Neuroprotection by (-)-epigallocatechin-3-gallate in a rat model of stroke is mediated through inhibition of endoplasmic reticulum stress

  • Authors:
    • Chengye Yao
    • Jiancheng Zhang
    • Gongping Liu
    • Fang Chen
    • Yun Lin
  • Corresponding author:
  • View Affiliations

  • Published online on: Wednesday, November 6, 2013
  • Pages: 69-72 DOI: 10.3892/mmr.2013.1778

Abstract

(-)-Epigallocatechin-3‑gallate (EGCG), the predominant constituent of green tea, has been demonstrated to be neuroprotective against stroke in rats. However, the precise mechanism of EGCG responsible for neuroprotective activity remains unclear and no established treatment for decreasing the resulting neurological damage of stroke exists. The present study was designed to investigate the neuroprotective mechanism of EGCG on transient focal cerebral ischemia in rats. EGCG, when applied immediately following ischemia, significantly decreased the expression of endoplasmic reticulum stress (ERS)‑related markers, [glucose‑regulated protein 78 (GRP78), C/EBP‑homologous protein (CHOP) and caspase‑12] and apoptosis 24 h following reperfusion. EGCG treatment also significantly reduced infarct volumes and increased neurological scores which was correlated with elevated levels of TRPC6 and phosphorylation of cAMP/Ca2+ response element‑binding protein (p‑CREB) activity, and decreased calpain‑specific aII‑spectrin breakdown product (SBDP145) activity. When mitogen‑activated protein kinase kinase (MEK) activity was specifically inhibited, the neuroprotective effect of EGCG was attenuated and a correlated decrease in CREB activity was observed. In conclusion, the results clearly demonstrated that intracerebroventricular injection of EGCG immediately following ischemia, inhibits ERS and improves the neurological status of rats that have undergone middle cerebral artery occlusion via the inhibition of calpain‑mediated TRPC6 proteolysis and the subsequent activation of CREB via the MEK/extracellular signal-regulated kinases (ERK) pathway.

Introduction

The endoplasmic reticulum (ER) is an organelle that folds and synthesizes transmembrane, intraorganellar and secretory proteins. The disequilibrium of ER homeostasis, including glucose deprivation, disturbance of the redox environment, perturbation of calcium homeostasis and exposure to free radicals disrupts the normal function of the ER and induces ER stress (1). Pro-apoptotic proteins are expressed under ER stress, including CHOP and caspase-12 and -3, which results in neuronal cell death. ER stress plays a role in the pathogenesis of a variety of human diseases, including neuronal degenerative diseases, ischemia/reperfusion injury, and heart diseases (24).

The transient receptor potential cation channels (TRPC) are a subfamily of nonselective cation channels permeable to Ca2+, which are present in numerous cell types including neurons (5,6). The TRPC6 channel is involved in the promotion of neuronal survival following focal cerebral ischemia. Activation of calpain leads to TRPC6 degradation and neuronal damage in ischemia (7). Previous studies have determined that the TRPC6 channel is essential in promoting neuronal survival and indicates that the activation of CREB is a key downstream effector for the neuronal protective effect of the TRPC6 channel in vitro and in vivo(7,8). Therefore, it may be a used as a novel therapeutic strategy to protect against ischemic brain damage as the inhibition of TRPC6 degradation preserves neuronal survival.

EGCG is the predominant constituent of green tea (9). It has been shown to promote neuronal plasticity (10) and to improve cognitive function and learning ability (11,12). In addition, EGCG has also been demonstrated to reduce delayed cell death near the hippocampus and the excitotoxic neuronal damage that occurs in ischemic lesions following transient ischemia (2,13,14). However, the precise mechanism of the neuroprotective activity of EGCG remains unclear. Previous studies have not fully demonstrated the effects of EGCG on TRPC6/CREB-mediated neuroprotection.

