丹參和葛根為我國民間常用的傳統藥材, 常用於心血管疾病的治療。本試驗主要研究丹參葛根複方(DG, 7:3)對大鼠基底動脈的舒張作用 及腦保護作用。 / 上述所有藥物對U46619預收縮的基底動脈環呈現濃度依賴性的舒張作用。一氧化氮合酶抑制劑L-NAME以及鳥苷酸環化酶抑制劑ODG部分抑制葛根素的舒張作用。在另一組去內皮試驗中, 腺苷酸環化酶抑制劑SQ22536, 鳥苷酸環化酶抑制劑ODG, 大電導鈣離子依賴型鉀通道抑制劑Iberiotoxin以及電壓門控型鉀通道抑制劑4-AP對所有藥物的舒張作用沒有影響。然而， ATP型鉀通道抑制劑格列本脲能夠抑制丹參葛根複方，丹參，葛根，丹參素，葛根素，大豆苷以及大豆苷元的最大舒張反應。內向整流型鉀通道抑制劑氯化鋇則降低丹參酚酸B和大豆苷元的最大反應值。非選擇性鉀通道抑制劑乙基氯化銨以及所有鉀通道抑制劑的混合物顯著抑制上述所有藥物的舒張作用。除了葛根素之外，所有的藥物動度依賴性的抑制氯化鈣所引起的血管收縮。 / 體內研究發現大鼠經歷10分鐘雙側頸總動脈夾閉合並低血壓，及24小時的複灌後，與假手術組動物相比，腦血流量顯著降低，氧化性損傷明顯可見。連續7天口服丹參葛根複方（0.3g/kg 和 3g/kg）, 丹參 (3g/kg)，或者葛根 (3g/kg)對血壓沒有影響。但是，高劑量的丹參葛根複方 (3g/kg) 能夠提高超氧化物歧化酶和過氧化氫酶的活性，抑制丙二醛和一氧化氮的產生。3g/kg的葛根可以提高超氧化物歧化酶的活性，3g/kg的丹參則能抑制一氧化氮的產生。在大鼠中動脈阻塞模型中，連續7天口服丹參葛根複方（3g/kg）能明顯降低腦部的梗死率，同時改善大鼠的神經行為學。 / 總體來說，研究發現丹參葛根複方，丹參，葛根，丹參酚酸B，大豆苷以及大豆苷元的血管舒張作用是通過開平滑肌細胞的通鉀離子通道以及抑制鈣離子內流而實現的。然而葛根素的血管舒張作用是內皮依賴性的，通過產生一氧化氮，開放平滑肌細胞的鉀離子通道而實現的。丹參葛根複方能起到一定的腦保護作用。總而言之，研究表明上述藥物可能會對阻塞性腦血管病的人群有益處。 / Danshen and gegen are used in traditional Chinese medicine for the treatment of cardiovascular diseases. In this study, the relaxant actions of a danshen and gegen formulation (DG; ratio 7:3) and its constituents were investigated on rat-isolated cerebral basilar artery. In addition, the neuroprotective effect of DG was explored in rats subjected to global and focal ischaemia. / DG and all its constituents produced concentration-dependent relaxation of the artery rings precontracted by U46619. Removal of the endothelium had no effect on their vasodilator actions except the maximum response (I[subscript max]) to puerarin was inhibited by 42%. The nitric oxide synthase (NOS) inhibitor L-NAME and guanylyl cyclase (GC) inhibitor ODQ but not the cyclo-oxygenase (COX) inhibitor flurbiprofen produced partial inhibition on the puerarin-induced effect. In a set of endothelium-denuded artery rings, adenylyl cyclase (AC) inhibitor SQ22536, GC inhibitor ODQ, KV channel inhibitor 4-aminopyridine (4-AP) and BK[subscript Ca) channel inhibitor iberiotoxin had no influence on their vasodilator actions. However, pretreatment with K[subscript ATP] channel inhibitor glibenclamide reduced Imax to DG, danshen, gegen, danshensu, puerarin, daidzein and daidzin. K[subscript IR] inhibitor barium chloride (BaCl₂) reduced II[subscript max] to salvianolic acid B and daidzein. The non-selective K⁺ channel inhibitor tetraethylammonium (TEA), or a combination of all the K⁺ channel inhibitors produced significant partial inhibitions on all the agents’ actions. Electrophysiological studies on smooth muscle cells isolated from rat basilar artery also confirmed that DG, danshen, gegen danshensu, puerarin, daidzein and daidzin elevated K[subscript ATP] currents. In addition, DG and all its constituents, except puerarin, produced concentration-dependent inhibition on CaCl₂-induced vasoconstrictions. These findings were confirmed by con-focal microscopy studies. / In vivo study on a rat model of global ischaemia showed that challenging the rats with 10 min bilateral common carotid artery occlusion combined with hypotension, and followed by 24 h reperfusion produced significant decrease in cerebral blood flow and oxidative damage compared to sham-operated animals. Administration of DG (0.3 g/kg and 3 g/kg, p.o.), danshen (3 g/kg, p.o.) or gegen (3 g/kg, p.o.) for 7 days had no effect on blood pressure. However, the 7 days treatment with DG (3 g/kg) restored superoxide dismutase (SOD) and catalase (CAT) activities, suppressed the production of maleic dialdehyde (MDA), and inhibited the production of nitric oxide (NO). In addition, gegen (3 g/kg) restored SOD enzyme activity, whereas, danshen (3 g/kg) inhibited NO production. In addition, treatment with DG (3 g/kg) showed a significant reduction in infarct weight and improved the neurological deficit in a rat model of focal cerebral ischaemia induced by middle cerebral artery occlusion (MCAO). / In conclusion, the vasorelaxant actions of DG, danshen, gegen, salvianolic acid B, danshensu, daidzein and daidzin were found to involve the opening of K⁺ channels and inhibition of Ca²⁺ influx in the vascular smooth muscle cells. In contrast, puerarin produced vasodilatation via an endothelium-dependent mechanism involving NO production and an endothelium-independent pathway mediated by the opening of K⁺ channels. DG may have some cerebro-protective effects. Overall, the present studies showed that DG and its constituents could be beneficial to patients with obstructive cerebrovascular diseases. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Deng, Yan. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2012. / Includes bibliographical references (leaves 164-178). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstract also in Chinese. / ABSTRACT --- p.v / 摘要 --- p.iviii / ACKNOWLEDGEMENTS --- p.x / PUBLICATIONS BASED ON THIS THESIS --- p.xii / ABBREVIATIONS --- p.xiv / Chapter CHAPTER 1 --- Introduction --- p.1 / Chapter 1.1 --- Chinese Medicines in treatment of cerebrovascular diseases --- p.2 / Chapter 1.2 --- Danshen --- p.4 / Chapter 1.2.1 --- Chemical constituents --- p.4 / Chapter 1.2.1.1 --- Hydrophilic compounds of danshen --- p.4 / Chapter 1.2.1.2 --- Lipophilic compounds of danshen --- p.5 / Chapter 1.2.1.3 --- Other compounds --- p.5 / Chapter 1.2.2 --- Pharmacological activities --- p.5 / Chapter 1.2.2.1 --- Vascular protection --- p.5 / Chapter 1.2.2.2 --- Anti-tumour --- p.7 / Chapter 1.2.2.3 --- Treatment of liver diseases --- p.8 / Chapter 1.2.2.4 --- Treatment of drug addiction --- p.9 / Chapter 1.2.2.5 --- Treatment of kidney diseases --- p.10 / Chapter 1.2.3 --- Pharmacokinetics --- p.10 / Chapter 1.3 --- Gegen --- p.12 / Chapter 1.3.1 --- Chemical constituents --- p.12 / Chapter 1.3.2 --- Pharmacology --- p.13 / Chapter 1.3.2.1 --- Vascular effects --- p.13 / Chapter 1.3.2.2 --- Anti-diabetes --- p.14 / Chapter 1.3.2.3 --- Anti-hypercholesterolaemia --- p.15 / Chapter 1.3.2.4 --- Anti-inflammation --- p.16 / Chapter 1.3.2.5 --- Anti-platelet aggregation --- p.17 / Chapter 1.3.3 --- Pharmacokinetics --- p.17 / Chapter 1.4 --- Danshen and gegen formulation --- p.19 / Chapter 1.5 --- Mechanisms of vasodilatation --- p.22 / Chapter 1.5.1 --- Endothelium derived relaxant factors (EDRFs) --- p.22 / Chapter 1.5.1.1 --- Nitric oxide (NO) --- p.22 / Chapter 1.5.1.2 --- Prostacyclin (PGI₂) --- p.23 / Chapter 1.5.1.3 --- Endothelium-derived hyperpolarizating factors (EDHFs)- --- p.23 / Chapter 1.5.2 --- Signal transduction pathways --- p.24 / Chapter 1.5.2.1 --- Guanylyl cyclase-cGMP pathway --- p.24 / Chapter 1.5.2.2 --- Adenylyl cyclase-cAMP pathway --- p.24 / Chapter 1.5.3 --- Ion channels --- p.25 / Chapter 1.5.3.1 --- Potassium channels (K⁺ channels) --- p.25 / Chapter 1.5.3.2 --- Calcium channel (Ca²⁺ channels) --- p.25 / Chapter 1.6 --- Aims of study --- p.27 / Chapter CHAPTER 2 --- Materials and method --- p.28 / Chapter 2.1 --- Herbal preparation --- p.28 / Chapter 2.1.1 --- DG, danshen and gegen preparation --- p.28 / Chapter 2.1.2 --- Identification and quantification of chemical markers in DG water extract --- p.29 / Chapter 2.2 --- Experiments on rat basilar artery --- p.30 / Chapter 2.2.1 --- Animals --- p.30 / Chapter 2.2.2 --- Chemicals --- p.30 / Chapter 2.2.3 --- Isolation and mounting of blood vessels --- p.33 / Chapter 2.2.4 --- Protocols --- p.34 / Chapter 2.2.4.1 --- Effects on U46619-precontracted tone --- p.34 / Chapter 2.2.4.2 --- Endothelium-dependent mechanism --- p.34 / Chapter 2.2.4.3 --- Endothelium-independent mechanism --- p.35 / Chapter 2.2.4.4 --- Calcium channels --- p.36 / Chapter 2.2.4.5 --- Positive control --- p.36 / Chapter 2.2.5 --- Statistical analysis --- p.37 / Chapter 2.3 --- Experiments on rat cerebral basilar artery smooth muscle cells K[subscript ATP] channals --- p.38 / Chapter 2.3.1 --- Animals --- p.38 / Chapter 2.3.2 --- Chemicals --- p.38 / Chapter 2.3.3 --- Isolation of