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Investigation of the anti-mycobacterial and cytotoxic effect of three medicinal plants used in the traditional treatment of tuberculosis in northern Mexico and the southwest U.S.Beltran, Oscar. January 2008 (has links)
Thesis (M.S.)--University of Texas at El Paso, 2008. / Title from title screen. Vita. CD-ROM. Includes bibliographical references. Also available online.
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A phytochemical study of Citrullus vulgaris Schroeder and A study of the reaction of theophylline with barbiturates /Higgins, Walter Mayo, January 1943 (has links)
Thesis (Ph. D.)--University of Wisconsin--Madison, 1943. / Typescript. Vita. eContent provider-neutral record in process. Description based on print version record. Bibliographies: leaves 60-89, 126-135.
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Avaliação da atividade anti-carie dos compostos bioativos isolados das plantas Mikania laevigata Schultz Bip. ex Baker e Mikania glomerata Sprengel / Evaluation of anitcarie activity of isolated bioactive compounds of Mikania laevigata Schultz Bip. ex Baker e Mikania glomerata Sprengel plantsYatsuda, Regiane 21 July 2006 (has links)
Orientadores: Pedro Luiz Rosalen, Hyun Koo / Tese (doutorado) - Universidade Estadual de Campinas, Faculdade de Odontologia de Piracicaba / Made available in DSpace on 2018-08-07T01:21:21Z (GMT). No. of bitstreams: 1
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Previous issue date: 2006 / Resumo: O uso de plantas medicinais vem crescendo nos últimos anos e isto é devido principalmente as descobertas de suas propriedades biológicas, e entre estas plantas destacam-se as do gênero Mikania. Assim, a procura pela descoberta de novos produtos naturais com atividade antibacteriana para a prevenção de doenças bucais e talvez com menores efeitos adversos quando comparados aos produtos industrializados seria muito importante para obtenção de um meio efetivo de controle da formação de um biofilme patogênico. Assim, em estudos anteriores, a fração hexânica das plantas Mikania laevigata e Mikania glomerata apresentaram atividade antimicrobiana e inibiram a aderência dos estreptococos do grupo mutans. Deste modo, o objetivo geral deste trabalho foi avaliar o efeito antimicrobiano dos compostos isolados das frações hexânicas das plantas M. laevigata e M. glomerata (guaco) sobre os estreptococos do grupo mutans. Para isso, foram realizados três estudos, sendo o objetivo do estudo 1 analisar a composição química e o efeito antimicrobiano dos compostos isolados e identificados das frações hexânicas das plantas M. laevigata e M. glomerata sobre o crescimento bacteriano e a aderência celular dos estreptococos do grupo mutans. O objetivo do estudo 2 foi determinar a influência destes compostos isolados na formação e na composição de polissacarídeos de biofilmes de Streptococcus mutans UA159 formados em discos de hidroxiapatita, na queda de pH gücolítico e seu efeito na translocação de prótons pela atividade da ATPase. Além disso, foram realizados estudos para analisar seu efeito na atividade de glucosiltransferase (GTF) B em solução (GTF-sol) e em superfície (GTF-sup). O objetivo do estudo 3 foi avaliar as atividades dos compostos isolados da Mikania utilizando modelo experimental de cárie em ratos. No estudo 1, os ácidos cupressênico, diterpênico e caurenóico foram os compostos que apresentaram atividade antimicrobiana (CIM entre 2,5-20 ng/mL e CBM entre 2,5-40 jjg/mL) e inibição da aderência celular entre 1,25-5 ug/mL, sendo que os compostos espatulenol, caurenol, ácido grandiflórico não apresentaram atividade biológica nas concentrações testadas. No estudo 2, somente os três compostos ativos isolados da Mikania foram avaliados na concentração 500 }ig/mL. Os resultados demonstraram que os ácidos cupressênico, diterpênico e caurenóico apresentaram efeito na viabilidade bacteriana dos biofilmes, reduziram a produção de ácidos orgânicos (pH final entre 6,4-5,8) e a atividade da ATPase (28-40%). Os compostos também foram potentes inibidores da atividade de GTFs, tanto em solução quanto em superfície de hidroxiapatita, sendo 50-60% de redução para GTF B-sol e 50-80% de redução para GTF B-surf. Nos biofilmes, o peso seco e a quantidade de polissacarídeos solúveis, insolúveis e intracelulares também foram significativamente reduzidos com o tratamento dos três compostos (p<0,05). No estudo 3, a aplicação tópica 2 vezes ao dia dos ácidos cupressênico, diterpênico e caurenóico (500 ug/mL) promoveu a redução na incidência de cárie em superfície lisa e sulcai (p<0,05), além da diminuição na porcentagem de infecção por S. mutans UA159 pelos ácidos diterpênico e caurenóico, não sendo afetada a microbiota total dessas ratas. Desta brma, concluímos que os ácidos cupressênico, diterpênico