Inhibition of Kupffer cell functions as an explanation for the hepatoprotective properties of silibinin

Abstract

The flavonoid silibinin, the main compound extracted perimental liver intoxication (carbon tetrachloride, from the milk thistle Silybum marianum, displays hepa-ethanol, acetaminophen, and phenylhydrazine). 1-6 The toprotective properties in acute and chronic liver injury. flavonoids from S. marianum were found to prevent To further elucidate the mechanisms by which it acts, liver lipid peroxidation, changes in the phospholipid we studied the effects of silibinin on different functions composition of the membranes, hepatic glutathione of isolated rat Kupffer cells, namely the formation of depletion, and improved functional markers of liver superoxide anion radical (O 0 2), nitric oxide (NO), tumor damage. Likewise, human clinical studies performed necrosis factor a (TNF-a), prostaglandin E 2 (PGE 2), and with patients with chronic alcoholic liver disease leukotriene B 4 (LTB 4). Production of O 0 2 and NO were showed beneficial effects of silymarin on the normaliza-inhibited in a dose-dependent manner, with an 50% in-tion of hepatic function markers. 7 From these results, hibitory concentration (IC 50) value around 80 mmol/L. No effect on TNF-a formation was detected. Opposite effects most investigators conclude that the crucial protective were found on the cyclooxygenase and 5-lipoxygenase mechanism is an inhibition of lipid peroxidation caused pathway of arachidonic acid metabolism. Whereas no by silymarin's free-radical-scavenging properties. A influence on PGE 2 formation was observed with silibinin membrane-stabilizing effect and an enhancement of concentrations up to 100 mmol/L, a strong inhibitory ef-protein synthesis have also been proposed. 8 However, fect on LTB 4 formation became evident. The IC 50-value the precise mechanism by which silymarin acts is still for inhibiting the formation of this eicosanoid was deter-unclear. mined to be 15 mmol/L silibinin. The strong inhibition of Despite the fact that silymarin would be expected to LTB 4 formation by silibinin was confirmed in experi-display its major effects on liver cells, its effect on Kupf-ments with phagocytic cells isolated from human liver. fer cells has not been studied so far. By liberating dif-Hence, while rather high concentrations of silibinin are necessary to diminish free radical formation by acti-ferent mediators, Kupffer cells not only play an im-vated Kupffer cells, significant inhibition of the 5-lipoxy-portant role in the body's defense system but may also genase pathway already occurs at silibinin concentra-contribute to liver damage. 9 Therefore, it was the aim tions which are achieved in vivo. Selective inhibition of of this study to determine the effect of silibinin on dif-leukotriene formation by Kupffer cells can at least partly ferent functions of Kupffer cells, namely the formation account for the hepatoprotective properties of silibinin. of superoxide anion radical (O 0 2), nitric oxide (NO), tu-(HEPATOLOGY 1996;23:749-754.) mor necrosis factor alpha (TNF-a), prostaglandin E 2 (PGE 2), and leukotriene B 4 (LTB 4). The naturally occurring flavonoid silibinin is the main compound extracted from the seeds of the milk MATERIALS AND METHODS thistle Silybum marianum (L), Gaertn. The whole ex-Animals. Male Wistar rats (250-300 g) were obtained from tract, silymarin, is composed of the three isomers sili-the Zentrales Tierlaboratorium (Universitä tsklinikum Es-binin, silidianin, and silichristin. A protective effect of sen). Animals were fed with a standard laboratory chow and silymarin has been described in different models of ex-water ad libitum. All animals received humane care in compliance with the institutional guidelines. Materials. Silibinin was kindly provided by Madaus AG (Cologne, Germany). Collagenase D, lucigenin (9,9-bis-N-Abbreviations: O 0 2 , superoxide anion radical; NO, nitric oxide; TNF-a, tumor methylacridinium nitrate), xanthine oxidase, superoxide dis-necrosis factor a; PGE2, prostaglandin E2; LTB4, leukotriene B4; PMA, phor-mutase, calcium ionophore A 23187, and RPMI 1640 medium bol-12-myristate-13-acetate; LPS, lipopolysaccharides. From the 1 Institut fü r Physiologische Chemie und 2 Abteilung fü r Allge-were purchased from Boehringer (Mannheim, Germany). Col-meine Chirurgie, Universitä tsklinikum, Hufelandstrasse 55, D-45122 Essen, lagenase V, phorbol-12-myristate-13-acetate (PMA), lipopoly-Germany. saccharides (LPS), and N-nitro-L-arginine were from Sigma