rat cerebral vascular smooth muscle cells --- p.40 / Chapter 2.3.4 --- Whole cell patch-clamp electrophysiology --- p.40 / Chapter 2.3.5 --- Statistical analysis --- p.44 / Chapter 2.4 --- Experiments on rat cerebral basilar artery smooth muscle cells calcium channels --- p.45 / Chapter 2.4.1 --- Animals --- p.45 / Chapter 2.4.2 --- Chemicals --- p.45 / Chapter 2.4.3 --- Isolation of rat cerebral vascular smooth muscle cells --- p.47 / Chapter 2.4.4 --- Dye loading and determination of [Ca²⁺]i --- p.47 / Chapter 2.4.5 --- Statistical analysis --- p.48 / Chapter 2.5 --- In vivo study of global ischaemia --- p.49 / Chapter 2.5.1 --- Animals --- p.49 / Chapter 2.5.2 --- Drugs and chemicals --- p.49 / Chapter 2.5.3 --- Experimental protocols for global ischaemia --- p.49 / Chapter 2.5.4 --- Induction of global ischaemia --- p.50 / Chapter 2.5.5 --- Blood pressure measurement --- p.52 / Chapter 2.5.6 --- Measurement of cerebral blood flow --- p.52 / Chapter 2.5.7 --- Biochemical assessment --- p.53 / Chapter 2.5.7.1. --- Dissection and homogenization --- p.53 / Chapter 2.5.7.2. --- Measurement of malondialdehyde (MDA) --- p.53 / Chapter 2.5.7.3. --- Estimation of nitrite --- p.53 / Chapter 2.5.7.4 --- Superoxide dismutase activity (SOD) --- p.54 / Chapter 2.5.7.5 --- Reduced glutathione (GSH) --- p.54 / Chapter 2.5.7.6 --- Catalase (CAT) --- p.55 / Chapter 2.5.7.7 --- NOS activity --- p.55 / Chapter 2.5.7.8 --- Protein --- p.56 / Chapter 2.5.8 --- Statistical analysis --- p.56 / Chapter 2.6 --- In vivo study of focal ischaemia --- p.57 / Chapter 2.6.1 --- Animals --- p.57 / Chapter 2.6.2 --- Drugs and chemicals --- p.57 / Chapter 2.6.3 --- Experimental protocols for global ischaemia --- p.57 / Chapter 2.6.4 --- Focal cerebral ischaemia-reperfusion model --- p.57 / Chapter 2.6.5 --- Assessment of neurobehavioural changes --- p.59 / Chapter 2.6.6 --- Assessment of cerebral infarction --- p.60 / Chapter 2.6.7 --- Statistical analysis --- p.60 / Chapter CHAPTER 3 --- Results --- p.61 / Chapter 3.1 --- Identification and quantification of chemical markers in DG water extract --- p.61 / Chapter 3.2 --- Effects of DG and its constituents on rat cerebral basilar artery --- p.64 / Chapter 3.2.1 --- Investigations on endothelium-dependent mechanisms --- p.64 / Chapter 3.2.2 --- Investigations on endothelium-independent mechanisms --- p.68 / Chapter 3.2.3 --- Positive control --- p.86 / Chapter 3.2.3 --- Investigations on calcium channels --- p.88 / Chapter 3.3 --- Effects of DG and its constituents on rat cerebral basilar artery smooth muscle cells --- p.91 / Chapter 3.3.1 --- Effects of water crude-extracts of DG, danshen, and gegen on K[subscript ATP] channels --- p.91 / Chapter 3.3.2 --- Effects of active constituents of danshen hydrophilic fraction on K[subscript ATP] channels --- p.100 / Chapter 3.3.3 --- Effects of the major isoflavonoids of gegen on K[subscript ATP] channels --- p.105 / Chapter 3.4 --- Effects of DG and its constituents on calcium channels of basilar artery smooth muscle cells --- p.112 / Chapter 3.5 --- Effects of DG, danshen and gegen on rat global ischaemia --- p.117 / Chapter 3.5.1 --- Effects of DG, danshen and gegen on rats’ blood pressure and cerebral blood flow --- p.117 / Chapter 3.5.2 --- Effects of DG, danshen and gegen on lipid peroxidation --- p.120 / Chapter 3.5.3 --- Effects of DG, danshen and gegen on SOD activity --- p.120 / Chapter 3.5.4 --- Effects of DG, danshen and gegen on CAT activity --- p.120 / Chapter 3.5.5 --- Effects of DG, danshen and gegen on reduced GSH level --- p.121 / Chapter 3.5.6 --- Effects of DG, danshen and gegen on NOS system --- p.126 / Chapter 3.6 --- Effect of DG on rat focal ischaemia --- p.129 / Chapter 3.6.1 --- Effect of DG on cerebral infarction --- p.129 / Chapter 3.6.2 --- Effect of DG on neurological deficits --- p.129 / Chapter CHAPTER 4 --- Discussion --- p.132 / Chapter 4.1 --- Studies of DG and its constituents on rat cerebral basilar artery --- p.133 / Chapter 4.1.1 --- Constituents of DG on U46619-precontracted tone --- p.133 / Chapter 4.1.2 --- Investigations on endothelium-dependent mechanisms --- p.133 / Chapter 4.1.3 --- Investigations on endothelium-independent mechanisms --- p.136 / Chapter 4.1.4 --- Investigations on calcium channels --- p.139 / Chapter 4.2 --- Effects of DG and its constituents on rat cerebral basilar artery smooth muscle cell K[subscript ATP] channels --- p.143 / Chapter 4.3 --- Effects of DG and its constituents on calcium influx in rat basilar artery smooth muscle cells --- p.147 / Chapter 4.4 --- Effects of DG, danshen and gegen on rat transient global ischaemia --- p.150 / Chapter 4.4.1 --- Effects of DG, danshen and gegen on rats’ blood pressure and cerebral blood flow --- p.150 / Chapter 4.4.2 --- Effects of DG, danshen and gegen on lipid peroxidation, SOD and CAT activity, and GSH level --- p.152 / Chapter 4.4.3 --- Effects of DG, danshen and gegen on NOS system --- p.155 / Chapter 4.5 --- Effects of DG on rat focal ischaemia --- p.157 / Chapter 4.6 --- Further studies --- p.160 / Chapter 4.7 --- Conclusion --- p.162 / REFERENCES --- p.164

Background. Danshen, the dried root of Salvia miltiorrhiza Bge, is used for treating coronary heart disease. In China, numerous pharmaceutical dosage forms of Danshen are commercially available. Although the pharmacological effects of different components of Danshen are well identified, its absorption as well as pharmacokinetics studies are still insufficient and inconsistent. The current study aims to: (1) screen for the major absorbable active components of Danshen; (2) interpret the absorption mechanism and pharmacokinetics characteristics of the identified components; (3) develop an improved oral drug delivery system for the identified components of Danshen. / Conclusion. Both danshensu and SAB have limited intestinal permeability and oral bioavailabilities. Our results demonstrated the usefulness of sodium caprate as a potential absorption enhancer for danshensu and SAB in Danshen product. / Methods. Six major active components in commercially available Danshen products were identified and quantified. In vitro human Caco-2 cell monolayer model, rat in situ intestinal perfusion model as well as rat in vivo pharmacokinetic model were used to investigate the intestinal absorption and pharmacokinetics profiles of the identified Danshen components. Effect of the absorption enhancer on the oral absorption and bioavailabilities of the studied Danshen components was further evaluated. / Results. Danshensu, salvianolic acid B (SAB) and protocatechuic aldehyde (PCA) were identified as the major components in Danshen products. Investigations using in vitro, in situ and in vivo model found that both danshensu and SAB had poor permeabilities and low bioavailabilities (Danshensu: 11.09%; SAB: 3.90%), which may be due to their absorption via the paracellular transport pathways. Studies of PCA suggested that it may have a intestinal first pass metabolism with an oral bioavailability of only 18.02%. It was found that the permeabilities of both danshensu and SAB were significantly increased upon addition of sodium caprate, a paracellular absorption enhancer. The oral bioavailabilities of both danshensu and SAB in pure compound form as well as Danshen extract form were also increased in the presence of sodium caprate in rats. / Zhou, Limin. / Advisers: Zuo Zhong; Moses S.S. Chow. / Source: Dissertation Abstracts International, Volume: 70-06, Section: B, page: 3457. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2008. / Includes bibliographical references. / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Electronic reproduction. [Ann Arbor, MI] : ProQuest Information and Learning, [200-] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstracts in English and Chinese. / School code: 1307.