e caurenóico isolados da M. laevigata e M. glomerata possuem potencial antimicrobiano, inibindo os fatores de virulência dos Streptococcus mutans e a cárie em modelo in vivo, demonstrando serem promissores agentes anti-cárie e antí-placa / Abstract: Considering the great use of plants as medicinal substances in the popular medicine, it is critical to investigate their biological and chemical properties in order to not anly help to enhance our understanding of the therapeutic potential of these natural products, but also how to make them more effective pharmacological agents. The development of therapeutic agents aimed at disrupting both colonization of the teeth by dental pathogens and the subsequent formation of dental plaque is one of the prime strategies to reduce the incidence of tooth decay. The hexanic fraction of the plants Mikania laevigata and Mikania glomerata showed antimicrobial activities and inhibit the adherence of mutans streptococci. The overall aim of this study was to evaluate the antimicrobial effect of the isolated compounds of the hexanic fractions of Mikania laevigata and Mikania glomerata (guaco) on mutans streptococci. Therefore, three studies were carried out. in study 1 ,the antimicrobial activity of the isolated compounds was assessed by determination of minimum inhibitory concentration (MIC), minimum bactericidal concentration (MBC) and inhibition of cell adherence (Adh) to glass surface of mutans streptococci. In study 2, the influence of the isolated compounds were evaluated on viability, development, polysaccharide composition and acidogenicity of S. mutans biofilms, on glucans production by purified glucosyltransferases (GTFs) adsorbed to hydroxyapatite beads, and on membrane-associated F-ATPase and glycolytic activities. In study 3, the influence of the isolated compounds on caries development in vivo was evaluated. The acids cupressenic, diterpenic and kaurenoic were the compounds most effective in inhibiting the growth of the bacterial strains tested (MIC 2.5-20 ug/ml and MBC 2.5-40 ug/ml) and adherence cells (1.25-5 ug/ml). These three compounds were tested on the concentration of 500 pg/ml on tne study 2, and showed antimicrobial activity reducing the viable cells and the dry-weight of the biofilm treated, and also reduced the soluble, insoluble extracellular and intracellular polysaccharide of this biofilms. The compounds were able to reduce the acid production on the biofilms (final pH between 6.4 and 5.8) and the activity of F-ATPase (28-40%). The activity of the GTF B on solution (50-60%) and on surface (50-80%) of hydroxyapatite was also reduced by the three compounds. In study 3, the topical application twice a day of cupressenic, diterpenic and kaurenoic acids (500 ug/ml) showed cariostatic effect on smooth-surface and sulcal caries, and also showing reduction of the percentage of Streptococcus mutans UA159 infection by diterpenic and kaurenoic acids (p<0.05), not showing differences on the total microbiology (p>0.05). In conclusion, the cupressenic, diterpenic and kaurenoic acids isolated from M. laevigata and M. glomerata have relevant antimicrobial activity and inhibit the virulence factors of mutans streptococci in vitro and in vivo, being promising anti-caries and anti-plaque agents / Doutorado / Farmacologia, Anestesiologia e Terapeutica / Doutor em Odontologia
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Potencial de ação antimicrobiana in vitro de extratos de plantas na inibição de Candida spp, Streptococcus mutans e Staphylococcus aureus / In vitro antimicrobial activity of plants extracts against Candida spp, Streptococcus mutans e Staphylococcus aureusAnibal, Paula Cristina 23 February 2007 (has links)
Orientador: Jose Francisco Hofling / Dissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Odontologia de Piracicaba / Made available in DSpace on 2018-08-08T12:53:19Z (GMT). No. of bitstreams: 1
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Previous issue date: 2007 / Resumo: A cavidade oral abriga uma complexa comunidade de microrganismos que podem estar aderidos aos dentes, à mucosa epitelial ou formando biofilme. A maioria desses microrganismos são comensais, com uma minoria causando infecções oportunistas. Dentre esses microrganismos, encontramos as leveduras do gênero Candida, principalmente em indivÃduos imunocomprometidos; o principal agente etiológico da cárie dental Streptococcus mutans e, apesar da presença controversa, o patógeno oportunista Staphylococcus aureus. Diante do desenvolvimento de resistência aos antimicrobianos, há uma grande preocupação relacionada a qual tipo de tratamento utilizar no combate à s infecções provocadas por esses microrganismos. O emprego de extratos brutos de plantas tem merecido a atenção de pesquisadores de vários paÃses, já que podem inibir o crescimento bacteriano e fúngico por diferentes mecanismos quando comparados aos antimicrobianos, justificando a pesquisa em questão. As plantas utilizadas nesta pesquisa