The aim of this study was to investigate whether the administration of EGCG immediately following ischemia exhibits a neuroprotective effect on ischemic neurons in a middle cerebral artery occlusion (MCAO) rat model. This study also aimed to determine whether EGCG inhibits ER stress (ERS) and improves the neurological status through the inhibition of calpain-mediated TRPC6 proteolysis and the subsequent activation of CREB via the mitogen-activated protein kinase kinase (MEK)/extracellular signal-regulated kinases (ERK) pathway.

Materials and methods

Middle cerebral artery occlusion (MCAO) model

Male Sprague-Dawley rats (weight, 200–250 g) were purchased from Hunan Weasleyg Scene of Experimental Animals Co., Ltd. (Hubei, China). The experiments were approved by the Committee of Experimental Animals of Tongji Medical College (Hubei, China) and conformed to internationally accepted ethical standards (Guide for the care and use of laboratory animals; NIH Publication 80–23, revised 1978). Briefly, rats were anesthetized with an intraperitoneal (i.p.) injection of chloral hydrate (400 mg/kg) and placed in the supine position with the limbs taped to the operation table. A midline skin incision was performed, the right external carotid artery was exposed and its branches were ligated. A 4-0 monofilament nylon suture (Beijing Shandong Industrial Corp., Beijing, China) with a rounded tip was introduced into the internal carotid artery through the common carotid artery and advanced until faint resistance was felt. Following 2 h of transient MCAO, blood flow was restored by the withdrawal of the nylon thread to allow reperfusion, which was confirmed by a laser Doppler flowmeter; Periflux system 5000, Perimed, Stockholm, Sweden. Sham-operated rats underwent the same procedure without the filament insertion. Throughout the experiments, body temperature was maintained at 37±0.5°C with a homeothermic (RWD Life Science Co., Ltd., Shenzhen, China). Regional cerebral blood flow (rCBF) was monitored by a laser-Doppler flowmeter prior to, during and following MCAO, as well as prior to death (Fig. 1). Animals that did not show a CBF reduction of ≥70% and animals that died following ischemia induction were excluded from the experimental group. Prior to reperfusion, rats with incomplete MCAO (~10%) were excluded from further study by a blinded observer.

Drug treatment

Rat intracerebroventricular (ICV) injection was performed under anesthesia using a stereotaxic instrument (RWD Life Science Co., Ltd.) with a microsyringe pump (Shanghai Guangzheng Medical Equipment Co., Ltd., Shanghai, China). A scalp incision was perfomed and a burr hole was made in the right parietal skull, 1.8 mm lateral and 1.0 mm posterior to the bregma. A syringe was inserted into the brain to a depth of 4.2 mm below the cortical surface. EGCG or PD98059 was dissolved in dimethyl sulfoxide (DMSO) (all obtained from Sigma-Aldrich, St. Louis, MO, USA). EGCG (1 mg/ml; 5 μl) or 1% DMSO (5 μl) was injected slowly (0.5 μl/min) into the right ventricle immediately following ischemia. PD98059 (0.75 mg/rat, i.p.) or 1% DMSO (0.5 ml, i.p.) was administered to rats 20 min prior to the operation.

The rats were randomly divided into four groups and each group was again divided into three subgroups (n=12 per subgroup) according to the time of reperfusion following ischemia. The experimental groups and subgroups were as follows: Sham-operation (group S; subgroup S6, S12 and S24); MCAO (group I; subgroup I6, I12 and I24); ischemia combined with EGCG treatment (group E; subgroup E6, E12 and E24) and ischemia combined with EGCG plus PD98059 [a mitogen-activated protein kinase kinase (MEK) inhibitor]treatment (group C; subgroup C6, C12 and C24). Another 27 rats were randomly divided into 3 groups (n=9 per group): Sham-operation; MCAO (Group I) and ischemia combined with PD98059 treatment (Group P).

