Fast track paperCurcumin, an antioxidant and anti-inflammatory agent, induces heme oxygenase-1 and protects endothelial cells against oxidative stress
Introduction
Curcumin is a major active component of the food flavor turmeric. It is extracted from the powdered dry rhizome of Curcuma longa Linn (Zingiberacee), a perennial herb widely cultivated in tropical regions of Asia. It has been used for centuries in indigenous medicine for the treatment of a variety of inflammatory conditions and other diseases [1]. Several studies in recent years have shown that curcumin is a potent inhibitor of tumor initiation in vivo [2], [3] and possesses antiproliferative activities against tumor cells in vitro [4], [5]. Besides its anticarcinogenic properties, relatively low concentrations of curcumin exhibit remarkable anti-inflammatory and antioxidant effects [1], [6], [7], [8]. Although the exact mechanism(s) by which curcumin promotes these effects remains to be elucidated, the antioxidant properties of this yellow pigment appear to be an essential component underlying its pleiotropic biological activities. In fact, curcumin has been reported to inhibit lipid peroxidation and to effectively scavenge superoxide anion and hydroxyl radicals [7]. In addition to its inherent ability to attenuate the reactivity of oxygen free radical species, curcumin has been shown in vivo to enhance the activities of detoxifying enzymes such as glutathione-S-transferase [9]. Whether induction of distinct antioxidant genes in mammalian tissues contributes to the variety of pharmacological actions mediated by curcumin has yet to be examined.
Among the various genes encoding for proteins that possess antioxidant characteristics, heme oxygenase-1 (HO-1) has attracted particular interest as it is finely up-regulated by stress conditions and generates products that might have important biological activities [10]. Heme oxygenase is a widely distributed enzyme in mammalian tissues and its main function is associated with the degradation of heme to iron, carbon monoxide (CO), and biliverdin [11], the latter being converted to bilirubin by the cytosolic enzyme biliverdin reductase. Both biliverdin and bilirubin possess antioxidant properties [12] and HO-1-derived CO has been implicated in vasoregulation and signal transduction mechanisms [13], [14]. Physiological concentrations of bilirubin have been shown to protect endothelial cells against hydrogen peroxide-mediated injury and prevent protein oxidation in human blood plasma [15], [16]. We have recently reported that increased bilirubin production following HO-1 stimulation is associated with decreased peroxynitrite-mediated cytotoxicity [17] and reduction of postischemic myocardial dysfunction in an isolated rat heart preparation [18]. Otterbein and coworkers have also demonstrated that exposure of rats to low concentration of CO increases tolerance to hyperoxic lung injury [19]. Collectively, the HO-1 pathway appears to: play a key role in the preservation of tissue integrity against oxidative stress [17], [20], [21]; contribute to the modulation of inflammatory responses in vivo [22,23]; and act in synchrony with other crucial enzymatic systems in the maintenance of cellular homeostasis [21].
In the present study we report the effect of curcumin on HO-1 gene regulation, protein expression, and heme oxygenase activity in vascular endothelial cells. In addition, we have examined the interrelation between curcumin and HO-1 in protection against oxidant-mediated injury under normoxic and hypoxic conditions. The biological relevance of these findings will be discussed.
Section snippets
Materials
Tin protoporphyrin IX (SnPPIX), an inhibitor of heme oxygenase activity, was obtained from Porphyrin Products Inc. (Logan, UT, USA). Curcumin and all other chemicals were purchased from Sigma-Aldrich (Poole, UK) unless otherwise specified. Stock solutions of curcumin (5 mM) were prepared freshly on the day of the experiment by dissolving the compound in ethanol. A cDNA probe for rat HO-1 (kindly provided by Prof. Shibahara, Japan) was used to detect HO-1 mRNA expression [24].
Experimental protocol
Bovine aortic
Effect of curcumin on heme oxygenase activity and HO-1 protein expression in vascular endothelial cells
The effect of various concentrations of curcumin (0–30 μM) on endothelial heme oxygenase activity and HO-1 protein expression is shown in Fig. 1. Exposure of endothelial cells to curcumin (1–15 μM) for 18 h resulted in a concentration-dependent increase in heme oxygenase activity (Fig. 1A). The increase was significantly different from control (untreated cells, P < 0.05), with a maximal enzymatic activity (12-fold increase) at 15 μM curcumin. The antioxidant N-acetylcysteine did not
Discussion
In the present study we report that low concentrations of curcumin, the active component of the food flavor turmeric, up-regulate endothelial HO-1 gene, protein expression and heme oxygenase activity in a concentration- and time-dependent manner. Interestingly, this effect was greatly potentiated when endothelial cells were exposed to curcumin under conditions of reduced oxygen tension (hypoxia). Prolonged treatment with curcumin resulted in high resistance to oxidant-mediated cell injury and
Acknowledgements
This work was supported by grants from the National Heart Research Fund, Leeds (R.M.), British Heart Foundation (PG99005, R.M.) and National Kidney Research Fund (R30/1/99, R.M.).
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