Research ArticleEnoxaparin reduces hepatic vascular resistance and portal pressure in cirrhotic rats
Graphical abstract
Introduction
Increased resistance to portal blood flow, derived from architectural alterations of the liver parenchyma and a dynamic increment in the hepatic vascular tone, is the primary factor in the development of portal hypertension [1], [2]. Architectural distortion of the cirrhotic liver is partly due to excessive synthesis of extracellular matrix components performed by dys-regulated fibrogenic cells such as hepatic stellate cells (HSC) and portal myofibroblasts [3]. Several evidences suggest that liver fibrogenesis is positively influenced by inflammation and thrombosis [4]. Indeed, it has been demonstrated that different pro-coagulation factors, such as the serine protease thrombin, dysregulate HSC phenotype through the stimulation of their protease activated receptors (PARs) [5], [6], [7], [8], [9]. Moreover, PARs expression is highly upregulated in human livers undergoing acute and chronic injury [10], and their inhibition results in a significant amelioration in HSC phenotype and hepatic fibrosis progression in experimental models of mild liver damage [11], [12]. Generation of thrombin, from its precursor prothrombin, is efficiently performed by the activated coagulation molecule factor Xa. It is nowadays recognized that either thrombin or factor Xa are agonists of PARs and PAR signaling represents one of the main pathways of HSC activation and collagen deposition [7], [8], [13], [14], [15].
In addition, thrombin-derived clot formation may promote flow disturbances and occlusion of small-sized intrahepatic veins and sinusoids, representing another triggering factor of liver tissue remodelling [16]. This mechanism, named parenchymal extinction, includes progressive vascular obliteration due to thrombosis or inflammation leading to both apoptosis and atrophy of the liver [17].
Heparins, through their interaction with antithrombin, which induces a conformational change in antithrombin molecule, greatly facilitates the interactions between antithrombin and serine protease targets such as thrombin, and also factor Xa, IXa, XIa and XIIa [18]. Potential beneficial effects of the low molecular weight heparin (LMWH)-induced inhibition of the coagulation cascade have been evaluated in liver fibrosis [19], [20]. Previous translational studies, performed in experimental models of mild liver damage induced by CCl4 and common bile duct ligation, have proposed that LMWH may improve hepatic regeneration, by the inhibition of HSC proliferation and/or by inducing a marked reduction in the hepatic cytonecrosis index respectively [19], [21]. Whether the anti-fibrotic effects of the drug is directly due to its anticoagulative effect preventing thrombosis or to the inactivation of cell-mediated fibrogenic mechanisms remains unknown. In addition, it has not yet been evaluated whether these beneficial effects are also observed in more severe liver damage (ie. established cirrhosis). Indeed, a possible beneficial role of LMWH in the treatment of advanced liver disease was recently suggested by a randomised controlled trial investigating the effect of continuous use of the LMWH enoxaparin to prevent portal vein thrombosis (PVT) in patients with cirrhosis. In this study, enoxaparin reduced the incidence of PVT and, although the underlying mechanisms remain largely unknown, also reduced the incidence of liver decompensation and improved survival [22].
In addition, heparins may also have beneficial effects on the vasculature via endothelium-dependent mechanisms. Indeed, LMWH has been shown to upregulate nitric oxide (NO) levels by activation of its synthesis [23], [24], [25].
The present study aimed to characterize the effects, and underlying mechanisms, of acute, short, long-term and preventive administration of the LMWH enoxaparin on the hepatic and systemic hemodynamics and on liver fibrosis in two experimental models of cirrhosis (CCl4 and thioacetamide).
Section snippets
Induction of cirrhosis by CCl4 and LMWH administration
Cirrhosis was induced in male Wistar rats (50–75 g) with CCl4 inhalation three times a week. Phenobarbital (0.3 g/L) was added to drinking water as previously described [26]. When animals developed ascites, after approximately 12–15 weeks of CCl4 inhalation, phenobarbital and CCl4 administration was discontinued. Rats were then randomized to receive either enoxaparin (1.8 mg/kg body weight, subcutaneously; Clexane, Sanofi-Aventis), or its vehicle (saline 0.9%), according to three different
Acute enoxaparin treatment in CCl4-cirrhotic rats
Acute administration of enoxaparin did not affect any systemic or hepatic hemodynamic parameters evaluated (Table 1A). Moreover, acute enoxaparin did not modify hepatic cGMP levels or hepatic oxidative stress content (data not shown).
Short-term enoxaparin administration in CCl4-cirrhotic rats
One-week enoxaparin-treated rats had significantly lower portal pressure (PP) than vehicle-treated rats (Table 1B) without significant changes in portal blood flow (PBF). However, probably due to high variability, we were unable to identify significant changes in
Discussion
Previous studies examining the natural history of liver disease in cohorts of HCV infected patients suggested a role of the coagulation cascade in hepatic fibrogenesis [36], [37]. This hypothesis has gained much more attention after the demonstration that patients with cirrhosis exhibit more frequently a prothrombotic state with increased generation of thrombin than an hypocoagulative state [38], [39]. Indeed, either by activating HSC with the ensuing increase in collagen secretion, and/or by
Conflict of interest
The authors who have taken part in this study declared that they do not have anything to disclose regarding funding or conflict of interest with respect to this manuscript.
