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The protective role of (-)-epigallocatechin-3-gallate in thrombin-induced neuronal cell apoptosis and JNK-MAPK activation.

He Q, Bao L, Zimering J, Zan K, Zhang Z, Shi H, Zu J, Yang X, Hua F, Ye X, Cui G - Neuroreport (2015)

Bottom Line: Flow cytometric analysis and western blotting demonstrated that thrombin-induced neuron degeneration occurs through apoptosis.These data suggest that EGCG may have protective effects against thrombin-induced neuroapoptosis by inhibiting the activation of JNK, leading to caspase 3 cleavage.EGCG is a novel candidate neuroprotective agent against intracerebral hemorrhage-induced neurotoxicity.

View Article: PubMed Central - PubMed

Affiliation: aDepartment of Neurology, The Affiliated Hospital of Xuzhou Medical University, Xuzhou, Jiangsu, China bUniversity of Rochester School of Medicine and Dentistry, Rochester, New York, USA.

ABSTRACT
(-)-Epigallocatechin-3-gallate (EGCG), the major polyphenolic component of green tea, has anti-inflammatory and antioxidant properties and provides neuroprotection against central nervous system diseases. Yet, it is not known whether EGCG may be neuroprotective against intracerebral hemorrhage. In this study, we used a simplified in-vitro model of thrombin neurotoxicity to test whether EGCG provides neuroprotection against thrombin-associated toxicity. Exposure of primary cortical neurons to thrombin (100 U/ml) caused dose-dependent and time-dependent cytotoxicity. Cell Counting Kit 8 and lactate dehydrogenase were used to monitor cell viability after exposure of neurons to thrombin or EGCG and after EGCG pretreatment. Flow cytometric analysis and western blotting demonstrated that thrombin-induced neuron degeneration occurs through apoptosis. A concentration of 25 μM EGCG significantly abolished thrombin-induced toxicity and prevented apoptosis by suppressing c-Jun-N-terminal kinase (JNK) phosphorylation, and the JNK inhibitor SP600125 reduced thrombin-induced caspase 3 activation and apoptosis. These data suggest that EGCG may have protective effects against thrombin-induced neuroapoptosis by inhibiting the activation of JNK, leading to caspase 3 cleavage. EGCG is a novel candidate neuroprotective agent against intracerebral hemorrhage-induced neurotoxicity.

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CCK8 and LDH assays were used to detect cell viability. (a, b) Neuron cell viability after exposure of neurons to thrombin or EGCG with a different concentration gradient (*P<0.05 vs. 5 U/ml thrombin, ***P<0.001 vs. control group). (c, d) Neuron cell damage increased in the thrombin group and EGCG pretreatment significantly increased cell viability (*P<0.05 vs. control group). Columns represent the mean±SE. n=4. CCK8, Cell Counting Kit 8; EGCG, (−)-epigallocatechin-3-gallate; LDH, lactate dehydrogenase; TM, thrombin.
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Figure 2: CCK8 and LDH assays were used to detect cell viability. (a, b) Neuron cell viability after exposure of neurons to thrombin or EGCG with a different concentration gradient (*P<0.05 vs. 5 U/ml thrombin, ***P<0.001 vs. control group). (c, d) Neuron cell damage increased in the thrombin group and EGCG pretreatment significantly increased cell viability (*P<0.05 vs. control group). Columns represent the mean±SE. n=4. CCK8, Cell Counting Kit 8; EGCG, (−)-epigallocatechin-3-gallate; LDH, lactate dehydrogenase; TM, thrombin.

Mentions: A dose gradient of thrombin or EGCG incubated with the cells revealed that neuron viability was dose-dependent. Figure 2a shows that the optimal concentration of thrombin for induction was 50 or 100 U/ml. To achieve the best effect, we chose 100 U/ml thrombin for the subsequent experiment. LDH released from neurons treated with 50 μM EGCG was remarkably higher than that released by the control cell group (Fig. 2b). When neurons were pretreated with EGCG for 24 h and then treated with thrombin for 48 h, EGCG inhibited the thrombin-induced decrease in neuron viability (Fig. 2c and d).