Danshen, the dried root of Salvia miltiorrhiza Bunge, is a herb listed in the Chinese Pharmacopoeia for the treatment of cardiovascular and cerebrovascular diseases. Danshen has been reported to have antiplatelet, cardioprotective, anti-inflammatory, hepatoprotective, and anti-HIV effects in preclinical studies. However, exaggerated anticoagulation and bleeding complications have also been observed during concurrent use of Danshen and warfarin in patients, although the mechanism(s) of the herb-drug interaction, pharmacodynamic and/or pharmacokinetic interactions, remained uncertain. Characterization of the cytochrome P450 isoforms responsible for the metabolism(s) of drugs and herbal constituents is important for the identification of potential drug-drug or drug-herb interactions. The present study investigated the effects of Danshen on the metabolism of probe substrates of specific CYP isoforms including CYP1A2, CYP3A and CYP2C9, the isoforms that are responsible for the metabolism of warfarin to assess the potential interactions of Danshen with drugs that utilize these isoforms for their biotransformation. / Firstly, the effects of Danshen and its tanshinone components on CYP1A2 activity were investigated in vitro and in vivo in the rat. Formulated Danshen extract, the ethanolic extract and the aqueous extract from Danshen root, and individual tanshinones inhibited phenacetin O-deethylation (CYP1A2) activity in vitro. Enzyme kinetic studies showed that tanshinone I, tanshinone IIA, cryptotanshinone, and dihydrotanshinone were competitive CYP1A2 inhibitors. Acute, sub-chronic and chronic pretreatments of formulated Danshen extract decreased the clearance (CL) of caffeine, with a concomitant increase in the area under concentration-time curve (AUC), and prolongation of the plasma half-life (T 1/2). These results suggested that Danshen inhibited rat CYP1A2 activity and altered the pharmacokinetics of the CYP1A2 probe substrates in vivo. / In conclusion, these results confirmed that Danshen-inhibited CYP activity, especially CYP1A2, then CYP2C9/11 (CYP2C9 in human, CYP2C11 in rats) in vitro. In vivo studies confirmed the clearance of the probe substrates was also decreased when co-administered with Danshen. Given that CYP1A2, CYP2C9 and CYP3A4 are responsible for the metabolism and disposition of a large number of drugs currently used in man, the concomitant use of Danshen with drugs which are substrates of CYP1A2, 2C9 and 3A4, especially CYP1A2, must be met with great caution. / Secondly, the effects of Danshen and its tanshinone components on CYP3A activity were investigated in vitro and in vivo in the rat. Formulated Danshen extract, the ethanolic extract from Danshen root, and tanshinones inhibited testosterone 6beta-hydroxylation (CYP3A) activity in vitro. Enzyme kinetic studies showed that tanshinone I, tanshinone IIA, and cryptotanshinone were competitive CYP3A inhibitors, whereas dihydrotanshinone was a noncompetitive CYP3A inhibitor. In vivo studies showed the pretreatments of formulated Danshen extract did not significantly change the pharmacokinetics of midazolam. / The effects of Danshen and its tanshinones on human CYP1A2 (phenacetin O-deethylation), CYP3A4 (testosterone 6beta-hydroxylation), and CYP2C9 (tolbutamide 4-hydroxylation) activities were also investigated in vitro using pooled human liver microsomes and human CYP isoforms. The ethanolic fraction of Danshen root was more effective than water-soluble fraction in inhibiting human CYP1A2, CYP3A4 and CYP2C9 activities. Enzyme kinetic studies showed that tanshinone I, tanshinone IIA, and cryptotanshinone were competitive inhibitors of CYP1A2, CYP3A4 and CYP2C9 with varying effectiveness. Dihydrotanshinone was not a competitive inhibitor of CYP1A2 and CYP2C9, but a noncompetitive CYP3A4 inhibitor. CYP1A2 was most affected and CYP3A4 was least affected by Danshen and tanshinones. Compared with the results obtained from rat and human, rat is a good animal model for predicting Danshen-drug interactions in humans, especially drugs which are substrates of CYP1A2. / Thirdly, the effects of Danshen and its tanshinone components on CYP2C11 activity were investigated in vitro and in vivo in the rat. Formulated Danshen extract, the ethanolic extract from Danshen root, and tanshinones inhibited testosterone 2alpha-hydroxylation (CYP2C11) activity in vitro. Enzyme kinetic studies showed that tanshinone I, tanshinone IIA, cryptotanshinone, and dihydrotanshinone were competitive CYP2C11 inhibitors. Sub-chronic pretreatment of formulated Danshen extract increased the AUC, T1/2 but decreased CL of tolbutamide. These results suggested that Danshen inhibited the CYP2C activity in the rat. In conclusion, these results confirmed the possible mechanism is enzyme inhibition, involved in the interaction of Danshen and warfarin previously observed in rats. / Wang, Xin. / "September 2007." / Source: Dissertation Abstracts International, Volume: 69-08, Section: B, page: 4699. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2007. / Includes bibliographical references (p. 284-302). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Electronic reproduction. [Ann Arbor, MI] : ProQuest Information and Learning, [200-] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstract in English and Chinese. / School code: 1307.

