O objetivo desse estudo foi avaliar o efeito clínico e microbiológico residual, na saúde bucal de 40 crianças, cujas mães participaram de um programa preventivo longitudinal, finalizado há sete anos. Elas foram divididas em grupo teste (GT, n=26) e placebo (GP, n=14). Também fizeram parte do estudo, crianças, de mães que deixaram de participar, no decorrer do programa grupo controle, (GC, n=9). Foram avaliados os índices de cárie (OMS, 1999), de placa visível, de sangramento gengival (AINAMO; BAY, 1975), profundidade do sulco gengival à sondagem (GOMES-FILHO et al., 2006), contagem de estreptococos do grupo mutans (EGM) em UFC/mL de saliva e, fatores indicadores de risco à cárie dentária (FIRC). Foram feitas comparações intergrupos, intragrupos e, da amostra total, em diferentes momentos. Considerou-se nessas comparações os dados coletados ao final do programa preventivo (data base) e no controle após sete anos. Das avaliações intergrupos realizadas após sete anos, as que obtiveram resultados significantes foram: a contagem de EGM em UFC/mL de saliva, maior para o GC em relação ao GP e GT (Kruskall-Wallis, p= 0,014); o FIRC relativo ao motivo da consulta, com maior procura por tratamento preventivo para o GC (Quiquadrado, p=0,04) e maior procura por tratamento restaurador para o GP (Quiquadrado, p=0,01). Das avaliações intragrupos realizadas entre a data base e após sete anos, a que obteve resultado significante foi: maior ceo-s para o GP (ANOVA e Tukey, p=0,00). Das avaliações da amostra como um todo, realizadas após sete anos, as que obtiveram resultados significantes foram: relacionadas aos FIRC (Pearson, p<0,05 e r= -1 a 1), envolvendo fator socioeconômico; procura pelo cirurgião-dentista (pela mãe, para a criança, por motivo de dor), frequência de escovação, frequência de visitas ao cirurgião-dentista, frequência de escovação após o jantar e, escolaridade do pai; relacionadas aos índices de saúde bucal (Pearson, p< 0,05 e r= 0,97 a 1), envolvendo índice de cárie, índice de placa visível e, mancha branca. A avaliação do efeito residual do programa preventivo de cárie aplicado às mães como objetivo proteger seus filhos, auxilia na determinação de melhores estratégias, para a implementação de programas preventivos no futuro. Conclui-se pelos dados analisados que o programa foi capaz de manifestar alguns resultados positivos na população diretamente envolvida, após sete anos do seu término / The aim of this study was to evaluate the clinical and microbiological residual effect of a longitudinal preventive program directed to mothers on the oral health of 40 children, after seven years. The children were divided into two groups: test group (TG, n = 26) and placebo (PG, n = 14). An additional control group (CG, n = 9) corresponded to children whose mothers dropped out of the preventive program. The clinical oral health indices evaluated were: dental caries (WHO, 1999); visible plaque and gingival bleeding (AINAMO; BAY, 1975) and depth of the gingival sulcus probing (GOMES-FILHO et al., 2006). The count of mutans streptococci (MS) in CFU / mL of saliva, and risk factors indicators associated with dental caries (RFIC) were also evaluated. Comparisons were made between groups, within groups and with the total sample, at different times. The data obtained at the end of the preventive program (baseline) as well as those after seven years were considered in these comparisons. The intergroup evaluations performed after seven years, showed significant higher MS count in CFU / mL of saliva for CG when compared PG and TG (Kruskal-Wallis, p = 0.014); concerning the RFIC, the higher reason for consultation was preventive treatment for CG (Chi-square, p = 0.04) and restorative treatment for the PG (Chi-square, p = .01). The intragroup evaluation for PG performed between baseline and after seven years, showed a significant increase in the dmfs (ANOVA and Tukey\'s test, p = .00). Evaluations of the total sample performed after seven years, showed significant results related to the RFIC (Pearson, p <0.05 and r = ±1), involving socioeconomic factor, searching for dentist (by mother to the child, by reason of pain), frequency of brushing, frequency of visits to the dentist, frequent brushing after dinner, and father\'s education. Besides, factors related to oral health indices (Pearson, p <0.05 and r = 0.97 to 1), involving caries, plaque, and visible index and, white spot. The evaluation of the residual effect of a caries preventive program applied to mothers, aiming to maintain oral health of their children, helps to determine better strategies for the implementation of prevention programs in the future. Considering the data of the present study it is concluded that the program was able to express some positive results in the population directly involved, after seven years of its completion.

A prática odontológica expõe pacientes e profissionais a numerosos microorganismos e infecções. A transmissão pode ocorrer via sangue, saliva ou partículas dispersas no ar e por meio de contato direto ou indireto via instrumentos ou superfícies contaminadas. Assim, é necessário se utilizar precauções universais e procedimentos de controle de infecção para proteger tanto profissionais quanto pacientes. Na radiologia odontológica, como não há produção de aerosóis, muitos profissionais menosprezam os cuidados com a biossegurança. Nosso objetivo neste trabalho foi de alertar os profissionais com relação ao risco de contaminação cruzada por meio de todos os equipamentos envolvidos nos procedimentos radiológicos e apresentar medidas efetivas para controle de infecção cruzada por meio de um protocolo de biossegurança para uso em radiologia odontológica. / Patients and dentists are exposed to many different microorganisms and infections in dental practice. Infection may occur through the blood, saliva or airborne particles, as well as direct or indirect contact with contaminated instruments or surfaces. Thus, it is necessary to follow universal precautions and infection control procedures to protect both the dentist and the patient. Because there is no production of aerosol particles in oral radiology, many specialists ignore biosafety procedures. The purpose of this study was to warn specialists of the risk of cross contamination through the equipment used in radiology, as well as to propose effective measures for the control of cross infection, developing a biosafety protocol to be adopted in oral radiology.

Liu Bonnie Pui-ling. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2004. / Includes bibliographical references (leaves 134-163). / Abstracts in English and Chinese. / Acknowledgments --- p.i / Abbreviations --- p.ii / Abstract / English --- p.1 / Chinese --- p.3 / Chapter Chapter 1 --- General Introduction / Chapter 1.1 --- Colorectal Cancer Statistics --- p.5 / Chapter 1.2 --- Colon Cancer --- p.5 / Chapter 1.3 --- Treatment --- p.6 / Chapter 1.4 --- Effects of Cytotoxic Drug Treatment --- p.7 / Chapter 1.5 --- Cell Cycle --- p.8 / Chapter 1.6 --- Oxidases --- p.9 / Chapter 1.7 --- Chemistry of Novel β-carboline: Tangutorine --- p.11 / Chapter 1.8 --- Aim of Study --- p.14 / Chapter Chapter 2 --- Cytotoxicity / Chapter 2.1 --- Introduction --- p.15 / Chapter 2.2 --- Materials and Methods --- p.18 / Chapter 2.3 --- Results --- p.23 / Chapter 2.4 --- Discussion --- p.44 / Chapter Chapter 3 --- Oxidase Activity and Protein Oxidation / Chapter 3.1 --- Introduction --- p.48 / Chapter 3.2 --- Materials and Methods --- p.54 / Chapter 3.3 --- Results --- p.60 / Chapter 3.4 --- Discussion --- p.80 / Chapter Chapter 4 --- Cell Cycle / Chapter 4.1 --- Introduction --- p.89 / Chapter 4.2 --- Materials and Methods --- p.93 / Chapter 4.3 --- Results --- p.96 / Chapter 4.4 --- Discussion --- p.118 / Chapter Chapter 5 --- General Discussion --- p.126 / References --- p.134

