Elsevier

Toxicology in Vitro

Volume 25, Issue 4, June 2011, Pages 839-847
Toxicology in Vitro

Double antioxidant activities of rosiglitazone against high glucose-induced oxidative stress in hepatocyte

https://doi.org/10.1016/j.tiv.2011.02.004Get rights and content

Abstract

Chronic hyperglycemia is the hallmark of diabetes and its complication. High glucose-induced excessive reactive oxygen species (ROS) production has been considered to play an important role in the development of diabetes. However, the influence of high glucose on the liver remains to be clarified. Rosiglitazone (RSG) is a member of thiazolidinediones (TDZs) family, which is the ligand of the of nuclear transcription factor peroxisome proliferator-activated receptor-γ (PPARγ), being used clinically for the treatment of type 2 diabetic patients through their insulin-sensitizing effect. In the present study, we investigated the cytotoxicity of high glucose in QZG hepatocytes and evaluated the protective effect of RSG. The results showed that high glucose significantly reduced cell viability through generation of ROS via activation of PKC, which was inhibited by RSG. On the one hand, RSG notably inhibited the activation of PKC induced by high glucose independent of PPARγ, leading to the decrease of ROS generation. On the other hand, RSG notably increased the expression of key antioxidant transcription factor Nrf2 and antioxidant enzyme HO-1 in a PPARγ-dependent manner, leading to the elimination of excessive ROS. In addition, RSG also inhibited the decrease of COX-2 expression induced by high glucose through activating PPARγ. Furthermore, the activation of Akt and MAPKs was involved in the effect of RSG on Nrf2, HO-1 and COX-2. In summary, our study supports the hypothesis that RSG protect hepatocytes from high glucose-induced toxicity through PPARγ-dependent and PPARγ-independent pathways.

Introduction

Over the past decades, there has been a significant increase in the prevalence of diabetes, and current estimates indicate that by the year 2030, over 350 million people worldwide will be afflicted with this disease (Seghrouchni et al., 2002). Diabetes increases the risk of microvascular and macrovascular complications and premature death in the general population and results in a huge economic burden for society. A higher prevalence of diabetes among urban residents than among rural residents has been observed in developing countries throughout the world (Yang et al., 2010b). Chronic hyperglycemia is the hallmark of diabetes and the complications, leading to protein glycation, systemic low grade inflammation, and endothelial dysfunction through the induction of reactive oxygen species (ROS) generation (Ceolotto et al., 2007, Gallo et al., 2005, Han et al., 2006, Zheng et al., 2009). Oxidative stress is believed to be involved in the development of diabetic complications in animal models and in humans (Baynes, 1999). In the cells exposed to high glucose, excessive generation of ROS is considered to be mainly attributed to the activation of PKC (Kwan et al., 2005) and the polyol pathways (Baynes and Thorpe, 1996), mitochondrial dysfunction (Williamson et al., 1993) and the formation of advanced glycation products (Bagi et al., 2004).

In recent years, the studies concerning the mechanism underlying the effect of high glucose-induced ROS in diabetes attracted extensive attention. The liver is a central regulator of glucose homeostasis and stores or releases glucose according to metabolic demands. So, our understanding of high glucose-induced damage to liver and approaches to protect this injury are of great importance. Kaplowitz and Tsukamoto, (1996) have reported that oxidative stress played an important role in the pathogenesis of non-alcoholic steatohepatitis (NASH), which is the main liver damage observed in type 1 and 2 diabetes. Increased oxidative stress in rat liver was discovered in streptozotocin-induced diabetes (Kakkar et al., 1998). In addition, high glucose stimulated hepatic stellate cells to proliferate and to produce collagen through ROS production (Sugimoto et al., 2005). Moreover, high-glucose diet induced significant oxidative stress in liver of C57BL/6J mice (Du D et al., 2010). Indeed, oxidative injury to the liver in diabetes plays a fundamental role in the development of hepatic diabetic complications (Albano et al., 2005, Lukivskaya et al., 2007, Mitsuyoshi et al., 2006, Refaie et al., 2009).

Thiazolidinediones (TDZs) are ligands of the family of nuclear transcription factor peroxisome proliferator-activated receptor-γ (PPARγ), being used clinically for the treatment of type 2 diabetic patients through their insulin-sensitizing effect. Upon ligand binding, PPARγ forms a heterodimer with the retinoic X receptor, which binds to PPAR response elements (PPREs) within the promoter region of target genes (Berger and Moller, 2002). PPARγ is involved in the regulation of energy homeostasis. It has been shown that PPARγ agonists play an important role in regulating adipocyte differentiation, lipid and glucose metabolism, and inflammation (Guan et al., 2002). Rosiglitazone (RSG) is one classic member of TDZs family. In endothelial cells, treatment with RSG attenuates high glucose-induced ROS generation through its action on PPARγ, which is linked to the PKC pathway (Whiteside et al., 2009). Activation of PPARγ is involved in the protective effects of several Chinese herbal medicines and chemicals, most of which have antioxidant activities (Bae et al., 2010, Qiao et al., 2010, Shimada and Hiraishi, 2010, Yang et al., 2010a). In addition, the members of PPARγ family are well established to be important in modulating the fibrogenic response to liver injury (Qiao et al., 2010) and in the improvements in hepatic insulin sensitivity (Bouskila et al., 2005). The pivotal role of PPARγ agonist in the liver, although important for the regulation of genes involved in glucose and lipid metabolism, has generally not been fully appreciated. In the present study, we aim to investigate the protective effects of RSG against high glucose-induced oxidative damage to hepatocyte and its underlying mechanism.

Section snippets

Materials and reagents

P-Akt, P-p38 MAPK, P-ERK, Akt, ERK, HO-1 and Nrf2 antibodies were purchased from Epitomics. P-PKC, PKC, COX-2, PPARγ and p38 MAPK antibodies were purchased from Bioworld Technology. βactin antibody was purchased from Santa Cruz Biotechnology (Santa Cruz, CA). Glucose, rosiglitazone (RSG), DCFH-DA, DMSO and bovine serum albumin (BSA, fraction V, and essentially fatty acids free, low endotoxin) were purchased from Sigma–Aldrich (St. Louis, MO). Most of the other chemicals and reagents used in

Cytoprotective effect of RSG against high glucose-induced toxicity in hepatocytes

QZG hepatocytes were used, in the present study, to investigate high glucose-induced cytotoxicity and the underlying mechanisms. QZG cells were incubated with various concentrations of glucose (10, 15, 20, 25, and 30 mM) for 48 h. As shown in Fig. 1A, the viability of cells treated with 10–30 mM glucose for 48 h decreased markedly compared with that of control. In our subsequent experiments, a fixed concentration (30 mM) of glucose was used. In order to examine whether RSG exhibit a protective

Discussion

Hyperglycemia is the key characteristic of diabetes and its related complications. There are numerous studies concerning the effect of high glucose on the structure and function of endothelial cell. However, literature about the influence of excessive glucose on hepatocyte is relatively limited. In the current study, we investigated the possible mechanism underlying the effect of high glucose on hepatocyte in vitro, and observed the protective effect of RSG against this damage. The results

Conflicts of interest

The authors declare that there are no conflicts of interest.

Acknowledgment

This study was supported by the National Natural Science Foundation No. 30872135 and the Natural Science Foundation of Shaanxi Province No. 2010JZ004.

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