丹参是一种著名的传统中药，富含丹酚酸和丹参酮。其中，丹参酮的潜在抗肿瘤作用近年来引起众多关注。本研究评价了主要的丹参酮及其衍生物对结肠癌细胞的细胞毒性。结果显示DHTS具有最强的抗结肠癌活性和显著的肿瘤特异性细胞毒性，其细胞毒性主要由于凋亡诱导而不是引起坏死。初步的构效关系分析提示丹参酮母环结构中的A环和B环增加的离域性有助于提高其对结肠癌细胞的细胞毒性，而非二维结构及较小的D环也是进行结构改造的可能方向。 / 基于以上发现，本研究进一步探讨了DHTS的体内外抗肿瘤活性及内在机制。本研究发现DHTS的促凋亡活性并不依赖于p53的表达，而依赖于caspase活性及线粒体介导的细胞质中氧自由基 ROS及钙离子的聚集。DHTS可引起浓度及时间依赖caspase-9/-3/-7的活化而并未显著引起caspase-8的活化，这一现象发生于同样以浓度及时间依赖方式进行的线粒体中cytochrome c及AIF转位之后。在DHTS诱导的结肠癌细胞凋亡中，cytochrome c及caspase介导的凋亡通路及AIF介导的凋亡通路均被激活并显示出两条通路之间的交叉调控。 / 此外，线粒体在DHTS的促凋亡活性中的作用在本研究中被深入探讨。本研究发现线粒体可能是DHTS的一个直接靶点, 而氧化磷酸化复合体III则更可能是其作用位点。DHTS可以引起迅速而明显的线粒体功能障碍，随之引起细胞质中大量的氧自由基及钙离子聚集，诱导凋亡的产生。 / 与体外结果一致，本研究证实了DHTS对免疫缺陷小鼠中的结肠癌移植廇也具有明显的抗肿瘤作用。与溶媒对照组比较，DHTS治疗组中移植廇的增长显著被减缓，在治疗终点时的廇体积与重量也显著被降低。TUNEL检测确认DHTS诱导移植廇中癌细胞的显著凋亡。免疫荧光检测也发现DHTS诱导caspase-3及caspase-7在移植廇中癌细胞的明显活化。 / 综上所述，本研究提供了丹参酮抗结肠癌活性的一些初步构效关系的信息，为提高丹参酮抗结肠癌活性的结构改造提供一定的参考。更重要的是，本研究证明了DHTS的体内外抗结肠癌活性并探讨了其作用机制及可能靶点，为DHTS作为新的应用于抗结肠癌药物或辅助治疗用药提供了临床前研究证据。 / Salvia miltiorrhiza Bunge, also known as Danshen, rich in phenolic acid and tanshinones, has been widely used to treat various kinds of diseases including heart diseases and hepatitis in China with minimal side effects. Among the tanshinones, tanshinone I, tanshinone IIA, cryptotanshinone and dihydrotanshinone I are the major bioactive constituents in this herb. In this study, the anti-colon cancer potential of five tanshinones and six derivatives of tanshinone IIA were evaluated in several colon cancer cell lines. It was found that apoptosis but not necrosis contributed significantly to the cytotoxicity of the tanshinones. Dihydrotanshinone I (DHTS) was confirmed to be the most potent and selective anti-cancer compound among the tanshinones tested in this study. Preliminary SAR (structure activity relationship) of tanshinones reveals that the increase of delocalizability of A and B rings in the chemical structure of the tanshinones enhances their cytotoxicity on cancer cells, while compounds with a non-planar and small sized D ring region are better choices for anti-cancer effect. / The underlying mechanisms of the anti-colon cancer activity of DHTS were further studied. It was found that apoptosis induced by DHTS was p53 independent but caspase dependent, which was closely related to intracellular accumulation of ROS (reactive oxidant stress) and calcium mediated by mitochondria. A concentration- and time-dependent activation of caspase-9,-3,-7 but not caspase-8 by DHTS in HCT116 cells was detected after the translocation of cytochrome c and AIF (apoptosis inducing factor) from mitochondria. In this process, the crosstalk between the caspase-dependent and caspase-independent pathways was firstly shown in the apoptotic mechanism of DHTS. To this end, the release of cytochrome c happened first and the translocation of apoptosis inducing factor (AIF) was prevented by a pan caspase inhibitor. In the meantime, the release of cytochrome c and activation of caspase-9 and PARP (poly-ADP-ribose polymerase) cleavage were decreased after AIF knockdown. Especially, mitochondrion was suggested to be the direct target of DHTS and OXPHOS complex III but not OXPHOS complex I was probably the acting site of DHTS. / In accordance with the results obtained in vitro, the potential anti-colon cancer activity of DHTS was also observed in nude mice with xenograft tumors and the compound did not produce any observable systemic toxicity. DHTS efficiently delayed tumor growth by decreasing the tumor size and weight through the induction of apoptosis in cancer cells but not by inhibition of cell proliferation. In the same tissues, a distinct activation of caspase-3 and caspase-7 in tumor cells was also detected by immunofluorescence assay. / Collectively, the present study provides preliminary information about the SAR of the anti-colon cancer activity for tanshinones. It also confirms that DHTS is a promising compound for anti-cancer action both in vitro and in vivo. In addition, this study gives us a better understanding regarding the mechanisms of how DHTS induces apoptosis in cancer cells. All these findings could provide solid pre-clinical evidence to propel the development and application of DHTS and perhaps its derivatives as novel therapeutic or adjuvant agents for the treatment of colon cancer. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Wang, Lin. / Thesis (Ph.D.) Chinese University of Hong Kong, 2013. / Includes bibliographical references (leaves 122-132). / Abstracts also in Chinese. / Wang, Lin.

Cheung, Wing Shing David. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2013. / Includes bibliographical references (leaves 201-217). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstract also in Chinese.

Chan Yin Ling. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2006. / Includes bibliographical references (leaves 147-167). / Abstracts in English and Chinese. / ABSTRACT --- p.III / ACKNOWLEDGEMENT --- p.X / TABLE OF CONTENTS --- p.XI / ABBREVIATIONS --- p.XV / LIST OF FIGURES --- p.XVII / LIST OF TABLES --- p.XXI / Chapter CHAPTER 1 --- INTRODUCTION --- p.1 / Chapter 1.1 --- Introduction to Cardiovascular Disease and Atherosclerosis --- p.1 / Chapter 1.1.1 --- Cardiovascular Disease --- p.1 / Chapter 1.1.2 --- A therosclerosis --- p.3 / Chapter 1.1.2.1 --- Structure of Arteries --- p.4 / Chapter 1.1.2.2 --- Pathophysiology of Atherosclerosis --- p.5 / Chapter 1.1.2.3 --- Endothelial Dysfunction --- p.8 / Chapter 1.1.3 --- Current Western Therapies --- p.11 / Chapter 1.1.3.1 --- Surgery --- p.11 / Chapter 1.1.3.2 --- Western Medications --- p.13 / Chapter 1.1.4 --- Traditional Chinese Medicine --- p.17 / Chapter 1.1.4.1 --- Long History --- p.17 / Chapter 1.1.4.2 --- As Alternative Medicine --- p.18 / Chapter 1.1.4.3 --- Modernization of Chinese Medicine --- p.19 / Chapter 1.2 --- Introduction and Selection of Chinese Medicine --- p.20 / Chapter 1.2.1 --- Selection ofTCM Formulation from Pharmacopoeia --- p.20 / Chapter 1.2.1.1 --- Compound Formulation --- p.20 / Chapter 1.2.2 --- Introduction to the Herbal Medicines --- p.21 / Chapter 1.2.2.1 --- Danshen (Salvia miltiorrhiza) --- p.21 / Chapter 1.2.2.2 --- Gegen (Puerariae thomsonii and Puerariae lobata) --- p.22 / Chapter 1.2.2.3 --- Yanhu (Corydalis yanhusuo) and its Exclusion --- p.24 / Chapter 1.2.3 --- Source and Authentication of the Herbal Medicines --- p.25 / Chapter CHAPTER 2 --- OPTIMIZATION OF DANSHEN-GEGEN FORMULA --- p.26 / Chapter 2.1 --- Project History --- p.26 / Chapter 2.2 --- aims for the present study --- p.27 / Chapter 2.3 --- Methods and Materials --- p.30 / Chapter 2.3.1 --- Extracts --- p.30 / Chapter 2.3.2 --- Extraction Process --- p.31 / Chapter 2.3.3 --- In vitro Antioxidation Model --- p.33 / Chapter 2.3.4 --- Ex vivo Vasodilation Model --- p.35 / Chapter 2.3.5 --- Statistical Analysis --- p.38 / Chapter 2.4 --- Results --- p.39 / Chapter 2.4.1 --- Vasodilation Results --- p.39 / Chapter 2.4.2 --- Antioxidation Results --- p.43 / Chapter 2.5 --- Discussion --- p.46 / Chapter 2.6 --- Further Modification of the Formula --- p.49 / Chapter 