冠狀動脈旁路移植術是治療合併左主幹及多隻冠狀動脈狹窄性病變患者的理想方法。然而靜脈橋失效極大地限制了冠脈搭橋手術的遠期療效。基於對靜脈橋失效潛在機制的研究，近年來開發出了多種針對性的防治手段。但是除了積極的降脂治療，目前尚未有其它療法獲得臨床證實可以有效改善靜脈橋遠期通暢率。所以，本研究旨在探索與防治靜脈橋再狹窄相關的新型生物標靶和防治策略。 / 我們應用豬大隱靜脈植入頸內動脈模型，觀察骨橋蛋白是否參與靜脈橋動脈化進程以及其與基質金屬蛋白酶功能活動的關係。我們發現骨橋蛋白表達在靜脈橋動脈化過程中顯著增加，並且與基質金屬蛋白酶2/9和增殖細胞數量的變化同步。此外，骨橋蛋白富集區域在靜脈橋內的再分佈與血管壁重構進程相關。這些結果表明, 骨橋蛋白積極參與了靜脈橋壁重構，而抑制骨橋蛋白表達作為防治靜脈橋失效的治療策略值得深入研究。 / 我們運用體外培養的方法研究了在高糖環境中骨成形蛋白4與靜脈內皮細胞舒張功能障礙的關係。我們發現，骨成形蛋白4在糖尿病患者的大隱靜脈與高糖培養的人臍靜脈內皮細胞中顯著增加；而骨成形蛋白4的高表達與靜脈血管內皮細胞依賴性舒張功能受損有關。本研究結果為解釋糖尿病患者有著較高的冠脈搭橋術後靜脈橋失效率提供了新證據，同時也為改善此類患者靜脈橋通暢率提出了潛在的治療靶點。 / 通過轉染金屬蛋白酶-3抑制物 (TIMP-3)基因來針對性地抑制血管中層平滑肌細胞的遷移和增殖，可以有效地減少靜脈橋新生內膜增生。基於前期研究，我們觀察了在豬模型中運用重組腺病毒轉載TIMP-3(RAdTIMP-3) 防治靜脈橋狹窄的遠期效果(3個月)。結果發現，即使在腺病毒載體已被清除的情況下，RAdTIMP-3對靜脈橋的良性保護作用仍持續存在。此外，我們通過比較術後7天與3個月獲取的橋血管中炎性標記物表達的差異，發現腺病毒轉染並未對靜脈橋造成長期的炎性損害。因此，我們認為RAd-TIMP3基因能夠安全有效地防治靜脈橋遠期狹窄。本研究結果為TIMP-3基因治療轉化至臨床實踐提供了可靠的前期證據。 / Coronary artery bypass grafting (CABG) remains the “gold standard“ for treating high-risk patients with unprotected left-main or multi-vessel coronary lesions. However, the long-term success of CABG is largely limited by an inadequate patency of saphenous vein grafts. To date, various therapeutic strategies targeting at the underlying mechanisms involved in the pathogenesis of vein graft failure (VGF) have been proposed and tested. However, apart from lipid-lowering therapy, no other intervention appears to have sustained benefits on improving vein graft patency in the clinical setting. Therefore, the aim of this study is to explore novel sets of molecular targets and effective therapeutic strategies to prevent VGF. / Novel molecules involved in the pathogenesis of vein graft failure / Using a porcine model, we assessed the involvement of osteopontin (OPN) in the venous arterialization and its relationship with the matrix metalloproteinases (MMPs). We found that the expression of OPN was significantly increased over the 3-month study period. Moreover, the expression of OPN at different time points well correlated with the fluctuating activities of MMP-2/9 and the number of proliferative cells. We also observed a time-dependent redistribution of OPN protein accumulating in different layers of the venous wall. These findings suggest a contributory role of OPN protein involved in the process of vein graft wall remodeling. / We used pig and human saphenous veins (SVs), as well as human umbilical endothelial cells (HUVECs), to investigate the changes of bone morphogenic protein-4 (BMP4) expression and its effects on endothelium-dependent relaxations (EDRs) under hyperglycemic conditions. Our results demonstrated a marked increase of BMP4 expression in SVs from diabetic patients and in HUVECs cultured with hyperglycemic medium. Moreover, such an increase of BMP4 contributes significantly to the impaired EDRs in venous conduits. Our findings add novel evidence that helps explain the high prevalence of VGF in diabetic patients undergoing CABG, and also suggest BMP4 as a potential therapeutic target to improve vein graft patency in this population. / Novel Therapeutic Strategy -- Gene Therapy / Aiming at blocking the development of neointima formation caused by vascular smooth muscle cells migration and proliferation, genetic transfection of tissue inhibitor of metalloproteinases-3 (TIMP-3) to vein grafts has shown promising results. Based on our previous study, we used recombinant adenoviruses that carry TIMP-3 (RAdTIMP-3) as a therapeutic gene to evaluate its long-term (3 months) effects on the pathological vein graft wall thickening in vivo. We found that the RAdTIMP-3-treated vein grafts had significantly reduced intimal and medial thickness compared with grafts from the control groups at 3 months, even after adenoviruses had already been cleared from transduced tissue. Furthermore, by assessing the amount of macrophages and the level of three inflammatory biomarkers within grafts harvested at 7 days and 3 months after implantation, we did not observe any detrimental effects of adenoviral transfection on the inflammatory status within the vein grafts. We therefore concluded that overexpression of TIMP-3 could effectively inhibit vein graft wall over-thickening in the longer-term. Our findings suggested the ex vivo RAdTIMP-3 gene therapy an attractive candidate for future clinical translation. / 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, Jia. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2012. / Includes bibliographical references (leaves 109-143). / 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.i / Declaration --- p.vii / Acknowledgement --- p.viii / Table of Contents --- p.x / List of Abbreviations --- p.xvi / List of Figures/Tables --- p.xviii / Chapter Chapter I --- INTRODUCTION --- p.1 / Chapter 1.1 --- SAPHENOUS VEIN GRAFTS IN CORONARY REVASCULARIZATION --- p.3 / Chapter 1.1.1 --- The use of venous conduits in CABG --- p.3 / Chapter 1.1.2 --- The long-term patency of saphenous vein grafts --- p.4 / Chapter 1.1.3 --- PCI for vein graft diseases --- p.6 / Chapter 1.1.4 --- Vein graft failure and adverse clinical outcomes --- p.7 / Chapter 1.2 --- MORPHOLOGY AND PHYSIOLOGY OF A NORMAL SAPHENOUS VEIN --- p.8 / Chapter 1.3 --- THE PATHOPHYSIOLOGY OF VEIN GRAFT FAILURE --- p.10 / Chapter 1.3.1 --- The quality of vein grafts prior to grafting --- p.10 / Chapter 1.3.1.1 --- Pre-existing endothelial dysfunction --- p.10 / Chapter 1.3.1.2 --- Surgical injuries --- p.11 / Chapter 1.3.2 --- Mechanisms of the pathological vein graft wall thickening --- p.12 / Chapter 1.3.2.1 --- Platelet activation and coagulant cascade --- p.13 / Chapter 1.3.2.2 --- Leukocytes recruitment and inflammation --- p.13 / Chapter 1.3.2.3 --- Hemodynamic forces --- p.14 / Chapter 1.3.2.4 --- Growth factors and VSMCs activation --- p.15 / Chapter 1.3.2.5 --- Contribution of adventitial and graft-extrinsic cells --- p.16 / Chapter 1.3.2.6 --- Oxidative stress --- p.17 / Chapter 1.3.2.7 --- Concomitant risk factors and vein graft atherosclerosis --- p.17 / Chapter 1.4 --- STRATEGIES FOR THE PREVENTION OF VEIN GRAFT FAILUR --- p.18 / Chapter 1.4.1 --- Minimizing surgical injuries --- p.18 / Chapter 1.4.2 --- Pharmacologic interventions --- p.19 / Chapter 1.4.3 --- External supports --- p.21 / Chapter 1.4.4 --- Genetic engineering of the vein graft --- p.23 / Chapter 1.4.4.1 --- Delivery systems --- p.23 / Chapter 1.4.4.2 --- Therapeutic strategies of the genetic modulation --- p.25 / Chapter 1.4.4.2.1 --- Antithrombotic and anticoagulant strategies --- p.25 / Chapter 1.4.4.2.2 --- Therapies for endothelial protection and regeneration --- p.27 / Chapter 1.4.4.2.3 --- Reducing inflammation and atherosclerosis --- p.28 / Chapter 1.4.4.2.4 --- Antioxidative therapy --- p.29 / Chapter 1.4.4.2.5 --- Therapies targeting at the cellular proliferation --- p.29 / Chapter 1.4.4.2.6 --- Inhibiting extracellular matrix reorganization --- p.31 / Chapter 1.5 --- CONCLUSIONS --- p.32 / Chapter Chapter II --- MATERIALS AND METHODS --- p.34 / Chapter 2.1 --- MATERIALS --- p.35 / Chapter 2.1.1 --- Reagents and equipment --- p.35 / Chapter 2.1.1.1 --- General materials and equipment for animal model --- p.35 / Chapter 2.1.1.2 --- General reagents and equipment for western blot --- p.35 / Chapter 2.1.1.3 --- General reagents and equipment for immunohistochemistry --- p.36 / Chapter 2.1.1.4 --- General reagents and equipment for venous ECs functional studies --- p.37 / Chapter 2.1.2 --- Buffers --- p.37 / Chapter 2.1.2.1 --- Buffers for human and animal samples --- p.37 / Chapter 2.1.2.2 --- Buffers for western blot --- p.38 / Chapter 2.1.2.3 --- Immunohistochemistry buffers --- p.39 / Chapter 2.1.2 --- Antibodies and adenoviral vectors --- p.41 / Chapter 2.2 --- METHODS --- p.41 / Chapter 2.2.1 --- Animal model --- p.41 / Chapter 2.2.2 --- Functional studies --- p.44 / Chapter 2.2.3 --- Human endothelial cells culture --- p.44 / Chapter 2.2.4 --- Western blot analysis --- p.45 / Chapter 2.2.5 --- Immunochemistry and immunofluorescence --- p.46 / Chapter Chapter III --- ROLE OF BMP4 IN VENOUS ENDOTHELIAL DYSFUNCTION --- p.47 / Chapter 3.1 --- INTRODUCTION --- p.48 / Chapter 3.2 --- MATERIALS AND METHODS --- p.49 / Chapter 3.2.1 --- Patient characteristics --- p.49 / Chapter 3.2.2 --- Preparation of human vein segments --- p.51 / Chapter 3.2.3 --- Porcine saphenous veins culture --- p.51 / Chapter 3.2.4 --- Functional studies of vein segments --- p.52 / Chapter 3.2.5 --- Cell culture --- p.53 / Chapter 3.3.6 --- Western blot analysis of BMP4 --- p.53 / Chapter 3.3.7 --- ROS measurement by dihydroethidium fluorescence imaging --- p.54 / Chapter 3.2.8 --- Statistical analysis --- p.54 / Chapter 3.3 --- RESULTS --- p.54 / Chapter 3.3.1 --- ACh-induced EDRs are impaired in diabetic veins --- p.54 / Chapter 3.3.2 --- The expression of BMP4 is upregulated under hyperglycemic condition --- p.55 / Chapter 3.3.3 --- BMP4 induces venous endothelial dysfunction in diabetes --- p.56 / Chapter 3.3.4 --- BMP4 impairs EDRs in cultured porcine saphenous veins --- p.58 / Chapter 3.4 --- DISCUSSION --- p.59 / Chapter 3.5 --- CONCLUSIONS --- p.62 / Chapter Chapter IV --- ROLE OF OSTEOPONTIN IN VEIN GRAFT REMODELING --- p.63 / Chapter 4.1 --- INTRODUCTION --- p.64 / Chapter 4.2 --- MATERIALS AND METHODS --- p.66 / Chapter 4.2.1 --- Surgical procedures --- p.66 / Chapter 4.2.2 --- Immunohistochemistry --- p.67 / Chapter 4.2.3 --- Western blot --- p.68 / Chapter 4.2.4 --- Gelatin zymography --- p.69 / Chapter 4.2.5 --- Cell proliferation --- p.69 / Chapter 4.2.6 --- Statistical analysis --- p.69 / Chapter 4.3 --- RESULTS --- p.70 / Chapter 4.3.1 --- Expression and redistribution of OPN protein within the venous wall --- p.70 / Chapter 4.3.2 --- The fluctuating expression of the matrix metalloproteinases --- p.72 / Chapter 4.3.3 --- Vascular smooth muscle cells proliferation --- p.74 / Chapter 4.4 --- DISCUSSION --- p.75 / Chapter 4.5 --- CONCLUIONS --- p.79 / Chapter Chapter V --- TIMP-3 GENE THERAPY FOR NEOINTIMA FORMATION --- p.81 / Chapter 5.1 --- INTRODUCTION --- p.82 / Chapter 5.2 --- MATERIALS AND METHODS --- p.84 / Chapter 5.2.1 --- Materials --- p.84 / Chapter 5.2.2 --- Grafting of pig saphenous veins and adenoviral transfection --- p.84 / Chapter 5.2.3 --- Histologic and morphometric analysis of the vein graft --- p.87 / Chapter 5.2.4 --- Immunocytochemistry --- p.87 / Chapter 5.2.5 --- Data analysis and statistics --- p.88 / Chapter 5.3 --- RESULTS --- p.89 / Chapter 5.3.1 --- Histologic and morphometric analysis of the vein graft --- p.89 / Chapter 5.3.2 --- Overexpression of TIMP-3 in porcine interposition grafts --- p.91 / Chapter 5.3.3 --- Endothelial cell coverage and VSMCs content --- p.92 / Chapter 5.3.4 --- Inflammation in vein grafts --- p.92 / Chapter 5.4 --- DISCUSSION --- p.97 / Chapter Chapter VI --- SUMMARY AND DISCUSSION OF MAJOR FINDINGS --- p.103 / Chapter 6.1 --- SUMMARY AND DISCUSSION --- p.104 / Chapter 6.1.1 --- The role of BMP4 in the pathogenesis of venous endothelial dysfunction --- p.104 / Chapter 6.1.2 --- The involvement of osteopontin in the process of vein graft remodeling --- p.105 / Chapter 6.1.3 --- Sustained benefits of adenoviruses-mediated TIMP-3 gene transfer in reducing vein graft neointima formation --- p.106 / Chapter 6.1.4 --- The inflammatory responses induced by adenoviral transfection --- p.106 / Chapter 6.1.5 --- Perspectives: novel therapeutic targets and clinical translation --- p.107 / Chapter 6.2 --- CONCLUSIONS --- p.108 / REFERENCES --- p.109 / PUBLICATION LIST --- p.144

Steroid-associated osteonecrosis (SAON) accounts for a major fraction in non-traumatic osteonecosis (ON) and generally has poor prognosis even after surgical interventions. This suggests that prevention is one of the important intervention strategies for SAON. So far, there is lacking of proven prevention modalities for SAON. / Study I was to establish an alternative SAON model. Based on the proposed pathogenesis of SAON that the intravascular thrombosis and extravascular marrow fat deposition are the two important contributors to the development of ON lesion, lipopolysaccharides (LPS) could induce vascular dysfunction and even thrombosis, and methylprednisolone (MPS) could induce the adipogenesis of marrow mesenchymal stem cells (MSCs). They were accordingly used in a combination for ON induction in animals. / Study II was to investigate the effect of Herbal Epimedium-derived formula for prevention of ON using the validated SAON animal model. Efficacy of the herbal Epimedium-derived formula was assessed for prevention of SAON using the animal model. Thirty adult male rabbits were used in this study. The ON incidence was set as the end-point for evaluation of the prevention efficay. For the potential intervention targets, the intravascular thrombosis and extravascular marrow fat formation were evaluated hematologically and histopathologically. The vascular structure and function were evaluated by advanced bioimaging modalities of micro-CT and MRI. / Study III was to investigate the bioactive compound(s) from the herbal Epimedium-derived herbal formula for prevention of SAON. Phytochemical analysis identified seven compounds in this efficacy-proven formula, with icariin as the major compound accounting for more than 80% in weight. Icariin was therefore tested for its prevention efficacy using the SAON animal model. / Study IV was to investigate the underlying mechanism(s) of bioactive compound Icariin in effective prevention of SAON using in vitro cell models. As activation of endothelial cells and adipogenesis of MSCs are suggested to be the two major events involving in vascular dysfunction and marrow fat formation in SAON animal model, Icariin were accordingly hypothesized to be able to prevent activation of endothelial cells and inhibit adipogenesis of MSCs. / Summary. After summarizing the major findings of these four logically interrelated studies, it was able to conclude that Icariin was the identified bioactive compound from the herbal Epimedium-derived formula, which was able to reduce the SAON incidence with inhibition of intravascular thrombosis and extravascular marrow fat formation in an established rabbit model. The underlying mechanisms might be related to its effects on protection of endothelial cells activation and inhibition of MSCs adipogenesis (This can be summarized in the following picture). This study provides a new bioactive agent Icariin for SAON prevention and potential future clinical application. (Abstract shortened by UMI.) / The following research questions were addressed in the present study: (1) Is there an alternative SAON animal model? (Study I); (2) Whether the herbal Epimedium-derived formula is able to prevent SAON in this animal model? (Study II); (3) What is the bioactive compound(s) in this herbal Epimedium-derived formula? (Study III); (4) How does this bioactive compound prevent SAON? (Study IV) / Sheng, Hui. / Adviser: Ling Qin. / Source: Dissertation Abstracts International, Volume: 70-06, Section: B, page: 3421. / 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.

