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postgraduate thesis: Molecular mechanisms of acacetin for the blockade of Kv1.5 and Kv4.3 channels and the cardio-protection against ischemia-reperfusion injury

TitleMolecular mechanisms of acacetin for the blockade of Kv1.5 and Kv4.3 channels and the cardio-protection against ischemia-reperfusion injury
Authors
Advisors
Advisor(s):Li, GRTse, HF
Issue Date2012
PublisherThe University of Hong Kong (Pokfulam, Hong Kong)
Citation
Wu, H. [巫慧鈞]. (2012). Molecular mechanisms of acacetin for the blockade of Kv1.5 and Kv4.3 channels and the cardio-protection against ischemia-reperfusion injury. (Thesis). University of Hong Kong, Pokfulam, Hong Kong SAR. Retrieved from http://dx.doi.org/10.5353/th_b4961749
AbstractAcacetin is a natural flavone compound, initially isolated from the traditional Chinese medicinal herb Tianshanxuelian, and has been recently demonstrated effective in anti-atrial fibrillation by blocking atrial-selective K+ currents, ultra-rapidly delayed rectifier K+ current (IKur), transient outward K+ current (Ito), and acetylcholine-activated K+ current (IKACh). However, the molecular determinants of acacetin blocking IKur and Ito are not fully understood. My PhD project was designed to investigate the cellular and molecular mechanisms of the channel blocking effects using HEK 293 cells expressing cloned hKv1.5 and hKv4.3 channels that encode IKur and Ito, respectively. In addition, I am interested to explore whether acacetin has potential cardioprotective effect against ischemia/reperfusion injury and the related cellular and molecular mechanisms. In the first part of my PhD project, I identified that acacetin blocked hKv1.5 channels in both open and closed states by binding to the S6 segment of the channel. The blockade effect was use- and frequency-dependent. IC50 of acacetin on hKv1.5 current was 3.5, 3.1, 2.9, 2.1 and 1.7 μM at 0.2, 0.5, 1.0, 3.0, and 4.0 Hz respectively. In site-directed mutagenesis studies, mutations in hKv1.5 channel S6-segment, V505A and I508A, remarkably reduced the channel sensitivity to acacetin with IC50 29.5μM for V505A and 19.1 μM for I508A. In the second part of my study, I demonstrated that acacetin blocked hKv4.3 channels by binding to the pore of the channel, decreased the recovery of the channel from inactivation and positively shifted the activation conductance. The blockade of hKv4.3 channels by acacetin was use- and frequency-dependent. IC50 of acacetin for inhibiting hKv4.3 was 7.9, 6.1, 3.9, and 3.2 μM at 0.2, 0.5, 1.0, and 3.3 Hz respectively. Site-directed mutagenesis study revealed that hKv4.3 mutations T366A, T367A in the P-loop helix and V392A, I395A and V399A in the S6-segment are the interaction binding sites of acacetin with hKv4.3 channel. In the third part of my study, I found that acacetin had cardiac protective effect against ischemia/reperfusion injury in isolated rat and guinea pig hearts. Acacetin significantly improved cardiac function recovery, decreased infarct size and prevented ventricular fibrillation induced by regional ischemia/reperfusion in rat hearts with the underlying molecular mechanisms of anti-apoptosis, anti-inflammation and anti-oxidation. The cardiac protection effect from acacetin is independent of its blockade of IKur and Ito channels, since similar cardioprotective effects via the same cellular and molecular mechanisms was also observed in guinea pig hearts, in which no IKur and Ito are expressed. Collectively, my PhD thesis demonstrates the novel information that acacetin blocks both closed and open hKv1.5 channels by binding to their S6 domain, and blocks the open state of hKv4.3 channels by binding to their P-loop filter helix and S6 domain. The use- and rate-dependent blocking effects on hKv1.5 and hKv4.3 channels by acacetin should be beneficial to prevent atrial fibrillation occurrence and reverse atrial fibrillation to sinus rhythm. In addition, my study provides the novel information that acacetin may have cardioprotection against regional ischemia/reperfusion injury.
DegreeDoctor of Philosophy
SubjectFlavonoids - Physiological effect
Reperfusion injury - Animal models
Potassium channels
Ischemia - Animal models
Dept/ProgramMedicine
Persistent Identifierhttp://hdl.handle.net/10722/222801

 