Measurements of infarct volume

At 24 h following reperfusion, rats were decapitated and the brains were rapidly removed and frozen at −20°C for 10 min. Sliced brain tissues were stained with 2% 2,3,5-triphenyltetrazolium chloride (TTC; Sigma-Aldrich) for 30 min at 37°C followed by overnight immersion in 4% paraformaldehyde. The extent of ischemic infarction was traced and the integrated volume was calculated using ImageJ 1.45 software (National Institutes of Health, Bethesda, MD, USA). The relative infarction volume was calculated by the following equation, giving a correction for edema: {[Total lesion volume - (ipsilateral hemisphere volume - contralateral hemisphere volume)] / contralateral hemisphere volume} ×100.

Neurological scoring

Neurological scores were evaluated by a blinded observer 24 h following reperfusion with a scoring system as described previously (15,16).

Western blot analysis

The rats were euthanized by decapitation at 6, 12 and 24 h following reperfusion and the infarct side of the cortex was harvested. Total protein extraction was performed according to the manufacturer’s instructions in the kit (KGP250; Keygen Biotech, Nanjing, China) for western blot analysis of TRPC6 and aII-spectrin. Nuclear protein extraction was performed according to the manufacturer’s instructions in the kit (Fisher Scientific, Pittsburgh, PA, USA) for p-CREB. Protein levels in the extracts were quantified using a bicinchoninic acid (BCA) assay. Equal quantities of total or nuclear protein extracts were separated by sodium dodecyl sulphate polyacrylamide gel electrophoresis and transferred to polyvinylidene difluoride membranes by electrophoresis. Membranes were incubated in Tris-buffered saline, TBS containing 1% Tween-20 (TBST) blocking buffer and 5% non-fat dry milk for 1 h at room temperature. Membranes were incubated overnight at 4°C with either a mouse monoclonal anti-aII-spectrin (dilution, 1:1000; Enzo Biochem, New York, NY, USA), rabbit polyclonal anti-TRPC6 (dilution, 1:1000; Abcam Cambridge, MA, USA), rabbit monoclonal anti-p-CREB (dilution, 1:1000) mouse monoclonal anti-CHOP (dilution, 1:1000), rabbit polyclonal anti-GRP78 (dilution, 1:1000; Cell Signaling Technology Inc., Beverly, MA, USA), rabbit polyclonal anti-caspase-12 (dilution, 1:200; Beijing Biosynthesis Biotechnology Co., Ltd, Beijing, China), rabbit polyclonal anti-Lamin B1 (dilution, 1:500; Bioworld Technology Inc., Harrogate, UK) or mouse monoclonal anti-glyceraldehyde 3-phosphate dehydrogenase (GAPDH) antibody (dilution, 1:1000; Proteintech Group, Inc., Hubei, China). This was followed by incubation with horseradish peroxidase-labeled secondary anti-mouse IgG or anti-rabbit IgG antibodies (dilution, 1:5000; Proteintech Group, Inc.), respectively. Labeled proteins were detected with the ChemiDocXRS+ chemiluminescence imaging system (Bio-Rad, Hercules, CA, USA) and bands were quantified using lab imaging software. The experiments were repeated in triplicate.

Quantum dot-based immunofluorescence and immunohistochemistry

At 24 h following reperfusion, the rats were perfused with 250 ml of 0.9% cold saline followed by 100 ml of 4% paraformaldehyde in phosphate-buffered saline (pH 7.4). The brains were then rapidly removed, blocked and embedded in paraffin. Paraffin-embedded brains were cut into 4-μm thick sections according to standard procedures. The paraffin sections (n=3 for each group) were incubated overnight with antibodies against TRPC6 (1:100; Abcam) at 4°C subsequent to blocking with bovine serum albumin (BSA). The samples were then incubated with a biotinylated secondary antibody at 37°C for 30 min. Subsequent to blocking with BSA, the paraffin sections were incubated with streptavidin-conjugated QDs605 (dilution, 1:100; Wuhan Jiayuan Quantum Dots Co., Ltd., Hubei, China). The cell nuclei were stained with 4′,6-diamidino-2-phenylindole (DAPI). TRPC6-positive cells were measured at a magnification of ×200 per visual field in the peri-infarct region; three visual fields per section and three brain sections per rat were analyzed. Fluorescent signals were detected by fluorescence microscopy (BX51; Olympus, Tokyo, Japan) and signal intensities were collected for statistical analysis. Images were captured with a Doppler imaging system (CRi Nuance Fx; Caliper Life Sciences, Hopkinton, MA, USA).