foram a Mentha piperita, Rosmarinus officinalis, Tabebuia avellanedae, Arrabidaea chica, Syzygium cumini e Punica granatum, extraÃdas com os solventes diclorometano e metanol, os quais foram eliminados em rotaevaporador, para a obtenção dos extratos brutos, a fim de se avaliar sua concentração inibitória mÃnima (CIM) frente a essas cepas de microrganismos. Os resultados obtidos revelaram-se promissores com as espécies de Candida testadas, principalmente com o extrato metanólico, cuja inibição apresentou atividade com alta sensibilidade das leveduras, porém essa mesma atividade não ocorreu entre as bactérias. Esses dados indicam a necessidade da ampliação do conhecimento sobre as espécies de plantas com potencial antimicrobiano, além de se testar extratos mais purificados e com propriedades farmacológicas ativas contra organismos microbianos. De qualquer forma, estes resultados nos levam a considerar esses extratos vegetais como fontes valiosas para a descoberta de novas moléculas bioativas empregadas para o tratamento alternativo no combate aos agentes microbianos, principalmente à queles resistentes à s drogas antifúngicas e antibacterianas convencionais / Abstract: The oral cavity is a source of a complex microorganisms community that may adhere to teeth, epithelial mucosa or form biofilms. Most of these strains are thought to be commensal, and a small number cause opportunistic infections. Among these microorganisms Candida yeast is the most commom in immunocompromised patients; Sretptococcus mutans is regarded as the primary etiologic agent of dental caries and, although considered controversial the presence, Staphylococcus aureus is also mentioned as an opportunistic pathogen in the oral cavity. Considering the frequent use of antifungal and antibiotics as antimicrobial agents, the development of resistance to these drugs has become an important and worrying factor for the treatment of the infections caused by these microorganisms. The use of crude plants extracts has been a researcherâ?¿s concern from all over the world, since they may inhibit the bacteria and fungi growing by different mechanisms present in the conventional antimicrobials, becoming an important source of investigation. The plants used in this research were Mentha piperita, Rosmarinus officinalis, Tabebuia avellanedae, Arrabidaea chica, Syzygium cumini and Punica granatum, extracted with dichloromethane and methanol solvents, followed by a minimal inhibitory concentration (MIC) evaluation. The data obtained with Candida strains were promising, specially with the methanolic extract that showed activity in a higher sensibility, but the same activity did not occur among the bacterias. This information increases our knowledge of the crude plants extracts effect against microbial organisms. Nevertheless, the results of this study indicated that these extracts are a promising source for the alternative treatment against the microbial agents, specially for those resistant to the conventional antimicrobial agents / Mestrado / Microbiologia e Imunologia / Mestre em Biologia Buco-Dental
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Studies on the pharmacological properties of resin from Protium heptaphyllum (Aubl.) March. and its major constituent, alpha-and beta-amyrin mixture / Estudo das propriedades farmacolÃgicas da resina de Protium heptaphyllum (Aubl) March e de seus principais constituintes, mistura de alpha e beta amirinaFrancisco de Assis Oliveira 24 June 2005 (has links)
CoordenaÃÃo de AperfeiÃoamento de Pessoal de NÃvel Superior / Protium heptaphyllum March (Burseraceae) populary known as âalmÃcegaâ is a popular medicinal plant largely encountered in the Amazon region, various States of Brazil and in several South American Countries. The oily amorphous exudate obtained from this plant is widely used in skin diseases, healing of ulcers, and as an analgesic and anti-inflamatory agent. Phytochemical studies reveled the presence of several monoterpenes and pentacyclic triterpenes such as a mixture of α- e β â amyrin, maniladilol and breine. The present study aimed to investigate the general toxicity and to establish the pharmacological activity of resin and is major triterpenoid mixture, the α and β â amyrin. In toxicity tests, both the resin and triterpene mixture exhibited low toxicity to mice. Resin at doses up 5 g/kg, (p.o.) or 2 g/kg (i.p.) and triterpene mixture up to 3 g/kg, (p.o.) or 2 g/kg (i.p.) failed to induce any mortality in mice. In Artemia-lethality test, the calculated (probit analysis) CL50 values for resin and triterpenes were in the order of 42,54  19,96 and 400  27,85 μg/mL, respectively. In pharmacological tests, the resin