2.6.1 --- Extracts --- p.49 / Chapter 2.6.2 --- Results --- p.49 / Chapter 2.7 --- discussion --- p.52 / Chapter CHAPTER 3 --- MARKER CHEMICAL CONTENTS OF HERBAL EXTRACTS AND THEIR PHARMACOLOGICAL PROPERTIES --- p.56 / Chapter 3.1 --- HPLC Analysis of Marker Contents --- p.56 / Chapter 3.1.1 --- Methods --- p.57 / Chapter 3.1.2 --- Results --- p.58 / Chapter 3.1.2.1 --- HPLC Chromatograms --- p.59 / Chapter 3.1.2.2 --- Content Percentage of Marker Compounds --- p.63 / Chapter 3.1.3 --- Discussion --- p.64 / Chapter 3.2 --- Studies on Marker Compounds --- p.65 / Chapter 3.2.1 --- Introduction --- p.65 / Chapter 3.2.2 --- Methods and Materials --- p.67 / Chapter 3.2.2.1 --- Source of Pure Compounds --- p.67 / Chapter 3.2.2.2 --- Purification and Identification of SAB --- p.68 / Chapter 3.2.2.3 --- Vasodilation model --- p.70 / Chapter 3.2.2.4 --- Antioxidation Model --- p.71 / Chapter 3.2.2.5 --- Structures of Pure Compounds --- p.72 / Chapter 3.2.3 --- Results --- p.73 / Chapter 3.2.3.1 --- Vasodilation Results --- p.73 / Chapter 3.2.3.2 --- Antioxidation Results --- p.76 / Chapter 3.3 --- Discussion --- p.79 / Chapter 3.4 --- Synergistic Effect Study --- p.85 / Chapter 3.4.1 --- Introduction --- p.85 / Chapter 3.4.2 --- Methods --- p.85 / Chapter 3.4.3 --- Results --- p.86 / Chapter 3.4.4 --- Discussion --- p.88 / Chapter 3.5 --- STUDY ON 3'-HYDROXYPlIERARIN AND 3'-METHOXYPUERARIN PURIFIED FROM YFGE --- p.90 / Chapter 3.5.1 --- 3 '-hydroxypuerarin and 3'-methoxypuerarin --- p.90 / Chapter 3.5.2 --- Methods and Materials --- p.91 / Chapter 3.5.2.1 --- Purification by HPLC semi-preparation --- p.91 / Chapter 3.5.2.2 --- Bioassays --- p.93 / Chapter 3.5.3 --- Results --- p.94 / Chapter 3.5.3.1 --- Vasodilation Study --- p.94 / Chapter 3.5.3.2 --- Antioxidative Effect of Yege --- p.95 / Chapter 3.5.4 --- Discussion / Chapter CHAPTER 4 --- MECHANISTIC STUDY --- p.98 / Chapter 4.1 --- Introduction --- p.98 / Chapter 4.1.1 --- Nitric Oxide-mediated Vasodilation --- p.99 / Chapter 4.1.2 --- Prostacyclin-mediated Vasodilation --- p.100 / Chapter 4.1.3 --- EDHF-mediated Vasodilation --- p.101 / Chapter 4.1.4 --- Endothelium-dependent and -independent Vasodilations --- p.103 / Chapter 4.2 --- Methods and Materials --- p.104 / Chapter 4.3 --- Results --- p.107 / Chapter 4.3.1 --- Danshen-Gegen Formula (DY80) --- p.107 / Chapter 4.3.2 --- Salvianolic acid B --- p.112 / Chapter 4.3.3 --- Daidzein --- p.117 / Chapter 4.4 --- Discussion --- p.121 / Chapter CHAPTER 5 --- STUDY ON LIPID PEROXIDATION AND UPTAKE BY MACROPHAGES --- p.128 / Chapter 5.1 --- Study of DY 80 and SAB on Copper-ion induced Low Density Lipoprotein Oxidation --- p.128 / Chapter 5.1.1 --- Pathologic Role of oxidized Low Density Lipoprotein --- p.128 / Chapter 5.1.2 --- Antioxidants in Low Density Lipoprotein and Role of Transition Metals --- p.129 / Chapter 5.1.3 --- Methods and Materials --- p.130 / Chapter 5.1.4 --- Results --- p.131 / Chapter 5.1.5 --- Discussion --- p.133 / Chapter 5.2 --- Study of Scavenger Receptor Regulation in Macrophages --- p.135 / Chapter 5.2.1 --- Introduction --- p.135 / Chapter 5.2.2 --- Methods and Materials --- p.136 / Chapter 5.2.3 --- Results --- p.139 / Chapter 5.2.4 --- Discussions --- p.140 / Chapter CHAPTER 6 --- General Discussion --- p.143 / REFERENCES --- p.147

心臟病目前仍然是最普遍的威脅人類生命安全的三大病因之一。同西藥相比, 傳統中醫藥具有多靶點，協同作用及小副作用等特性。在中藥歷史中， 丹參和葛根這兩種草藥經常出現在中藥方劑用於治療心血管相關的疾病，已有幾千年的歷史。 我們實驗室發現了一個丹參葛根湯劑具有保護動脈粥樣硬化病人心臟功能的作用，並且可以使收縮的大鼠大動脈舒張的作用。 本研究主要通過舒張豬冠狀動脈，提高大鼠對抗過氧化和離子擾動能力以及提高血管增生四個方面探討丹參葛根複方水提物 (質量比7:3) (DG配方)對血管的作用以提供其治療心血管疾病的藥理基礎。 / 在本研究的第一部， 我們主要探討了DG配方對缺血再灌注損傷的心臟及其心肌細胞的保護作用。我們發現DG配方明顯抑制了心臟損傷相關的肌酸激酶和乳酸脫氫的釋放。同時DG配方顯著促進了再灌注後冠狀動脈內血流量速度和收縮力度的恢復。這些結果說明DG配方可以保護缺血再灌注心臟並且有效促進其功能恢復。我們還觀察了長期給大鼠用DG配方14天後其心臟在缺血再灌注中的表現。類似於再灌注時給藥的結果，DG配方同樣抑制了損傷酶的釋放並且有效促進了冠狀動脈內血流量速度和收縮力度的恢復。 / 同時，在缺氧再灌注離體細胞模型中，我們發現DG配方明顯抑制了缺氧再灌注損傷帶來的細胞死亡。流式細胞儀分析結果表明，藥物處理組中的凋亡類的細胞明顯比對照組中少主要通過抑制促凋亡的caspase3表達明以及促進抗凋亡的Bcl2表達升高。DG配方減少了心肌細胞內細胞色素c從線粒體中釋放明顯以及抑制了線粒體去極化。這說明DG配方也保護了線粒體的膜的完整性，從而確保線粒體功能進而保證細胞的能量系統穩定。最有意思的是DG配方可以直接抑制缺氧再灌注相關的兩條通路, 它不僅抑制活性氧化物質的釋放, 同時也抑制了再灌注後鈣離子的累積。總之，DG配方以抗氧化和抗離子擾動的方式保護了在缺血缺氧再灌注損傷中心臟和心肌細胞的結構和功能。 / 第二部分的研究是關於DG配方對從豬心臟上分離的左冠狀動脈前室間支 (左前降支) 血管的作用及其內在的機制，我們的結果表明對由U46619引起的冠狀動脈血管收縮DG配方表現了濃度依賴的舒血管作用。而該作用並非依賴于內皮細胞及其釋放的舒張血管因數一氧化氮和前列腺素類似物環素和大部分的鉀離子通道。其中只有內向整合鉀離子通道部分參與了舒血管的過程。肌球蛋白輕鏈的磷酸化明顯被DG配方抑制，但是RhoA 的活性並沒有受其影響。鈣離子引發的血管收縮則被DG配方濃度依賴性的受到抑制。這部分的研究證明瞭DG配方主要通過類似鈣離子通道拮抗劑作用抑制鈣離子進入到血管平滑肌細胞減少肌球蛋白磷酸化達到舒張血管的作用。結果說明DG配方可以作為一種安全的藥物用於治療心血管疾病特別是高血壓和心絞痛。 / 本研究的第三部分是關於DG配方的促血管增生的作用。我們發現DG配方可以明顯促進斑馬魚的腸下動脈的出芽並且促進血管增生相關基因的表達，血管內皮細胞生長因數及其受體的mRNA表達。內皮細胞是血管增生的基礎。所以我們利用人源微血管內皮細胞檢測了DG配方在細胞的增生，遷移，分化和形成血管方面的影響以解釋它在斑馬魚中促進血管增生的作用機理。結果發現，DG配方明顯促進了該種內皮細胞的增殖，遷移和形成管狀結構。 / 綜上所述，DG配方可以通過舒張血管，抗氧化，抗離子紊亂和促進血管增生提供心血管保護功能。DG配方通過螯合活性氧化物質和抑制鈣離子的累積保護了因缺血再灌注引起的心臟損傷，說明DG配方可以作為手術的輔助藥物減少心臟病人在缺血再灌注過程中受到的損傷。它以拮抗L型鈣離子通道方式減少鈣離子進入到血管平滑肌細胞來舒張收縮的冠狀動脈血管。說明DG配方可以用於治療高血壓和心絞痛等心臟病。另外DG配方也可以促進血管增生，可用于心肌梗死病人促進其心臟血管系統重建，本研究對於未來臨床實驗具有重要的參考價值。 / Coronary heart diseases (CHD) are one of the most prevalent causes of premature death all over the world. In contrast to western medicine, traditional Chinese medicine (TCM) has shown the benefit of multi-targeting and synergism to treat CHD. Two kinds of Chinese herbs, Danshen (Radix Salviae Miltiorrhiza) (D) and Gegen (Radix Puerariae Lobatae) (G) always present on the TCM formula for treating heart disease. We found a useful formula of Danshen and Gegen decoction with weight ratio of 7:3 (DG) exerting properties of improving the heart function in patient with atheroslcerosis and providing vasodiation and antioxidant protection on the rat cardiovascular system. The present study was designed to evaluate the effects of DG on the vascular activity by its properties on antioxidant and anti-ion stunning to inhibiting the ischemia and reperfusion injury, vasodilation effect on pig coronary artery and angiogenesis effect on zebrafish model. / In the first part of the study, we explored protective effect of DG on rat hearts and cardiomyocytes after ischemia-reperfusion and hypoxia-reoxygenation injury. Comparing to control group, the release of creatine kinase (CK) and lactate dehydrogenase (LDH) significantly decreased in the DG treated groups in a dose-dependent manner. The recovery percentage of coronary flow and contractile force in the DG was higher than that in the control group. These results suggested that DG dose-dependently improved the heart function after ischemia and reperfusion injury in a dose-dependent manner. We also examined chronic effect of DG (14 days pretreatment) on rat heart with ischemia and reperfusion injury. DG induced rat heart with high potential to deal with I/R injury, less damaged enzymes release and high recovery percentage of heart function recovery. / In the cell hypoxia and reoxygenation model, DG significantly inhibited the cell death after H/R treatment with bcl2 expression increase and caspase3 expression decrease. DG also reversed the H/R-induced mitochondrial depolarization and inhibited cytochrome c diffusing out of mitochondria, which confirmed DG anti-apoptosis activity. DG also was