Financial support
This work was funded by the Ministerio de Economía y Competitividad, SAF (2013-44723-R) and Fondo Investigaciones Sanitarias (PI14/00029 and PI13/00341), and the European Union (Fondos FEDER, “una manera de hacer Europa”). FC and DMT have a Sheila Sherlock Fellowship from the European Association for the Study of the Liver. JG-S has a Ramón y Cajal contract from the Ministerio de Economía y Competitividad. CIBEREHD is funded by the Instituto de Salud Carlos III.
Authors’ contributions
FC and MV designed the study, performed experiments, analyzed data and wrote the manuscript; HG, DMT, EL and OG-I performed experiments and analyzed data; MA analyzed data and wrote the manuscript; JB conceived ideas, critically reviewed the manuscript and obtained funding; JG-S designed the study, conceived ideas, wrote the manuscript and obtained funding; JCG-P designed the study, conceived ideas, wrote the manuscript, obtained funding and directed the study.
Acknowledgments
Authors are indebted to Montse Monclús and Sergi Vila for their excellent technical assistance.
References (46)
- et al.
Complications of cirrhosis I. Portal hypertension
J Hepatol
(2000) - et al.
Functional aspects on the pathophysiology of portal hypertension in cirrhosis
J Hepatol
(2012) Thrombin is a potent inducer of connective tissue growth factor production via proteolytic activation of protease-activated receptor-1
J Biol Chem
(2000)- et al.
Parenchymal extinction: coagulation and hepatic fibrogenesis
Clin Liver Dis
(2009) - et al.
Regulation of hepatic stellate cell proliferation and collagen synthesis by proteinase-activated receptors
J Hepatol
(2002) - et al.
Factor Xa stimulates proinflammatory and profibrotic responses in fibroblasts via protease-activated receptor-2 activation
Am J Pathol
(2008) - et al.
Role of thrombosis in the pathogenesis of congestive hepatic fibrosis (cardiac cirrhosis)
Hepatology
(1995) - et al.
Hepatic and portal vein thrombosis in cirrhosis: possible role in development of parenchymal extinction and portal hypertension
Hepatology
(1995) - et al.
A study of unfractionated and low molecular weight heparins in a model of cholestatic liver injury in the rat
Pharmacol Res
(2005) - et al.
Low molecular weight heparin prevents hepatic fibrogenesis caused by carbon tetrachloride in the rat
J Hepatol
(2007)
Resveratrol improves intrahepatic endothelial dysfunction and reduces hepatic fibrosis and portal pressure in cirrhotic rats
J Hepatol
Simvastatin treatment improves liver sinusoidal endothelial dysfunction in CCl(4) cirrhotic rats
J Hepatol
A novel model of occlusive thrombus formation in mice
Lab Invest
Recombinant human manganese superoxide dismutase reduces liver fibrosis and portal pressure in CCl4-cirrhotic rats
J Hepatol
Natural history of liver fibrosis progression in patients with chronic hepatitis C. The OBSVIRC, METAVIR, CLINIVIR, and DOSVIRC groups
Lancet
An imbalance of pro- vs anti-coagulation factors in plasma from patients with cirrhosis
Gastroenterology
Bacterial lipopolysaccharide decreases thrombomodulin expression in the sinusoidal endothelial cells of rats – A possible mechanism of intrasinusoidal microthrombus formation and liver dysfunction
J Hepatol
Fibrin(ogen)-independent role of plasminogen activators in acetaminophen-induced liver injury
Am J Pathol
Efficacy and safety of anticoagulation on patients with cirrhosis and portal vein thrombosis
Clin Gastroenterol Hepatol
Biology of hepatic stellate cells and their possible relevance in the pathogenesis of portal hypertension in cirrhosis
Semin Liver Dis
Inflammatory pathways in liver homeostasis and liver injury
Clin Rev Allergy Immunol
Thrombin stimulates fibroblast procollagen production via proteolytic activation of protease-activated receptor 1
Biochem J
Cited by (91)
Recent advances in promising drugs for primary prevention of gastroesophageal variceal bleeding with cirrhotic portal hypertension
2024, Hepatobiliary and Pancreatic Diseases InternationalActivation of cGAS-STING signaling pathway promotes liver fibrosis and hepatic sinusoidal microthrombosis
2023, International ImmunopharmacologyCandesartan cilexetil ameliorates NOSTRIN-NO dependent portal hypertension in cirrhosis and ACLF
2023, European Journal of PharmacologyMechanobiology of portal hypertension
2023, JHEP Reports
- †
These authors contributed equally as joint first authors.