Shock/sepsis/trauma/critical care
Silymarin, the Antioxidant Component of Silybum marianum, Prevents Sepsis-Induced Acute Lung and Brain Injury

https://doi.org/10.1016/j.jss.2007.03.072Get rights and content

Background

Sepsis is associated with enhanced generation of reactive oxygen species, which leads to multiple organ dysfunctions. Based on the potent antioxidant effects of silymarin, we investigated the putative protective role of silymarin against sepsis-induced oxidative damage in lung and brain tissues.

Materials and methods

Sepsis was induced by cecal ligation and perforation (CLP). Sham and CLP groups received either vehicle or silymarin (50 mg/kg, p.o.) or 150 mg/kg i.p. N-acetylcysteine (NAC) for 10 days prior and immediately after the operation. Six hours after the surgery, rats were decapitated and blood was collected for the measurement of proinflammatory cytokines (tumor necrosis factor-alpha, interleukin-1β [IL-1β], and IL-6) levels, lactate dehydrogenase activity, and total antioxidant capacity. Lung and brain samples were taken for the measurement of malondialdehyde and glutathione levels, myeloperoxidase activity, thromboplastic activity, and also for histological assessment. Formation of reactive oxygen species in tissue samples was monitored by using chemiluminescence technique with luminol and lusigenin probe.

Results

Sepsis increased serum TNF-α, IL-1β, IL-6 levels, and lactate dehydrogenase activity and decreased total antioxidant capacity. On the other hand, tissue glutathione levels were decreased while malondialdehyde levels and myeloperoxidase activity were increased in both the lung and the brain tissues due to CLP. Furthermore, luminol and lucigenin chemiluminescence were significantly increased in the CLP group, indicating the presence of the oxidative damage. Silymarine and NAC treatment reversed these biochemical parameters and preserved tissue morphology as evidenced by histological evaluation.

Conclusions

Silymarin, like NAC, reduced sepsis-induced remote organ injury, at least in part, through its ability to balance oxidant–antioxidant status, to inhibit neutrophil infiltration, and to regulate the release of inflammatory mediators.

Introduction

Sepsis is a generalized inflammatory response, which involves organ systems remote from the locus of the initial infectious insult [1]. Recent studies have shown that sepsis is associated with enhanced generation of reactive oxygen metabolites, which lead to multiple organ dysfunctions. Activation of macrophages and cytokines by endotoxin and the subsequent formation of reactive oxygen and nitrogen species are of central pathogenic importance in various inflammatory diseases including sepsis. However, whether different tissues behave the same in the pathological changes still remains to be evaluated [2].

The release of endotoxin (lipopolysaccharide, LPS) from bacteria is generally believed to be the initial event in the development of sepsis. LPS activates inflammatory cells of the myeloid lineage that subsequently amplify the inflammatory response by releasing various cytokines, such as tumor necrosis factor-α (TNF-α) and interleukin-1β (IL-1β). This systemic inflammatory cascade results in polymorphonuclear leukocytes (PMNs) sequestration in the various systemic organs, e.g., lungs, heart. Subsequent PMN extravasation can lead to vascular dysfunction as well as parenchymal cell dysfunction [1].

While antioxidants could be used to counteract the toxicity of reactive oxygen metabolites, free radical ablation for the treatment of sepsis was proposed to be useful in the clinical setting of sepsis-induced multiple organ failure [3].

Flavonoids are naturally occurring substances that possess various pharmacological actions and therapeutic applications. Some, due to their phenolic structures, have antioxidant effect and inhibit free-radical-mediated processes [4]. The extracts of the flowers and leaves of Silybum marianum (St. Mary’s thistle, milk thistle) have been used for centuries to treat liver, spleen, and gallbladder disorders [5]. In the 1960s the biologically active principles of the seed and fruit extracts were isolated, and the chemical structures were elucidated. The isolation led first to a mixture that was named silymarin, and it was with this flavonolignan mixture that most of the clinical studies were carried out. The main constituents are silibinin, isosilibinin, silicristin, and silidianin [6].