found to significantly decrease the intracellular calcium accumulation and reactive oxygen species release within H9c2. / In the second part of present study, results revealed that DG elicited a concentration-dependent relaxation of U46619-preconstricted porcine coronary artery. DG-induced relaxation responses were not altered by the presence of endothelium-related dilator inhibitors, most potassium channel blockers, GMP and AMP pathway inhibitors and endothelium removal. Ba²⁺ (an inward rectifier K⁺ channel blocker) slightly attenuated DG-induced relaxation. The protein expression of phosphorylated myosin light chain (MLC) was inhibited by DG in a concentration-dependent manner whereas the activity of RhoA was not modified. Ca²⁺-induced contraction of coronary artery was inhibited by DG in a concentration-dependent fashion. DG acted as an antagonist of calcium channel inducing the porcine artery dilation. / The third part of the present study is about the pro-angiogenic effect of DG. We found that DG dose-dependently induced zebrafish sub-intestinal vessel sprouting and increased the mRNA expression of vascular endothelial growth factor (VEGF) and its receptors. To explore the underlying mechanism, we also examined the proangiogenic effect of DG on the angiogenesis of endothelial cells. The results showed that DG induced the HMEC-1 proliferation, migration and forming tube. / In conclusion, we found that DG could provide cardiac and cardiovascular protection by its multiple targets. It prevented heart injuries after ischemia or hypoxia and reperfusion through scavenging ROS and inhibiting calcium accumulation. Moreover, it mainly acts as an antagonist of L-type calcium channel to relax the contracted LAD vessel. It also exerted property of inducing angiogenesis in vivo and in vitro. Therefore, DG would be useful for treating coronary artery disease depending on its multiple targets. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Hu, Fan. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2012. / Includes bibliographical references (leaves 170-215). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstract also in Chinese. / Chapter 1 --- Intorduction --- p.1 / Chapter 1.1 --- Cardiovascular system and coronary artery diseases --- p.1 / Chapter 1.1.1 --- The cardiovascular system --- p.1 / Chapter 1.1.2 --- Contraction and relaxation of the vascular myocyte in arteries --- p.4 / Chapter 1.1.2.1 --- Ultrastructure of the vascular myocyte --- p.4 / Chapter 1.1.2.2 --- Contraction mechanisms of vascular myocyte --- p.5 / Chapter 1.1.2.3 --- Relaxation mechanisms of vascular myocyte --- p.7 / Chapter 1.1.3 --- Chronic coronary heart disease --- p.9 / Chapter 1.2 --- The way to treat chronic CAD --- p.11 / Chapter 1.2.1 --- Angiogenesis --- p.11 / Chapter 1.2.2 --- Clinical surgery for treating CAD --- p.13 / Chapter 1.2.2.1 --- Three common surgeries for treating CAD --- p.13 / Chapter 1.2.2.2 --- Ischemia and reperfusion (I/R) injury in surgeries --- p.15 / Chapter 1.2.3 --- Drugs for treating CAD --- p.19 / Chapter 1.2.3.1 --- Western medicine therapy in CAD --- p.19 / Chapter 1.2.3.2 --- Traditional Chinese Medicine treatment in CAD --- p.20 / Chapter 1.3 --- Aims of studies --- p.28 / Chapter 2 --- Materials and Methods --- p.29 / Chapter 2.1 --- Solutions and Materials --- p.29 / Chapter 2.1.1 --- Solutions --- p.29 / Chapter 2.1.2 --- Chemicals and enzymes --- p.36 / Chapter 2.2 --- Methods --- p.38 / Chapter 2.2.1 --- Herbal preparation --- p.38 / Chapter 2.2.2 --- Identification and quantification of chemical markers in Danshen and Gegen decoction (DG) --- p.38 / Chapter 2.2.3 --- Assay development for the determination of the DG marker compounds in rat plasma --- p.40 / Chapter 2.2.4 --- Isolation of pig left anterior descending coronary artery --- p.44 / Chapter 2.2.5 --- Isometric tension measurement --- p.45 / Chapter 2.2.6 --- Langendorff related experiment --- p.50 / Chapter 2.2.7 --- Cell culture of H9c2 cells --- p.54 / Chapter 2.2.8 --- Cell viability assay (MTT assay) --- p.56 / Chapter 2.2.9 --- Cell proliferation measurement --- p.57 / Chapter 2.2.10 --- Hypoxia and reperfusion cell model (H9c2) --- p.58 / Chapter 2.2.11 --- Determination of cell apoptosis with Annexin VFITC and PI double staining --- p.59 / Chapter 2.2.12 --- Measurement of mitochondria depolarization --- p.61 / Chapter 2.2.13 --- Measurement of ROS release --- p.63 / Chapter 2.2.14 --- Measurement of calcium localization in H9c2 cells by fluo4 dye and confocal microscopy --- p.64 / Chapter 2.2.15 --- Extraction of proteins from tissue, cell and subcellular fractions --- p.65 / Chapter 2.2.16 --- Western blot assay --- p.67 / Chapter 2.2.17 --- Human microvascular endothelial cells (HMEC1) cell culture --- p.68 / Chapter 2.2.18 --- Cell cycle analysis by PI staining --- p.69 / Chapter 2.2.19 --- Scratch assay for HMEC1cells migration --- p.70 / Chapter 2.2.20 --- Tube formation assay --- p.71 / Chapter 2.2.21 --- Vessel sprouting of Zebrafish --- p.72 / Chapter 2.2.22 --- Real time PCR --- p.74 / Chapter 2.2.23 --- Statistical analysis --- p.76 / Chapter 3 --- Chapter 3 Cardiac protection of Danshen and Gegen decoction in hypoxia/ischemia and reperfusion induced injury --- p.77 / Chapter 3.1 --- Introduction --- p.77 / Chapter 3.2 --- Results --- p.81 / Chapter 3.2.1 --- Cytotoxicity of DG --- p.81 / Chapter 3.2.2 --- The morphology alteration of H9c2 after H/R treatment --- p.83 / Chapter 3.2.3 --- Effect on H H9c2 cell survival after H/R treatment --- p.84 / Chapter 3.2.4 --- Effect on membrane skeleton of H9c2 cells with H/R injury --- p.86 / Chapter 3.2.5 --- Effect on the apoptosis in H9c2 cells induced by H/R injury --- p.88 / Chapter 3.2.6 --- Effect on cytochrome c release from mitochondria of damaged H9c2 cells --- p.92 / Chapter 3.2.7 --- Effect on mitochondria depolarization of H9c2 after H/R treatment --- p.94 / Chapter 3.2.8 --- Effect on reactive oxidant species (ROS) release --- p.96 / Chapter 3.2.9 --- Effect on calcium accumulation within H9c2 in the reperfusion phase --- p.98 / Chapter 3.2.10 --- Effect on heart functions of rat hearts with I/R injury (acute effect) --- p.101 / Chapter 3.2.11 --- Effect on heart function in rats with I/R injury (chronic effect) --- p.107 / Chapter 3.3 --- Discussion --- p.113 / Chapter 4 --- Chapter 4 Vasodilation effects of Danshen and Gegen decoction in porcine coronary artery and its underlying mechanism --- p.118 / Chapter 4.1 --- Introduction --- p.118 / Chapter 4.2 --- Results --- p.121 / Chapter 4.2.1 --- Investigations of endothelium dependent and independent mechanisms --- p.121 / Chapter 4.2.2 --- Effects on cAMP and cGMP pathway --- p.121 / Chapter 4.2.3 --- Effects on potassium channel opening --- p.121 / Chapter 4.2.4 --- Effects on calcium induced contraction and calcium sensitization --- p.122 / Chapter 4.2.5 --- Effects on MLC phosphorylations --- p.123 / Chapter 4.3 --- Discussion --- p.132 / Chapter 5 --- Chapter 5 In vitro and in vivo angiogenic effects of DG --- p.138 / Chapter 5.1 --- Introduction --- p.138 / Chapter 5.2 --- Results --- p.140 / Chapter 5.2.1 --- Effect on subintestinal vessels sprouting in the zebrafish embryo --- p.140 / Chapter 5.2.2 --- Effect on the transcription and expression of VEGFA and VEGF receptors -- Flt1 and KDR/Flk2 --- p.143 / Chapter 5.2.3 --- Effect on HMEC1 proliferation --- p.145 / Chapter 5.2.4 --- Effect on cell cycle of HMEC1 --- p.148 / Chapter 5.2.5 --- Effect on cell migration of HMEC1 --- p.151 / Chapter 5.2.6 --- Effect on tube formation of HMEC1 --- p.154 / Chapter 5.3 --- Discussion --- p.157 / Chapter 6 --- Chapter 6 Conclusions and future work --- p.160 / Chapter 6.1 --- Cardiac protection of DG in the I/R and H/R injury --- p.160 / Chapter 6.2 --- Vasodilation effect of DG on the porcine coronary artery --- p.165 / Chapter 6.3 --- Angiogenic effect of DG in vivo and in vitro --- p.167 / Chapter 6.4 --- Overall conclusion of the study --- p.169 / Chapter 7 --- References --- p.170