激素性骨壞死是由於經常使用脈衝性激素處理非骨科性問題引起的一種常見的骨科疾病。在組織病理學上，激素性骨壞死指骨死亡，血管內血栓閉塞和血管外骨髓脂肪沉積會引起缺血導致骨修復不足。上游的分子細胞病理學機制研究表明間充質幹細胞細胞池活性下降，成骨細胞凋亡和骨小梁基質退變導致的不充分修復是激素性骨壞死發生的重要因素。 / 間充質幹細胞是骨髓的基質組成部分，具有分化成多種細胞的潛能。最近的研究表明，激素性骨壞死可能是骨細胞和/或間質幹細胞病變引起的一種疾病。研究發現，在接受類固醇治療而發生骨壞死的病人中，骨髓間充質幹細胞活性下降和分化潛能發生改變。在骨髓細胞中，激素能夠誘導脂肪發生。盛輝等發現來源於激素性骨壞死兔子中的間充質幹細胞成脂分化增強，謝新薈等進一步發現發生激素性骨壞死的兔子，骨缺損修復延遲，這可能是由激素導致的間充質幹細胞潛能發生改變引起的。綜合以上研究表明，間充質幹細胞在骨壞死發生和修復過程中起著重要作用。我們之前報導過淫羊藿黃酮（EFs）的腸代謝產物淫羊藿素Icaritin通過抑制血栓的形成和脂肪沉澱預防激素性骨壞死。最近，我們把Icaritin整合到聚乳酸聚乙醇酸共聚物/磷酸三鈣（PLGA/TCP）支架材料中，形成PLGA/TCP/Icaritin複合支架材料。我們發現PLGA/TCP/Icaritin複合材料可以促進激素性骨壞死骨缺損的修復，肌肉移植發現PLGA/TCP/Icaritin也能促進新生血管的發生。我們也發現單純PLGA/TCP複合材料也能夠促進激素性骨壞死骨缺損的修復，但是潛在的機制尚不清楚。 / 骨是一個高度血管化的組織，依賴於血管和骨細胞密切的時空連結維持骨骼的完整性。因此，血管生成在骨骼發育和骨折修復過程中發揮著舉足輕重的作用。血管為骨的發育和再生提供氧氣，為基質輸送刺激間充質細胞特異性成骨的重要信號，另一方面，骨為血管生成輸送生長因數和細胞。 / 本論文分為以下四個主要部分： / 第一部分: 研究Icaritin對人源間充質幹細胞分化的作用及其機制。流式細胞分選鑒定結果表明我們使用的人源間充質幹細胞能夠特異表達間充質幹細胞表面標誌物。MTT實驗結果顯示Icaritin不影響間充質幹細胞的增殖；分化實驗表明Icaritin在沒有成骨誘導試劑存在的情況下無法影響間充質幹細胞的分化。在成骨誘導試劑存在的情況下，Icaritin促進間充質幹細胞成骨分化，抑制其成脂分化；即時螢光實時定量聚合酶鏈式擴增（RT-PCR）結果顯示Icaritin在間充質幹細胞分化過程中上調成骨基因的表達，下調成脂基因表達。進一步研發發現在成骨分化過程中，Icaritin能夠促進BMP2和beta-catenin 蛋白的表達，而BMP2抑制劑Noggin能夠能夠逆轉Icaritin促進的成骨發生。這些發現表明Icaritin能夠促進而非誘導間充質幹細胞的成骨分化，Icaritin調解間充質幹細胞成骨分化具有BMP2信號通路依賴性。 / 第二部分: 評估激素性骨壞死兔源間充質幹細胞的分化潛能及Icaritin 對異常分化的間充質幹細胞分化潛能的影響。結果表明Icaritin促進正常兔源間充質幹細胞的成骨分化，抑制其成脂分化。激素性骨壞死兔源間充質幹細胞的成骨分化潛能降低，成脂分化升高；而Icaritin能夠劑量依賴性地部分恢復降低的成骨分化潛能，抑制升高的成脂分化活性。激素性骨壞死兔源間充質幹細胞的增殖活性也下降但是不能被Icaritin恢復。Icaritin對激素性骨壞死兔源間充質幹細胞中下降的VEGF的表達無影響。這些發現顯示間充質幹細胞的分化潛能在激素性骨壞死發生過程中遭到破壞，但是能夠被Icaritin部分恢復。 / 第三部分: 評估Icaritin對體外成血管的影響。我們對Icaritin對人臍帶靜脈內皮細胞（HUVECs）的增殖、遷移、管狀結構形成及成血管相關基因的表達的影響進行了檢測。結果表明Icaritin不影響HUVECs的增殖、遷移和管狀結構的形成；RT-PCR結果顯示Icaritin對HUVECs中的VEGF, HIF1a, FGF2 and TGF-beta表達也沒有影響。這些發現表明Icaritin在體外並不能直接作用于血管生成。結果謝新薈和陳詩慧等人的體內研究結果可以推測在骨缺損修復過程中，Icaritin通過促進成骨間接促進血管生成。 / 第四部分: 主要研究Icaritin及複合生物材料在體外體內對間充質幹細胞歸巢的影響。結果表明Iaritin能夠促進間充質幹細胞的遷移並上調血管細胞黏附分子1（VCAM1）的表達。複合材料PLGA/TCP和PLGA/TCP/Icaritin在體外培養的條件下能夠募集間充質幹細胞到材料周圍及進入材料。間充質幹細胞體外用修飾性超順磁性氧化鐵（SPIO@SiO₂-NH₂）納米顆粒標記後，其分化潛能依然保留，增殖和潛能能力稍微下降。兔激素性骨壞死造模完成後，股骨遠端髓芯減壓壞死骨缺損手術，PLGA/TCP和PLGA/TCP/Icaritin複合材料植入缺損孔道，同時把SPIO@SiO₂-NH₂標記的間充質幹細胞注射到距離缺損區20毫米的骨髓腔內。結果顯示只有標記的間充質幹細胞植入而沒有材料植入時，缺損區被脂肪細胞充滿，並沒有標記的間充質幹細胞出現，而在缺損區附近和遠離缺損區的部位有標記的間充質幹細胞出現。同時植入PLGA/TCP複合材料和標記的間充質幹細胞時，標記的間充質幹細胞出現在缺損區的材料中，在缺損區附近沒有標記的間充質幹細胞出現，而在遠離缺損區的部位，有標記的間充質幹細胞出現。同時植入PLGA/TCP/Icaritin和標記的間充質幹細胞時，得到跟植入PLGA/TCP複合材料和標記的間充質幹細胞相似的結果，但是在缺損區域，SPIO陽性的間充質幹細胞數目在PLGA/TCP和PLGA/TCP/Icaritin組別中並未發現有顯著性差異。以上發現表明Icaritin和PLGA/TCP複合材料能夠在體外和體內促進間充質幹細胞的歸巢。 / 綜上所述，複合支架材料PLGA/TCP/Icaritin通過調節間充質幹細胞的歸巢和分化促進激素性骨壞死骨缺損的修復。Icaritin通過BMP2和Wnt/beta-catenin通路調解間充質幹細胞的成骨分化。這是首次研究發現Icaritin及PLGA/TCP支架材料影響骨缺損修復過程中幹細胞歸巢，但是分子細胞生物學機制還需要進一步的研究。 / Steroid-associated osteonecrosis (SAON) is a common orthopaedic problem as the pulsed steroids are frequently prescribed for the treatment of non-orthopaedic medical conditions. Histopathologically, SAON refers to death of bone. Intravascular thrombus occlusion and extravascular marrow lipid deposition cause ischemia, which leads to an inadequate repair of the bone. Recent study revealed upstream pathological mechanism at cellular and molecular level. The decrease in activity of mesenchymal stem cell (MSC) pool, apoptosis of osteocytes, and trabecular bone matrix degeneration may cause bone inadequate repair, a key pathological feature found in SAON. / MSCs are the stromal component of bone marrow (BM) and have the potential to differentiate into several cell types. Recent studies have suggested that SAON may be a disease of bone cells and/or MSCs. With corticosteroid therapy in patients, the MSCs activity decreased and differentiation potential changed. Steroids have been also shown to produce adipogenesis in bone-marrow cells. It has been found adipogenesis of MSCs from SAON rabbits elevated (Sheng et al., 2007a) and bone defect repair was delayed in rabbits with SAON (Xie et al., 2011), this may be caused by altered MSCs potentials. All these findings imply MSCs play a vital role in SAON development and bone defect repair. It had been reported that Icaritin, an intestinal metabolite of Epimedium-derived avonoids (EF) reduced SAON incidence with inhibition of both thrombosis and lipid deposition (Zhang et al., 2009a). More recently, we found integrating Icaritin into PLGA/TCP to form PLGA/TCP/Icaritin composite scaffold could promote SAON bone defect repair and more neovascularization formed in an intramuscular implantation model, and further found PLGA/TCP scaffold only also could promote SAON bone defect repair in rabbits (Wang et al., 2012a). But the underlying mechanism remains unclear. / Bone is a highly vascularized tissue reliant on the close spatial and temporal connection between blood vessels and bone cells to maintain skeletal integrity. Angiogenesis thus plays a pivotal role in skeletal development and bone fracture repair. The vasculature supplies oxygen to developing and regenerating bone and also delivers critical signals to the stroma that stimulate MSC specification to promote bone formation and repair. On the other hand, bone also supplies growth factors and cells for angiogenesis. The content of this thesis is divided into the following four major parts: / Part I: to study the effect and molecular mechanism of Icaritin on the differentiation of human bone marrow-derived MSCs. Human MSC was identified first by flow cytometery and result showed our cultured human MSC expressed standard surface markers of MSCs. 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay showed that the proliferation ability of MSCs was not affected by Icaritin. Differentiation assay showed that without oseteogenic supplements (OS), Icaritin had no effect on osteogenic differentiation of MSCs. With presence of OS, Icaritin promoted osteogenic differentiation while inhibited adipogenic differentiation of MSCs. Real- time polymerase chain reaction (RT-PCR) showed that Icaritin up-regulated osteoblastic marker genes expression during osteogenic differentiation of MSCs and inhibited adipogenic gene expression. Further studies showed that Icaritin enhanced the protein expression of BMP2 and beta-catenin, while BMP2 inhibitor Noggin reversed the Icaritin-enhanced osteogenesis. All these findings indicated Icaritin possessed osteopromotive but not osteoinductive potentials during the differentiation of MSCs. Icaritin regulated osteogenic differentiation of MSCs in BMP2 pathway dependent manner. / Part II: to evaluate the differentiation potential of MSCs derived from rabbit with SAON and the effect of Icaritin on the altered differentiation of MSCs. The results showed that Icaritin promoted osteogenic differentiation while inhibited adipogenic differentiation of MSCs derived from normal rabbit. Osteogenic differentiation potential of mesenchymal stem cells derived from rabbit with SAON declined and Icaritin partly rescued the declined osteogenic differentiation potential in dose-dependent manner. Adipogenic differentiation potential of MSCs derived from rabbit with SAON enhanced