding site.
The role of endothelium in cholestasis-induced vasorelaxation was first reported by Utkan et al. . They demonstrated that the blunted response of isolated femoral arteries of dogs with obstructive jaundice to norepinephrine and serotonin disappeared when endothelium was removed. In endothelium-intact rings precontracted with phenylephrine, endothelium derived relaxing factor (EDRF) relaxation responses to acetylcholine were increased significantly as compared with controls. In 2001, we showed that there was a significant reduction in acetylcholine-induced relaxation of the aortic rings by the second day after the bile-duct ligation operation, compared with those of sham-operated rats. Further reduction occurred in 5- and 7-day bile-duct ligated groups, reaching a plateau by the seventh day. The relaxation response to sodium nitroprusside in the aortic rings of sham and 7-day bile-duct ligated rats did not differ, implying the intact smooth muscle component of the relaxation pathway. l-NAME, a nitric oxide synthase inhibitor, attenuated the acetylcholine-induced relaxation in both groups (unoperated and bile-duct ligated), while l-arginine prevented this inhibitory effect. Therefore, we concluded that reduced acetylcholine-induced relaxation in the cholestatic aortic rings may be because of the decreased acetylcholine-induced nitric oxide release from endothelium or increased nitric oxide inactivation . We also demonstrated that both phenylephrine-induced vasoconstriction and acetylcholine-induced vasorelaxation decreased in the mesenteric vascular bed of bile duct-ligated rats; both of these changes were restored by chronic treatment of these subjects with l-NAME . Another study about the role of nitric oxide in the vascular dysfunction in cholestasis was done by Atucha et al. . They indicated that in smooth muscle cells isolated from the abdominal aorta of bile duct-ligated rats, the entry of calcium from extracellular space and its mobilization from internal stores in response to purinergic agonists were defective. They also proposed that nitric oxide, of an inducible origin, is involved in this altered calcium regulation. More in-depth in vivo investigations, especially chronic manipulation of nitrergic system is needed, particularly to help develop potential therapeutic tools.
Endogenous opioids peptides
There is now increasing evidence for a role of opioidergic system in the pathophysiology of cholestasis [33, 40, 67]. With the aid of their widely distributed receptors, opioids modulate many functions of the cardiovascular system. Their effects on the cardiovascular system are mediated either centrally, such as a role in the pathogenesis of hypotension during blood loss , or peripherally, such as presynaptic inhibition of noradrenaline release in the portal vein , or a direct effect on contractility of rat aorta at high concentrations .
We showed that chronic treatment with naltrexone, a non-selective opioid receptor antagonist, partially restored the impaired acetylcholine-induced vasorelaxation and phenylephrine-induced vasoconstriction in bile duct-ligated rats. These findings suggest a role for opioidergic system in reduced response of mesenteric vascular bed of cholestatic rats to vasorelaxant or vasoconstrictor agents . In another study, we also showed that decreased responsiveness of mesenteric vascular bed of cholestatic rats to clonidine was enhanced by 7-day treatment with naltrexone . These data suggest that increased activity of opioidergic system might contribute to impaired vascular response in cholestasis.
Chronic blockade of opioid receptors is also associated with attenuation of hepatic damage as assessed by plasma transaminases . We also showed that naltrexone markedly attenuated the development of hepatic fibrosis as well as matrix metalloproteinase 2 activity in bile-duct ligated rats. Moreover, we reported that the development of biliary cirrhosis altered the redox state with a decreased hepatic GSH/GSSG ratio and increased concentrations of hepatic S-nitrosothiols, which were partially or completely normalized by treatment with naltrexone respectively [72, 73]. Therefore, either direct action on cardiovascular tissue or protection against liver damage may be responsible for recovery of cardiovascular parameters after naltrexone administration in bile-duct ligated rats. It seems that the effect of opioid receptor blockade on cardiovascular abnormalities in liver disease does not necessarily reflect a direct action of endogenous opioids on the heart or vascular bed. This is important since some studies failed to report any alterations in haemodynamic parameters following acute administration of naloxone in subjects with liver disease .
Endogenous cannabinoids represent a class of lipid ligands that share receptor binding sites with plant-derived cannabinoids such as ∆9 -tetrahydrocannabinol . In addition to having their well-known neurobehavioral effects, anandamide, the endogenous ligand of cannabinoid receptors and ∆9-tetrahydrocannabinol influence a number of other physiological functions, including cardiovascular variables . Anandamide has been shown to be a vasorelaxant, particularly in the resistance vasculature. This vasorelaxation has been reported to be both endothelium-independent and -dependent, depending on the vascular bed .
Batkai et al., for the first time showed that anandamide levels were increased in cirrhotic monocytes and overactivation of CB1 receptors within the mesenteric vasculature might contribute to the development of splanchnic arterial vasodilatation and portal hypertension . The blockade of CB1 receptors by the antagonist SR141716A increases mean arterial pressure [78, 79] and peripheral resistance  in rats with CCl4-induced cirrhosis. We also showed that AM251, a selective CB1 antagonist, increased blood pressure and systemic vascular resistance of bile duct ligated-cirrhotic rats .
In our first study about the role of endocannabinoids in cholestasis, we showed that anandamide-induced relaxation was significantly potentiated in mesenteric vascular beds of cholestatic rats. Chronic treatment of bile duct-ligated animals with l-NAME, a non selective nitric oxide synthase inhibitor, and aminoguanidine, a selective inducible nitric oxide synthase inhibitor, blocked this hyperresponsiveness. On the contrary, chronic treatment of these subjects with naltrexone further potentiated the vasorelaxant effect of higher doses of anandamide. Although acute l-NAME treatment of mesenteric beds completely blocked the anandamide-induced vasorelaxation in sham-operated rats, this vasorelaxation still was present in bile duct-ligated animals. Anandamide-induced vasorelaxation remained unaffected after acute naltrexone treatment of mesenteric beds in both bile duct-ligated and sham-operated rats. Our results indicated that  there was enhanced anandamide- induced vasorelaxation in cholestatic rats, probably because of altered function of cannabinoid receptors and  Nitric oxide overproduction might be involved in cholestasis-induced vascular hyperresponsiveness to anandamide (81).
As we discussed in this review among several mediators, bile acids, nitric oxide, opioid peptides and endocannabinoids seem to play more prominent roles in development of cardiovascular abnormalities in obstructive cholestasis. Recent studies about the modulatory actions of nitric oxide and opioid peptides in cholestasis mainly utilized non-specific opioid antagonists and nitric oxide synthase inhibitors. Further investigations are needed to further clarify the role of different opioid receptor subtypes and nitric oxide synthase isoforms.
Considering the role of endocannabinoid system in the mesenteric beds of cholestatic animals, it is plausible to hypothesize that this system might also play a role in cardiac abnormalities in cholestasis. However, the data supporting vascular effects of endocannabinoid system in cholestasis are also preliminary and further studies are needed.
Finally, another area that needs further investigation is the role of interaction between endocannabinoid and nitrergic systems in cardiovascular abnormalities of cholestasis. We demonstrated for the first time such an interaction in mesenteric beds of cholestatic rats, however further studies are needed to shed light on the nature of this interaction.
The authors are grateful for helpful editing and critical comments of Dr Seyed Ali Gaskari.