Cheung Ho Yan. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2005. / Includes bibliographical references (leaves 164-175). / Abstracts in English and Chinese. / Abstract --- p.i / Acknowledgements --- p.vi / Publications based on the work in this thesis --- p.vii / Table of content --- p.viii / Abbreviations --- p.xii / Chapter CHAPTER 1 --- INTRODUCTION --- p.1 / Chapter 1.1 --- Traditional Chinese Medicine --- p.1 / Chapter 1.1.1 --- Danshen --- p.2 / Chapter 1.1.2 --- Chemical constituents --- p.5 / Chapter 1.1.3 --- Pharmacological effects --- p.7 / Chapter 1.1.3.1 --- On blood vessels --- p.7 / Chapter 1.1.3.2 --- On blood pressure --- p.8 / Chapter 1.1.3.3 --- On heart --- p.8 / Chapter 1.1.3.4 --- On myocardial ischaemia and reperfusion --- p.9 / Chapter 1.1.3.5 --- On platelet activity --- p.10 / Chapter 1.1.3.6 --- Other actions --- p.11 / Chapter 1.1.4 --- Clinical studies --- p.12 / Chapter 1.2 --- The Vascular System --- p.13 / Chapter 1.2.1 --- The circulation network --- p.13 / Chapter 1.2.2 --- Physiology of blood vessels --- p.13 / Chapter 1.2.3 --- Control of vascular lone --- p.14 / Chapter 1.3 --- Mechanisms of Vasodilatation --- p.16 / Chapter 1.3.1 --- Endothelium derived relaxant factors (EDRFs) --- p.16 / Chapter 1.3.1.1 --- Nitric oxide (NO) --- p.16 / Chapter 1.3.1.2 --- Prostacyclin (PGI：) --- p.17 / Chapter 1.3.1.3 --- Endotheliun-derived hyperpolarization factors (EDHFs) --- p.18 / Chapter 1.3.1.3.1 --- Epoxyeicosatrienoic acids (EETs) --- p.19 / Chapter 1.3.1.3.2 --- Potassium ion (IC) --- p.20 / Chapter 1.3.1.3.3 --- Gap junction --- p.20 / Chapter 1.3.2 --- Signal transduction pathways --- p.21 / Chapter 1.3.2.1 --- Guanylyl cyclase-cGMP pathway --- p.21 / Chapter 1.3.2.2 --- Adenylyl cyclase-cAMP pathway --- p.22 / Chapter 1.3.3 --- Ion channels in vascular smooth muscle cell --- p.24 / Chapter 1.3.3.1 --- Potassium channels (K+ channels) --- p.24 / Chapter 1.3.3.2 --- Calcium channels (Ca2+ channels) --- p.24 / Chapter 1.3.3.3 --- Chloride channel (Cl channel) --- p.25 / Chapter 1.3.4 --- Receptor-operated mechanisms --- p.27 / Chapter 1.3.4.1 --- Muscarinic receptors --- p.27 / Chapter 1.3.4.2 --- Adrenoceptors --- p.27 / Chapter 1.3.4.3 --- Histamine receptors --- p.28 / Chapter 1.3.4.4 --- CGRP receptors --- p.29 / Chapter 1.3.4.5 --- Tachykinin receptors --- p.30 / Chapter 1.4 --- Aims of the studies --- p.31 / Chapter CHAPTER 2 --- MATERIALS AND METHODS --- p.32 / Chapter 2.1 --- Extraction of Water and Lipid-solubie Fractions from Danshen --- p.32 / Chapter 2.1.1 --- Preparation of water-soluble and lipid-soluble fractions --- p.33 / Chapter 2.2 --- Experiments on Rat Knee Joint --- p.35 / Chapter 2.2.1 --- Animals --- p.35 / Chapter 2.2.2 --- Materials --- p.35 / Chapter 2.2.3 --- Preparatory protocols --- p.37 / Chapter 2.2.3.1 --- Anaesthesia of animals --- p.37 / Chapter 2.2.3.2 --- Cannulation of trachea --- p.37 / Chapter 2.2.3.3 --- Cannulation of carotid artery --- p.38 / Chapter 2.2.3.4 --- Blood pressure measurement --- p.38 / Chapter 2.2.4 --- Measurement of knee joint blood flow --- p.39 / Chapter 2.2.4.1 --- Preparation for measurement of knee joint blood flow --- p.41 / Chapter 2.2.5 --- Experimental protocols --- p.41 / Chapter 2.2.5.1 --- Danshen on knee joint blood flow --- p.41 / Chapter 2.2.5.2 --- Antagonists on Danshen --- p.41 / Chapter 2.2.5.3 --- Positive controls --- p.43 / Chapter 2.2.6 --- Image analysis --- p.44 / Chapter 2.2.7 --- Data analysis --- p.44 / Chapter 2.3 --- Experiments on Rat Femoral Artery --- p.45 / Chapter 2.3.1 --- Animals --- p.45 / Chapter 2.3.2 --- Materials --- p.45 / Chapter 2.3.2.1 --- Chemicals --- p.45 / Chapter 2.3.2.2 --- Physiological salt solution --- p.48 / Chapter 2.3.3 --- Preparatory protocols --- p.48 / Chapter 2.3.3.1 --- Small vessel myograph --- p.48 / Chapter 2.3.3.2 --- Isolation and mounting of tissue --- p.49 / Chapter 2.3.4 --- Experimental protocols --- p.50 / Chapter 2.3.4.1 --- Studies on the vasodilator response to Danshen --- p.50 / Chapter 2.3.4.2 --- Studies of antagonists on Danshen --- p.50 / Chapter 2.3.4.2.1 --- Endothelium-dependent mechanisms --- p.51 / Chapter 2.3.4.2.2 --- Endothelium-independent mechanisms --- p.54 / Chapter 2.3.4.2.3 --- K+ channel blockers --- p.54 / Chapter 2.3.4.2.4 --- Positive controls --- p.55 / Chapter 2.3.4.3 --- Danshen on Ca2+-induced contraction --- p.56 / Chapter 2.3.5 --- Data analysis --- p.57 / Chapter CHAPTER 3 --- RESULTS --- p.58 / Chapter 3.1 --- Danshen on Rat Knee Joint Blood Flow --- p.58 / Chapter 3.1.1 --- Topical administration of Danshen --- p.58 / Chapter 3.1.2 --- Antagonists on Danshen --- p.59 / Chapter 3.1.2.1 --- Muscarinic receptor antagonist --- p.59 / Chapter 3.1.2.2 --- β-adrenoceptor antagonist --- p.60 / Chapter 3.1.2.3 --- Histamine receptor antagonists --- p.60 / Chapter 3.1.2.4 --- Nitric oxide synthase inhibitor --- p.61 / Chapter 3.1.2.5 --- Cyclo-oxygenase inhibitors --- p.62 / Chapter 3.1.2.6 --- CGRPi receptor antagonist --- p.62 / Chapter 3.1.2.7 --- NK1 receptor antagonist --- p.63 / Chapter 3.1.2.8 --- Potassium channel inhibitor --- p.64 / Chapter 3.1.2.9 --- "Combination of cyclo-oxygenase inhibitor, nitric oxide synthase inhibitor and CGRP1 receptor antagonist" --- p.64 / Chapter 3.1.3 --- Antagonists on water-soluble fraction of Danshen --- p.91 / Chapter 3.1.3.1 --- Nitric oxide synthase inhibitor --- p.91 / Chapter 3.1.3.2 --- Cyclo-oxygenase inhibitors --- p.91 / Chapter 3.1.3.3 --- CGRP1 receptor antagonist --- p.92 / Chapter 3.1.3.4 --- NK1 receptor antagonist --- p.92 / Chapter 3.1.3.5 --- Potassium channel inhibitor --- p.92 / Chapter 3.2 --- Danshen on Rat Femoral Artery --- p.99 / Chapter 3.2.1 --- Danshen on precontracted arterial ring --- p.99 / Chapter 3.2.2 --- Endothelium-dependent mechanisms --- p.106 / Chapter 3.2.3 --- Endothelium-independent mechanisms --- p.114 / Chapter 3.2.4 --- K+ channel blockers --- p.119 / Chapter 3.2.4.1 --- Effect on Danshen --- p.119 / Chapter 3.2.4.2 --- Effect on water-soluble and lipid-soluble fractions of Danshen --- p.121 / Chapter 3.2.4.3 --- Effect on Danshensu --- p.122 / Chapter 3.2.5 --- Danshen on Ca2+-induced contractions --- p.133 / Chapter CHAPTER 4 --- DISCUSSION --- p.138 / Chapter 4.1 --- In Vivo Studies of Danshen on Rat Knee Joint Blood Flow --- p.139 / Chapter 4.2 --- In Vitro Studies of Danshen on Isolated Rat Femoral Artery --- p.148 / Chapter 4.2.1 --- Comparisons of the use of different precontractors --- p.148 / Chapter 4.2.2 --- Investigations on endothelium-dependent mechanisms --- p.151 / Chapter 4.2.3 --- Investigations on endothelium-independent mechanisms --- p.152 / Chapter 4.2.4 --- Effects of K+ channel blockers --- p.154 / Chapter 4.2.5 --- Inhibition of Ca2+ influx in vascular smooth muscle --- p.157 / Chapter 4.3 --- Comparisons of Results from In Vivo and In Vitro Studies --- p.159 / Chapter 4.4 --- Future Studies --- p.161 / Chapter 4.5 --- Conclusion --- p.162 / REFERENCES --- p.164

