Authors
D. VAN DER GRAAFF (1), W. KWANTEN (1), J. DE MAN (2), B. DE WINTER (2), P. MICHIELSEN (1), S. FRANCQUE (1) / [1] UZA, Universitair Ziekenhuis Antwerpen, Edegem, Belgium, Department of Gastroenterology and Hepatology, [2] Laboratory of Experimental Medicine and Pediatrics, 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. Similar to the mechanisms involved in portal hypertension during cirrhosis, hyperreactivity to vasoconstrictive agents is a potential mechanism of the increased IHVR in steatosis.
Aim
The present study seeks to elucidate the vasoconstrictive effects of alpha 1-adrenergic mediators and endothelin-1 (ET-1) 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=6-8/group) on a methionine-choline-deficient diet, inducing severe steatosis after 4 weeks, and compared to rats on a control diet. The effects of the alpha 1-adrenergic agonist methoxamine (Mx, 10-6 - 3x10-4 M), the alpha 1-adrenergic antagonist prazosin (PRZ, 10-7 - 10-4 M) and the vasoconstrictor ET-1 (10-12 - 3x10-10 M) were studied in dose-response experiments, increasing the doses by 0.5 log M every 5 minutes at a constant flow of 30 mL/min. Subsequently, flow-pressure curves were constructed to investigate the effects of 10-5 M Mx and 10-10 M ET-1 by increasing the flow by 5 mL/min every 5 minutes (10 – 50 mL/min).
Results
The basal THPG in steatotic livers was significantly increased compared to controls at all flows (10 – 50 mL/min); at 30 mL/min 5.4 ± 0.3 mmHg versus 4.4 ± 0.2 mmHg [p<0.001] respectively. Dose-response curves showed a significantly increased vascular sensitivity and responsiveness of steatotic livers to Mx (EC50=10-5.0 M, Tmax=9.8 mmHg) compared to controls (EC50=10-4.8 M, Tmax=8.2 mmHg [p<0.05]). In flow-pressure experiments, the THPG of steatotic livers showed a significantly increased reactivity to Mx at all flows (10 – 50 mL/min). At 30 mL/min, Mx (10-5 M) significantly increased the THPG in control rats from 4.4 ± 0.2 mmHg to 7.5 ± 0.4 mmHg (70.5% increase; p<0.001). In steatotic animals, Mx (10-5 M) increased the THPG after even more pronounced from 5.4 ± 0.3 mmHg to 10.5 ± 0.6 mmHg at 30 mL/min (94.4% increase; p<0.001). PRZ did not alter the THPG in both controls and steatosis. ET-1 induced a dose-dependent increase of the THPG in both controls and steatosis, with significantly increased sensitivity and responsiveness to ET-1 in steatosis (20.3 ± 1.3 mmHg at 3x10-10 M) compared to controls (14.9 ± 1.4 mmHg at 3x10-10 M, p<0.001). Flow-pressure experiments confirmed the hyperreactivity to ET-1 in steatotic livers with a more rapid and higher increase in THPG and the maximum THPG was reached at significantly lower flows compared to controls (controls 15.3 ± 1.1 mmHg at 30 mL/min, increase of 247.7% compared to Krebs; steatosis 23.8 ± 0.6 mmHg at 30 mL/min, increase of 340.7% compared to Krebs, p<0.001).
Conclusions
Steatotic rat livers demonstrate an increased reactivity to both Mx and ET-1 induced vasoconstriction, as compared to controls. The reactivity to the vasoconstrictor ET-1 was relatively larger compared to Mx. These results suggest that increased reactivity to vasoconstrictors, in particular to ET-1, can potentially be held responsible for the increased THPG as reported in NAFLD.
