Basic Study
Copyright ©The Author(s) 2018. Published by Baishideng Publishing Group Inc. All rights reserved.
World J Gastroenterol. Oct 14, 2018; 24(38): 4356-4368
Published online Oct 14, 2018. doi: 10.3748/wjg.v24.i38.4356
Analysis of the nitric oxide-cyclic guanosine monophosphate pathway in experimental liver cirrhosis suggests phosphodiesterase-5 as potential target to treat portal hypertension
Denise Schaffner, Adhara Lazaro, Peter Deibert, Peter Hasselblatt, Patrick Stoll, Lisa Fauth, Manfred W Baumstark, Irmgard Merfort, Annette Schmitt-Graeff, Wolfgang Kreisel
Denise Schaffner, Adhara Lazaro, Peter Deibert, Manfred W Baumstark, Institute for Exercise-und Occupational Medicine, Faculty of Medicine, University of Freiburg, Freiburg 79106, Germany
Denise Schaffner, Irmgard Merfort, Department of Pharmaceutical Biology and Biotechnology, University of Freiburg, Freiburg 79104, Germany
Peter Hasselblatt, Wolfgang Kreisel, Department of Medicine II, Gastroenterology, Hepatology, Endocrinology, and Infectious Diseases, Faculty of Medicine, University of Freiburg, Freiburg 79106, Germany
Patrick Stoll, Anaesthesiological Practice, Freiburg 79104, Germany
Lisa Fauth, Annette Schmitt-Graeff, Institute of Clinical Pathology, Faculty of Medicine, University of Freiburg, Freiburg 79106, Germany
Author contributions: Kreisel W and Deibert P designed the concept of the study; Schaffner D performed the experiments and statistical calculations and provided additional ideas; Lazaro A assisted in the laboratory experiments and in writing the manuscript; Hasselblatt P supervised the PCR study and helped in the interpretation of results; Stoll P supervised Schaffner D and Kreisel W in the initial performance the operative procedure for hemodynamic measurements; Fauth L performed the histological assessment of the degree of liver damage; Baumstark MW supervised the statistical calculations; Merfort I supervised the advanced training of Schaffner D; Schmitt-Graeff A performed and interpreted the immunohistochemical data; all authors contributed in writing the manuscript.
Institutional animal care and use committee statement: The animal research protocol was approved by the local institutional animal care and use committee (Regierungspräsidium Freiburg, Germany, ref. No. G-13/89). Animal care was performed in accordance to the rules of the German animal protection law and the animal care guidelines of the European community (2010/63/EU).
Conflict-of-interest statement: There was no conflict of interest.
Data sharing statement: No additional data are available.
ARRIVE guidelines statement: The ARRIVE Guidelines have been adopted.
Open-Access: This article is an open-access article which was selected by an in-house editor and fully peer-reviewed by external reviewers. It is distributed in accordance with the Creative Commons Attribution Non Commercial (CC BY-NC 4.0) license, which permits others to distribute, remix, adapt, build upon this work non-commercially, and license their derivative works on different terms, provided the original work is properly cited and the use is non-commercial. See:
Correspondence to: Wolfgang Kreisel, MD, Emeritus Professor, Department of Medicine II, Gastroenterology, Hepatology, Endocrinology, and Infectious Diseases, Faculty of Medicine, University of Freiburg, Hugstetter Str. 55, Freiburg 79106, Germany.
Telephone: +49-761-27034010 Fax: +49-761-27074880
Received: July 16, 2018
Peer-review started: July 16, 2018
First decision: August 1, 2018
Revised: August 3, 2018
Accepted: August 24, 2018
Article in press: August 24, 2018
Published online: October 14, 2018
Processing time: 90 Days and 8.5 Hours
Research background

Cirrhotic portal hypertension is partly caused by sinusoidal constriction, wherein the nitric oxide-cyclic guanosine monophosphate (NO-cGMP) pathway plays a pivotal role. Whereas in systemic circulation there is excess NO formation, NO availability is reduced inside the liver (“NO-paradox”). Currently non-selective beta-blockers (NSBB) are mainly used as medical therapy for portal hypertension. β1 receptor blockade reduces portal flow by decreasing cardiac output while β2 blockade allows unopposed α1-adrenergic activity resulting in splanchnic vasoconstriction and decreased portal inflow. However, their therapeutic efficacy is limited by frequent side effects such as circulatory dysregulation, which often prevent sufficient dosing, particularly in decompensated cirrhosis.