Yam Wing Sze. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2005. / Includes bibliographical references (leaves 154-160). / Abstracts in English and Chinese. / Abstract English --- p.i / Chinese --- p.iii / Acknowledgments --- p.v / Table of contents --- p.vii / List of Tables --- p.x / List of Figures --- p.xi / List of Abbreviations --- p.xvi / Chapter Chapter 1 --- Introduction --- p.1 / Chapter 1.1 --- Chinese Medicine and Western Medication --- p.1 / Chapter 1.2 --- Chinese Medicine and Compound Formula --- p.2 / Chapter 1.3 --- Cardiovascular disease (CVD) and atherosclerosis --- p.6 / Chapter 1.4 --- General Research Objectives --- p.19 / Chapter Chapter 2 --- Establishment of compound formulation and Extract Preparation --- p.21 / Chapter 2.1 --- Formulation searched from Chinese Pharmacopoeia --- p.21 / Chapter 2.2 --- Aqueous extract preparation --- p.25 / Chapter 2.2.1 --- Materials and Methods --- p.25 / Chapter 2.2.2 --- Discussion --- p.27 / Chapter Chapter 3 --- Vasodilation study --- p.28 / Chapter 3.1 --- Vascular Smooth Muscle Contraction and Relaxation --- p.28 / Chapter 3.2 --- Endothelium and Vasodilation --- p.30 / Chapter 3.3 --- Vasodilation in organ bath --- p.32 / Chapter 3.3.1 --- Materials and Methods --- p.32 / Chapter 3.3.2 --- Results --- p.35 / Chapter 3.3.3 --- Discussion --- p.40 / Chapter 3.4 --- Endothelium dependent vasodilation --- p.40 / Chapter 3.4.1 --- Materials and Methods --- p.43 / Chapter 3.4.2 --- Results --- p.45 / Chapter 3.4.3 --- Discussion --- p.54 / Chapter 3.5 --- Adrenoceptor and vasodilation --- p.55 / Chapter 3.5.1 --- Materials and Methods --- p.57 / Chapter 3.5.2 --- Results --- p.58 / Chapter 3.5.3 --- Discussion --- p.62 / Chapter 3.6 --- Potassium Channels and Vasodilation --- p.63 / Chapter 3.6.1 --- Materials and Methods --- p.65 / Chapter 3.6.2 --- Results --- p.67 / Chapter 3.6.3 --- Discussion and Summary --- p.77 / Chapter 3.7 --- Potential active components from Fenge and Danshen --- p.82 / Chapter 3.7.1 --- Materials and Methods --- p.82 / Chapter 3.7.2 --- Results --- p.83 / Chapter 3.7.3 --- Discussion --- p.87 / Chapter Chapter 4 --- Comparison of Fenge and Yege --- p.88 / Chapter 4.1 --- Vasodilative effects of Fenge and Yege --- p.89 / Chapter 4.1.1 --- Materials and Methods --- p.89 / Chapter 4.1.2 --- Results --- p.89 / Chapter 4.1.3 --- Discussion --- p.101 / Chapter 4.2 --- The comparison of antioxidative effect between Yege and Fenge --- p.104 / Chapter 4.2.1 --- Red blood cell hemolysis model --- p.106 / Chapter 4.2.1.1 --- Materials and Methods --- p.106 / Chapter 4.2.1.2 --- Results --- p.108 / Chapter 4.2.1.3 --- Discussion --- p.110 / Chapter 4.2.2 --- Ischemia-reperfusion on Langendroff --- p.112 / Chapter 4.2.2.1 --- Materials and Methods --- p.114 / Chapter 4.2.2.2 --- Results --- p.117 / Chapter 4.2.2.3 --- Discussion --- p.125 / Chapter Chapter 5 --- Comparison of Chemical Profiles of Fenge and Yege --- p.127 / Chapter 5.1 --- The application of HPLC --- p.127 / Chapter 5.2 --- HPLC standardization --- p.129 / Chapter 5.2.1 --- Materials and Methods --- p.132 / Chapter 5.2.2 --- Results --- p.133 / Chapter 5.2.3 --- Discussion --- p.144 / Chapter Chapter 6 --- "Summaries, Discussion and prospects" --- p.146 / Chapter 6.1 --- Summaries and Discussion --- p.146 / Chapter 6.2 --- Prospects --- p.148 / Chapter 6.2.1 --- "Cardiovascular tonic effect of pure compounds, extracts with difference solvents and their vasodilative mechanism." --- p.148 / Chapter 6.2.2 --- Macrophage Foam Cell and Atherosclerosis --- p.149 / Chapter 6.2.3 --- The D:F (7:3) and D:Y (7:3) compound formulae capsule with GMP --- p.152 / References --- p.154

Alzhemier's disease (AD) is a common form of dementia which is characterized by the deposition of amyloids in affected neurons and a cholinergic neurotransmission deficit in the brain. Current therapeutic intervention for AD is primarily based on inhibition of brain acetylcholinesterase (AChE) to restore the brain acetylcholine level. Cryptotanshinone (CT) is a diterprene which is extracted from the root of Salvia miltiorrhiza, an herb that is commonly prescribed in Chinese medicine to treat cardiovascular disease. The present study is aimed at verifying CT's property as an AChE inhibitor using different models. By AChE activity assay, CT was found to be a dual inhibitor which inhibits both human acetylcholinesterase (AChE) and butylcholinesterase (BuChE) with similar IC50. CT inhibited human AChE in a reversible manner, and the inhibition showed the characteristics of mixed-type. To human BuChE, CT is an uncompetitive inhibitor. CT can also inhibit AChE from rat cortical neurons. Apart from AChE inhibition, CT was demonstrated to have ameliorating effect on glutamate excitotoxicity, which is a cause of neuron death in AD. Further study showing that CT treatment can reduce cellular tau phosphorylation, which is the downstream effector of glutamate-induced excitotoxicity. In animal model, the effect of CT on learning impairment in scopolamine-treated rats was also evaluated by the acquisition protocol of Morris water maze. The task learning ability of scopolamine-treated rats was significantly reversed by CT, and the CT-fed rats were able to develop spatial searching strategy comparable to the control animals. Chronic administration of CT at effective doses did not cause significant hepatotoxicity. Cholinergic side effect of muscle weakness was not observed in CT treated rats. On the contrary CT was found to increase the locomotor activity of NIH mice in forced swimming test through reducing the lactic acid in the circulation. Data in this study gives further support on CT's potential as a therapeutic drug for treating AD. / by Wong, Kin Kwan Kelvin. / Source: Dissertation Abstracts International, Volume: 73-01, Section: B, page: . / Thesis (Ph.D.)--Chinese University of Hong Kong, 2009. / Includes bibliographical references (leaves 144-167). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Electronic reproduction. [Ann Arbor, MI] : ProQuest Information and Learning, [201-] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstract also in Chinese.