while the enhanced adipogenesis could be depressed by Icaritin. The proliferation ability of MSCs derived from rabbit with SAON declined while could not be rescued by Icaritin. VEGF expression decreased in MSCs derived from rabbit with SAON but its expression could not be influenced by Icaritin. These findings showed that the differentiation potential of MSCs destroyed during SAON development and this potential could be partially restored by Icaritin. / Part III: to evaluate the in vitro angiogenic effect of Icaritin. The proliferation, migration and tube formation ability of human umbilical vein cells (HUVECs) were detected. The results showed that Icaritin did not affect HUVECs proliferation, migration and tube-like structure formation of HUVECs. Real time PCR showed that VEGF, HIF1a, FGF2 and TGF-beta expression in HUVECs was not changed when HUVECs were treated by Icaritin. These data indicated Icaritin did not directly impact angiogenesis in vitro. Combined with in vivo findings, we supposed Icaritin promoted angiogenesis through its enhanced osteogenesis during bone defect repair. / Part IV: to study Icaritin and scaffold impact on stem cell homing in vitro and in vivo. It was found Icaritin promoted the migration of rabbit MSCs and increased vascular cell adhesion molecule 1 (VCAM1) expression. Composite scaffolds PLGA/TCP and PLGA/TCP/Icaritin could recruit rabbit MSCs under in vitro culture condition. When labeled with SPIO@SiO₂-NH₂, the differentiation potential of rabbit MSCs retained while proliferation and migration ability of rabbit MSCs declined. Two weeks after SAON establishment, PLGA/TCP and PLGA/TCP/Icaritin scaffolds were implanted into the bone tunnel after core-decompression in initial necrotic bone defect in rabbits with SAON, immediately with SPIO@SiO₂-NH₂ labeled MSCs injected into bone marrow cavity locally. The results showed that without scaffold implantation, the tunnel was filled with fat cells and fibrotic tissues and there was no label MSC in the tunnel while there were more labeled cells appeared in bone marrow near the tunnel than far away the tunnel, with both PLGA/TCP and PLGA/TCP/Icaritin implantation, the labeled MSCs migrated into scaffold after its implantation into the bone tunnel while there was no labeled cell next to the tunnel but some were shown away from the tunnel. No significant difference was found in SPIO positive MSCs in bone tunnel between PLGA/TCP and PLGA/TCP/Icaritin group. The findings indicated that at least PLGA/TCP scaffold itself promoted MSCs homing in vitro and in vivo where the released icaritin could execute its osteopromotive effects. / In summary, the composite scaffold PLGA/TCP/Icaritin enhanced bone defect repair in rabbit with SAON by promoting homing and osteogenesis of MSCs. Icaritin promoted osteogenic differentiation of MSCs through BMP2 mediated signal pathway, such as Wnt/beta-catenin signal pathway. It is first time to report that PLGA/TCP scaffold promoted MSCs homing during bone defect repair, but underlying molecular and cellular mechanism need to be further studied. / 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. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Yao, Dong. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2012. / Includes bibliographical references (leaves 137-158). / Abstract also in Chinese; some appendixes also in Chinese. / ACKNOWLEDGEMENTS --- p.i / TABLE OF CONTENTS --- p.iii / ABSTRACT (IN ENGLISH) --- p.x / ABSTRACT (IN CHINESE) --- p.xiv / FLOWCHART --- p.xviii / LIST OF PUBLICATIONS --- p.xix / LIST OF ABBREVIATIONS --- p.xxi / LIST OF FIGURES --- p.xxiv / Chapter CHAPTER 1: --- Introduction --- p.1 / Chapter 1 --- Osteonecrosis --- p.2 / Chapter 1.1. --- Etiology --- p.2 / Chapter 1.2. --- Anatomy of femoral head --- p.3 / Chapter 1.3. --- Pathogenesis --- p.4 / Chapter 1.3.1. --- Intraosseous hypertension (Compartment Syndrome of Bone --- p.4 / Chapter 1.3.2. --- Intraosseous hypertension (Compartment Syndrome of Bone) --- p.4 / Chapter 1.3.3. --- Coagulation --- p.5 / Chapter 1.4. --- Development stages of osteonecrosis --- p.5 / Chapter 2. --- Steroids-associated osteonecrosis --- p.11 / Chapter 2.1. --- Epidemiology --- p.12 / Chapter 2.2. --- Histopathology --- p.12 / Chapter 2.3. --- Etiopathogenesis --- p.13 / Chapter 2.3.1. --- Steroid and fat metabolism --- p.14 / Chapter 2.3.2. --- Steroid and endothelial cells --- p.15 / Chapter 2.3.3. --- Steroid and coagulation --- p.16 / Chapter 2.3.4. --- Steroid and angiogenesis --- p.17 / Chapter 2.4. --- Steroid and mesenchymal stem cells (MSCs) --- p.18 / Chapter 2.5. --- Treatment strategies for SAON --- p.18 / Chapter 2.5.1. --- Prevention --- p.19 / Chapter 2.5.2. --- Nonoperative treatment --- p.19 / Chapter 2.5.3. --- Operative treatment --- p.19 / Chapter 2.5.3.1. --- Core decompression strategy --- p.20 / Chapter 2.5.3.2. --- Tissue engineering approach --- p.22 / Chapter 3. --- Epimedium-derived flavonoids (EFs) --- p.22 / Chapter 3.1. --- Icaritin -Intestinal metabolism of EFs --- p.24 / Chapter 3.1.1. --- Anti-tumor activity --- p..25 / Chapter 3.1.2. --- Neuroprotective effects --- p.25 / Chapter 3.1.3. --- Embryonic stem cells differentiation --- p.25 / Chapter 3.1.4. --- Osteogenic differentiation --- p.26 / Chapter 4. --- Poly lactic-co-glycolic acid / tricalcium phosphate (PLGA/TCP) scaffold --- p.26 / Chapter 5. --- PLGA/TCP/Icaritin --- p.28 / Chapter 6. --- Hypothesis of this study --- p.28 / Chapter 7. --- Objective --- p.29 / Chapter CHAPTER 2: --- The effect of phytomolecule Icaritin on differentiation of human mesenchymal stem cells in vitro --- p.30 / Chapter 1. --- Introduction --- p.31 / Chapter 2. --- Material and Methods --- p.33 / Chapter 2.1. --- Ethics --- p.33 / Chapter 2.2. --- Reagents and cell culture --- p.33 / Chapter 2.3. --- Surface phenotypes of human BM-MSCs --- p.33 / Chapter 2.4. --- Osteogenic and adipogenic differentiation of human BM-MSCs treated with Icaritin --- p.34 / Chapter 2.5. --- MTT assay for proliferation of BM-MSCs --- p.34 / Chapter 2.6. --- ALP staining --- p.35 / Chapter 2.7. --- ALP activity assay --- p.35 / Chapter 2.8. --- Alizarin Red S staining --- p.35 / Chapter 2.9. --- Oil Red O staining --- p.35 / Chapter 2.10. --- Ribonucleic acid (RNA) isolation --- p.36 / Chapter 2.11. --- Reverse transcription --- p.36 / Chapter 2.12. --- Real time polymerase chain reaction (RT-PCR) --- p.37 / Chapter 2.13. --- Western blotting --- p.37 / Chapter 2.14. --- Osteogenetic analysis of human MSCs after the addition of BMP2 inhibitor Noggin --- p.39 / Chapter 2.15. --- Statistical analysis --- p.39 / Chapter 3. --- Results --- p.40 / Chapter 3.1. --- Characterization of surface phenotypes of human BM-MSCs --- p.40 / Chapter 3.2. --- Icaritin had no effect on human mesenchymal stem cells (MSCs) proliferation --- p..41 / Chapter 3.3. --- Icaritin promoted osteogenic differentiation of MSCs in presence of osteogenic supplement --- p.42 / Chapter 3.4. --- Icaritin