TdT mediated dUTP nick-end labeling (TUNEL) assay

TUNEL staining analysis was used to detect apoptotic cell death 24 h following reperfusion. TUNEL staining was conducted with a kit (Roche Diagnostics GmbH, Mannheim, Germany). TUNEL-positive nuclei with chromatin condensation and fragmented nuclei were considered as probable apoptotic cells. The total number of TUNEL-positive neurons in the ipsilateral hemisphere was counted in three different fields for each section (by an investigator who was blinded to the studies) by light microscopy at a magnification of ×400.

Statistical analysis

GraphPad Prism software (version 5 for Windows, GraphPad software Inc., La Jolla, CA, USA) was used for all statistical analyses. Values are presented as the mean ± standard error of the mean. The neurological score data comparison was analyzed using the Kruskal-Wallis test followed by a post hoc Dunn’s test. For all other measurements, one-way analysis of variance followed by Newman-Keuls multiple comparison test was used. P<0.05 was considered to indicate a statistically significant difference.

Results

EGCG blocks calpain-specific aII-spectrin breakdown product (SBDP145) formation

Compared with the sham-operated group the SBDP145 level in the MCAO group was significantly increased after 12 h (P<0.05), an increase that was greatest at 24 h (P<0.01; Fig. 2A and B). When MCAO rats were treated with EGCG the SBDP145 level was significantly decreased at 12 h (P<0.01) and the decrease was greatest at 24 h (P<0.01; Fig. 2C and D).

EGCG inhibits calpain-mediated TRPC6 channel degradation

Compared with the sham-operated group, TRPC6 levels in the MCAO group were significantly decreased at 6 (P<0.05), 12 and 24 h (P<0.01; Fig. 2E and F). When MCAO rats were treated with EGCG, the protein level of TRPC6 was significantly increased at 12 and 24 h (P<0.05 and P<0.01, respectively). Immunofluorescence analysis showed a cytomembrane staining pattern of TRPC6 in neurons of the cerebral cortex (Fig. 2G).

PD98059 exhibits no effect on ischemic stroke in rats at 24 h following reperfusion

To determine the effect of PD98059 in the stroke rats, PD98059 was administered 20 min prior to the operation. Notably, with the application of PD98059 alone, no statistical significance was identified in the protein levels of p-CREB in MCAO rats (Fig. 3A). There was no significant difference between the two groups in the measurements of the the infarct volumes and neurological scores (Fig. 3B and C).

EGCG maintains phosphorylation of CREB by blocking TRPC6 degradation

Compared with the sham-operated group, the level of p-CREB in the MCAO group was significantly decreased at 12 and 24 h (P<0.05 and P<0.01, respectively; Fig. 4A and B). Compared with the MCAO group, the level of p-CREB in the EGCG-treated group was significantly increased at 12 and 24 h (P<0.05 and P<0.01, respectively). In the rats treated with PD98059, the level of p-CREB was significantly decreased at 12 and 24 h (P<0.01 and P<0.01, respectively) compared with that of the EGCG-treated group.

EGCG inhibits ERS and apoptosis 24 h following reperfusion

In the MCAO group, the protein levels of ERS-related markers GRP78, CHOP and caspase-12 were significantly increased compared with that in the sham-operation group (P<0.01, P<0.01 and P<0.01, respectively; Fig. 5Aa–c). When MCAO rats were treated with EGCG, the protein levels of CHOP, GRP78 and caspase-12 were significantly decreased (P<0.01, P<0.01 and P<0.01, respectively). Subsequent to the administration of PD98059, the protein levels were significantly increased compared with that of the EGCG-treated group.

In the TUNEL assay, EGCG significantly reduced apoptotic cell death in the right cortex compared with that in the MCAO group (P<0.01; Fig. 5D and E). Following treatment with PD98059, the apoptotic cell death was significantly increased compared with that of the EGCG-treated group (P<0.01).

