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Conference Paper: Click chemistry-assisted identification of caffeic acid propargylamide (CAPA)-modified protein Keap1 demonstrates a general mechanism underlying the neuroprotective and neuritogenic activities of o-catechol-bearing natural products
Title | Click chemistry-assisted identification of caffeic acid propargylamide (CAPA)-modified protein Keap1 demonstrates a general mechanism underlying the neuroprotective and neuritogenic activities of o-catechol-bearing natural products |
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Authors | |
Issue Date | 2014 |
Citation | The 8th Pong Ding Yuen International Symposium on Traditional Chinese Medicine cum, The 2nd International Chinese Symposium on Free Radical Research & The 6th Symposium for Three Districts of Cross-straits on Free Radical Research, The University of Hong Kong, Hong Kong, 15-16 November 2014. How to Cite? |
Abstract | Objectives: Caffeic acid and its derivatives bear a typical o-catechol structural element and exhibit a broad spectrum of pharmacological activities including antioxidant, anti-inflammatory, chemopreventive, anticancer, antibacterial and neuroprotective effects. The present study was designed to characterize the neuritogenic and neuroprotective activities of caffeic acid derivatives and the underlying molecular mechanisms. Methods: We synthesized caffeic acid propargylamide (CAPA) as a probe to label the intracellular protein targets in a covalent manner. Caffeic acid-modified proteins are extracted from CAPA-treated cells and incubated with Azido-propyl biotin through a Click chemistry procedure. The biotin tag was detected by streptavidin-HRP conjugate. For neurite outgrowth assay, the neurites were visualized by neurite outgrowth staining kit and neuronal biomarkers (e.g. β3-tubulin and MAP2) were detected by Western blotting and immunostaining. Cell viability was assayed by a colorimetric MTT assay. ROS and RNS were determined by fluorescent probes, DCFH-DA and DAF-FM DA, respectively. Results: We isolated a predominant protein with a size of 72 kDa according to the detection with streptavidin-HRP conjugate. We further verified that CAPA covalently bound to Keap1, a key protein to regulate the activation of transcription factor Nrf2 and antioxidant enzyme heme oxygenase-1 (HO-1) pathway. CAPA was unexpectedly found to be a potent neuritogenic agent. CAPA potentiated nerve growth factor (NGF)-induced neurite outgrowth in PC12 cells in a concentration- and time-dependent manner. Moreover, CAPA could also protect neuronal cells against 6-hydroxydopamine (6-OHDA) toxicity via suppressing 6-OHDA-induced production of reactive oxygen species (ROS)/reactive nitrogen species (RNS) and preserving the integrity of mitochondrial membrane. Importantly, HO-1 inhibitor almost abolished the neuritogenic and neuroprotective activities of CAPA. Conclusion: These results suggest that CAPA and related caffeic acid derivatives exhibit neuroprotective and neuritogenic activities via the modification of Keap1 and subsequent activation of Nrf2/HO-1 pathway. |
Description | Conference Theme: Free Radical, Chinese Medicine and Translational Medicine - 自由基, 中醫藥, 轉化醫學 Parallel Session - Session 2: Chinese Medicine - Chinese Medicine & Antioxidative Therapy: no. PS-CM-06 |
Persistent Identifier | http://hdl.handle.net/10722/213637 |
DC Field | Value | Language |
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dc.contributor.author | Rong, J | - |
dc.contributor.author | Yang, C | - |
dc.date.accessioned | 2015-08-07T08:26:45Z | - |
dc.date.available | 2015-08-07T08:26:45Z | - |
dc.date.issued | 2014 | - |
dc.identifier.citation | The 8th Pong Ding Yuen International Symposium on Traditional Chinese Medicine cum, The 2nd International Chinese Symposium on Free Radical Research & The 6th Symposium for Three Districts of Cross-straits on Free Radical Research, The University of Hong Kong, Hong Kong, 15-16 November 2014. | - |
dc.identifier.uri | http://hdl.handle.net/10722/213637 | - |
dc.description | Conference Theme: Free Radical, Chinese Medicine and Translational Medicine - 自由基, 中醫藥, 轉化醫學 | - |
dc.description | Parallel Session - Session 2: Chinese Medicine - Chinese Medicine & Antioxidative Therapy: no. PS-CM-06 | - |
dc.description.abstract | Objectives: Caffeic acid and its derivatives bear a typical o-catechol structural element and exhibit a broad spectrum of pharmacological activities including antioxidant, anti-inflammatory, chemopreventive, anticancer, antibacterial and neuroprotective effects. The present study was designed to characterize the neuritogenic and neuroprotective activities of caffeic acid derivatives and the underlying molecular mechanisms. Methods: We synthesized caffeic acid propargylamide (CAPA) as a probe to label the intracellular protein targets in a covalent manner. Caffeic acid-modified proteins are extracted from CAPA-treated cells and incubated with Azido-propyl biotin through a Click chemistry procedure. The biotin tag was detected by streptavidin-HRP conjugate. For neurite outgrowth assay, the neurites were visualized by neurite outgrowth staining kit and neuronal biomarkers (e.g. β3-tubulin and MAP2) were detected by Western blotting and immunostaining. Cell viability was assayed by a colorimetric MTT assay. ROS and RNS were determined by fluorescent probes, DCFH-DA and DAF-FM DA, respectively. Results: We isolated a predominant protein with a size of 72 kDa according to the detection with streptavidin-HRP conjugate. We further verified that CAPA covalently bound to Keap1, a key protein to regulate the activation of transcription factor Nrf2 and antioxidant enzyme heme oxygenase-1 (HO-1) pathway. CAPA was unexpectedly found to be a potent neuritogenic agent. CAPA potentiated nerve growth factor (NGF)-induced neurite outgrowth in PC12 cells in a concentration- and time-dependent manner. Moreover, CAPA could also protect neuronal cells against 6-hydroxydopamine (6-OHDA) toxicity via suppressing 6-OHDA-induced production of reactive oxygen species (ROS)/reactive nitrogen species (RNS) and preserving the integrity of mitochondrial membrane. Importantly, HO-1 inhibitor almost abolished the neuritogenic and neuroprotective activities of CAPA. Conclusion: These results suggest that CAPA and related caffeic acid derivatives exhibit neuroprotective and neuritogenic activities via the modification of Keap1 and subsequent activation of Nrf2/HO-1 pathway. | - |
dc.language | eng | - |
dc.relation.ispartof | 8th Pong Ding Yuen International Symposium on Traditional Chinese Medicine, 2nd International Chinese Symposium on Free Radical Research & the 6th Symposium for Three Districts of Cross-straits on Free Radical Research | - |
dc.relation.ispartof | 第八屇龐鼎元國際中醫藥研討會暨第二屇世界華人自由基生物學與自由基醫學學術大會及第六屇海峽兩岸三地自由基生物學與自由基醫學研討會 | - |
dc.title | Click chemistry-assisted identification of caffeic acid propargylamide (CAPA)-modified protein Keap1 demonstrates a general mechanism underlying the neuroprotective and neuritogenic activities of o-catechol-bearing natural products | - |
dc.type | Conference_Paper | - |
dc.identifier.email | Rong, J: jrong@hku.hk | - |
dc.identifier.authority | Rong, J=rp00515 | - |
dc.identifier.hkuros | 247196 | - |