Grape seed extract has superior beneficial effects than vitamin E on oxidative stress and apoptosis in the hippocampus of streptozotocin induced diabetic rats

Highlights

• Diabetes leads to functional and structural changes in the central nervous systems.

• Grape seed extract (GSE) and vitamin E have anti-oxidative and -apoptotic properties.

• GSE and vitamin E decrease oxidative stress occurring in the diabetic rats.

• GSE and vitamin E exert anti-apoptotic effects in the hippocampus of diabetic rats.

• GSE is more useful in preventing diabetes induced neuronal complications.

Abstract

We aimed to investigate the effects of grape seed extract (GSE) and vitamin E (Vit E) on oxidative stress and apoptosis in the hippocampus of streptozotocin-induced diabetic rats. In Control, Diabetic, and Diabetic treated with GSE (Diabetic + GSE) and vitamin E (Diabetic + Vit E) groups, oxidative stress index (OSI), TUNEL staining and Bcl-2, Bcl-XL, Bax, caspase-3, -9, and -8, Cyt-c, TNF-α, and NF-κB gene expressions were evaluated. OSI was significantly increased in the plasma and hippocampus of the Diabetic compared to Control group and decreased in Diabetic + GSE and Diabetic + Vit E groups compared to Diabetic. TUNEL positive neurons significantly increased in the hippocampus of the Diabetic group compared to Control and decreased in Diabetic + GSE (more prominently) and Diabetic + Vit E groups compared to Diabetic. In the hippocampus of the Diabetic group, Bcl-2 and Bcl-XL gene expressions were significantly decreased; Bax, caspase-3, -9, and -8, Cyt-c, TNF-α, and NF-κB gene expressions were significantly increased compared to Control. In Diabetic + GSE and Diabetic + Vit E groups, Bcl-2 gene expressions were significantly increased; Bcl-XL gene expressions did not differ compared to the Diabetic group. The expression of Bax, caspase-3, -9, and -8, Cyt-c, TNF-α, and NF-κB genes in the Diabetic + GSE group and the expression of caspase-3 and -9, TNF-α, and NF-κB genes in the Diabetic + Vit E group were significantly decreased compared to Diabetic. In conclusion, GSE (more prominently) and vitamin E decreased oxidative stress and neuronal apoptosis occurring in the hippocampus of diabetic rats.

Introduction

Diabetes mellitus is an important metabolic disorder that causes functional and structural changes in the central nervous systems. Moderate cerebral atrophy and increased subcortical and brain stem lesions have been reported in uncontrolled diabetic patients. Previous studies have stated that memory, learning, and cognitive impairments are more common in diabetics than in nondiabetics (Reagan, 2007, Muriach et al., 2006, Li et al., 2002). The hippocampus is a critical integration center for cognitive functions such as learning and memory in the mammalian brain. Since the hippocampus is susceptible to hyperglycemic damage, it is considered to be a specific target for diabetes-related changes within the central nervous system (Alvarez et al., 2009).

Oxidative stress increased by hyperglycemia leads to numerous cellular reactions in the central nervous system. At the cellular level, mechanisms of high-glucose (HG)-induced cell death are sustained by oxidative stress in many cell types. Excessive formation of reactive oxygen-free radicals (ROS) as well as insufficient antioxidant capacity may damage cellular components. HG-induced free radical attack increases neuron death by causing damage to lipids, proteins and DNA (Bournival et al., 2012). At least, programmed cell death (apoptosis) may be a possible mechanism of glucose neurotoxicity (Piotrowski, 1999, Sharifi et al., 2007, Hernández-Fonseca et al., 2009, Bournival et al., 2012). Apoptosis occurs with gene regulation and is characterized by the induction of the expression of a number of transcription factors and genes. Increased Bax expression, alteration in Bcl-2 expression, increased cytochrome c (Cyt-c) release and increased caspase-3, -9, and -8 expressions have been shown to play a role in HG-mediated toxicity which induces apoptosis of neurons (Sharifi et al., 2007, Jafari Anarkooli et al., 2008, Hao et al., 2012, Fu et al., 2012). Additionally, induction of the apoptosis by increased activation of pro-inflammatory cytokines such as nuclear factor kappaB (NF-κB) (Alvarez-Nölting et al., 2012, Muriach et al., 2006, Jing et al., 2013) and tumor necrosis factor-α (TNF-α) (Marfella et al., 2006, Jing et al., 2013) in streptozotocin (STZ) diabetic rats has been previously reported. Consistent with the abovementioned mechanisms, increased numbers of TUNEL positive cells have been observed in various brain regions of diabetic animals (Li et al., 2002, Hernández-Fonseca et al., 2009, Huang et al., 2012, Liu et al., 2013).