EGCG significantly reduces infarct volumes and promotes functional recovery in ipsilateral ischemic hemispheres 24 h following reperfusion

Following ischemia/reperfusion injury, a white-stained infarct area was observed in the MCAO group. By contrast, treatment with EGCG significantly reduced infarct volumes compared with that of the MCAO group 24 h following reperfusion (P<0.01). Following the application of PD98059, the infarct volumes were significantly increased compared with that of the EGCG-treated group (P<0.01; Fig. 5E).

There was also significant improvement in the neurological score 24 h following reperfusion with EGCG treatment (P<0.01). When treated with PD98059, the neurological scores were significantly decreased compared with that of the EGCG-treated group (P<0.01; Fig. 5F).

Discussion

The results clearly demonstrated that ICV injection of EGCG at low doses immediately following ischemia improved the outcome as measured by TTC staining and neurological scoring. EGCG treatment also significantly inhibited ERS and apoptosis. EGCG improved the neurological status and inhibited ERS, which was correlated with elevated TRPC6 and p-CREB activity and decreased SBDP145 activity. When MEK activity was inhibited, the neuroprotective effect of EGCG was attenuated and a correlated decrease in CREB activity was observed. These results demonstrated that EGCG, is important in the prevention of cerebral ischemic injury (17), and may be used as a therapeutic intervention for stroke during the acute or subacute period similar to the drug edaravone (18).

Calpains are intracellular calcium-dependent cysteine endopeptidases that are activated by cytosolic Ca2+ overload (19). The most studied target of calpain is aII-spectrin, a 280-kDa neuronal protein that localizes to axons and functions in cortical cytoskeleton matrix support. The aII-spectrin breakdown product, SBDP145, results from the sequential calpain cleavage of aII-spectrin which generates SBDP150 followed by cleavage to remove the additional 5 kDa (20,21). In the present study, MCAO rats exhibited elevated levels of SBDP145 in the cortical regions of the ipsilateral hemisphere in the first 24 h following ischemic injury. EGCG treatment significantly reduced SBDP145 formation at 12 and 24 h. The results clearly demonstrated that EGCG, when applied immediately following ischemia, inhibited calpain activation and induced resistance to ischemia/reperfusion injuries.

TRPC channels are non-selective cation channels that are expressed in numerous multicellular organisms with different functions (5). TRPC6 is involved in brain-derived neurotrophic factor (BDNF)-mediated growth cone turning, neuron survival and spine formation (8,22). TRPC6 was specifically degraded in transient ischemia and this degradation occurred prior to and during neuronal cell death. In addition, TRPC6 protein in neurons in ischemia was specifically downregulated by calpain proteolysis. Inhibition of calpain proteolysis of TRPC6 protected animals from ischemic brain damage. In the present study, the protein levels of TRPC6 were markedly decreased at 6 h and the reduction in TRPC6 protein levels remained at 12 and 24 h in the MCAO group, these results were consistent with a previous study (7). EGCG treatment significantly enhanced the protein levels of TRPC6 at 12 and 24 h. In addition, EGCG-treated rats exhibited significantly lower infarct volumes and also increased functional recovery compared with that of MCAO rats at 24 h. Therefore, the results indicated that EGCG treatment protected rats from ischemic brain damage through the inhibition of calpain proteolysis of TRPC6.

A modest level of Ca2+ influx through TRPC6 channels leads to the activation of ERK, which activates CREB to promote neuronal survival (8). Inhibition of TRPC6 channel degradation maintained the phosphorylation of CREB and prevented ischemic brain damage (7). CREB activation is a critical event in the neuroprotection from ischemic injury (23,24). In the present study, the protein levels of p-CREB were significantly increased in the EGCG-treated group at 12 and 24 h. When MEK activity was inhibited, the neuroprotective effect of EGCG was attenuated and correlated with decreased CREB activity levels. The results demonstrated that EGCG, when administered immediately following ischemia, stimulated the MEK/ERK pathway that ultimately induced CREB activation and contributed to neuroprotection at 24 h. In addition, EGCG significantly reduced TRPC6 degradation induced by ischemia at 24 h. Therefore, the results suggested that EGCG blocked calpain-mediated TRPC6 channel degradation which activated CREB through the MEK/ERK pathway and contributed to neuroprotection.