was analysed for anti-inflamatory (carrageenan-induced edema, cotton pellet-induced granuloma, and vascular permeability increase induced by i.p. acetic acid) and gastroprotective (absolute ethanol and acidified ethanol) effects, whereas the triterpene mixture was examined in assays that demostrate gastroprotective (against lesions induced by absolute ethanol), antipruritus (against pruritus induced by Dextran T40 and compound 48/80), acute and visceral antinociceptive (test of subplantar and intracolonic capsaicin) and hepatoprotective (against acetaminophen- and Ga1N/LPS-induced models of hepatitis) effects. In anti-inflammatory test, the resin (200 e 400 mg/kg, p.o.) although failed to modify the carrageenan-induced acute rat paw-edema response, it caused signficant inhibitions at a dose of 400mg/kg on the formation of cotton pellet-induced granulomas and on the vascular permeability increase induced by i.p. acetic acid in mice. In addition, the (200 e 400 mg/kg) showed gastroprotective potential against absolute- and acidified ethanol- induced gastric lesions as evidenced from siginificant diminution in lesion scores, restoration of the ethanol-induced depletion of non-protein sulfhydryl content More over, the resin demonstrated an antisecretory effect on gastric acid secretion induced in 4-h pylorus ligated rats. The triterpene mixture also produced similar gastroprotection against ethanol-induced lesions in a manner similar to capsaicin, a pungent principle from hot peppers. This protection possibly involves capsaicin-sencitive primary afferents since it was abolished in mice pretreated with a neurotoxic dose of capsaicin. The α and β â amyrin mixture (100 mg/kg) manifested antipruritus effect as evidenced from suppression of scratching behaviour in the mouse model of prurits induced by s.c. injections of dextran T40 and compound 48/80. Besides, it also produced an antiedematogenic effect in model of hind paw edema induced by histamine, compound 48/80 and dextran T40 and markedly depressed the compound 48/80-elicited rat mast cell degranulation (ex vivo). An antinociceptive effect of triterpenoid mixture (3-100 mg/kg) was observed in capsaicin-evoked somatic (1.6 μg/site, suplantar) and visceral (149 μg, intracolonic) models of nociception in mice. Greater suppression of nociceptive behaviors were evidenced at a dose of 10 mg/kg α and β â amyrin mixture, which mimicked the effect produced by ruthenium red, a non-competitive capsaicin antagonist. The antinociceptive effect of triterpenoid mixture was found to be naloxone (2 mg/kg)- sensitive, suggesting an opioid mechanism. A blockade by triterpene mixture was also evidenced on the hyperthermic but not the hypothermic response of subcutaneously administered capsaicin (10 mg/kg) suggesting possible incolvement of TRPV1 receptor. In open-field and rota-rod tests, the triterpene mixture did not manifest signs of either sedation or motor abnormality in mice that could account for the observed antinociception. In the model of acetaminophen (500 mg/kg)-induced hepatotoxicity, the triterpenoid mixture (50 and 100 mg/kg) effectively reduced the elevated serum AST and ALT levels, restored the depleted GSH and markedly diminished the histopathological alterations. Potentation of pentobarbital-sleeping time was, however observed at these doses of triterpenoid, incidating a probable suppression of cytochrome P450 and thus a diminished metabolite formation that may account for reduced acetaminophen toxicity. The α- and β â amyrin mixture offered complete protection against the mortality associated with Ga1N/LPS , but caused only a moderate diminution of serum enzymes and histopathological alterations. Taken together, these findings show that the resin and α- and β â amyrin mixture possess low toxicity and have a wide therapeutic potential with anti-inflammatory, antinociceptive, antipruritus, and gastro- and hepato-protective actions. Most of the effects of triterpenoid mixture appear to involve in part the participation of primary sensory afferents in their actions. / A espÃcie Protium heptaphyllum (Aubl.) March (Burseraceae) popularmente conhecida como almÃcega, à encontrada na regiÃo AmazÃnica, em vÃrios Estados do Brasil e paÃses da AmÃrica do Sul. Esta espÃcie exsuda uma resina oleosa e amorfa, usada na medicina popular como analgÃsico, cicatrizante e expectorante. Estudos fitoquÃmicos demonstraram a presenÃa de monoterpenos e triterpenos pentacÃclicos, tais como α - amirina e β - amirina, maniladilol e breina. O presente trabalho teve como objetivo investigar os efeitos tÃxicos e farmacolÃgicos da resina e de seus constituintes majoritÃrios, a mistura de triterpenos α e β â amirina. Na avaliaÃÃo dos efeitos tÃxicos observamos a toxicidade aguda destes produtos em camundongos e Artemia sp. Analisando os efeitos sistÃmicos, avaliamos