Authors
D. VAN DER GRAAFF (1), W. KWANTEN (2), J. DE MAN (3), B. DE WINTER (4), P. MICHIELSEN (2), S. FRANCQUE (2) / [1] Antwerp University Hospital, Edegem, Belgium, Gastro-enterology hepatology, [2] UZA, Universitair Ziekenhuis Antwerpen, Edegem, Belgium, Department of Gastroenterology and Hepatology, [3] Laboratory of Experimental Medicine and Pediatrics, University of Antwerp, Antwerp, Belgium, Department of Gastroenterology and Hepatology, [4] Laboratory of Experimental Medicine and Pediatrics, Division of Gastroenterology, University of Antwerp, Antwerp, Belgium, Department of Gastroenterology and Hepatology
Introduction
Non-alcoholic fatty liver disease (NAFLD) has become the most prevalent chronic liver disease of the Western world. Prior to development of inflammation or fibrosis, the intrahepatic vascular resistance (IHVR) is increased both in animals and in patients, impairing hepatic blood flow and potentially causing disease progression. Altered reactivity to or decreased production of nitric oxide (NO) are potential mechanisms, although previous data have shown conflicting results.
Aim
The present study seeks to elucidate the role of NO-mediated mechanisms on the IHVR in severe steatosis without inflammation or fibrosis.
Methods
The IHVR was studied by measuring the transhepatic pressure gradient (THPG) in an in situ ex vivo rat perfusion model, in which the liver is isolated, connected to a circuit with a pump and perfused by Krebs solution, with or without addition of drugs. The THPG was studied in Wistar rats (n=5-7/group) on a methionine-choline-deficient diet, inducing severe steatosis after 4 weeks, and compared to rats on a control diet. First, pre-constriction was induced by adding Methoxamine (alpha 1-adrenergic agonist, 10-5 M) to the perfusion fluid. Subsequently, after 10 minutes the effects of acetylcholine (ACh, stimulator of endothelial NO production, 10-6 - 3x10-4M) and sodium nitroprusside (SNP, NO donor, 10-5 - 3x10-3 M) in presence of Mx were studied in dose-response experiments, in which the dose was increased by 0.5 log M every 5 minutes at a constant flow of 30 mL/min.
Results
The basal THPG in steatotic livers was significantly increased compared to controls at all flows (10 – 50 mL/min), with respectively 5.4 ± 0.3 mmHg and 4.4 ± 0.2 mmHg at 30 mL/min [p<0.001]. Mx induced an increase of the THPG in both controls and steatosis, which was, as expected, more pronounced in steatosis. Ach did not alter THPG significantly in control rats nor in steatotic rats. Adding SNP to the perfusion fluid decreased the THPG in both groups. The decrease of the THPG appeared to be delayed in steatosis, but at higher doses of SNP (10-4 – 3x10-3 M) the magnitude of the decrease was equal, as shown after correction for the more pronounced increase in THPG by Mx in steatotic livers (controls + saline -3.05 ± 0.67 mmHg vs. controls + SNP -6.57 ± 1.59 mmHg [p<0.001]; steatosis + saline -2.95 ± 1.68 mmHg vs. steatosis + SNP -6.24 ± 1.20 mmHg [p<0.05]).
Conclusions
These experiments reconfirm an increased IHVR and hyperreactivity to Mx in steatosis compared to controls, as observed previously by our group. ACh did not significantly decrease the THPG in steatosis nor controls, which is conflicting with results of other research groups. The sensitivity to NO-mediated relaxation was unaltered, as shown by the equal reactivity to SNP in both controls and steatosis.