The ER is an organelle that is important in the maintenance of intracellular calcium homeostasis and proper folding of newly synthesized secretory and membranous proteins (25). ER functions are disturbed by different insults, such as the accumulation of unfolded proteins and the disruption of intracellular calcium homeostasis (26,27), which result in ER stress. However, if ER stress is too severe, the unfolded protein response initiates the apoptotic pathway (28). Increased expression of GRP78 is a marker of ER stress (29,30). In addition, CHOP participates in apoptosis signaling pathways and serves as a hallmark of ER stress (31,32). ER stress-induced neuronal cell death is important in stroke pathophysiology (33) and involves the activation of caspase-12 (34,35), which is specific to apoptosis mediated by ER stress (9). In the present study, EGCG treatment significantly decreased the infarct volumes and improved functional recovery at 24 h. In addition, the ER stress was enhanced in the brain following ischemia/reperfusion as demonstrated by the significant elevation of the ER stress-related markers GRP78, CHOP and caspase-12 in the cortex and EGCG was demonstrated to inhibit this. However, the effect of EGCG was attenuated by the MEK inhibitor PD98059. In addition, when treated with PD98059, the apoptotic cell death was also significantly increased. Thus, this study has demonstrated that ICV injection of EGCG inhibited ER stress and apoptosis through the MEK/ERK/CREB pathway, which contributed to its neuroprotective effects. The results indicated that EGCG exerted its neuroprotective effects by activating ERK/CREB pathways and the subsequent inhibition of ERS.

In conclusion, the present study has demonstrated that administration of EGCG immediately following ischemia inhibited ER stress and improved the neurological status through the inhibition of calpain proteolysis of TRPC6 and the subsequent activation of CREB via the MEK/ERK pathway.

Acknowledgements

This study was supported by the National Natural Science Foundation of China (grant no. 30901984).

References

1 

Kaufman RJ: Stress signaling from the lumen of the endoplasmic reticulum: coordination of gene transcriptional and translational controls. Genes Dev. 13:1211–1233. 1999. View Article : Google Scholar : PubMed/NCBI

2 

Zhang B, Rusciano D and Osborne NN: Orally administered epigallocatechin gallate attenuates retinal neuronal death in vivo and light-induced apoptosis in vitro. Brain Res. 1198:141–152. 2008. View Article : Google Scholar : PubMed/NCBI

3 

Vivien D, Gauberti M, Montagne A, Defer G and Touzé E: Impact of tissue plasminogen activator on the neurovascular unit: from clinical data to experimental evidence. J Cereb Blood Flow Metab. 31:2119–2134. 2011. View Article : Google Scholar : PubMed/NCBI

4 

Martinou JC, Dubois-Dauphin M, Staple JK, et al: Overexpression of BCL-2 in transgenic mice protects neurons from naturally occurring cell death and experimental ischemia. Neuron. 13:1017–1030. 1994. View Article : Google Scholar : PubMed/NCBI

5 

Montell C, Birnbaumer L and Flockerzi V: The TRP channels, a remarkably functional family. Cell. 108:595–598. 2002. View Article : Google Scholar : PubMed/NCBI

6 

Harteneck C, Plant TD and Schultz G: From worm to man: three subfamilies of TRP channels. Trends Neurosci. 23:159–166. 2000. View Article : Google Scholar : PubMed/NCBI

7 

Du W, Huang J, Yao H, Zhou K, Duan B and Wang Y: Inhibition of TRPC6 degradation suppresses ischemic brain damage in rats. J Clin Invest. 120:3480–3492. 2010. View Article : Google Scholar : PubMed/NCBI