DC FieldValueLanguage
dc.contributor.advisorLi, GR-
dc.contributor.advisorTse, HF-
dc.contributor.authorWu, Huijun-
dc.contributor.author巫慧鈞-
dc.date.accessioned2016-01-29T23:12:49Z-
dc.date.available2016-01-29T23:12:49Z-
dc.date.issued2012-
dc.identifier.citationWu, H. [巫慧鈞]. (2012). Molecular mechanisms of acacetin for the blockade of Kv1.5 and Kv4.3 channels and the cardio-protection against ischemia-reperfusion injury. (Thesis). University of Hong Kong, Pokfulam, Hong Kong SAR. Retrieved from http://dx.doi.org/10.5353/th_b4961749-
dc.identifier.urihttp://hdl.handle.net/10722/222801-
dc.description.abstractAcacetin is a natural flavone compound, initially isolated from the traditional Chinese medicinal herb Tianshanxuelian, and has been recently demonstrated effective in anti-atrial fibrillation by blocking atrial-selective K+ currents, ultra-rapidly delayed rectifier K+ current (IKur), transient outward K+ current (Ito), and acetylcholine-activated K+ current (IKACh). However, the molecular determinants of acacetin blocking IKur and Ito are not fully understood. My PhD project was designed to investigate the cellular and molecular mechanisms of the channel blocking effects using HEK 293 cells expressing cloned hKv1.5 and hKv4.3 channels that encode IKur and Ito, respectively. In addition, I am interested to explore whether acacetin has potential cardioprotective effect against ischemia/reperfusion injury and the related cellular and molecular mechanisms. In the first part of my PhD project, I identified that acacetin blocked hKv1.5 channels in both open and closed states by binding to the S6 segment of the channel. The blockade effect was use- and frequency-dependent. IC50 of acacetin on hKv1.5 current was 3.5, 3.1, 2.9, 2.1 and 1.7 μM at 0.2, 0.5, 1.0, 3.0, and 4.0 Hz respectively. In site-directed mutagenesis studies, mutations in hKv1.5 channel S6-segment, V505A and I508A, remarkably reduced the channel sensitivity to acacetin with IC50 29.5μM for V505A and 19.1 μM for I508A. In the second part of my study, I demonstrated that acacetin blocked hKv4.3 channels by binding to the pore of the channel, decreased the recovery of the channel from inactivation and positively shifted the activation conductance. The blockade of hKv4.3 channels by acacetin was use- and frequency-dependent. IC50 of acacetin for inhibiting hKv4.3 was 7.9, 6.1, 3.9, and 3.2 μM at 0.2, 0.5, 1.0, and 3.3 Hz respectively. Site-directed mutagenesis study revealed that hKv4.3 mutations T366A, T367A in the P-loop helix and V392A, I395A and V399A in the S6-segment are the interaction binding sites of acacetin with hKv4.3 channel. In the third part of my study, I found that acacetin had cardiac protective effect against ischemia/reperfusion injury in isolated rat and guinea pig hearts. Acacetin significantly improved cardiac function recovery, decreased infarct size and prevented ventricular fibrillation induced by regional ischemia/reperfusion in rat hearts with the underlying molecular mechanisms of anti-apoptosis, anti-inflammation and anti-oxidation. The cardiac protection effect from acacetin is independent of its blockade of IKur and Ito channels, since similar cardioprotective effects via the same cellular and molecular mechanisms was also observed in guinea pig hearts, in which no IKur and Ito are expressed. Collectively, my PhD thesis demonstrates the novel information that acacetin blocks both closed and open hKv1.5 channels by binding to their S6 domain, and blocks the open state of hKv4.3 channels by binding to their P-loop filter helix and S6 domain. The use- and rate-dependent blocking effects on hKv1.5 and hKv4.3 channels by acacetin should be beneficial to prevent atrial fibrillation occurrence and reverse atrial fibrillation to sinus rhythm. In addition, my study provides the novel information that acacetin may have cardioprotection against regional ischemia/reperfusion injury.-
dc.languageeng-
dc.publisherThe University of Hong Kong (Pokfulam, Hong Kong)-
dc.relation.ispartofHKU Theses Online (HKUTO)-
dc.rightsThe author retains all proprietary rights, (such as patent rights) and the right to use in future works.-
dc.rightsCreative Commons: Attribution 3.0 Hong Kong License-
dc.subject.lcshFlavonoids - Physiological effect-
dc.subject.lcshReperfusion injury - Animal models-
dc.subject.lcshPotassium channels-
dc.subject.lcshIschemia - Animal models-
dc.titleMolecular mechanisms of acacetin for the blockade of Kv1.5 and Kv4.3 channels and the cardio-protection against ischemia-reperfusion injury-
dc.typePG_Thesis-
dc.identifier.hkulb4961749-
dc.description.thesisnameDoctor of Philosophy-
dc.description.thesislevelDoctoral-
dc.description.thesisdisciplineMedicine-
dc.description.naturepublished_or_final_version-
dc.identifier.doi10.5353/th_b4961749-
dc.date.hkucongregation2013-

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