a atividade antiinflamatÃria da resina (edema de pata induzido por carragenina, granuloma induzido por âpelletsâ de algodÃo e permeabilidade vascular induzida por Ãcido acÃtico) e da mistura de α e β â amirina (edema induzido por histamina, serotonina, dextrana T40 e composto 48/80). Examinamos ainda as atividades gastroprotetora e antisecretÃria da resina (lesÃes gÃstricas induzidas pelo etanol absoluto e etanol acidificado e secreÃÃo Ãcida induzida pela ligaÃÃo pilÃrica) e as atividades gastroprotetora (lesÃes gÃstricas induzidas pelo etanol absoluto, com animais dessensibilizados por capsaicina), antipruriginosa (prurido induzido pelo dextrana T40 e composto 448/80 e desgranulaÃÃo de mastÃcitos ex vivo) antinociceptiva (nocicepÃÃo induzida pela administraÃÃo subplantar e intracolÃnica de capsaicina, resposta hipotÃrmica induzida por capsaicina) e hepatoprotetora (lesÃes hepÃticas induzidas por acetaminofeno e Ga1N/LPS) da mistura de α e β â amirinas. NÃo foi possÃvel estabelecer as DL50 da resina (atà 5 g/kg, v.o. e 1 g/kg, i.p.) e da mistura de α e β â amirina (atà 3 g/kg, v.o. e atà 2 g/kg, i.p.) em camundongos. A mistura de α e β â amirina, mas nÃo a resina, mostrou toxicidade para Artemisa sp, sendo as CL50 de 42,54  19,96 e 400  27,85 μg/mL, respectivamente. Nos modelos de permeabilidade vascular induzido por Ãcido acÃtico (camundongo) e granuloma induzido por âpelletâ de algodÃo (ratos), a resina demonstrou efeito antiinflamatÃrio significativo na dose de 400mg/kg, reduzindo a permeabilidade vascular e o peso seco do granuloma. Contudo, a reina nÃo apresentou atividade sobre edema induzido por carragenina (ratos). Adicionalmente, a resina preveniu as lesÃes gÃstricas induzidas por etanol absoluto e etanol acidificado, alÃm de impedir a depleÃÃo dos grupos sulfidrilas produzida pelo etanol absoluto nas doses de 200 e 400 mg/kg. Um efeito antisecretÃrio da resina (200 e 400mg/kg) foi observado no modelo de secreÃÃo Ãcida induzida pela ligaÃÃo pilÃrica em ratos. A mistura de α e β â amirina tambÃm exibiu atividade gastroprotetora inibindo as lesÃes gÃstricas por etanol absoluto, cujo mecanismo parece envolver os neurÃnios sensoriais primÃrios sensÃveis à capsaicina. A administraÃÃo oral dos triterpenos α e β â amirina (100 mg/kg), apresentou atividade antiedematogÃnica, nos modelos de edema de pata induzidos por histamina, composto 48/80 e dextrana T40, mas nÃo sobre o edema induzido por serotonina. A atividade antipruriginosa tambÃm foi observada com as α e β â amirina nas doses variando de 50 a 200 mg/kg, em modelos de prurido induzido por dextrana T40 e pelo composto 48/80 e na reduÃÃo (100 mg/kg) da degranulaÃÃo de mastÃcitos peritoneais ex vivo pelo composto 48/80. O efeito antinociceptivo da mistura, nas doses de 3 a 100 mg/kg, foi verificado atravÃs da inibiÃÃo dos comportamentos de nocicepÃÃo induzidos pela administraÃÃo subplantar ou intracolÃnica de capsaicina em camundongos. A antinocicepÃÃo produzida por estes triterpenos (10 mg/kg) sobre o tempo de lambedura induzido pela capsaicina (1,6 μg/20 μL) nÃo foi potencializada nem revestida pelo vermelho de rutÃnio (1,5 mg/kg), mas foi significativamente inibida pela naloxona (2 mg/kg), sugerindo mecanismo opiÃide. A participaÃÃo dos receptores α2 - adrenÃrgicos neste efeito tambÃm foi eliminada, tendo em vista que a ioimbina nÃo reverteu o efeito antinociceptivo das amirinas no modelo de nocicepÃÃo visceral induzida pela capsaicina. Estes triterpenos bloquearam ainda a hipertermia induzida pela capsaicina (10 mg/kg), mas nÃo reverteram a resposta hipotÃrmica induzida por este agente, sugerindo a participaÃÃo do receptor vanilÃide (TRPV1) no efeito antinociceptivo das amirinas. Nos modelos de hepatoxidade, a mistura de α e β â amirina (50 e 100 mg/kg) reduziu o aumento dos nÃveis sÃricos de ALT e AST e restabeleceu os nÃveis de GSH hepÃticos, diminuindo as alteraÃÃes histopatolÃgicas induzidas pelo acetaminofeno (500 mg/kg), alÃm de potencializar o tempo de sono induzido por pentobarbital sÃdico (50 mg/kg), indicando que este efeito hepatoprotetor envolve a inibiÃÃo do citocromo P â 450. A mistura ofereceu ainda completa proteÃÃo contra a mortalidade induzida por Ga1N/LPS, reduzindo as lesÃes hepÃticas em camundongos e reduzindo os nÃveis sÃricos de ALT, mas nÃo de AST ou GSH hepÃticos, sugerindo um possÃvel feito neuroimunomodulatÃrio neste modelo. Os triterpenos α e β â amirina nas doses variando de 3 a 30 mg/kg, nÃo manifestam efeitos sedativos ou incoordenaÃÃo motora em camundongos. A resina e mistura de α e β â amirina possuem baixa toxicidade e atividades antiinflamatÃria e gastroprotetora. Os triterpenos α e β â amirina exibiram atividade antipruriginosa, antinociceptiva e hepatoprotetora, cujos efeitos envolvem, pelo menos em parte, a participaÃÃo dos neurÃnios aferentes sensoriais primÃrios.