enhanced mineralization in osteogenic differentiation of MSCs only in presence of osteogenic supplement --- p.44 / Chapter 3.5. --- Icaritin upregulated mRNA expression of osteoblastic marker genes during osteogenic differentiation of MSCs --- p.45 / Chapter 3.6. --- Icaritin enhanced the protein expression of BMP2 and beta-catenin, while BMP2 inhibitor Noggin reversed the Icaritin-enhanced osteogenesis --- p..48 / Chapter 3.7. --- Icaritin inhibited fat droplets formation during adipogenic differentiation of MSCs --- p.50 / Chapter 4. --- Discussion --- p.52 / Chapter 5. --- Conclusion --- p.56 / Chapter CHAPTER 3: --- Icaritin rescued abnormal differentiation potential of MSCs derived from rabbit with SAON --- p.57 / Chapter 1. --- Introduction --- p.58 / Chapter 2. --- Methods and materials --- p.59 / Chapter 2.1. --- SAON model establishment --- p.59 / Chapter 2.2. --- Primary bone mesenchymal stem cells (BMSCs) isolation and culture --- p.60 / Chapter 2.3. --- Osteogenic and adipogenic differentiation of rabbit BM-MSCs treated with Icaritin --- p.61 / Chapter 2.4. --- MTT Assay for Proliferation of BM-MSCs --- p.62 / Chapter 2.5. --- ALP Staining --- p.62 / Chapter 2.6. --- ALP Activity Assay --- p.62 / Chapter 2.7. --- Alizarin Red S Staining --- p.62 / Chapter 2.8. --- Oil Red O Staining --- p.63 / Chapter 2.9. --- RNA Isolation --- p.63 / Chapter 2.10. --- Reverse transcription --- p.64 / Chapter 2.11. --- Real time Polymerase chain reaction (RT-PCR) --- p.64 / Chapter 2.12. --- Western blotting performance --- p.65 / Chapter 2.13. --- Statistical analysis --- p.65 / Chapter 3. --- Results --- p.66 / Chapter 3.1. --- The osteogenic differentiation potential declined while adipogenic differentiation ability elevated of MSCs derived from SAON rabbits --- p.66 / Chapter 3.2. --- The dose-dependent effect of Icaritin on osteogenic differentiation enhancement of MSCs from normal and SAON rabbits --- p.68 / Chapter 3.3. --- Icaritin inhibited adipogenic differentiation of MSCs both derived from normal and SAON rabbits --- p..71 / Chapter 3.4. --- PPAR-γ and aP2 proteins expression increased in SAON rabbit while inhibited by Icaritin both in normal and SAON rabbit --- p.74 / Chapter 3.5. --- Proliferation ability of MSCs derived from SAON rabbit declined and Icaritin had no effect on proliferation both derived from normal and SAON rabbit --- p.75 / Chapter 3.6. --- Icaritin had no effect on the expression of VEGF which decreased in MSCs derived SAON --- p.76 / Chapter 4. --- Discussion --- p.76 / Chapter 5. --- Conclusion --- p.81 / Chapter CHAPTER 4: --- The effect of Icaritin on angiogenesis in vitro --- p.82 / Chapter 1. --- Introduction --- p.83 / Chapter 2. --- Material and Methods --- p.85 / Chapter 2.1. --- Cell culture --- p.85 / Chapter 2.2. --- Proliferation assay --- p.85 / Chapter 2.3. --- Scratch-wound healing assay --- p..86 / Chapter 2.4. --- Migration Assay --- p.86 / Chapter 2.5. --- In vitro Angiogenesis Assay --- p.87 / Chapter 2.6. --- RNA Isolation and Real-time PCR Performance --- p.87 / Chapter 2.7. --- Statistical Analysis --- p.88 / Chapter 3. --- Results --- p.88 / Chapter 3.1. --- Icaritin did not affect HUVECs migration --- p.88 / Chapter 3.2. --- Icaritin had no effect on tube formation on growth factors reduced Matrigel --- p.92 / Chapter 3.3. --- Icaritin had no effect on HUVECs proliferation --- p.94 / Chapter 3.4. --- Icaritin did not change the angiogenesis related gene expression --- p.95 / Chapter 4. --- Discussion --- p.96 / Chapter 5. --- Conclusion --- p.100 / Chapter CHAPTER 5: --- Effect of PLGA/TCP and PLGA/TCP/Icaritin composite scaffolds on stem cell homing during bone defect repair with SAON --- p.101 / Chapter 1. --- Introduction --- p.102 / Chapter 2. --- Material and Methods --- p.106 / Chapter 2.1. --- Preparation of porous PLGA/TCP/Icaritin composite scaffolds --- p.106 / Chapter 2.2. --- Primary bone mesenchymal stem cells (BMSCs) isolation and culture --- p.106 / Chapter 2.3. --- Wound healing assay --- p.107 / Chapter 2.4. --- In vitro MSCs recruitment assay of scaffolds --- p.107 / Chapter 2.5. --- MSCs labeling with SPIO@SiO2-NH2 nanoparticle --- p.108 / Chapter 2.6. --- Prussian blue staining --- p.108 / Chapter 2.7. --- MTT assay for SPIO@SiO2-NH2 labeled MSCs --- p.108 / Chapter 2.8. --- Osteogenic and adipogenic differentiation of SPIO@SiO2-NH2 labeled MSCs --- p.109 / Chapter 2.9. --- Real time PCR --- p.109 / Chapter 2.10. --- Animal model establishment --- p.109 / Chapter 2.11. --- Descriptive histology and histomorphometry --- p.110 / Chapter 2.12. --- In vivo magnetic resonance imaging (MRI) of nanoparticle-labeled MSCs --- p.112 / Chapter 2.13. --- Statistical analysis --- p.112 / Chapter 3. --- Results --- p.112 / Chapter 3.1. --- Icaritin promoted MSCs migration in vitro --- p.112 / Chapter 3.2. --- PLGA/TCP and PLGA/TCP/Icaritin recruited MSCs when incubated in vitro --- p.114 / Chapter 3.3. --- Stem cell potentials of MSC after SPIO@SiO2-NH2 labeling --- p.118 / Chapter 3.4. --- PLGA/TCP and PLGA/TCP/Icaritin promoted MSCs homing in vivo --- p.122 / Chapter 4. --- Discussion --- p.126 / Chapter 5. --- Conclusion --- p.136 / Chapter CHAPTER 6: --- Summary of the study and future research --- p.137 / Chapter 1. --- Summary of the study --- p.138 / Chapter 2. --- Limitations and further studies --- p.139 / APPENDIXES --- p.142 / REFERENCES --- p.147

Chan Sieu Gaen. / Year shown on spine: 1997. / Questionnaries in Chinese and English. / Thesis (M.Phil.)--Chinese University of Hong Kong, 1996. / Includes bibliographical references. / Acknowledgement / List of tables --- p.i / Abstract --- p.vi / Chapter Chapter 1 --- Introduction --- p.1 / Chapter 1.0 --- Demographic changes of the population / Chapter 2.0 --- Falls / Chapter 2.1 --- Significance of the problem / Chapter 2.2 --- Physical consequences of falls / Chapter 2.3 --- Psychological consequences of falls / Chapter 2.4 --- Impact on health services / Chapter 2.5 --- Mortality / Chapter 2.6 --- Economic consequences / Chapter Chapter 2 --- Risk factors for falls --- p.12 / Chapter 2.1 --- Environmental hazards and circumstances of falls / Chapter 2.2 --- Social demographic factors / Chapter 2.3 --- Health status / Chapter 2.4 --- History of falls / Chapter 2.5 --- Mental/Depression / Chapter 2.6 --- Health behavior / Chapter 2.7 --- Bone mass and anthropometric factors / Chapter 2.8 --- Risk factors for multiple falls / Chapter 2.9 --- Fall injuries / Chapter 2.10 --- Additive effects of risk factors / Chapter Chapter 3 --- "Limitation of previous studies, objectives and study methods" --- p.31 / Chapter 3.1 --- Hospital reports and instiutional-based studies / Chapter 3.2 --- Study design / Chapter 3.3 --- Selection of subjects / Chapter 3.4 --- Classification and definition of falls / Chapter 3.5 --- Studies on falls in Hong Kong / Chapter 3.6 --- Hong Kong study / Chapter 3.7 --- Study methods / Chapter 3.8 --- Data collection / Chapter 3.9 --- Data analysis / Chapter Chapter 4 --- Characteristics of respondents and distribution of falls --- p.48 / Chapter 4.1 --- Characteristics of respondents / Chapter 4.2 --- Proportion of fallers and rate of falls / Chapter 4.3 --- Discussion / Chapter Chapter 5 --- Circumstances and consequences of falls --- p.65 / Chapter 5.1 --- Time and place of falls / Chapter 5.2 --- Activity during and reasons for falls / Chapter 5.3 --- Predisposing factors for falls / Chapter 5.4 --- Consequences of falls / Chapter 5.5 --- Discussion / Chapter Chapter 6 --- Characteristics of fallers and nonfallers --- p.85 / Chapter 6.1 --- Sociodemographic characteristics / Chapter 6.2 --- Health status / Chapter 6.3 --- Health behavior / Chapter 6.4 --- Bone mass and body measurements / Chapter Chapter 7 --- Risk factors associated with fallers --- p.99 / Chapter 7.1 --- Risk factors for fallers (including single and multiple) / Chapter 7.2 --- Multiple fallers / Chapter 7.3 --- Risk factors for single and multiple fallers (vs. non- fallers) / Chapter 7.4 --- Comparison of risk factors associated with major injurious falls vs. those with non-major injurious falls / Chapter 7.5 --- Independent predictors of fallers / Chapter 7.6 --- The additive effects of independent risk factors / Chapter Chapter 8 --- Discussion on risk factors associated with falls --- p.131 / Chapter 8.1 --- Social demographic factors and falls / Chapter 8.2 --- Acute illnesses/chronic diseases / Chapter 8.3 --- Medical consultation and hospitalization / Chapter 8.4 --- History of falls / Chapter 8.5 --- Perceived health status and ADL / Chapter 8.6 --- Mental status / Chapter 8.7 --- Health behavior / Chapter 8.8 --- Physical measurement and neuromuscular impairment / Chapter 8.9 --- Vision / Chapter 8.10 --- Medications / Chapter 8.11 --- Fallers with occasional falls/multiple falls / Chapter 8.12 --- Additive effect of risk factors / Chapter Chapter 9 --- Conclusion and recommendations --- p.147 / Chapter 9.1 --- Summary of findings / Chapter 9.2 --- Confirmation of risk factors found in baseline cross-sectional study / Chapter 9.3 --- Limitation of the present study / Chapter 9.4 --- Strength of the present study / Chapter 9.5 --- Recommendations on areas for further research / Chapter 9.6 --- Recommendations on prevention and intervention measures / References --- p.155 / Appendix / Calculation of sample size --- p.1 / Pooled logistic regression analysis on risk factors associated with falls --- p.2 / Baseline questionnaire --- p.3 / First follow-up questionnaire --- p.4 / Fall ascertainment questionnaire --- p.5

by Cheung Ngai Man, Edmund. / Thesis (M.Phil.)--Chinese University of Hong Kong, 1997. / Includes bibliographical references (leaves 73-83). / Acknowledgments --- p.i / Abstract --- p.iii / List of Figures --- p.viii / List of Tables --- p.xi / Chapters / Chapter 1 --- Introduction --- p.1 / Chapter 1.1 --- Intramedullary nailing --- p.1 / Chapter 1.2 --- Reaming technique for intramedullary nailing --- p.3 / Chapter 1.3 --- The relationship between pulmonary fat embolism and reaming technique --- p.7 / Chapter 1.4 --- Objectives --- p.10 / Chapter 2 --- Methodology --- p.12 / Chapter 2.1 --- The measurement of the intramedullary pressure --- p.12 / Chapter 2.1.1 --- Animal model --- p.12 / Chapter 2.1.2 --- Intramedullary pressure measurement device --- p.12 / Chapter 2.1.3 --- Operative procedure --- p.14 / Chapter 2.1.4 --- Intramedullary pressure measurement --- p.16 / Chapter 2.2 --- The measurement of the plasma lipids and marrow lipids --- p.19 / Chapter 2.2.1 --- Samples collection --- p.19 / Chapter 2.2.2 --- Lipid extraction --- p.19 / Chapter 2.2.3 --- Thin layer chromatography --- p.20 / Chapter 2.2.4 --- Methylation --- p.24 / Chapter 2.2.5 --- Gas chromatographic analysis --- p.24 / Chapter 2.3 --- The measurement of the pulmonary lipids and fat emboli --- p.27 / Chapter 2.3.1 --- Pulmonary tissue collection --- p.27 / Chapter 2.3.2 --- Preparation for measurement of pulmonary lipids --- p.27 / Chapter 2.3.3 --- Fat emboli staining --- p.27 / Chapter 2.3.4 --- Image analysis --- p.28 / Chapter 2.4 --- Statistical analysis --- p.31 / Chapter 3 --- Results --- p.32 / Chapter 3.1 --- Intramedullary pressure measurement --- p.32 / Chapter 3.2 --- The analysis of bone marrow lipids --- p.34 / Chapter 3.3 --- The changes of the plasma lipids during reaming --- p.39 / Chapter 3.4 --- The measurement of the pulmonary fat emboli --- p.44 / Chapter 3.5 --- The relationship between the intramedullary pressure and plasma lipids and pulmonary fat intravasation --- p.52 / Chapter 4 --- Discuss --- p.55 / Chapter 4.1 --- The experimental design --- p.55 / Chapter 4.2 --- The change of the intramedullary pressures --- p.57 / Chapter 4.3 --- The application of the gas chromatography --- p.59 / Chapter 4.4 --- The composition of bone marrow lipids --- p.62 / Chapter 4.5 --- The changes of plasma lipids --- p.63 / Chapter 4.6 --- The pulmonary fat embolisation --- p.65 / Chapter 5 --- Conclusion --- p.69 / Chapter 6 --- Future direction on this study --- p.71 / References --- p.73 / Appendix --- p.84 / Chapter 1 --- The operation of the IM Press device --- p.84 / Chapter 2 --- The calibration of the IM Press --- p.85 / Chapter 3 --- The preparation of the internal standards for the lipid analysis --- p.89 / Chapter 4 --- The composition of the bone marrow lipids --- p.91 / Chapter 5 --- The composition of plasma lipids --- p.95 / Chapter 6 --- The composition of pulmonary lipids --- p.101 / Chapter 7 --- The measurement of the pulmonary fat emboli --- p.105

A Atenção Domiciliar (AD) é uma modalidade de assistência que deve ser concebida como a capacidade de mobilizar, articular e colocar em prática conhecimentos, habilidades e atitudes necessárias ao desempenho efetivo das competências dos enfermeiros. Em se tratando do risco de infecção as pesquisas nessa área são escassas no Brasil, principalmente na perspectiva das ações preventivas. O estudo representa uma contribuição para com a atenção domiciliar, no que concerne às competências do enfermeiro em atuar nos fatores de risco para infecção e, assim, avançar nas diretrizes de prevenção e controle da infecção. Trata-se de um estudo de natureza quase-experimental considerando a primeira etapa e, pesquisa metodológica na construção das diretrizes. Utilizou-se como referencial teórico os princípios de Durand (2000) e, a avaliação de consenso foi subsidiada na técnica Delphi. O cenário envolveu as Unidades Básicas de Saúde do município de Teresina - PI. Os participantes foram divididos em dois grupos sendo um com 19 enfermeiros atuantes nas equipes da ESF e, o outro grupo com 15 pesquisadores/enfermeiros recrutados pelo método snow-ball (representando as quatro diferentes regiões do País). Para a organização dos dados utilizou-se o software Alceste 4.8, e a análise estatística descritiva foi ordenada em quartis. A lista de competências, referente à primeira rodada da Técnica Delphi totalizou 48 