8 

Jia Y, Zhou J, Tai Y and Wang Y: TRPC channels promote cerebellar granule neuron survival. Nat Neurosci. 10:559–567. 2007. View Article : Google Scholar : PubMed/NCBI

9 

Graham HN: Green tea composition, consumption, and polyphenol chemistry. Prev Med. 21:334–350. 1992. View Article : Google Scholar : PubMed/NCBI

10 

Xie W, Ramakrishna N, Wieraszko A and Hwang YW: Promotion of neuronal plasticity by (−)-epigallocatechin-3-gallate. Neurochem Res. 33:776–783. 2008.

11 

Haque AM, Hashimoto M, Katakura M, Tanabe Y, Hara Y and Shido O: Long-term administration of green tea catechins improves spatial cognition learning ability in rats. J Nutr. 136:1043–1047. 2006.PubMed/NCBI

12 

van Praag H, Lucero MJ, Yeo GW, et al: Plant-derived flavanol (−)epicatechin enhances angiogenesis and retention of spatial memory in mice. J Neurosci. 27:5869–5878. 2007.

13 

Nagai K, Jiang MH, Hada J, et al: (−)-Epigallocatechin gallate protects against NO stress-induced neuronal damage after ischemia by acting as an anti-oxidant. Brain Res. 956:319–322. 2002.

14 

Sutherland BA, Shaw OM, Clarkson AN, Jackson DN, Sammut IA and Appleton I: Neuroprotective effects of (−)-epigallocatechin gallate following hypoxia-ischemia-induced brain damage: novel mechanisms of action. FASEB J. 19:258–260. 2005.

15 

Garcia JH, Wagner S, Liu KF and Hu XJ: Neurological deficit and extent of neuronal necrosis attributable to middle cerebral artery occlusion in rats. Statistical validation. Stroke. 26:627–635. 1995. View Article : Google Scholar

16 

Tsubokawa T, Jadhav V, Solaroglu I, Shiokawa Y, Konishi Y and Zhang JH: Lecithinized superoxide dismutase improves outcomes and attenuates focal cerebral ischemic injury via antiapoptotic mechanisms in rats. Stroke. 38:1057–1062. 2007. View Article : Google Scholar : PubMed/NCBI

17 

Arab L, Liu W and Elashoff D: Green and black tea consumption and risk of stroke: a meta-analysis. Stroke. 40:1786–1792. 2009. View Article : Google Scholar : PubMed/NCBI

18 

Zhang N, Komine-Kobayashi M, Tanaka R, Liu M, Mizuno Y and Urabe T: Edaravone reduces early accumulation of oxidative products and sequential inflammatory responses after transient focal ischemia in mice brain. Stroke. 36:2220–2225. 2005. View Article : Google Scholar : PubMed/NCBI

19 

Goll DE, Thompson VF, Li H, Wei W and Cong J: The calpain system. Physiol Rev. 83:731–801. 2003.

20 

Nath R, Raser KJ, Stafford D, et al: Non-erythroid alpha-spectrin breakdown by calpain and interleukin 1 beta-converting-enzyme-like protease(s) in apoptotic cells: contributory roles of both protease families in neuronal apoptosis. Biochem J. 319:683–690. 1996.

21 

Wang KK: Calpain and caspase: can you tell the difference?, by kevin KW Wang Vol 23, pp 20–26. Trends Neurosci. 23:592000.

22 

Li Y, Jia YC, Cui K, et al: Essential role of TRPC channels in the guidance of nerve growth cones by brain-derived neurotrophic factor. Nature. 434:894–898. 2005. View Article : Google Scholar : PubMed/NCBI

23 

Walton MR and Dragunow I: Is CREB a key to neuronal survival? Trends Neurosci. 23:48–53. 2000. View Article : Google Scholar : PubMed/NCBI

24 

Finkbeiner S: CREB couples neurotrophin signals to survival messages. Neuron. 25:11–14. 2000. View Article : Google Scholar : PubMed/NCBI

