Mak Chun-yin. / "February 2001." / Thesis (Ph.D.)--Chinese University of Hong Kong, 2001. / Includes bibliographical references (p. 156-169). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Mode of access: World Wide Web. / Abstracts in English and Chinese.

何首烏為寥科多年生藤本植物，而中藥何首烏則為該植物的乾燥塊根。常用於治療白髮及與老年化相關的疾病。然而，何首烏的治療機理卻少有報導。本文對何首烏的粗提物及其主要生物活性成份2,3,5,4' -四羥基二萃乙烯-2-0-ß-D-葡萄糖苷(THSG) 在老鼠及人類的黑色素細胞中影響黑色素生成機理及細胞毒性進行了深入研究。 / 利用高效液相層析連接質譜儀的測定， THSG 在水及乙醇粗提物中含量分別為0.064 及0.75 的百分比。由於THSG 在水及乙醇粗提物中佔有一定份量，所以它對粗提物所產生的生物及生化反應有著重要的影響。在低於細胞毒範圍的劑量內，何首烏粗提物及THSG 能降低老鼠黑色素細胞株melan-a 的左旋多巴(L-DOPA)的轉化反應。在細胞毒性檢測中，水粗提物及THSG 在<100 μg/ml的劑量下均未有對至少5 種黑色素細胞株及黑色素瘤細胞造成傷害。然而，乙醇粗提物的細胞毒卻是水粗提物或THSG 的3-4 倍。 / 在無細胞系統中， THSG 在可逆轉的情況下抑制酪氨酸酶將左旋多巴轉化成黑色素。在細胞系統中，它也能阻止由蛋白激醋A (PKA)引發的黑色素生成反應。THSG 在老鼠及人類黑色素細胞中的中位值抑制率(lC₅₀)分別為123.0 μM及61.5 μM。 THSG 抑制酪氮酸醋的能力展現在黑色素細胞/角質細胞的共培養比在單黑色素細胞培養中更明顯。 / 調控酪氯酸臨可以在脫氧核糖核酸(DNA)轉錄及翻譯後修飾兩方面達成。在DNA 轉錄中，小眼球相關轉錄因數(MITF)的減少導致酪氨酸酶的表達隨著THSG 濃度而減少。翻譯後酪氮酸臨主要依靠蛋白激臨C-ß (PKC-ß)使其磷酸化，從而增加酪氯酸酶/酪氯酸酶相關蛋白-1(TRP-1 )組成複合蛋白。然而THSG 均減少蛋白激酶 C-ß的表達及酪氮酸酪/酪氮酸醋相關蛋白-1 所組成的複合蛋白。 另一方面， THSG 卻沒有影響酪氯酸酶蛋白在內質網/高爾基氏複合體內的糖基化及內涵體與溶酶體間的運輸。 / 總而言之，本文首次展示何首烏粗提物及THSG 在單細胞培養及共培養細胞的系統下抑制黑色素生成。THSG 能在可逆轉的抑制機制下阻止酪氯酸酶作出反應。而在PKA 引發的黑色素生成反應中， THSG 也能在DNA 轉錄及翻譯後修飾等過程中減低酪氯酸酶的活性。 / Radix Polygoni Multiflori, the dried root of Polygonum multiflorum (PM), is well documented for its clinical effects in treating various diseases associated with aging and hair graying, but the evidence based-mechanisms remain largely unknown. In this study, PM was extracted with water and 70% ethanol and a major constituent, 2,3,5,4'-tetrahydroxystilbene-2-0-I3-D-glucoside (THSG) of about 0.064% and 0.75%, respectively, were found in these extracts as analyzed by high-performance liquid chromatography (HPLC) coupled to mass spectrometry. The melanogenic properties and cytotoxicity of the two extracts and THSG were evaluated using murine and human melanocytes. / Both water and ethanol extracts of PM and THSG showed a dose-dependent anti-melanogenic activity in an in vitro murine melan-a melanocyte assay for reduction of L-DOPA conversion by tyrosinase. Of at least 5 melanoma and melanocyte cell lines tested, both water PM extract and THSG were relatively safe, which at doses <100 μg/ml did not demonstrate any significant cytotoxic effects. On the other hand, ethanol PM extract was about 3-4 folds more cytotoxic. / Tyrosinase is the rate-limiting enzyme for melanogenesis. In a cell-free kinetic analysis, THSG inhibited tyrosinase activity in a reversible and non-competitive manner. At the cellular level, this inhibition is mediated through a PKA-dependent melanogenic pathway, as well as in a dose-dependent manner, with IC₅₀ = 123.0 μM and 61.5 μM for murine and human melanocytes, respectively. Tyrosinase was much more sensitive to the inhibitory effect of THSG in the melanocytelkeratinocyte co-culture system than in the melanocyte mono-culture system. / Functional tyrosinase is regulated at both transcriptional and post-translational modification levels. At the transcription level, THSG reduced expression of microphthalmia-associated transcription factor (MITF) esulted in a down-regulation of tyrosinase expression. At the post-translational modification level, THSG inhibited expression of PKC-β which is responsible for tyrosinase phosphorylation, and enhanced tyrosinase/TRP-1 complex formation. On the hand, THSG did not affect glycosylation of tyrosinase nor its trafficking from ER/Golgi to endosomal/ lysosomal compartments. / Taken our results together, the anti-melanogenic property of PM extracts and THSG were firstly demonstrated in both mono- and co-culture system using murine or human melanocytes and keratinocytes. THSG is a reversible and competitive inhibitor, which lowered the tyrosinase activity at both transcription and post-translational modification levels via PKA-mediated melanogenesis. / 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. / Cheung, Wing Ki. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2011. / Includes bibliographical references (leaves 160-174). / 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 / General introduction / Chapter 1.1 --- Anatomy and Physiology of the Skin --- p.5 / Chapter 1.1.1 --- Epidermis --- p.5 / Chapter 1.1.1.1 --- Stratum basale --- p.6 / Chapter 1.1.1.2 --- Stratum spinosum --- p.6 / Chapter 1.1.1.3 --- stratum granulosum --- p.7 / Chapter 1.1.1.4 --- Stratum corneum --- p.7 / Chapter 1.1.2 --- Dermis --- p.8 / Chapter 1.2 --- Melanogenesis of the Skin --- p.9 / Chapter 1.2.1 --- History of melanogenesis study --- p.9 / Chapter 1.2.2 --- Today's melanogenesis study --- p.10 / Chapter 1.3 --- Hyperpigmentary Disorders --- p.14 / Chapter 1.3.1 --- Malasma --- p.14 / Chapter 1.3.2 --- Lentigines --- p.15 / Chapter 1.3.2.1 --- Lentigo simplex --- p.16 / Chapter 1.3.2.2 --- Lentigo senilis etActinicus --- p.16 / Chapter 1.3.3 --- Post-inflammatory hyperpigmentation --- p.17 / Chapter 1.4 --- Current Available Treatment for Hyperpigmentation --- p.18 / Chapter 1.4.1 --- Topical treatment and their strategies --- p.18 / Chapter 1.4.1.1 --- Inhibition of tyrosinase activity --- p.18 / Chapter 1.4.1.2 --- Antioxidation --- p.21 / Chapter 1.4.1.3 --- Melanosome transfer inhibition --- p.22 / Chapter 1.4.1.4 --- Stimulation of desquamation --- p.22 / Chapter 1.4.2 --- Laser treatment and their strategies --- p.23 / Chapter 1.4.3 --- Sunscreen --- p.24 / Chapter 1.5 --- Theories and the Treatment of Hyperpigmentation with Chinese Herbal Medicine --- p.25 / Chapter 1.6 --- Testing Systems --- p.26 / Chapter 1.7 --- Aims and Objectives of Study --- p.27 / Chapter 2 / Chemical Properties of THSG / Chapter 2.1 --- Introduction --- p.30 / Chapter 2.1.1 --- Radix Polygoni multiflori --- p.30 / Chapter 2.1.2 --- 2,3,5,4'-tetrahydroxystilbene glucoside (THSG) --- p.31 / Chapter 2.1.2.1 --- Stilbene --- p.32 / Chapter 2.1.2.2 --- Chemical properties of THSG --- p.34 / Chapter 2.1.3 --- Objectives --- p.35 / Chapter 2.2 --- Materials and Methods --- p.36 / Chapter 2.2.1 --- Plant materials --- p.36 / Chapter 2.2.2 --- Extraction --- p.36 / Chapter 2.2.3 --- High performance liquid chromatography (HPLC) analysis --- p.36 / Chapter 2.2.4 --- Enzymatic hydrolysis of THSG and salicin --- p.37 / Chapter 2.2.5 --- HPLC/MS analysis --- p.38 / Chapter 2.2.6 --- Benedict's test --- p.38 / Chapter 2.2.7 --- Enzymatic oxidation --- p.38 / Chapter 2.2.8 --- Thin layer chromatography (TLC) analysis --- p.39 / Chapter 2.3 --- Results / Chapter 2.3.1 --- The THSG content in water and alcohol extracts of PM --- p.40 / Chapter 2.3.2 --- The stability of THSG against oxidation --- p.41 / Chapter 2.3.3 --- Enzymatic hydrolysis ofTHSG --- p.42 / Chapter 2.4 --- Discussion --- p.46 / Chapter 3 / The Melanogenic inhibitory mechanisms of Radix Polygonum multiflorum (PM) extracts and THSG in murine melanocyte / Chapter 3.1 --- Introduction --- p.47 / Chapter 3.1.1 --- Murine melanocyte --- p.47 / Chapter 3.1.2 --- Melanogenesis --- p.48 / Chapter 3.1.2.1 --- Factors affecting Melanogenesis --- Oxidation --- p.49 / Chapter 3.1.2.2 --- Factors affecting Melanogenesis --- UV radiation --- p.50 / Chapter 3.1.2.3 --- Factors affecting melanogenesis---- Cellular regulation --- p.51 / Chapter 3.1.2.3.1 --- The regulation of transcription, translation, and post-translational modification of tyrosianse --- p.51 / Chapter 3.1.2.3.2 --- PKA-, PKC-, and PKG- melangenic pathways --- p.56 / Chapter 3.1.3 --- Kinetic analysis of tyrosinase --- p.57 / Chapter 3.1.4 --- Objectives --- p.58 / Chapter 3.2 --- Materials and Methods --- p.59 / Chapter 3.2.1 --- Cell culture --- p.59 / Chapter 3.2.2 --- Kinetic analysis of tyrosinase activity inhibition --- p.60 / Chapter 3.2.3 --- SRB assay --- p.61 / Chapter 3.2.4 --- L -DOPA conversion assay --- p.62 / Chapter 3.2.5 --- Melanin production measurement --- p.62 / Chapter 3.2.6 --- ROS detection by flow cytometer --- p.63 / Chapter 3.2.7 --- In-situ tyrosinase activity assay --- p.63 / Chapter 3.2.8 --- Western blotting (WB) analysis --- p.64 / Chapter 3.2.9 --- Immunofluorescence microscopy --- p.66 / Chapter 3.2.10 --- Glycosylation analysis --- p.67 / Chapter 3.2.11 --- Co-Immunoprecipitation --- p.68 / Chapter 3.2.12 --- Radix Polygonum Multiflorum and THSG metabolite collection from rat serum --- p.69 / Chapter 3.3 --- Results --- p.71 / Chapter 3.3.1 --- Enzyme kinetic study of the catalysis of L-DOPA by murine melanocyte lysate --- p.71 / Chapter 3.3.2 --- Inhibitory effect of crude PM preparations and THSG on tyrosinase activity and melanin synthesis in murine melan-a melanocytes --- p.74 / Chapter 3.3.3 --- Effect of THSG on H₂0₂- induced oxidation --- p.77 / Chapter 3.3.4 --- THSG inhibits PKA-induced melanogenesis --- p.78 / Chapter 3.3.5 --- Reduction of in situ tyrosinase activity in PKA-induced melanogenesis --- p.82 / Chapter 3.3.6 --- Alternation of melanogenic proteins --- p.85 / Chapter 3.3.7 --- THSG does not alter the tyrosinase trafficking in ER and Golgi --- p.89 / Chapter 3.3.8 --- THSG does not alter the tyrosinase trafficking in endosomal lysosomal compartments --- p.93 / Chapter 3.3.9 --- Glycosylation analysis --- p.95 / Chapter 3.3.10 --- Reduction of interaction between tyrosinase and TRP-1 to form heterodimeric complexes --- p.97 / Chapter 3.3.11 --- The metabolite of PM water extract and THSG maintained the in vitro tyrosinase activity --- p.101 / Chapter 3.4 --- Discussion --- p.103 / Chapter 4 / The Inhibitory Effect of THSG on Melanogenesis in Monolayer Culture of Human Melanocytes and in Co-culture of Melanocyte-Keratinocyte / Chapter 4.1 --- Introduction --- p.110 / Chapter 4.1.1 --- Human melanocyte --- p.110 / Chapter 4.1.1.1 --- The origin and the development of melanocyte --- p.110 / Chapter 4.1.1.2 --- Morphology, body site distribution and histological location --- p.111 / Chapter 4.1.1.3 --- In vitro growth of human melanocyte --- p.112 / Chapter 4.1.1.3.1 --- Lifespan vs. culture conditions --- p.113 / Chapter 4.1.1.3.2 --- Lifespan vs. donor age and skin type --- p.114 / Chapter 4.1.1.4 --- Modulation of pigmentation in response to stress --- p.114 / Chapter 4.1.1.5 --- Difference between human and murine TRPs --- p.115 / Chapter 4.1.2 --- Keratinocyte-Melanocyte interaction --- p.117 / Chapter 4.1.2.1 --- Release of melanogenic factors --- p.117 / Chapter 4.1.2.2 --- Release of survival and proliferating factors --- p.118 / Chapter 4.1.2.3 --- Melanosome transfer determines the cutaneous pigmentation --- p.118 / Chapter 4.1.2.3.1 --- Molecular events during melanosome transfer --- p.119 / Chapter 4.1.2.4 --- Others --- p.121 / Chapter 4.1.3 --- Objectives --- p.121 / Chapter 4.2 --- Materials and Methods --- p.122 / Chapter 4.2.1 --- Cell Culture --- p.122 / Chapter 4.2.1.1 --- Human melanocytes isolation and cultivation --- p.122 / Chapter 4.2.1.2 --- Immortalized keratinocytes - HaCaT cells --- p.123 / Chapter 4.2.1.3 --- Co-culture of melanocytes and HaCaT cells --- p.124 / Chapter 4.2.1.3.1 --- Monolayer co-culture --- p.124 / Chapter 4.2.1.3.2 --- Two-layer co-culture --- p.124 / Chapter 4.2.2 --- SRB assay --- p.125 / Chapter 4.2.3 --- L-DOPA conversion assay --- p.125 / Chapter 4.2.4 --- Western blotting (WB) analysis --- p.125 / Chapter 4.2.5 --- Light microscopy and immunofluorescent microscopy --- p.126 / Chapter 4.2.6 --- cAMP immunoassay --- p.126 / Chapter 4.3 --- Results --- p.128 / Chapter 4.3.1 --- The isolation and purification of human melanocytes --- p.128 / Chapter 4.3.2 --- Aging vs. Tyrosinase activity and melanin content --- p.131 / Chapter 4.3.3 --- Inhibitory effect of THSG in tyrosinase activity in human melanocyte --- p.133 / Chapter 4.3.4 --- Alternation of melanogenic proteins --- p.135 / Chapter 4.3.5 --- Sensitization of melanocytes to THSG treatment in co-culture system --- p.138 / Chapter 4.3.6 --- Induction of melanocyte dendricity in co-culture system --- p.140 / Chapter 4.3.7 --- THSG inhibited cAMP induction by forskolin and paracrine factors from keratinocytes --- p.141 / Chapter 4.4 --- Discussion --- p.143 / Discussion / Chapter 5.1 --- Discussion --- p.149 / References --- p.160

by Lau Chun Hong. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2004. / Includes bibliographical references (leaves 179-195). / Abstracts in English and Chinese. / Abstract --- p.i / Abstract in Chinese 摘要 --- p.iv / Acknowledgements --- p.vi / Table of Contents --- p.vii / List of Abbreviations --- p.xiii / List of Figures --- p.xvi / List of Tables --- p.xviii / Publications --- p.xix / Chapter Chapter1 --- Introduction --- p.1 / Chapter 1.1 --- Epidemiology of Diabetes Mellitus --- p.1 / Chapter 1.2 --- Definition of Diabetes Mellitus --- p.2 / Chapter 1.3 --- Glucose Homeostasis and Diabetes Mellitus --- p.2 / Chapter 1.4 --- Classification of Diabetes Mellitus --- p.5 / Chapter 1.4.1 --- Type 1 Diabetes Mellitus --- p.5 / Chapter 1.4.2 --- Type 2 Diabetes Mellitus --- p.6 / Chapter 1.4.3 --- Other Specific Types --- p.7 / Chapter 1.4.4 --- Gestational Diabetes --- p.9 / Chapter 1.4.5 --- Clinical Stages of Diabetes --- p.9 / Chapter 1.5 --- Diagnostic Criteria of Diabetes Mellitus --- p.10 / Chapter 1.6 --- Complications of Diabetes Mellitus --- p.12 / Chapter 1.7 --- Pharmacological Treatment of Diabetes --- p.13 / Chapter 1.7.1 --- Treatment of Type 1 Diabetes --- p.13 / Chapter 1.7.2 --- Treatment of Type 2 Diabetes --- p.14 / Chapter 1.7.2.1 --- Sulphonylureas --- p.17 / Chapter 1.7.2.2 --- Biguanides --- p.18 / Chapter 1.7.2.3 --- α-Glucosidase Inhibitors --- p.19 / Chapter 1.7.2.4 --- Thiazolidinediones --- p.20 / Chapter 1.8 --- Diabetes and Traditional Chinese Medicine --- p.21 / Chapter 1.9 --- Project Objective --- p.27 / Chapter Chapter2 --- Botanical and Phytochemical Studies --- p.28 / Chapter 2.1 --- Introduction --- p.28 / Chapter 2.2 --- Materials --- p.31 / Chapter 2.3 --- Authentication of Herbal Material --- p.41 / Chapter 2.3.1 --- Materials --- p.41 / Chapter 2.3.2 --- Phytochemical Studies --- p.43 / Chapter 2.3.2.1 --- Sample Preparation --- p.43 / Chapter 2.3.2.2 --- Thin Layer Chromatography --- p.46 / Chapter 2.3.3 --- Results --- p.51 / Chapter 2.4 --- Extraction of Herbal Material --- p.56 / Chapter 2.4.1 --- Materials and Methods --- p.56 / Chapter 2.4.2 --- Results --- p.46 / Chapter 2.5 --- Quantification of Sugar Content in Herbal Extracts --- p.58 / Chapter 2.5.1 --- Introduction --- p.58 / Chapter 2.5.2 --- Materials and Methods --- p.58 / Chapter 2.5.3 --- Results --- p.61 / Chapter 2.6 --- Discussion --- p.65 / Chapter Chapter3 --- In vitro Studies on Formula 2 and its Individual Herbs --- p.68 / Chapter 3.1 --- Introduction --- p.68 / Chapter 3.2 --- Intestinal Glucose Absorption Studies --- p.69 / Chapter 3.2.1 --- Introduction --- p.69 / Chapter 3.2.2 --- Materials and Methods --- p.70 / Chapter 3.2.2.1 --- Preparation of BBMV --- p.71 / Chapter 3.2.2.2 --- BBMV Glucose Uptake Assay --- p.72 / Chapter 3.2.2.3 --- Bicinchoninic Acid (BCA) Protein Assay --- p.73 / Chapter 3.2.2.4 --- Preparation of Herbal Chloroform Extract --- p.74 / Chapter 3.2.2.5 --- Glucose Uptake Assay with Herbal Extracts --- p.75 / Chapter 3.2.3 --- Results --- p.76 / Chapter 3.3 --- Hepatic