For protection from HG-related neurodegeneration, increasing antioxidant levels (such as vitamins A, E, and C and glutathione) has been reported to be as important as preventing the production of excessive free radicals (Zaltzberg et al., 1999). Vitamin E has been reported to reduce the generation of reactive oxygen-free radicals, damage oxidative substances, and maintain membrane fluidity in diabetic rat brains (Hong et al., 2004). Vitamin E prevents lipid peroxidation, elevates glutathione levels in the hippocampus, and ameliorates learning and memory performance in STZ diabetic rats (Tuzcu and Baydas, 2006). The neuroprotective action of vitamin E involves reduction of oxidative stress as well as partial inhibition of apoptosis (Demiralay et al., 2008). In fact, the inhibitory effect of vitamin E on caspase-3 and some cytokines has been alleged (such as TNF-a, NF-κB, IL-1b, TGF-1b) in diabetic rats (Kuhad and Chopra, 2009a, Kuhad and Chopra, 2009b).

Grape seed extract (GSE) is another powerful free radical scavenger (Balu et al., 2006, Devi et al., 2006, Lu et al., 2010). Grape seeds contain flavonoids (catechin, epicatechin, procyanidins and anthocyanins) and phenolic acids (gallic acid and ellagic acid) (Yilmaz and Toledo, 2006). GSE has been shown to provide a superior anti-oxidant effect compared to vitamin E at equal doses by weight (Bagchi et al., 1997, Ariga, 2004). Recent studies have reported that GSE has a potential therapeutic effect via reducing oxidative stress on diabetes and its related complications (Sato et al., 2001, Pinent et al., 2004, Cui et al., 2008, Chis et al., 2009, Kiyici et al., 2010, Mansouri et al., 2011, Ding et al., 2013). However, it has also been alleged that it has limited effects in this context (Belviranli et al., 2012). Besides some of its positive effects on the hippocampus (Xu et al., 2008) and cerebral cortex (Lu et al., 2010) of STZ-induced diabetic rats, we have not encountered any study about its effects on diabetes-related cerebral neurodegeneration. Yet, it has been argued in recent years that it has neuroprotective effects on many individual diseases. It has been shown that grape seed extract suppresses lipid peroxidation and reduces brain injury in hypoxic–ischemic rat models (Feng et al., 2007). It protects from MeHg-induced (Yang⁎ et al., 2012b) and O-ethyl-S,S-dipropyl phosphorodithioate (MOCAP)-induced neurotoxicity (Ray et al., 2000, Bagchi et al., 2001) and from Alzheimer's disease (Ksiezak-Reding et al., 2012), inhibits glutamate excitotoxicity (Ahn et al., 2011), and enhances neurogenesis in the dentate gyrus (Yoo et al., 2011). Although previous studies have already presented antiapoptotic effects of GSE in various tissues, including reduction of apoptotic cell death and suppression of proapoptotic proteins (Sato et al., 2001, Ozkan et al., 2012, Bayatli et al., 2013, Cedó et al., 2013, Chen et al., 2013, Katiyar and Athar, 2013), there are limited studies about its effects on neurons (Ray et al., 2000, Bagchi et al., 2001, Balu et al., 2006, Feng et al., 2007, Yang et al., 2012a, Yang et al., 2012b). Moreover, limited information is available about the effects of GSE on hippocampal neuronal loss in STZ-induced diabetic rats.

We attempted in this study to assess the anti-oxidative, anti-inflammatory, and anti-apoptotic effects of GSE that was obtained from red grapes of Calkarasi (Vitis vinifera L. cv.) from the vicinity of Denizli and vitamin E on neuronal loss occurring in the hippocampus of STZ-induced diabetic rats.