25 

Yin XM, Oltvai ZN and Korsmeyer SJ: BH1 and BH2 domains of Bcl-2 are required for inhibition of apoptosis and heterodimerization with Bax. Nature. 369:321–323. 1994. View Article : Google Scholar : PubMed/NCBI

26 

He B: Viruses, endoplasmic reticulum stress, and interferon responses. Cell Death Differ. 13:393–403. 2006. View Article : Google Scholar : PubMed/NCBI

27 

Ginsberg MD: Neuroprotection for ischemic stroke: past, present and future. Neuropharmacology. 55:363–389. 2008. View Article : Google Scholar : PubMed/NCBI

28 

Love S: Apoptosis and brain ischaemia. Prog Neuropsychopharmacol Biol Psychiatry. 27:267–282. 2003. View Article : Google Scholar

29 

Yoshida H, Matsui T, Yamamoto A, Okada T and Mori K: XBP1 mRNA is induced by ATF6 and spliced by IRE1 in response to ER stress to produce a highly active transcription factor. Cell. 107:881–891. 2001. View Article : Google Scholar : PubMed/NCBI

30 

Groenendyk J, Sreenivasaiah PK, Kim do H, Agellon LB and Michalak M: Biology of endoplasmic reticulum stress in the heart. Circ Res. 107:1185–1197. 2010. View Article : Google Scholar : PubMed/NCBI

31 

DeGracia DJ and Montie HL: Cerebral ischemia and the unfolded protein response. J Neurochem. 91:1–8. 2004. View Article : Google Scholar : PubMed/NCBI

32 

Tajiri S, Oyadomari S, Yano S, et al: Ischemia-induced neuronal cell death is mediated by the endoplasmic reticulum stress pathway involving CHOP. Cell Death Differ. 11:403–415. 2004. View Article : Google Scholar : PubMed/NCBI

33 

Zhu DY, Lau L, Liu SH, Wei JS and Lu YM: Activation of cAMP-response-element-binding protein (CREB) after focal cerebral ischemia stimulates neurogenesis in the adult dentate gyrus. Proc Natl Acad Sci USA. 101:9453–9457. 2004. View Article : Google Scholar : PubMed/NCBI

34 

Zinkstok SM, Vergouwen MD, Engelter ST, et al: Safety and functional outcome of thrombolysis in dissection-related ischemic stroke: a meta-analysis of individual patient data. Stroke. 42:2515–2520. 2011. View Article : Google Scholar : PubMed/NCBI

35 

Paschen W and Mengesdorf T: Cellular abnormalities linked to endoplasmic reticulum dysfunction in cerebrovascular disease - therapeutic potential. Pharmacol Ther. 108:362–375. 2005. View Article : Google Scholar : PubMed/NCBI

Journal Cover

January 2014
Volume 9 Issue 1

Print ISSN: 1791-2997
Online ISSN:1791-3004

2013 Impact Factor: 1.484
2014 I.F. (Expected) ≥ 1.777 Ranked #59/122 Medicine Research and Experimental
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APA
Yao, C., Zhang, J., Liu, G., Chen, F., & Lin, Y. (2014). Neuroprotection by (-)-epigallocatechin-3-gallate in a rat model of stroke is mediated through inhibition of endoplasmic reticulum stress. Molecular Medicine Reports, 9, 69-72. http://dx.doi.org/10.3892/mmr.2013.1778
MLA
Yao, C., Zhang, J., Liu, G., Chen, F., Lin, Y."Neuroprotection by (-)-epigallocatechin-3-gallate in a rat model of stroke is mediated through inhibition of endoplasmic reticulum stress". Molecular Medicine Reports 9.1 (2014): 69-72.
Chicago
Yao, C., Zhang, J., Liu, G., Chen, F., Lin, Y."Neuroprotection by (-)-epigallocatechin-3-gallate in a rat model of stroke is mediated through inhibition of endoplasmic reticulum stress". Molecular Medicine Reports 9, no. 1 (2014): 69-72. http://dx.doi.org/10.3892/mmr.2013.1778