Gluconeogenesis Studies --- p.79 / Chapter 3.3.1 --- Introduction --- p.79 / Chapter 3.3.2 --- Materials and Methods --- p.82 / Chapter 3.3.2.1 --- Cell Culture --- p.83 / Chapter 3.3.2.2 --- Glucose Production Assay --- p.83 / Chapter 3.3.2.3 --- PEPCK Assay --- p.85 / Chapter 3.3.3 --- Results --- p.86 / Chapter 3.4 --- Cellular Glucose Uptake Studies --- p.88 / Chapter 3.4.1 --- Introduction --- p.88 / Chapter 3.4.2 --- Materials and Methods --- p.89 / Chapter 3.4.2.1 --- Cell Culture --- p.89 / Chapter 3.4.2.2 --- Differentiation of 3T3-L1 --- p.90 / Chapter 3.4.2.3 --- 2-Deoxy-D-glucose Uptake Assay --- p.91 / Chapter 3.4.3 --- Results --- p.92 / Chapter 3.5 --- Discussion --- p.96 / Chapter 3.5.1 --- Intestinal Glucose Absorption Studies by BBMV --- p.96 / Chapter 3.5.2 --- Hepatic Gluconeogenesis Studies by H4IIE Cells --- p.97 / Chapter 3.5.3 --- Cellular Glucose Uptake Studies by Hs68 and 3T3-L1 Cells --- p.99 / Chapter 3.5.4 --- Conclusions --- p.100 / Chapter Chapter4 --- In vivo Studies on Selected Herbs --- p.103 / Chapter 4.1 --- Introduction --- p.103 / Chapter 4.1.1 --- Animal Models of Type 2 Diabetes --- p.103 / Chapter 4.1.2 --- Chemically-induced Diabetic Models --- p.104 / Chapter 4.1.3 --- Neonatal-STZ Diabetic Rats --- p.107 / Chapter 4.2 --- Basal Glycaemia Test --- p.109 / Chapter 4.2.1 --- Animals --- p.109 / Chapter 4.2.2 --- Testing Method --- p.110 / Chapter 4.2.3 --- Results --- p.112 / Chapter 4.3 --- Oral Glucose Tolerance Test --- p.114 / Chapter 4.3.1 --- Animals --- p.114 / Chapter 4.3.2 --- Testing Method --- p.114 / Chapter 4.3.3 --- Results --- p.116 / Chapter 4.4 --- Discussion --- p.119 / Chapter Chapter5 --- Bioassay-guided Fractionation of Cortex Moutan --- p.125 / Chapter 5.1 --- Introduction --- p.125 / Chapter 5.1.1 --- Phytochemical Studies of Cortex Moutan --- p.125 / Chapter 5.2 --- Organic Extraction of Cortex Moutan --- p.128 / Chapter 5.2.1 --- Extraction Method --- p.128 / Chapter 5.2.2 --- Results --- p.129 / Chapter 5.3 --- BBMV Glucose Uptake Assay with Fraction CM C --- p.131 / Chapter 5.3.1 --- Materials and Methods --- p.131 / Chapter 5.3.2 --- Results --- p.131 / Chapter 5.4 --- In vivo Studies of Fraction CM-C --- p.133 / Chapter 5.4.1 --- Materials and Methods --- p.133 / Chapter 5.4.2 --- Results --- p.133 / Chapter 5.5 --- Fractionation of Fraction CM-C --- p.137 / Chapter 5.5.1 --- Materials and Methods --- p.137 / Chapter 5.5.2 --- Results --- p.139 / Chapter 5.6 --- BBMV Glucose Uptake Assay with CM-C Sub-fractions --- p.142 / Chapter 5.6.1 --- Results --- p.142 / Chapter 5.7 --- Isolation of Active Compound in Fraction CM-C4 --- p.144 / Chapter 5.7.1 --- Materials and Methods --- p.145 / Chapter 5.7.2 --- Results --- p.146 / Chapter 5.8 --- Structure Elucidation of CM-C4a --- p.148 / Chapter 5.8.1 --- Materials and Methods --- p.148 / Chapter 5.8.2 --- Results --- p.149 / Chapter 5.9 --- Effect of Paeonol in Oral Glucose Tolerance Test --- p.152 / Chapter 5.9.1 --- Materials and Methods --- p.152 / Chapter 5.9.2 --- Results --- p.153 / Chapter 5.10 --- Discussion --- p.155 / Chapter Chapter6 --- General Discussion --- p.163 / Chapter 6.1 --- Introduction --- p.163 / Chapter 6.2 --- Summary of Research Findings --- p.164 / Chapter 6.3 --- Limitations and Improvements --- p.167 / Chapter 6.4 --- Future Directions --- p.169 / Chapter 6.5 --- Conclusion --- p.170 / Appendices --- p.172 / Appendix 1 Low Resolution EI Mass Spectrum of Paeonol Reference --- p.173 / Appendix 2 Low Resolution EI Mass Spectrum of CM-C4a --- p.174 / Appendix 3 High Resolution EI Mass Spectrum of Paeonol Reference --- p.175 / Appendix 4 High Resolution EI Mass Spectrum of CM-C4a --- p.176 / Appendix 5 1H-NMR Spectrum of Paeonol Reference --- p.177 / Appendix 6 1H-NMR Spectrum of CM-C4a --- p.178 / References --- p.179

Further mechanistic studies demonstrated that both Radix Rubiae and realgar were capable of inducing cellular apoptosis on HaCaT cells in a dose- and time-dependent manner as shown by morphological inspection, DNA fragmentation, TUNEL assay, cell cycle analysis, annexin V---PI staining and Western blot analysis. HPLC fingerprintings were constructed for quality control of the Radix Rubiae extract using mollugin as the chemical marker. Further phytochemical study found that ethyl acetate fraction of this herb possessed potent growth inhibition on HaCaT cells, with IC50 of 0.9 microg/ml. However, the chemical compounds obtained from commercial sources including mollugin, alizarin, purpurin, and quinizarin failed to induce growth inhibition. Meanwhile, arsenic trioxide, arsenic pentoxide and arsenic iodide, three arsenic salts presented in realgar, had significant anti-proliferative effect on HaCaT cells, with IC50 values of 2.4, 16 and 6.8 microM, respectively; and cellular apoptosis was found to be the underlying mechanism for the observed growth inhibitory activity. Furthermore, Radix Rubiae, realgar and arsenic compounds were also revealed to possess growth inhibition when evaluated in a PHA-activated PBMC model, and all of the substances except arsenic pentoxide significantly attenuated the release of inflammatory cytokines such as IFN-y, TNF-alpha and IL-2 in PBMC, indicating an anti-inflammatory effect. The in vivo mouse tail model experiments demonstrated that arsenic trioxide, arsenic pentoxide and arsenic iodide were able to markedly induce mouse tail keratinocyte differentiation, while such differentiation-modulating effect observed in the fraction of Radix Rubiae was only marginal. / In summary, Radix Rubiae and realgar extracts and three arsenic compounds have been identified and characterized as potential anti-psoriatic agents. The discoveries from the present PhD project not only help put the traditional use of these medicinal substances for psoriasis treatment on a scientific footing, but also open up new opportunities for their development into novel anti-psoriatic therapies. / Psoriasis, a chronic inflammatory skin disorder affecting approximately 2-3% of the population worldwide, is characterized histologically by hyperproliferation and aberrant differentiation of epidermal keratinocytes. Many conventional therapies are offered for psoriasis treatment but there exist problems such as unsatisfactory efficacy, side effects and drug resistance. Many patients therefore turn to alternative and complementary medicines for help. Traditionally, Chinese herbal medicine has been extensively used to treat psoriasis and produced promising clinical results. The present PhD study was conducted to investigate psoriasis-treating Chinese herbal medicines with an aim to identify effective anti-psoriatic agents. Sixty Chinese medicinal materials were selected for the screening project based on their ethnomedical use in psoriasis. The ethanolic extracts of these medicinal substances were evaluated for their anti-proliferative action on cultured HaCaT human keratinocytes using microplate SRB and MTT assays. Among them, the root of Rubia cordifolia L. (Radix Rubiae) and realgar were found to have significant anti-proliferative effects, with IC50 values of 1.4 and 6.6 microg/ml, respectively as measured by MTT assay, while they exerted mild significant cytotoxicity on the human fibroblast Hs-68 cell line. / Tse, Wai Pui. / Advisers: C. T. Che; Z. X. Lin. / Source: Dissertation Abstracts International, Volume: 70-09, Section: B, page: . / Thesis submitted in: October 2008. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2009. / Includes bibliographical references (leaves 298-340). / 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.