Many approaches have been proposed to specifically target the reduced NO availability in diseased liver. Liver specific NO delivering drugs (e.g., NCX-1000) yielded disappointing results. Statins lead to enhanced activity of endothelial NO synthase. Although statins reduced portal hypertension in a clinical setting, the effects on clinical outcome were modest. As vasodilating cGMP is converted to inactive 5’-GMP by phosphodiesterase-5 (PDE-5), PDE-5 inhibitors could exert a beneficial effect in portal hypertension therapy. However, preclinical and clinical studies on PDE-5 inhibition yielded promising but variable results.

Research motivation

Novel medical therapies of cirrhotic portal hypertension are clearly needed, because NSBB have only limited efficacy. Targeting the altered NO-cGMP pathway in liver cirrhosis may be an intriguing approach to solve this problem. The imbalance between vasodilating (cGMP) and vasoconstricting (e.g., endothelins) compounds may be positively influenced by inhibiting the conversion of cGMP to inactive 5’-GMP by PDE-5 inhibitors. In this preclinical study, liver disease-induced alterations of biochemical components of the NO-cGMP pathway, in which cGMP represents the most potent vasodilating compound, were investigated. The current study aimed to demonstrate a potential rationale for therapy of cirrhotic portal hypertension using PDE-5 inhibitors.

Research objectives

The study investigated alterations of the NO-cGMP in an animal model of liver fibrosis/cirrhosis. The aim was to show whether or not the data yielded a rationale for therapy of cirrhotic portal hypertension using inhibitors of PDE-5.

Research methods

Gene expression of key components of the NO-cGMP pathway was investigated using PCR. Expression and localization of PDE-5 in healthy and cirrhotic livers was demonstrated by immunohistochemistry. The vasodilating compound cGMP was measured by ELISA. Systemic and portal hemodynamic parameters were monitored after application of NaCl, low-dose Sildenafil, and high-dose Sildenafil.

Research results

Key enzymes of the NO-cGMP system were overexpressed in cirrhosis: endothelial NO synthase (2.2-fold), soluble guanylate cyclase subunit α1 (1.7-fold) and soluble guanylate cyclase subunit β1 (3.0-fold), and PDE-5 (11-fold). Inducible NO synthase was expressed only in diseased livers. Immunohistochemistry showed a zonation of PDE-5 protein in healthy livers: Pronounced expression in zone 3 hepatocytes and slight expression in perisinusoidal cells (probably hepatic stellate cells). This zonation of PDE-5 was abrogated in cirrhotic livers. The enzyme was overexpressed in perisinusoidal cells. The vasodilating compound cGMP was reduced in liver cirrhosis. Inhibition of PDE-5 by Sildenafil could at least partly correct these disturbances: It led to a significant increase of cGMP and a reduction of portal venous pressure.

Research conclusions

The present study provided evidence that alterations of the NO-cGMP pathway may play an important role in the pathogenesis of cirrhotic portal hypertension. In healthy liver a zonation of PDE-5 was found, which may have a significant function in maintaining an appropriate level of the vasodilating compound cGMP in the sinusoids. This zonation was lost in cirrhosis and PDE-5 was overexpressed in perisinusoidal cells resulting in decreased cGMP serum levels. This disturbance was at least partly counteracted by inhibition of PDE-5. Sildenafil induced a normalization of cGMP levels and a reduction of portal venous pressure.

Research perspectives

The present study provides the rationale for the use of inhibitors of PDE-5 in the therapy of cirrhotic portal hypertension. Clinical studies are needed to test whether patients with portal hypertension in liver cirrhosis will benefit from application of PDE-5 inhibitors in terms of clinical endpoints such as prevention of (re)-bleeding from esophageal varices or prolonged survival.