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Estudo do potencial antiinflamatÃrio do Ãleo-resina da Copaifera langsdorffii Desf. (COPAÃBA) e de seu constituinte diterpÃnico Ãcido KaurenÃico nos modelos experimentais de inflamaÃÃo intestinal / Studies on the anti-inflammatory potential of copaiba oil-resin from copaifera langsdorffii and its diterpene constituent kaurenoic acid in experimental models of intestinal inflammationLaura AndrÃa Farias Paiva 26 November 2004 (has links)
CoordenaÃÃo de AperfeiÃoamento de Pessoal de NÃvel Superior / O Ãleo-resina da Copaifera langsdorffii (Leguminaceae) Ã utilizado popularmente no tratamento de processos inflamatÃrios e na cicatrizaÃÃo de feridas e Ãlceras. Estudos prÃvios estabeleceram as propriedades gastroprotetora e antiinflamatÃria em modelos animais. O presente estudo avaliou o potencial antiinflamatÃrio do Ãleo-resina da Copaifera langsdorffii (ORCL) e de seu constituinte diterpÃnico, Ãcido kaurenÃico (AK) nos modelos experimentais de colite induzida por Ãcido acÃtico (UC-AA), por Ãcido trinitrobenzÃnico sulfÃnico (UC-TNBS), e ainda, indometacina e isquemia-reperfusÃo induzindo inflamaÃÃo intestinal (II-IND e II-I/R). E tambÃm, o potencial de cicatrizaÃÃo de feridas foi avaliado em ratos com feridas abertas e com incisÃo. Ratos foram prÃ-tratados via oral (15 e 2 horas antes) ou retal 2 horas apÃs a induÃÃo da colite, com ORCL (200 e 400mg/Kg), AK (50 e 100mg/Kg) ou veÃculo (1mL, 2% Tween 80 ou 1mL, 2% DMSO). A colite foi induzida pela aplicaÃÃo de 2mL de Ãcido acÃtico 4% (v/v) ou TNBS (0,25 mL com 20mg) e 24 ou 72 horas depois, os danos na mucosa do cÃlon foram avaliados, medidos os nÃveis de mieloperoxidase e malonaldeÃdo. No modelo de (CU-AA), houve uma importante reduÃÃo no escore da lesÃo e peso Ãmido nos animais tratados com as substÃncias teste, quando comparado ao controle veÃculo. Os efeitos foram confirmados na bioquÃmica pela significante reduÃÃo da atividade da mieloperoxidase (MDA), o marcador da infiltraÃÃo de neutrÃfilos e pela marcada diminuiÃÃo nos nÃveis de malonaldeÃdo, um indicador da lipoperoxidaÃÃo. AlÃm de o Ãcido acÃtico aumentar os nÃveis de nitrito e da enzima catalase no cÃlon, onde o tratamento com ORCL diminuiu significativamente. A anÃlise microscÃpica revelou uma diminuiÃÃo da infiltraÃÃo de cÃlulas inflamatÃrias e do edema da submucosa, nos segmentos do cÃlon tratados com ORCL ou AK. De maneira similar, no modelo de UC-TNBS, houve uma reduÃÃo do escore da lesÃo e peso Ãmido do cÃlon de animais prÃ-tratados com ORCL (400mg/Kg, v.o. ou retal) 2, 24 e 48 horas apÃs a injeÃÃo intracolÃnica de TNBS. A atividade da MPO, mas nÃo MDA e catalase foram significativamente afetados pelo prÃ-tratamento com ORCL. As observaÃÃes histolÃgicas indicam uma proteÃÃo parcial do ORCL, como UC-TNBS Ã um modelo crÃnico, talvez houvesse necessidade de uma terapia mais prolongada. No modelo de II-I/R, quarenta e cinco minutos de isquemia seguida de uma hora de reperfusÃo da artÃria mesentÃrica superior causou significante aumento nos nÃveis de MPO, catalase, MDA e nitrito, com uma significante diminuiÃÃo dos grupos sulfidrÃlicos nÃo-protÃicos (NP-SH/GSH) indicando um estresse oxidativo. Estes valores foram sigificamente revertido pelo prÃ-tratamento via oral com ORCL (200 e 400mg/Kg), sugerindo que ORCL anula o estresse oxidativo. Animais prÃ-tratados com ORCL (200 e 400mg/Kg, v.o.) ou AK (100mg/Kg, v.o.), 12 e 2 horas antes da administraÃÃo de 20mg/Kg de indometacina causando toxicidade intestinal, foi capaz de ser evidenciado pela diminuiÃÃo nos nÃveis de MPO e nitrito. Diferente da indometacina, ORCL mas nÃo AK falharam em induzir o aumento signifativo na permeabilidade intestinal. Este efeito do ORCL foi similar ao inibidor seletivo de COX-2, rofecoxibe. Estas observaÃÃes sugerem que ORCL Ã isento de toxicidade intestinal diferente da toxicidade intestinal dos inibidores clÃssicos nÃo seletivos da COX. AlÃm disso, ORCL promove cicatrizaÃÃo de feridas aberta ou com incisÃo em ratos evidenciada pela contraÃÃo e tensÃo da pele. Os dados indicam um potencial antiinflamatÃrio do Ãleo-resina da copaÃba e do seu diterpeno Ãcido kaurenÃico, possivelmente mediado pelos mecanismos antioxidantes