Antioxidant Activity of Selected Stilbenoid Derivatives in a Cellular Model System

Abstract

The stilbenoids, a group of naturally occurring phenolic compounds, are found in a variety of plants, including some berries that are used as food or for medicinal purposes. They are known to be beneficial for human health as anti-inflammatory, chemopreventive, and antioxidative agents. We have investigated a group of 19 stilbenoid substances in vitro using a cellular model of THP-1 macrophage-like cells and pyocyanin-induced oxidative stress to evaluate their antioxidant or pro-oxidant properties. Then we have determined any effects that they might have on the expression of the enzymes catalase, glutathione peroxidase, and heme oxygenase-1, and their effects on the activation of Nrf2. The experimental results showed that these stilbenoids could affect the formation of reactive oxygen species in a cellular model, producing either an antioxidative or pro-oxidative effect, depending on the structure pinostilbene (2) worked as a pro-oxidant and also decreased expression of catalase in the cell culture. Piceatannol (4) had shown reactive oxygen species (ROS) scavenging activity, whereas isorhapontigenin (18) had a mild direct antioxidant effect and activated Nrf2-antioxidant response element (ARE) system and elevated expression of Nrf2 and catalase. Their effects shown on cells in vitro warrant their further study in vivo.

Keywords: Nrf2; antioxidant; macrophages; pro-oxidant; pyocyanin; stilbenoid.

Figure 1

The effects of stilbenoids 1–19 (at a concentration of 15 μM) on the lipid peroxidation of linoleic acid caused by 2,2´-azobis(2-amidinopropane)dihydrochloride (AAPH), and measured as the production of malondialdehyde (MDA) using the thiobarbituric acid reactive substances (TBARS) assay. Quercetin was used as a standard (15 μM) and AAPH alone served as the positive control (PC). The negative control (NC) contained linoleic acid alone, and thus, no lipid peroxidation occurred. The effects of the vehicle were subtracted from that of each stilbenoid. * = p < 0.05; ** = p < 0.01; *** = p < 0.001; and **** = p < 0.0001.

Figure 2

Antioxidant and pro-oxidant effects of stilbenoids 1–19 (at a concentration of 2 μM) on the formation of ROS after 1 h of incubation. In the THP-1-XBlue-CD14-MD2 cell model, the formation of ROS was triggered by adding 100 μM pyocyanin; quercetin was used as the standard (2 μM), pyocyanin alone served as the positive control (PC; 100 μM) and the vehicle alone was the negative control (NC). ** = p < 0.01; **** = p < 0.0001.

Figure 3

Antioxidant or pro-oxidant effects of stilbenoids 1–19 (at a concentration of 2 μM) on the formation of ROS after 24 h of incubation. In the THP-1-XBlue-CD14-MD2 cell model, the formation of ROS was triggered by adding 100 μM pyocyanin; quercetin was used as the standard (2 μM), pyocyanin alone served as the positive control (PC; 100 μM) and the vehicle alone was the negative control (NC). * = p < 0.05; ** = p < 0.01; and **** = p < 0.0001.

Figure 4

Antioxidant and pro-oxidant effects of stilbenoids 1–19 alone (at a concentration of 2 μM) on the formation of ROS after 2 h of incubation. In the THP-1-XBlue-CD14-MD2 cell model, the formation of ROS was triggered by stilbenoids alone; quercetin was used as the standard (2 μM), pyocyanin alone served as the positive control (PC; 100 μM), and the vehicle alone was the negative control (NC). * = p < 0.05; ** = p < 0.01; *** = p < 0.001; and **** = p < 0.0001.

Figure 5

Antioxidant and pro-oxidant effects of stilbenoids 1–19 alone (at a concentration of 2 μM) on the formation of ROS after 24 h of incubation. In the THP-1-XBlue-CD14-MD2 cell model, the formation of ROS was triggered by stilbenoids alone; quercetin was used as the standard (2 μM), pyocyanin alone served as the positive control (PC; 100 μM), and the vehicle alone was the negative control (NC).

Figure 6

Effects of compounds 2, 4, and 18 (at a concentration of 2 µM) on the levels of selected antioxidant enzymes CAT, GPx, HO-1, and SOD-1 and -2, and on the expression of Nrf2 after 6 h of incubation. The THP-1-XBlue-CD14-MD2 cell model was used with quercetin as the standard (2 μM), pyocyanin alone (100 µM) as the positive control (PC), and the vehicle alone as the negative control (NC). ** = p < 0.01; *** = p < 0.001; and **** = p < 0.0001.

Figure 7

The effects of selected stilbenoids 2, 4, and 18 (at a concentration of 2 μM) on the activation of Nrf2-ARE system. The HepG2 cell model was transiently transfected with the ARE luciferase reporter vector firefly luminescence and a constitutively expressing Renilla vector. The results are expressed as the ratio of firefly to Renilla luminescence. Quercetin was used as the standard (2 μM), DL-sulforaphane was used as a positive control at a concentration of 10µM (PC), and the vehicle alone served as the negative control (NC). * = p < 0.05; *** = p < 0.001.