Xing Hongtao. / "February 2003." / Thesis (Ph.D.)--Chinese University of Hong Kong, 2003. / Includes bibliographical references (p. 149-176). / 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 Company, [200-] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Mode of access: World Wide Web. / Abstracts in English and Chinese.

Cao Hui. / Publication date from spine. / Thesis (Ph.D.)--Chinese University of Hong Kong, 1995. / Includes bibliographical references (leaves 180-194). / Acknowledgments --- p.v / Abstract --- p.vii / List of Tables --- p.xv / List of Figures --- p.xvii / Abbreviations and symbols --- p.xx / Chapter Chapter 1. --- General introduction / Chapter 1.1. --- Historical background --- p.1 / Chapter 1.2. --- Pharmacognostical development --- p.2 / Chapter 1.3. --- Importance of herb authentication --- p.3 / Chapter 1.4. --- Objective of study --- p.5 / Chapter Chapter 2. --- Literature review / Chapter 2.1. --- Botanical and taxonomic aspects --- p.9 / Chapter 2.1.1. --- Morphology --- p.9 / Chapter 2.1.2. --- Scientific names --- p.11 / Chapter 2.2. --- Chemical aspects --- p.13 / Chapter 2.3. --- Pharmacological aspects --- p.14 / Chapter 2.3.1. --- Antibacterial effect --- p.14 / Chapter 2.3.2. --- Antiphlogistic effect --- p.14 / Chapter 2.3.3. --- Antipyretic effect --- p.15 / Chapter 2.3.4. --- Effect in gastrointestinal propulsion --- p.15 / Chapter 2.3.5. --- Antineoplastic activity --- p.15 / Chapter 2.3.6. --- Hepatoprotective effect --- p.15 / Chapter 2.3.7. --- Inhibitory activity on enzymes --- p.17 / Chapter 2.3.8. --- Cardiovascular effect --- p.17 / Chapter 2.3.9. --- Acute toxicity (LD50) --- p.18 / Chapter 2.4. --- Pharmacognostical aspects --- p.18 / Chapter Chapter 3. --- Kudidan in Ben-cao literature / Chapter 3.1. --- Introduction --- p.23 / Chapter 3.2. --- Name evolution --- p.23 / Chapter 3.3. --- Natural distribution --- p.24 / Chapter 3.4. --- Characteristics --- p.25 / Chapter 3.5. --- Substitutions investigation --- p.26 / Chapter 3.6. --- Summary --- p.29 / Chapter Chapter 4. --- Morphological differences / Chapter 4.1. --- Plant identification --- p.36 / Chapter 4.1.1. --- Introduction --- p.36 / Chapter 4.1.2. --- Collection of voucher materials --- p.36 / Chapter 4.1.3. --- Plant morphology --- p.36 / Chapter 4.2. --- Macroscopical identification --- p.46 / Chapter 4.2.1. --- Introduction --- p.46 / Chapter 4.2.2. --- Materials and methods --- p.46 / Chapter 4.2.2.1. --- Commercial samples --- p.46 / Chapter 4.2.2.2. --- Macroscopical characteristics --- p.46 / Chapter 4.2.3. --- Results --- p.49 / Chapter Chapter 5. --- Histological identification / Chapter 5.1. --- Introduction --- p.58 / Chapter 5.2. --- Materials and methods --- p.59 / Chapter 5.2.1. --- Commercial samples --- p.59 / Chapter 5.2.1.1. --- Kudidan --- p.59 / Chapter 5.2.1.2. --- Pugongying --- p.59 / Chapter 5.2.1.3. --- Substitutes --- p.59 / Chapter 5.2.2. --- Authentic plant materials for comparison --- p.60 / Chapter 5.2.3. --- Methods --- p.60 / Chapter 5.2.3.1. --- Paraffin method --- p.60 / Chapter 5.2.3.2. --- Light microscopy --- p.62 / Chapter 5.2.3.3. --- Quantitative microscopy --- p.63 / Chapter 5.2.3.4. --- Scanning electron microscopy --- p.64 / Chapter 5.3. --- Results --- p.64 / Chapter 5.3.1. --- Microscopical characters of comparative plants --- p.64 / Chapter 5.3.2. --- Internal structures of herbs --- p.83 / Chapter 5.4. --- Discussion --- p.83 / Chapter Chapter 6. --- Chemical analysis / Chapter 6.1. --- Introduction --- p.99 / Chapter 6.2. --- Materials and methods --- p.100 / Chapter 6.2.1. --- Authentic samples --- p.100 / Chapter 6.2.2. --- Commercial samples --- p.100 / Chapter 6.2.3. --- Methods --- p.100 / Chapter 6.2.3.1. --- Isolation and characterization of standard substances --- p.100 / Chapter 6.2.3.2. --- Extraction of plant materials --- p.102 / Chapter 6.2.3.3. --- Thin layer chromatography --- p.102 / Chapter 6.3. --- Results and discussion --- p.104 / Chapter 6.3.1. --- TLC synopsis --- p.104 / Chapter 6.3.2. --- TLC analysis --- p.105 / Chapter 6.3.2.1. --- Qualitative evaluation of authentic plants --- p.105 / Chapter 6.3.2.2. --- Qualitative evaluation of commercial samples --- p.107 / Chapter 6.4. --- Summary --- p.107 / Chapter Chapter 7. --- Molecular fingerprinting / Chapter 7.1. --- Introduction --- p.115 / Chapter 7.2. --- Materials and methods --- p.120 / Chapter 7.2.1. --- Plant materials --- p.121 / Chapter 7.2.2. --- Herbal materials --- p.121 / Chapter 7.2.3. --- Total genomic DNA preparation --- p.121 / Chapter 7.2.3.1. --- CsCl/EtBr gradient method --- p.121 / Chapter 7.2.3.2. --- CTAB/CsCl gradient method --- p.123 / Chapter 7.2.3.3. --- CTAB miniprep method --- p.124 / Chapter 7.2.4. --- Qualitative analysis of genomic DNA --- p.125 / Chapter 7.2.5. --- Quantitative analysis of genomic DNA --- p.126 / Chapter 7.2.6. --- Genomic DNA fingerprinting --- p.126 / Chapter 7.2.6.1. --- DNA amplification --- p.126 / Chapter 7.2.6.1.1. --- AP-PCR --- p.127 / Chapter 7.2.1.1.2. --- RAPD --- p.128 / Chapter 7.2.6.2. --- Data analysis --- p.129 / Chapter 7.3. --- Results --- p.129 / Chapter 7.3.1. --- Studies on extraction of genomic DNA --- p.129 / Chapter 7.3.2. --- Genomic DNA fingerprinting by AP-PCR --- p.130 / Chapter 7.3.3. --- Genomic DNA fingerprinting by RAPD --- p.131 / Chapter 7.4. --- Discussion --- p.131 / Chapter 7.4.1. --- DNA extraction --- p.132 / Chapter 7.4.2. --- DNA fingerprinting of Kudidan --- p.136 / Chapter 7.4.3. --- Phylogenetic relationship between two genera Elephantopus and Pseudo-elephantopus of by DNA fingerprinting --- p.141 / Chapter Chapter 8. --- General summary and conclusion / Chapter 8.1. --- General summary --- p.165 / Chapter 8.1.1. --- Ben-cao investigation --- p.166 / Chapter 8.1.2. --- Investigation of commercial samples --- p.166 / Chapter 8.1.3. --- Histological characteristics --- p.167 / Chapter 8.1.4. --- Chemical analysis --- p.168 / Chapter 8.1.5. --- DNA fingerprinting --- p.168 / Chapter 8.2. --- Conclusion --- p.169 / Appendices / Chapter A) --- Solutions --- p.171 / Chapter B) --- Chinese characters cited in this Thesis --- p.173 / Chapter a) --- Herbal names --- p.173 / Chapter b) --- Book names --- p.175 / Chapter c) --- Personal names --- p.176 / Chapter d) --- Place names --- p.177 / Chapter e) --- Miscellaneous names --- p.179 / Bibliography --- p.180

Li, Yanhong. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2011. / Includes bibliographical references (leaves 192-212). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstract also in Chinese.