e anti-lipoperoxidativo / Copaiba oil-resin from Copaifera langsdorffii (Leguminaceae) is a reputed traditional remedy for the treatment of inflammatory conditions and to promote healing of ulcers and wounds. Previous studies established its anti-inflammatory and gastroprotective properties through animal experimentation. The present study extended these earlier studies to analyse the intestinal anti-inflammatory potential of oil-resin Copaifera langsdorffii (ORCL) and its diterpene constituent, kaurenoic acid (KA) in rat models of ulcerative colitis induced by acetic acid (AA-UC), and trinitribenzene sulfonic acid (TNBS-UC), and in indomethacin -and ischemia-reperfusion-induced intestinal inflammation (IND-II and I/R-II). Further, its wound healing potential was evaluated in rats on open and incision wounds. Rats were pretreated orally (15 hrs and 2 hrs before) or rectally 2 hrs before the induction of colitis with ORCL (200 and 400 mg/kg), KA (50 and 100 mg/kg) or vehicle (1 ml, 2% Tween 80 or 1 ml, 2% DMSO). Colitis was induced by intracolonic instillation of a 2 ml of 4% (v/v) acetic acid solution or TNBS (0.25 ml of 20 mg) and 24 hrs or 72 hrs latter, the colonic mucosa was analysed for the severity of macroscopic colonic damage, the myeloperoxidase and the malondialdehyde levels. In AA-UC model, a marked reduction in Gross damage score and in wet weight/length ratio of colonic tissue were evident in animals pretreated orally or rectally with test substances, as compared to vehicle alone-treated controls. This effect was confirmed biochemically by a significant reduction in colonic myeloperoxidase (MPO) activity, the marker of neutrophilic infiltration and by a marked decrease in malondialdehyde (MDA) level, an indicator of lipoperoxidation. Besides, AA elevated increase in the levels of nitrite and catalase activity in colon tissue was also significantly decreased by ORCL treatment. Furthermore, microscopical examination revealed the diminution of inflammatory cell infiltration, and the submucosal edema in colon segments of rats pretreated with ORCL or KA. In a similar manner, in TNBS-UC, a marked reduction in Gross damage score and in wet weight/length ratio of colonic tissue was evident by ORCL pretreatment (400 mg/kg, p.o. or intra-rectal) at 2, 24 and 48 hrs after intracolonic injection of TNBS. MPO activity but not the MDA and catalase levels were significantly affected by ORCL treatment. Histological observations also indicated only a partial protection by ORCL, suggesting that TNBS-UC being a chronic model, a more prolonged therapy may be needed. In the model of I/R-II, five forty minute of ischemia followed by one hour reperfusion of superior mesenteric artery caused significant elevations of MPO, catalase, MDA and nitrite levels with a significant decrease in non-protein sulfhydryls (NP-SH/ GSH) indicating an oxidative stress. These changes were significantly reversed by oral pretreatment with ORCL (200 and 400 mg/kg), suggesting that ORCL obliterates oxidative stress. Pretreatment of animals with ORCL (200 and 400 mg/kg, p.o.) or KA (100 mg/kg, p.o.), 12 and 2 hrs before the administration of 20 mg/kg indomethacin mitigated the intestinal toxicity as evidenced by decreases in tissue levels of MPO and nitrite. Unlike indomethacin, ORCL but not KA at either dose failed to induce a significant increase in intestinal permeability. This effect of ORCL simulated that of a selective COX-2 inhibitor, rofecoxib. These observations suggest that ORCL is devoid of intestinal toxicity unlike the classical non-selective COX inhibitors. Also, ORCL promoted wound healing in rats on experimental open or incision wounds as evidenced by an early wound contraction and increased wound tensile strength. The data indicate a significant anti-inflammatory potential of copaiba oil-resin and its diterpenoid, kaurenoic acid possibly mediated through an antioxidant/anti-lipoperoxidative mechanism(s)
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In vitro and in vivo study of pyrrolizidine alkaloids-induced hepatotoxicity. / CUHK electronic theses & dissertations collectionJanuary 2011 (has links)
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.