One of the important issues about silymarin is that it may be accepted as a safe herbal product, since no health hazards or side effects are known in conjunction with the proper administration of designed therapeutic dosages [4]. Recently oxidized derivatives of silybin (the major component forming 70–80% of silymarin) and their antiradical and antioxidant activity was studied by Gazak et al. [7]. Furthermore, its antioxidant, anti-inflammatory, and anticarcinogenic properties were demonstrated in the studies conducted with silymarin against oxidative stress, inflammatory responses, and benzoil peroxide-induced tumor promotion in mice [8, 9].

N-acetylcysteine (NAC) is a small molecule containing a thiol group, which has antioxidant properties, and is freely filterable with ready access to intracellular compartments. The diversity of pharmacological applications of NAC is due mainly to the chemical properties of the cysteinyl thiol group of its molecule, since the ability of reduced thiol groups to scavenge oxygen free radicals is well established [3]. Because of these properties, NAC is widely used in clinical practice as an antioxidant [10].

Based on the potent antioxidant and anti-inflammatory effects of silymarin, we investigated the putative protective role of oral silymarin treatment against sepsis-induced oxidative damage in the lung and brain in comparison with the reference antioxidant NAC.

Section snippets

Animals and Protocol for the Induction of Sepsis

Wistar albino rats of both sexes, weighing 250 to 300 g, were fasted for 12 h, but allowed free access to water before the experiments. The animals were kept in individual wire-bottom cages, in a room with a constant temperature of 22 ± 2°C with 12-h light and dark cycles, and fed standard rat chow. The study was approved by the Marmara University School of Medicine Animal Care and Use Committee (Istanbul, Turkey).

The rats were divided into the following six groups of eight rats (four males and

Survival Rates

Eight animals from each group were decapitated at the 6th hour and eight animals from each group were followed up for survival. The survival rates at 72 h post-CLP were as follows: 75% in both silymarin and NAC groups and 50% in the vehicle-treated sepsis group. Long-term survival (at week 1 post-CLP) was 62.5% in silymarin and 75% in the NAC-treated groups, while only 25% of rats survived in sepsis group.

Biochemical Analysis in the Serum

As shown in Table 1, TNF-α levels of the vehicle-treated CLP group were significantly

Discussion

In this study, we investigated the early time point of sepsis and observed a significant oxidative damage in lung and brain tissues, as evidenced by increased lipid peroxidation with a concomitant decrease in endogenous antioxidant glutathione level. Moreover, oxidative injury of the tissues studies was accompanied by neutrophil infiltration, as evidenced by high-tissue MPO levels. The oxidative damage and tissue neutrophil accumulation due to sepsis was totally abolished by NAC or silymarin

Conclusion

In conclusion, this study demonstrates that silymarin, a phenolic compound, reduces sepsis-induced oxidative organ injury and that the protective effect of silymarin can be attributed, at least in part, to its ability to balance oxidant–antioxidant status, to inhibit neutrophil infiltration, and to regulate the inflammatory mediators, suggesting a future role in the treatment of multiorgan failure due to sepsis.

Acknowledgment

The authors are grateful to MIKROGEN Pharmaceuticals, for supplying the silymarin.

References (37)

  • J.S. Kang et al.

    Protection against lipopolysaccharide-induced sepsis and inhibition of interleukin-1beta and prostaglandin E2 synthesis by silymarin

    Biochem Pharmacol

    (2004)
  • T. Ito et al.

    High-mobility group box 1 protein promotes development of microvascular thrombosis in rats

    J Thromb Haemost

    (2007)
  • C. Lupu et al.

    Tissue factor dependent coagulation is preferentially up-regulated within arterial branching areas in a baboon model of Escherichia coli sepsis

    Am J Pathol

    (2005)
  • R.A. Balk

    Severe sepsis and septic shockDefinitions, epidemiology, and clinical manifestations

    Crit Care Clin

    (2000)
  • Neviere RR, Ceepinskas G, Madorin WS, et al. LPS pretreatment ameliorates peritonitis-induced myocardial inflammation...
  • A.R. Heller et al.

    Acetylcysteine reduces respiratory burst but augments neutrophil phagocytosis in intensive care unit patients

    Crit Care Med

    (2001)
  • Milk Thistle (Silybum marianum) in PDR for Herbal Medicines

  • F. Rainone

    Milk thistle

    Am Family Physician

    (2005)
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