itens, sendo 26 classificadas como competências gerais e 22 específicas. Na segunda rodada, exclui-se apenas um item após análise de concordância entre os sujeitos. E, na terceira rodada foram mantidas as 47 competências, das quais as classificaram em três dimensões: conhecimentos, habilidades e atitudes. Esta organização tomou como base o referencial teórico, bem como as justificativas dos participantes. Dessa forma, 61 competências foram identificadas quando analisadas a luz do referencial teórico, e, articulada as praticas de prevenção e controle da infecção no ambiente domiciliar. Observa-se que, embora de modos heterogêneos, o conceito de competência para a prevenção da infecção tem sido apropriado de modo fértil pelos enfermeiros. Por fim, os resultados desse estudo mostram a relevância da atuação dos enfermeiros no manejo dos riscos de infecção, na manutenção do ambiente domiciliar biologicamente seguro, e no enfrentamento dos desafios da utilização de políticas públicas direcionadas para esse foco. Conclui-se que as práticas de prevenção e controle da infecção na atenção domiciliar são competências fundamentais reconhecidas pelos enfermeiros e, precisam ser incrementadas na formação em Enfermagem. / Home Care (AD) is a form of assistance that should be conceived as the ability to mobilize, articulate and apply knowledge, skills and attitudes necessary for the effective performance of the competences of nurses. In what concerns the risk of infection, research in this area is scarce in Brazil, especially in the perspective of preventive actions. This study represents a contribution to home care, in relation to the skills of nurses to act on risk factors for infection and thus advance the guidelines for prevention and control of infections. This is a study of quasi-experimental nature and, considering the second step in constructing the guidelines, a methodological research. Durand (2000) principles were used as theoretical references and the consensus evaluation was supported in Delphi techniques. The scenario involved Basic Health Units in the city of Teresina - PI. The subjects were divided into two groups being the first with 19 nurses working in the ESF teams for at least two years. The second group was attended by 15 researcher nurses who were recruited by the snow-ball method, until the establishment of a network representing four different regions of the country. For data organization Alceste4.8 software, and statistical, descriptive analysis sorted in quartiles were utilized. The list of skills on the first round of the Delphi totaled 48 items, with 26 classified as general and 22 specific competence. In the second round, after the analysis of agreement among subjects one competence was excluded, for not reaching the level of consensus determined. And in the third and final round of the Delphi technique were maintained 47 skills from the analysis of consensus, which were divided among the three dimensions, namely: knowledge, skills and attitudes. This organization was based on the theoretical reference as well as the justifications of the subjects, as a result, 61 competences were developed. The respective items were analyzed in light of the theoretical references, and we proceeded to the recovery of such knowledge, skills and attitudes for the prevention and control of infection in home environment. It is observed that although in heterogeneous ways, the concept of competence for the prevention of infection has been appropriated in a fertile way by nurses. Finally, the results of this study show the importance of the role of nurses in the management of the risks of infection, the maintenance of biologically safe home environment and in facing the challenges to contribute to the formulation and use of public policies directed to this focus. We conclude that the practices of prevention and infection control in home care are a core competence recognized by nurses and needs to be improved in Nursing graduation.

nÃo hà / A introduÃÃo na prÃtica clÃnica dos agentes bloqueadores do TNF-&#945; representou uma revoluÃÃo no tratamento das doenÃas inflamatÃrias crÃnicas: artrite reumatÃide (AR), espondilite anquilosante (EA), doenÃa de Crohn (DC) e psorÃase. Desde entÃo, seu uso tem sido ampliado, contudo essa terapia de alto custo econÃmico elevou o risco de desenvolvimento das doenÃas infecciosas entre elas, à tuberculose (TB). Por este motivo recomenda-se a investigaÃÃo da Tuberculose latente (TBL) antes do inÃcio da terapÃutica com os bloqueadores do TNF-&#945;. O objetivo do nosso estudo foi traÃar o perfil clÃnico-epidemiolÃgico dos pacientes nessa condiÃÃo clÃnica e avaliar os desfechos de tratamento de pacientes submetidos ao tratamento da TBL por indicaÃÃo do uso de bloqueadores do TNF-&#945; no AmbulatÃrio de Tisiologia do Hospital UniversitÃrio Walter CantÃdeo (HUWC/UFC). MÃtodo: Foi realizada no perÃodo de 2008-2009 uma anÃlise descritiva prospectiva desse grupo de pacientes seguido de um estudo analÃtico observacional do tipo transversal do desfecho. Resultados: Foram estudados 45 pacientes classificados como portadores de TBL por apresentarem teste tuberculÃnico (TT) &#8805; 5 mm e radiografia de tÃrax normal ou com lesÃes residuais mÃnimas. A maioria era do sexo feminino (56%), cerca da metade estava na faixa etÃria de 40 a 49 anos (45%). Em relaÃÃo à doenÃa de base tivemos diagnÃsticos reumatolÃgicos â artrite reumatÃide e espondilite (49,0%), dermatolÃgicos â psorÃase (37,7%) e doenÃa de Chron (13,3%). A presenÃa de 01 comorbidades foi observada em15/45 (33,3%) e em 6/45 (13,3%) pacientes apresentavam mais de uma doenÃa associada. Contato com paciente de TB foi relatado em 9/45 (20%) dos indivÃduos estudados. Dois pacientes referiram TB pulmonar tratada anteriormente. A maioria (88,9%) era de assintomÃticos respiratÃrios. O resultado mÃdio do TT foi 14,6mm, variando de 5 a 30 mm, sendo que 30/45 (66,7%) apresentou TT >10 mm. O uso de medicaÃÃo antiinflamatÃria nÃo influiu no resultado do TT. A anÃlise radiolÃgica do tÃrax foi normal em 64,4% e em 35,6 % foram observadas alteraÃÃes radiolÃgicas mÃnimas. Dos 45 pacientes estudados 37 utilizaram fÃrmacos para tratamento da doenÃa de base, 20 (54,0%) usaram corticosterÃides (prednisona) e 36 (97,3%) relataram o uso de imunossupressores, principalmente o Metrotexato. A isoniazida 300mg/dia foi utilizada no tratamento da TBL e os efeitos colaterais ocorreram em 15,6 % dos pacientes. Em relaÃÃo ao desfecho foi observado que 41 completaram o tratamento. Houve 01 abandono, 01 transferÃncia e 02 suspensÃes de tratamento por hepatite medicamentosa. ConclusÃo: A determinaÃÃo do perfil do paciente candidato ao tratamento da TBL pode contribuir para a uniformizaÃÃo dos procedimentos de rastreio e prevenÃÃo da TB, bem como para estabelecer os protocolos clÃnicos de uso e acompanhamento dos fÃrmacos anti-TNF. Todos os pacientes continuam em acompanhamento no ambulatÃrio de tisiologia do HUWC/UFC por um perÃodo mÃnimo de 5 anos.