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Model study and partial synthesis of prehispanolone and derivatives from hispanolone.January 1994 (has links)
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
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Authentication by molecular method of dendrobium used in Chinese medicine.January 2000 (has links)
by Lau Tai Wai. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2000. / Includes bibliographical references (leaves 117-127). / Abstracts in English and Chinese. / Table of Content --- p.i / Abbreviations --- p.iv / Abstract --- p.v / List of Figures --- p.ix / List of Tables --- p.xii / Chapter 1. --- Chapter One: Introduction --- p.1 / Chapter 1.1 --- Background on orchids --- p.2 / Chapter 1.2 --- Background on Dendrobium --- p.7 / Chapter 1.3 --- Background and history on Herba Dendrobii --- p.9 / Chapter 1.4 --- Reasons for study of Herba Dendrobii --- p.12 / Chapter 1.4.1 --- Demand --- p.12 / Chapter 1.4.2 --- Adulteration --- p.13 / Chapter 1.4.3 --- CITES --- p.13 / Chapter 1.5 --- Scientific researches on Herba Dendrobii --- p.14 / Chapter 1.5.1 --- Morphological studies --- p.15 / Chapter 1.5.2 --- Anatomical and microscopic studies --- p.16 / Chapter 1.5.3 --- Phytochemistry --- p.20 / Chapter 1.5.3.1 --- Chemicals identified --- p.20 / Chapter 1.5.3.2 --- Chemical authentication of Herba Dendrobii --- p.23 / Chapter 1.5.3.3 --- Effect of treatment on chemical composition --- p.23 / Chapter 1.5.4 --- Phylogenetic study of Dendrobium --- p.25 / Chapter 1.5.4.1 --- Phylogenetic analysis by molecular methods --- p.25 / Chapter 1.5.4.2 --- Phylogenetic analysis by anatomical methods --- p.27 / Chapter 1.5.5 --- Pharmacological effect --- p.29 / Chapter 2. --- Chapter two: Objectives and strategies --- p.30 / Chapter 3. --- Chapter Three: Materials and Methods --- p.33 / Chapter 3.1 --- Source of samples and their treatment --- p.34 / Chapter 3.1.1 --- Fresh materials --- p.34 / Chapter 3.1.2 --- Dry materials --- p.34 / Chapter 3.1.3 --- Outgroup species --- p.35 / Chapter 3.2 --- Experimental protocol --- p.40 / Chapter 3.2.1 --- Rationale of the experiment --- p.40 / Chapter 3.2.2 --- DNA extraction --- p.41 / Chapter 3.2.2.1 --- Cetyltrimethylammonium bromide extraction method --- p.41 / Chapter 3.2.2.1a --- Reagents and buffers --- p.41 / Chapter 3.2.2.1b --- Procedures of CTAB extraction method --- p.42 / Chapter 3.2.2.2 --- Modified DNA isolation protocol for dry samples --- p.43 / Chapter 3.2.2.2a --- Reagents and buffers --- p.43 / Chapter 3.2.2.2b --- Procedures of modified DNA isolation protocol for dry plant samples --- p.44 / Chapter 3.2.3 --- Agarose gel electrophoresis of genomic DNA or PCR products --- p.45 / Chapter 3.2.3a --- Reagents and buffers --- p.45 / Chapter 3.2.3b --- Procedures of agarose gel electrophoresis of genomic DNA or PCR products --- p.45 / Chapter 3.2.4 --- Qualitative and quantitative analysis of DNA --- p.46 / Chapter 3.2.5 --- Amplification of the internal transcribed spacer 2 (ITS 2) region by Polymerase Chain Reaction --- p.47 / Chapter 3.2.5a --- Internal transcribed spacer 2 (ITS 2) region --- p.47 / Chapter 3.2.5b --- Procedures of polymerase chain reaction of ITS 2 region --- p.48 / Chapter 3.2.6 --- Purification of PCR products or cycle sequencing products --- p.48 / Chapter 3.2.6.1 --- Ethanol precipitation --- p.48 / Chapter 3.2.6.2 --- GENECLEAN® protocols --- p.49 / Chapter 3.2.6.3 --- Spin Column Purification --- p.49 / Chapter 3.2.7 --- Cycle Sequencing --- p.50 / Chapter 3.2.8 --- Sample Electrophoresis --- p.51 / Chapter 3.2.8a --- Equipment and reagents --- p.51 / Chapter 3.2.8b --- Procedures of sample electrophoresis --- p.52 / Chapter 3.2.9 --- Sequence analysis --- p.52 / Chapter 4. --- Results --- p.53 / Chapter 4.1 --- Fresh materials --- p.54 / Chapter 4.1.1 --- Genomic DNA --- p.54 / Chapter 4.1.2 --- PCR products --- p.59 / Chapter 4.1.3 --- Sequence alignment --- p.66 / Chapter 4.1.4 --- Comparison of the sequences --- p.94 / Chapter 4.1.5 --- Percentage difference among Dendrobium --- p.96 / Chapter 4.1.6 --- Intra-specific variation of orchid species --- p.96 / Chapter 4.1.7 --- Phylogenetic analysis --- p.99 / Chapter 4.2 --- Dry materials --- p.101 / Chapter 4.2.1 --- Genomic DNA --- p.101 / Chapter 4.2.2 --- PCR products --- p.101 / Chapter 4.2.3 --- Sequencing result --- p.101 / Chapter 5. --- Discussion and Conclusion --- p.107 / Chapter 5.1 --- Reasons for authentication of Herba Dendrobii --- p.108 / Chapter 5.2 --- Fresh materials of Herba Dendrobii --- p.109 / Chapter 5.2.1 --- Authentication --- p.109 / Chapter 5.2.2 --- Phylogenetic analysis --- p.111 / Chapter 5.3 --- Dry materials of Herba Dendrobii --- p.114 / Chapter 5.4 --- Evaluation of the experimental method --- p.115 / Chapter 5.5 --- Conclusion --- p.116 / Chapter 6. --- Reference --- p.117 / Chapter 7. --- Appendix / Appendix 1: Number of species in each medicinal orchid geneus --- p.128 / Appendix 2: Photographs showing 15 of the 17 species of orchids used in this research project --- p.131
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In vivo and in vitro mechanistic studies of the wound healing effects of Rehmanniae Radix. / CUHK electronic theses & dissertations collectionJanuary 2012 (has links)
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
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