The protective role of (-)-epigallocatechin-3-gallate in thrombin-induced neuronal cell apoptosis and JNK-MAPK activation.

He Q, Bao L, Zimering J, Zan K, Zhang Z, Shi H, Zu J, Yang X, Hua F, Ye X, Cui G - Neuroreport (2015)

CCK8 and LDH assays were used to detect cell viability. (a, b) Neuron cell viability after exposure of neurons to thrombin or EGCG with a different concentration gradient (*P<0.05 vs. 5 U/ml thrombin, ***P<0.001 vs. control group). (c, d) Neuron cell damage increased in the thrombin group and EGCG pretreatment significantly increased cell viability (*P<0.05 vs. control group). Columns represent the mean±SE. n=4. CCK8, Cell Counting Kit 8; EGCG, (−)-epigallocatechin-3-gallate; LDH, lactate dehydrogenase; TM, thrombin.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4390119&req=5

Figure 2: CCK8 and LDH assays were used to detect cell viability. (a, b) Neuron cell viability after exposure of neurons to thrombin or EGCG with a different concentration gradient (*P<0.05 vs. 5 U/ml thrombin, ***P<0.001 vs. control group). (c, d) Neuron cell damage increased in the thrombin group and EGCG pretreatment significantly increased cell viability (*P<0.05 vs. control group). Columns represent the mean±SE. n=4. CCK8, Cell Counting Kit 8; EGCG, (−)-epigallocatechin-3-gallate; LDH, lactate dehydrogenase; TM, thrombin.
Mentions: A dose gradient of thrombin or EGCG incubated with the cells revealed that neuron viability was dose-dependent. Figure 2a shows that the optimal concentration of thrombin for induction was 50 or 100 U/ml. To achieve the best effect, we chose 100 U/ml thrombin for the subsequent experiment. LDH released from neurons treated with 50 μM EGCG was remarkably higher than that released by the control cell group (Fig. 2b). When neurons were pretreated with EGCG for 24 h and then treated with thrombin for 48 h, EGCG inhibited the thrombin-induced decrease in neuron viability (Fig. 2c and d).

Bottom Line: Flow cytometric analysis and western blotting demonstrated that thrombin-induced neuron degeneration occurs through apoptosis.These data suggest that EGCG may have protective effects against thrombin-induced neuroapoptosis by inhibiting the activation of JNK, leading to caspase 3 cleavage.EGCG is a novel candidate neuroprotective agent against intracerebral hemorrhage-induced neurotoxicity.

View Article: PubMed Central - PubMed

Affiliation: aDepartment of Neurology, The Affiliated Hospital of Xuzhou Medical University, Xuzhou, Jiangsu, China bUniversity of Rochester School of Medicine and Dentistry, Rochester, New York, USA.

ABSTRACT
(-)-Epigallocatechin-3-gallate (EGCG), the major polyphenolic component of green tea, has anti-inflammatory and antioxidant properties and provides neuroprotection against central nervous system diseases. Yet, it is not known whether EGCG may be neuroprotective against intracerebral hemorrhage. In this study, we used a simplified in-vitro model of thrombin neurotoxicity to test whether EGCG provides neuroprotection against thrombin-associated toxicity. Exposure of primary cortical neurons to thrombin (100 U/ml) caused dose-dependent and time-dependent cytotoxicity. Cell Counting Kit 8 and lactate dehydrogenase were used to monitor cell viability after exposure of neurons to thrombin or EGCG and after EGCG pretreatment. Flow cytometric analysis and western blotting demonstrated that thrombin-induced neuron degeneration occurs through apoptosis. A concentration of 25 μM EGCG significantly abolished thrombin-induced toxicity and prevented apoptosis by suppressing c-Jun-N-terminal kinase (JNK) phosphorylation, and the JNK inhibitor SP600125 reduced thrombin-induced caspase 3 activation and apoptosis. These data suggest that EGCG may have protective effects against thrombin-induced neuroapoptosis by inhibiting the activation of JNK, leading to caspase 3 cleavage. EGCG is a novel candidate neuroprotective agent against intracerebral hemorrhage-induced neurotoxicity.

Show MeSH
Related in: MedlinePlus