En Si Wang. / Thesis (Ph.D.)--Chinese University of Hong Kong, 1994. / Includes bibliographical references (leaves 126-140 (2nd gp.)). / Acknowledgements --- p.i / Contents --- p.ii / Abstract --- p.iv / List of Acronyms and Abbreviations --- p.vi / introduction --- p.1 / Chapter I. --- "Platelet Activating Factor (PAF)´ؤPast, Present, and Future" --- p.1 / Chapter I-1. --- What is PAF? --- p.1 / Chapter I-2. --- Biochemistry of PAF --- p.2 / Chapter I-2-1. --- Metabolic Cycle of PAF --- p.3 / Chapter I-2-1-A. --- Biosynthesis of PAF --- p.4 / Chapter I-2-1 -B. --- Inactivation of PAF --- p.6 / Chapter I-2-2. --- Role of Endogenous PAF in Cell --- p.7 / Chapter I-3. --- Chemistry of PAF --- p.8 / Chapter I-4. --- Pathobiology of PAF --- p.9 / Chapter II. --- PAF Receptor --- p.10 / Chapter II-1. --- Presence and Characteristics of PAF Receptor --- p.10 / Chapter II-l-l. --- Solubilization of PAF Receptor --- p.10 / Chapter II-1-2. --- G-Protein Involvement --- p.11 / Chapter II-1-3. --- Species Differences --- p.11 / Chapter II-1-4. --- Multiple Conformational States of PAF Receptor --- p.12 / Chapter II-1-5. --- PAF Receptor Heterogeneity --- p.12 / Chapter II-2. --- Putative Conformation of PAF Membrane Binding Sites --- p.13 / Chapter II-3. --- Recent Progress in PAF Receptor Research --- p.15 / Chapter III. --- PAF Receptor Antagonist --- p.18 / Chapter III-1. --- Classification of PAF Antagonists --- p.18 / Chapter III-2. --- Inhibition Types of PAF Receptor Antagonists --- p.19 / Chapter III-2-1. --- Nonspecific Inhibition of the Effects of PAF --- p.21 / Chapter III-2-2. --- Specific Inhibition of PAF --- p.22 / Chapter III-3. --- Recent Progress in PAF Receptor Antagonist Research --- p.22 / Chapter IV. --- Pharmacology and Syntheses of Spiro-Ether Structural Units --- p.26 / Chapter IV-1. --- Natural Products Containing Spiro-Ether and Related Structural Units --- p.30 / Chapter IV-1-1. --- Labdane Diterpenoids Containing Spiro-Ether Structural Units --- p.30 / Chapter IV-1-2. --- Leucodrin and Related Derivatives --- p.32 / Chapter IV-2. --- Synthetic Methods of Spiro-Ethers and Related Derivatives --- p.34 / Chapter V. --- Aim of the Present Work --- p.45 / RESULTS AND DISCUSSION --- p.47 / Chapter I. --- Isolation and Structure Elucidation of Prehispanolone (1) and Preleoheterin (3) --- p.47 / Chapter I-1. --- Material and Isolation --- p.47 / Chapter I-2. --- Structure Elucidation of Prehispanolone (1) and Preleoheterin (3) --- p.47 / Chapter II. --- Synthesis of Model Compounds --- p.53 / Chapter II-l. --- "Synthesis of 2-Methyl-1,7-dioxaspiro[4.4]nonane (137)" --- p.53 / Chapter II-2. --- "Synthesis of 2,2-Dimethyl-l,7-dioxaspiro[4.4]nonane (139)" --- p.68 / Chapter II-3. --- "Synthesis of 2,2-Diphenyl-1,7-dioxaspiro[4.4]nonane (141) and 2,2-Diphenyl-l,7-dioxaspiro[4.4]non-8-ene (142)" --- p.72 / Chapter III. --- "Partial Synthesis of 13R, 14,15-Dihydroprehispanolone (5)，13S,14,15-Di- hydroprehispanolone (135) and prehispanolone (1)" --- p.76 / CONCLUSION --- p.89 / EXPERIMENTAL SECTION --- p.91 / REFERENCES --- p.126 / APPENDIX --- p.141

影響全球數百萬的患者的慢性傷口，以其持續性過度發炎，纖維細胞增殖放緩，及血管生成受損為表徵。藥用草藥地黃已證明在大鼠糖尿足模型上有顯著傷口癒合作用。然而，關於地黃的炮製及其活性成分對此等傷口癒合的活動主要是未知的。 / 我們首先在地黃的炮製中，以抗一氧化氮（NO）和纖維細胞增殖實驗，確定了乾地黃表現出有效的傷口癒合活動。採用多方位生物活性導引分離（BGF），我們進一步研究乾地黃在抗炎，纖維細胞增殖，血管生成的活性成分，分別以抗NO產生，纖維細胞增殖，和TG(fli1:EGFP)y1/+(AB)斑馬魚芽血管生成模型為生物測定。此等具傷口癒合效果的活性成分將會作進一步研究。此外，我們會以電子細胞基質阻抗判斷(ECIS)的技術，對名為NF3(含RR的中草藥配方)在人類血管內皮細胞(HECV)上作體外血管生成及其信息的研究。 / 通過抗NO測定導引分離，我們證明活性萃取物C3比其粗提物擁有100倍更有效的抗炎效果。C3中含地黃苦甙元。地黃苦甙元可顯著地抑制NO的產生。C3中地黃苦甙元的存在可能是其具抗炎的原因。C3進一步以抑制一氧化氮合酶(iNOS)，環氧合酶-2(COX-2)和細胞白介素第六因子(IL-6)的基因，蛋白質，及/或中介物的表達，證明其舒緩炎症的作用。因此，C3可能對慢性傷口癒合中的炎症有治療作用。此外，我們發現先從水提的RR粗提物，再以乙酸乙酯提取的活性萃取物C2-B4，證明此萃取物在纖維細胞增殖中具最有效及劑量依存作用。 / 斑馬魚芽誘導模型導引分離顯示，C1-1萃取物在RR中具有最有效的血管生成作用。為斑馬魚芽誘導度身設計的30個血管生成相關基因顯示，C1-1廣泛地引發血管生成中的基因差異表達，特別在生長因子和血管穩定方面。就促進血管生成，C1-1進一步以體外人類微血管內皮細胞的血管生成檢測進行研究，結果發現C1-1具細胞移動及類血管生成能力。此外，降毛荚醛(norviburtinal)，在地黃提取物中首次被發現和具有血管生成作用。如此，C1-1和降毛荚醛為RR中血管生成作用的活性成分。 / 此外，應用ECIS技術，含RR配方的NF3能通過磷酯肌醇激酶(PI3K)及WiskottAldrich氏症候群神經元蛋白 (N-WASP) 路徑在人類血管內皮細胞（HECV）上誘導內皮細胞粘附，遷移及類血管生成。西方墨點法分析表明，NF3 在HECV上激活Akt和有絲分裂活化蛋白質激酶(MAPKs)的表達。這些誘導在各方面促進血管生成。因此，這顯示NF3能在分子及功能上激活血管生成作用的複雜性。此外，我們進一步支持ECIS在血管內皮細胞篩選傷口癒合劑中的高靈敏度。 / 總括而言，通過靶向抗炎，纖維細胞增殖增長和改善血管生成，各地黃生物活性導引化合物和萃取物，及其含RR的配方，可以在治療慢性傷口癒合上發揮功效。 / Chronic wounds, which influence millions of patients worldwide, are manifested with its sustained hyperinflammation, slackened fibroblast proliferation, and impaired angiogenesis. Agents retrieving these activities could facilitate the healing. Medicinal herb, Rehmanniae Radix (RR) demonstrated profound wound healing effect in rat diabetic foot model. However, the subtypes and the active components behind RR for such wound healing activities were largely unknown. / Here we firstly identified that dried RR, among its subtypes, exhibited potent wound healing activities through nitric oxide (NO) anti-inflammatory and fibroblast proliferation assays. Using multi-directional bioassay-guided fractionation (BGF), we further studied the active component(s) of dried RR in anti-inflammation, fibroblast proliferation, and angiogenesis, respectively, by anti-NO production, fibroblast proliferation, and TG(fli1:EGFP)[superscript y]¹/+(AB) zebrafish sprout angiogenesis model. Active component(s) of such wound healing effects were further characterized. Furthermore, with a RR-containing herbal formula, NF3, the in vitro angiogenic activities and its underlying signaling of NF3-treated human vascular endothelial cells (HECV) were studied using electric cell-substrate impedance sensing (ECIS) technology. / Via anti-NO assay-guided fractionation of dried RR, we demonstrated that the sub-fraction C3, possessed 100-fold more potent anti-inflammatory effect than that of the crude extract. Characterization of C3 showed that the anti-inflammatory activity could be partly due to the presence of rehmapicrogenin, which could significantly inhibit NO production. C3 was further demonstrated in blocking inflammation by inhibiting gene, protein, and/or mediator expression of inducible NO synthase, COX-2 and IL-6. Hence, C3 could be useful in treating inflammation in chronic wound healing. Additionally, we revealed that an active sub-fraction, C2-B4, from the ethyl acetate extract of the aqueous extract of RR, demonstrated the most potent and dose-dependent fibroblast proliferative effect. / Zebrafish sprout-inducing model-guided fractionation suggested C1-1 sub-fraction possessed the most potent angiogenesis effect in RR. A 30 tailor-made angiogenesis-associated gene panel designed for zebrafish sprout angiogenesis revealed that C1-1 triggered differential gene expression across wide angiogenic events, particularly concerned with those of growth factors and vessel stabilization. The pro-angiogenic activity was further supported by in vitro human microvascular endothelial cell-based angiogenesis assays, with C1-1 being pro-motogenic and tubule inducing. Also, norviburtinal was, for the first time, found in the extract of RR and possessed novel angiogenesis effect. Thus, C1-1 and norviburtinal were the active components responsible for the pro-angiogenesis effect of RR. / Moreover, RR-containing formula (NF3), which induced endothelial cell attachment, migration, and tubule formation in human vascular endothelial cell (HECV), could be mediated through PI3K and N-WASP pathways. Activated Akt and MAPK kinases expression in western blot analysis were also demonstrated in NF3-treated HECV. These inductions would promote angiogenesis at various levels. Hence, the complexity of angiogenesis effect activated by the NF3 treatment molecularly and functionally was shown, and we further supported the high sensitivity of ECIS in the screening of wound healing agents with endothelial cells. / In conclusion, through targeting anti-inflammation, elevated fibroblast proliferation and improved angiogenesis, our respective bioassay-guided active fractions and compounds in Rehmanniae Radix, and the RR-containing formula, could play beneficial uses in treating chronic wound healing. / 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. / Liu, Cheuk Lun. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2012. / Includes bibliographical references (leaves 221-249). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstract also in Chinese. / Table of Contents --- p.i / Abstract (in English) --- p.vi / Abstract (in Chinese) --- p.ix / Statement of Originality --- p.xiii / Acknowledgements --- p.xiv / Publications --- p.xiv / List of Tables --- p.xvii / List of Figures --- p.xviii / List of Abbreviations --- p.xxi / Chapter Chapter 1: --- Literature Review and Study Objectives / Chapter 1.1 --- Overview on wound healing / Chapter 1.1.1 --- Normal wound healing --- p.1 / Chapter 1.1.2 --- Chronic wound healing / Chapter 1.1.2.1 --- Venous ulcers --- p.8 / Chapter 1.1.2.2 --- Diabetic ulcers --- p.11 / Chapter 1.1.2.3 --- Mechanism of chronic wound healing --- p.13 / Chapter 1.1.2.4 --- Current treatments for chronic wound healing --- p.19 / Chapter 1.2 --- Rehmanniae Radix (RR) overview / Chapter 1.2.1 --- RR and its subtypes --- p.29 / Chapter 1.2.2 --- Chemistry of RR --- p.34 / Chapter 1.2.3 --- Pharmacology of RR --- p.38 / Chapter 1.2.3.1 --- RR and chronic wound healing / Chapter 1.3 --- Study objectives --- p.42 / Chapter Chapter 2: --- comparison of wound healing effect of the subtypes of rr / Chapter 2.1 --- Introduction --- p.43 / Chapter 2.2 --- Methods / Chapter 2.2.1 --- Preparation and authentication of subtypes of RR --- p.46 / Chapter 2.2.2 --- RAW 264.7 murine macrophage culture and sample treatment protocol, and cell viability test --- p.47 / Chapter 2.2.3 --- Nitric oxide inhibitory assay --- p.47 / Chapter 2.2.4 --- Hs27 human fibroblast culture and sample treatment protocol --- p.48 / Chapter 2.2.5 --- Fibroblast proliferation assay --- p.48 / Chapter 2.2.6 --- Statistical analysis --- p.49 / Chapter 2.3 --- Results --- p.63 / Chapter 2.3.1 --- Nitric oxide anti-inflammatory effect of the subtypes of RR --- p.49 / Chapter 2.3.2 --- Fibroblast proliferative effect of the subtypes of RR --- p.52 / Chapter 2.4 --- Discussion --- p.54 / Chapter Chapter 3: --- Fibroblast proliferative effect of RR / Chapter 3.1 --- Introduction --- p.57 / Chapter 3.2 --- Methods / Chapter 3.2.1 --- Preparation of aqueous extracts of RR --- p.61 / Chapter 3.2.2 --- Hs27 human fibroblast culture and sample treatment protocol --- p.61 / Chapter 3.2.3 --- Hs27 human fibroblast proliferation assay --- p.61 / Chapter 3.2.4 --- Bioassay-guided fractionation of RR --- p.61 / Chapter 3.2.5 --- LC-MS analysis of bioassay-guided fraction, C2-B4 --- p.65 / Chapter 3.2.6 --- Statistical analysis --- p.66 / Chapter 3.3 --- Results / Chapter 3.3.1 --- Fibroblast proliferative effect of RR aqueous crude extract and its bioassay-guided fractions --- p.67 / Chapter 3.3.2 --- Chemical structure of the isolated compounds --- p.70 / Chapter 3.3.3 --- LC-MS analysis of bioassay-guided fraction, C2-B4 --- p.72 / Chapter 3.4 --- Discussion --- p.73 / Chapter Chapter 4: --- Anti-inflammatory effect and its underlying mechanism of RR / Chapter 4.1 --- Introduction --- p.77 / Chapter 4.2 --- Methods / Chapter 4.2.1 --- Preparation of aqueous extracts of RR --- p.82 / Chapter 4.2.2 --- RAW 264.7 murine macrophage culture and sample treatment protocol, and cell viability test --- p.82 / Chapter 4.2.3 --- Assay for nitric oxide inhibitory effect using RAW264.7 cells --- p.83 / Chapter 4.2.4 --- Bioassay-guided fractionation of RR --- p.83 / Chapter 4.2.5 --- Ultra-high performance liquid chromatography/quadrupole time-of-flight mass spectrometry (UHPLC/QTOF-MS) analysis of sub-fraction, C3 --- p.85 / Chapter 4.2.6 --- Prostaglandin E2 (PGE2) and interleukin-6 (IL-6) assay --- p.87 / Chapter 4.2.7 --- Real-time PCR --- p.87 / Chapter 4.2.8 --- Western blot analysis --- p.89 / Chapter 4.2.9 --- Statistical analysis --- p.91 / Chapter 4.3 --- Results / Chapter 4.3.1 --- Nitric oxide inhibitory effects of L-NMMA, RR aqueous crude extract and its bioassay guided fractions --- p.92 / Chapter 4.3.2 --- LC-MS analysis of bioassay-guided fraction, C3 --- p.96 / Chapter 4.3.3 --- Suppression of inflammation-related mRNA expression level in macrophages by C3 --- p.97 / Chapter 4.3.4 --- Suppression of protein expression of inducible nitric oxide synthase and COX-2 in macrophages by C3 --- p.99 / Chapter 4.3.5 --- Inhibition of release of inflammatory cytokine in macrophages by C3 --- p.102 / Chapter 4.4 --- Discussion --- p.103 / Chapter Chapter 5: --- Angiogenesis effect and its underlying mechanism of RR / Chapter 5.1 --- Introduction --- p.111 / Chapter 5.2 --- Methods / Chapter 5.2.1 --- Preparation of aqueous extracts of RR --- p.113 / Chapter 5.2.2 --- Zebrafish culture --- p.113 / Chapter 5.2.3 --- Collection of zebrafish embryos and herbal treatment protocol --- p.115 / Chapter 5.2.4 --- Microinjection of vascular endothelial growth factor (VEGF) to zebrafish embryos --- p.116 / Chapter 5.2.5 --- Screening of zebrafish embryos using fluorescence microscopy --- p.116 / Chapter 5.2.6 --- Bioassay-guided fractionation of RR / Chapter 5.2.7 --- Isolation and structure elucidation of compound C2A --- p.120 / Chapter 5.2.8 --- Gas chromatographymass spectrometry (GC-MS) analysis of bioassay-guided fraction of RR, C1-1 --- p.120 / Chapter 5.2.9 --- Detection of zebrafish mRNA expression level by real-time PCR (RT-PCR) --- p.122 / Chapter 5.2.10 --- Human microvascular endothelial cell (HMEC-1) culture and sample treatment protocol --- p.125 / Chapter 5.2.11 --- HMEC-1 proliferation assay --- p.125 / Chapter 5.2.12 --- HMEC-1 scratch assay --- p.126 / Chapter 5.2.13 --- HMEC-1 tubule formation assay --- p.126 / Chapter 5.2.14 --- Statistical analysis --- p.127 / Chapter 5.3 --- Results / Chapter 5.3.1 --- Angiogenesis effects of RR aqueous crude extract, its bioassay-guided fractions and isolated compound, in zebrafish model --- p.128 / Chapter 5.3.2 --- Chemical structure and angiogenesis effect of the isolated compound C2A, norviburtinal --- p.133 / Chapter 5.3.3 --- Components of C1-1 from GC-MS analysis --- p.135 / Chapter 5.3.4 --- Angiogenesis effect of C1-1 in angiogenesis-related mRNA expression level in zebrafish --- p.137 / Chapter 5.3.5 --- Effect of endothelial cell proliferation of C1-1 in HMEC-1 cell --- p.142 / Chapter 5.3.6 --- Cell migration effect of C1-1 in HMEC-1 cell --- p.143 / Chapter 5.3.7 --- Tubule formation of C1-1 in HMEC-1 cell --- p.145 / Chapter 5.4 --- Discussion --- p.147 / Chapter Chapter 6: --- Angiogenesis effect and its underlying mechanism of RR AND AR-containing two-herbs formula, NF3 using ecis model / Chapter 6.1 --- Introduction --- p.165 / Chapter 6.2 --- Methods / Chapter 6.2.1 --- Preparation and authentication of aqueous extracts of NF3 --- p.176 / Chapter 6.2.2 --- Human vascular endothelial cells (HECV) culture --- p.177 / Chapter 6.2.3 --- HECV cell proliferation assay --- p.178 / Chapter 6.2.4 --- Scratch assay --- p.178 / Chapter 6.2.5 --- Tubule formation assay --- p.179 / Chapter 6.2.6 --- Electric cell-substrate impedance sensing (ECIS)-based cell attachment and motility assay --- p.180 / Chapter 6.2.7 --- Western blot analysis --- p.181 / Chapter 6.2.8 --- Statistical analysis --- p.182 / Chapter 6.3 --- Results / Chapter 6.3.1 --- LC-MS analysis of NF3 --- p.184 / Chapter 6.3.2 --- Effects of NF3, AR and RR on cell viability and migration of HECV --- p.185 / Chapter 6.3.3 --- Tubule formation effect of NF3, AR and RR in HECV cell --- p.188 / Chapter 6.3.4 --- Effects of NF3, AR, and RR on HECV cell attachment using ECIS model --- p.190 / Chapter 6.3.5 --- Effects of NF3, AR and RR on HECV cell migration using ECIS model --- p.191 / Chapter 6.3.6 --- Effects of NF3 on HECV for MAPK and Akt protein activation --- p.195 / Chapter 6.4 --- Discussion --- p.197 / Chapter Chapter 7: --- General Discussion and Conclusions / Chapter 7.1 --- General discussion and conclusions --- p.206 / Chapter 7.2 --- Limitation of the study --- p.215 / Chapter 7.3 --- Future work --- p.215 / Appendix --- p.218 / References --- p.221

銀屑病是一種免疫相關的慢性炎症性皮膚病，其發病率約占世界人口的1-3%，而現今仍然缺乏有效安全的根治方法。國內外使用中草藥治療銀屑病取得較好的療效，但目前缺少對其進行系統研究和開發。我們研究小組之前對61種常用治療銀屑病中藥進行篩選, 發現中藥茜草根和南蛇藤的乙醇提取物具有強大的抑制表皮細胞增生的作用，本博士研究課題的目的是確定新的安全有效的用于治療銀屑病的中藥化學成分, 並闡明其作用機制。 / 本研究篩選了28種存在于這兩種中藥中的化學單體成分，采用體外培養永生化的人類皮膚良性角質形成細胞株HaCaT, 應用MTT法, 繪制細胞生長曲線，獲得抑制50%細胞生長所需藥物濃度(IC50)。實驗結果發現1-羟基-3-甲基蒽醌（HMA）， 1，4-二氨基-2，3-（2-苯氧基乙氧基）蒽醌 （DBA）和南蛇藤表現了強大的抗表細胞生長作用，其48小時培養後的IC50分別爲17.9，15.8，1.1 μM. 值得一提的是這些化合物對正常人表皮角質細胞HEK和人類成纖維細胞Hs68只有相對輕微細胞毒性。 / 隨後進行的機理研究，通過熒光染色，DNA凝膠電泳，細胞周期檢測，流式細胞計檢測及Western blot 分析結果表明， HMA和南蛇藤素是通過誘導細胞凋亡作用抑制HaCaT細胞生長。其中南蛇藤素通過線粒體凋亡和死亡受體介導的兩種通路誘導細胞凋亡， 其誘導細胞凋亡作用與其抑制核因子-κB在HaCaT細胞中的表達和活化有關。 / 另一方面，DBA 抑制人體表皮角質細胞生長的作用機理在于其對角質細胞終末分化的誘導作用。DBA與HaCaT和HEK細胞共同培養96小時後，能顯著促進細胞角質化外膜形成，同時上調角蛋白K1/10，人體套膜蛋白，轉谷氨酰胺酶-1表達和下調角蛋白K5/14表達。而利用小鼠尾部鱗片表皮模型對HMA的外用制劑進行測試，結果顯示HMA誘導角質細胞終末分化能力較弱。 / 總而言之，本研究課題從兩種中藥中成功發現三個具有較強的抗銀屑病活性的化學單體成分，這些來自中藥的天然産物具有很好的開發成新的銀屑病治療外用制劑的應用前景。 / Psoriasis is an immunologically-mediated chronic inflammatory disease of the skin and joints affecting approximately 1-3% of the world’s population. Traditionally, Chinese medicine has been extensively used both inside and outside China for treating psoriasis with promising clinical results. Based on the promising findings in our previous screening project on 61 psoriasis-treating Chinese medicines which showed the root of Rubia cordifolia L. (Rubiae Radix) to have potent anti-psoriatic action, the present study aimed to identify active anti-psoriatic chemical constituents derived from Rubiae Radix and another Chinese herb namely Celastrus orbiculatus Thunb. and to elucidate the underlying mechanisms of action. / Microplate MTT assay was performed to evaluate the anti-proliferative actions of 28 selected Rubiae Radix-derived anthraquinones and other chemical ingredients on cultured HaCaT keratinocytes. Among them, 1-hydroxy-3-methyl-anthraquinone (HMA) and 1,4-diamino-2,3-bis(2-phenoxyethoxy)anthraquinone (DBA), as well as celastrol, a Celastrus orbiculatus-derived triterpene, were found to possess significant anti-proliferative action on HaCaT cells, with IC₅₀ value of 17.9, 15.8 and 1.1 μM, respectively. All DBA, HMA and celastrol showed only mild to moderate toxic effects on normal human keratinocyte HEK cells and human fibroblast Hs68 cells. / Mechanistically, celastrol and HMA was found to induce apoptosis in a dose-dependent manner in HaCaT cells as characterized by DNA fragmentation, phosphatidyl-serine externalization and activation of caspase 3. Further studies by flow cytometric and western blot analyses demonstrated that the celastrol-induced apoptosis on HaCaT cells was associated with the inhibition of NF-κB pathway and through caspase-related apoptotic pathway as characterized by activation of caspase proteins, regulation of Bcl-2 family proteins and depolarization of mitochondrial potential. / On the other hand, DBA showed an ability to induce terminal differentiation in cultured human keratinocytes and this capability is believed to be responsible for its growth inhibitory effects. DBA significantly accentuated the cornified envelope formation in HEK and HaCaT keratinocytes together with the augmentation of K1/K10, involucrin and transglutaminase 1 protein levels and decrease of expression of K5/K14 protein in DBA-treated cells. However, the subsequent in vivo study using a mouse tail model showed that HMA did not have significant effects on modulating keratinocyte terminal differentiation. / Taken together, our present PhD project successfully identified DBA, HMA and celastrol to have potent anti-psoriatic action on in vitro models, and the experimental findings render these naturally-occurring chemicals to be promising candidates for further development into anti-psoriatic pharmaceutical agents. / 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. / Zhou, Linli. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2012. / Includes bibliographical references (leaves 213-244). / 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 (English) --- p.i / Abstract (Chinese) --- p.iii / Publications --- p.v / Acknowledgements --- p.vii / Table of Contents --- p.viii / List of Figures --- p.xvii / List of Tables --- p.xxi / List of Abbreviations --- p.xxii / Chapter Chapter One --- General Introduction / Chapter 1.1 --- Psoriasis --- p.2 / Chapter 1.1.1 --- Structure of skin --- p.2 / Chapter 1.1.2 --- Epidemiology of psoriasis --- p.3 / Chapter 1.1.3 --- Pathogenesis --- p.5 / Chapter 1.1.4 --- Classification --- p.8 / Chapter 1.1.4.1 --- Nonpustular (plaque type) psoriasis --- p.9 / Chapter 1.1.4.2 --- Guttate psoriasis --- p.9 / Chapter 1.1.4.3 --- Pustular psoriasis --- p.9 / Chapter 1.1.4.4 --- Erythrodermic psoriasis --- p.10 / Chapter 1.1.4.5 --- Nail psoriasis --- p.11 / Chapter 1.1.4.6 --- Psoriatic arthritis --- p.11 / Chapter 1.1.5 --- Comorbidities --- p.13 / Chapter 1.2 --- Treatment of Psoriasis --- p.16 / Chapter 1.2.1 --- Conventional treatment for psoriasis --- p.16 / Chapter 1.2.1.1 --- Topical therapy --- p.16 / Chapter 1.2.1.2 --- Phototherapy --- p.19 / Chapter 1.2.1.3 --- Systemic therapy --- p.21 / Chapter 1.2.2 --- Targeted immunotherapy --- p.24 / Chapter 1.2.3 --- Combination, rotational and sequential therapy --- p.25 / Chapter 1.2.4 --- Complementary treatment --- p.26 / Chapter 1.3 --- Traditional Chinese Medicine for Psoriasis --- p.30 / Chapter 1.3.1 --- Prescriptions for psoriasis based on pattern differentiation --- p.30 / Chapter 1.3.2 --- Clinical and experimental study of TCM for psoriasis --- p.34 / Chapter 1.3.3 --- Possible action mechanisms of Chinese herbs for psoriasis --- p.34 / Chapter 1.3.4 --- Previous studies on TCM for psoriasis conducted by our research group --- p.35 / Chapter 1.4 --- Aims and Objectives of the Present Study --- p.38 / Chapter Chapter Two --- Phytochemical and Apoptotic Studies of Rubiae Radix-derived Anthraquinones and Other Related Compounds / Chapter 2.1 --- Introduction --- p.41 / Chapter 2.2 --- Selection and Screening of Rubiae Radix-derived anthraquinones and Other Related Compounds for Anti-proliferative Action on Cultured HaCaT Human Keratinocytes --- p.43 / Chapter 2.2.1 --- Introduction --- p.43 / Chapter 2.2.2 --- Materials and methods --- p.45 / Chapter 2.2.2.1 --- Procurement of Rubiae Radix-derived anthraquiones and other related compounds --- p.45 / Chapter 2.2.2.2 --- Purification of anthraquinones from Rubiae Radix --- p.50 / Chapter 2.2.2.3 --- General cell culture --- p.54 / Chapter 2.2.2.4 --- SRB assay --- p.55 / Chapter 2.2.2.5 --- MTT assay --- p.56 / Chapter 2.2.2.6 --- Assessment of synergistic or antagonistic effects between two active anthraquiones --- p.56 / Chapter 2.2.2.7 --- Statistical analysis --- p.57 / Chapter 2.2.3 --- Results --- p.57 / Chapter 2.2.3.1 --- Anti-proliferative effects of 35 Rubiae Radix fractions on HaCaT cells by SRB assay --- p.57 / Chapter 2.2.3.2 --- Anti-proliferative effects of the 27 anthraquinones and related compounds on HaCaT cells by SRB assay --- p.59 / Chapter 2.2.3.3 --- Confirmation of the anti-proliferative action of 8 active pure compounds using MTT assay --- p.61 / Chapter 2.2.3.4 --- Cytotoxic effects of 1-hydroxy-3-methyl-anthraquinone and1,4-diamino-2,3-bis(2-phenoxyethoxy)anthraquinone on the growth of HEK and Hs68 cells --- p.64 / Chapter 2.2.3.5 --- Drug interactions between different active anthraquinones --- p.67 / Chapter 2.2.4 --- Discussion --- p.69 / Chapter 2.3 --- Investigations of the Apoptotic Effects of DBA and HMA on HaCaT cells --- p.71 / Chapter 2.3.1 --- Introduction --- p.71 / Chapter 2.3.2 --- Materials and methods --- p.76 / Chapter 2.3.2.1 --- Chemicals --- p.76 / Chapter 2.3.2.2 --- General cell culture methods --- p.76 / Chapter 2.3.2.3 --- Cell cycle analysis with PI staining --- p.76 / Chapter 2.3.2.4 --- Hoechst fluorescence staining for morphological evaluation --- p.77 / Chapter 2.3.2.5 --- DNA fragmentation assay --- p.77 / Chapter 2.3.2.6 --- Detection of apoptosis by flow cytometry --- p.78 / Chapter 2.3.2.7 --- Prepare cytosol fraction of HaCaT cells --- p.79 / Chapter 2.3.2.8 --- Western blot analysis --- p.79 / Chapter 2.3.2.9 --- Statistical analysis --- p.80 / Chapter 2.3.3 --- Results --- p.76 / Chapter 2.3.3.1 --- Action of DBA and HMA on cell cycle progression --- p.80 / Chapter 2.3.3.2 --- Alteration of cellular morphology --- p.84 / Chapter 2.3.3.3 --- Detection of DNA fragmentation --- p.86 / Chapter 2.3.3.4 --- Quantitative analysis of apoptotic cells by annexin V-PI staining --- p.88 / Chapter 2.3.3.5 --- Activation of procaspase-3 and release of cytochrome c protein --- p.91 / Chapter 2.3.4 --- Discussion --- p.94 / Chapter 2.4 --- General Discussion --- p.97 / Chapter Chapter Three --- Effects of Rubiae Radix and Its-derived Anthraquinones on Keratinocyte Terminal Differentiation / Chapter 3.1 --- Introduction --- p.100 / Chapter 3.2 --- Materials and Methods --- p.105 / Chapter 3.2.1 --- Chemicals --- p.105 / Chapter 3.2.2 --- General cell culture --- p.105 / Chapter 3.2.3 --- Cornified envelope (CE) formation assay --- p.106 / Chapter 3.2.4 --- Western blot analysis --- p.107 / Chapter 3.2.4 --- Statistical analysis --- p.107 / Chapter 3.3 --- Results --- p.108 / Chapter 3.3.1 --- EA fraction of Rubiae Radix, DBA and HMA stimulates CE formation --- p.108 / Chapter 3.3.2 --- EA fraction of Rubiae Radix, DBA and HMA regulated TG1 expression and involucrin production in cultured human keratinocytes --- p.112 / Chapter 3.3.3 --- Regulation of cytokeratins by EA fraction of Rubiae Radix, DBA and HMA --- p.118 / Chapter 3.4 --- Discussion --- p.128 / Chapter Chapter Four --- Anti-psoriatic Action of Celastrol from Celastrus orbiculatus / Chapter 4.1 --- Introduction --- p.136 / Chapter 4.2 --- Anti-proliferative Action of Celastrol on Cultured Human Keratinocytes and Other Cell Types --- p.138 / Chapter 4.2.1 --- Introduction --- p.138 / Chapter 4.2.2 --- Materials and methods / Chapter 4.2.2.1 --- Chemicals --- p.138 / Chapter 4.2.2.2 --- General cell culture --- p.139 / Chapter 4.2.2.3 --- MTT assay --- p.139 / Chapter 4.2.2.4 --- Statistical analysis --- p.139 / Chapter 4.2.3 --- Results --- p.142 / Chapter 4.2.3.1 --- Anti-proliferative effect of celastrol on cultured cells --- p.142 / Chapter 4.2.4 --- Discussion --- p.145 / Chapter 4.3 --- Induction of Apoptosis by Celastrol on Human Keratinocytes --- p.146 / Chapter 4.3.1 --- Introduction --- p.146 / Chapter 4.3.2 --- Materials and methods --- p.146 / Chapter 4.3.2.1 --- Chemicals --- p.146 / Chapter 4.3.2.2 --- General cell culture --- p.147 / Chapter 4.3.2.3 --- Cell cycle analysis with PI staining --- p.147 / Chapter 4.3.2.4 --- Detection of apoptosis by flow cytometry --- p.147 / Chapter 4.3.2.5 --- Measurement of the mitochondrial membrane potential (ΔΨm) --- p.148 / Chapter 4.3.2.6 --- Western blot analysis --- p.148 / Chapter 4.3.2.7 --- Statistical analysis --- p.148 / Chapter 4.3.3 --- Results --- p.149 / Chapter 4.3.3.1 --- Induction of sub-G1 phase by celastrol on HaCaT cells --- p.149 / Chapter 4.3.3.2 --- Quantitative analysis of apoptotic cells by Annexin V-PI staining --- p.151 / Chapter 4.3.3.3 --- Alteration of ΔΨm --- p.153 / Chapter 4.3.3.4 --- Activation of caspase family protein --- p.155 / Chapter 4.3.3.5 --- Celastrol regulates the Bcl-2 family members --- p.159 / Chapter 4.3.4 --- Discussion --- p.161 / Chapter 4.4 --- Inhibition of NF-κB Transcription Factor Activation by Celastrol --- p.164 / Chapter 4.4.1 --- Introduction --- p.164 / Chapter 4.4.2 --- Materials and methods --- p.165 / Chapter 4.4.2.1 --- Chemicals --- p.165 / Chapter 4.4.2.2 --- General cell cultrue --- p.165 / Chapter 4.4.2.3 --- Western blot analysis --- p.165 / Chapter 4.4.2.4 --- Detect nuclear p65 by ELISA assay --- p.166 / Chapter 4.4.2.5 --- Statistical analysis --- p.166 / Chapter 4.4.3 --- Results --- p.167 / Chapter 4.4.3.1 --- Celastrol inhibited the NF-κB activation --- p.167 / Chapter 4.4.4 --- Discussion --- p.170 / Chapter 4.5 --- Induction of Terminal Differentiation by Celastrol --- p.173 / Chapter 4.5.1 --- Introduction --- p.173 / Chapter 4.5.2 --- Materials and methods --- p.174 / Chapter 4.5.2.1 --- Chemicals --- p.174 / Chapter 4.5.2.2 --- General cell culture --- p.174 / Chapter 4.5.2.3 --- CE formation assay --- p.174 / Chapter 4.5.2.4 --- Western blot analysis --- p.174 / Chapter 4.5.2.5 --- Statistical analysis --- p.174 / Chapter 4.5.3 --- Results --- p.175 / Chapter 4.5.3.1 --- Regulation of CE formation by celastrol --- p.175 / Chapter 4.5.3.2 --- Modulation of terminal differentiation markers by celastrol --- p.178 / Chapter 4.5.4 --- Discussion --- p.181 / Chapter 4.6 --- General Discussion --- p.183 / Chapter Chapter Five --- In vivo Anti-psoriatic Effects of Topical Preparation of 1-hydroxy-3-methyl-anthraquinone / Chapter 5.1 --- Introduction --- p.187 / Chapter 5.2 --- Material and Methods --- p.191 / Chapter 5.2.1 --- Chemicals --- p.191 / Chapter 5.2.2 --- Formulation of topical preparation containing HMA --- p.191 / Chapter 5.2.3 --- Mouse tail model --- p.192 / Chapter 5.2.4 --- Histopathological evaluation --- p.193 / Chapter 5.2.5 --- Statistical analysis --- p.194 / Chapter 5.3 --- Results --- p.195 / Chapter 5.3.1 --- Body weight profile --- p.195 / Chapter 5.3.2 --- Histological resutls --- p.197 / Chapter 5.4 --- Discussion --- p.201 / Chapter Chapter Six --- General Conclusions and Future Perspectives / Chapter 6.1 --- General Conclusions --- p.205 / Chapter 6.2 --- Future Perspectives --- p.210 / References / References by alphabetical order --- p.213