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  • About
  • The Global ETD Search service is a free service for researchers to find electronic theses and dissertations. This service is provided by the Networked Digital Library of Theses and Dissertations.
    Our metadata is collected from universities around the world. If you manage a university/consortium/country archive and want to be added, details can be found on the NDLTD website.
1

Efeito da fração aquosa das folhas de Costus spiralis (Jacq.) Roscoe sobre a função contrátil do coração de mamíferos / EFFECT OF AQUEOUS FRACTION OF LEAVES DE COSTUS SPIRALIS (JACQ.) ROSCOE CONTRACTILE FUNCTION ON HEART MAMMALS.

Britto, Raquel Moreira de 25 March 2011 (has links)
Teas and infusions from C. spiralis leaf have largely been used by folk medicine as diuretic, hypotensor, cytotoxic, immunomodulator, antilithiasic, antidiarrheic, antispasmodic, antiurolitic, antimicrobian, antifungic, antioxidant, antileishmania activity, antiinflamatory, and antiedematogenic activity. In spite of these biological effects attributed to the extracts of C. spiralis, nothing so far could be found in the scientific literature dealing with its effects on the mammalian myocardium.The present study aimed to describe the inotropic effects produced by extracts from the C. spiralis leaf on isolated guinea pig atrium, as well as to contribute for a better understanding about its mechanism of action in that tissue. In isolated mouse cardiomyocytes, the effect produced by those extracts on the intracellular calcium transient and on the sarcolemal L-type calcium current were also measured. Experiments performed to evaluate the contractile effects were carried out on isolated atrium from guinea pig (Cavia porcellus). Firstly, our purpose was to determine the most potent fraction obtained from the C. spiralis leaf. This was done by comparing the hydroalchoolic crude extract with the following ones: aqueous, chloroform, and ethyl acetate. A phytochemical analysis was performed on the fraction exhibiting the greater potency. This evaluation followed the procedures proposed by Matos (1997). The content of sodium and potassium in the most potent fraction was determined by flame photometry. In the contractile experiments, the atrial force was measured isometrically. Biological signals were captured, amplified, and then stored in computer to be processed off line. Intracellular calcium transients were studied by confocal microscopy with laser scanning by using the fluorescent dye FLUO 4AM. Calcium inward currents were measured in mouse cardiomyocytes by using patch clamp technique in the whole cell configuration. Yield percentage of the aqueous fraction (AqF) was 69,40%. This fraction showed the most potent depressor effect on the myocardial contractility (EC50 = 305 ± 41,00 mg/L, Hill constant = 1,46 ± 0,19). The following metabolites were found in the AqF: tannins, saponins, and polifenols (flavonol, flavononol, flavone, xanthone, phenol, and flavonoid). The potassium and sodium contents in 1 g/L of AqF were 1,91 and 0,15 mM, respectively. This was not enough to change the myocardial inotropism, even in the highest concentration of AqF used in the experiments. The contraction and the relaxation time, as well as the time related to the excitation-contraction coupling (stimulus-response) were not modified by adding AqF to the organ bath. However, AqF reduced the Efficiency Index for the contraction and relaxation phases. The Neyler & Merrillees protocol was employed to evaluate the AqF effect on the calcium inward current in myocardial cells. Our results showed that AqF is able to completely abolish the Bowditch phenomenon, suggesting that it could be acting by reducing the sarcolemal calcium current. Supported by those experimental evidences, experiments were proposed to better understand the relationship between AqF and calcium mechanisms in cardiac cells. The following results were obtained with 1,5 g/L AqF: 1) AqF completely abolished the positive inotropic effect induced by isoproterenol (10-1 to 103 pM); 2) AqF shifted rightwardly the concentration-effect curve for CaCl2 (0.5 to 7.0 mM) and increased the EC50 from 1.12 ± 0.07 (Hill = 1.5) to 7.23 ± 0.47 mM (Hill = 7.4) (n = 3; p < 0.05); 3) AqF completely abolished the positive inotropic effect of (-) BAY K8644 (5 to 2000 nM); 4) AqF reduced the intracellular fluorescence from 4.66 ±1.17 to 3.74 ± 1.0 a.u. (n = 30 cells, 4 mice, p < 0.05); 5) AqF did not modify the decay rate of the fluorescent signal (892 ± 37 to 930 ± 30 ms, n = 30 cells, 4 mice, p > 0.05), indicating that it does not interfiere with the calcium removal from the sarcoplasm; 6) AqF reduced the calcium inward current through L-type calcium channels from 6,29 ± 0,34 to 4,9 ± 0,2 A/F (23% , n = 5 animals, p < 0,05). This study brought us unto the following conclusions: 1) AqF is the most potent fraction obtained from C. spirallis leaves; 2) AqF contains the following secondary metabolites: tannins, saponins, and poliphenols; 3) AqF reduces the contraction force of the guinea pig left atrium; 4) AqF acts on the myocardium contractility by reducing the calcium entry in myocardial cells during contraction. / Preparados de Costus spiralis têm sido usados pela medicina popular (diurético, hipotensor, citotóxico, imunomodulador, antilitiásico, antidiarréico, antiespasmódico, antiurolítico, antimicrobiano, antifúngico, antioxidante, antileishmânia, anti-inflamatório e antiedematogênico). Apesar da gama de ações a eles atribuídas, nada pôde ser encontrado na literatura científica com respeito ao possível efeito dos Este trabalho visou determinar os efeitos inotrópicos obtidos das folhas de C. spiralis, que apresentava maior potência, bem como contribuir para o mecanismo de ação desse preparado no miocárdio de mamíferos. Os experimentos sobre contração foram realizados em átrio esquerdo de cobaia (Cavia porcellus), enquanto que as medidas de transiente de cálcio intracelular e de corrente de membrana foram feitas em cardiomiócitos de camundongo. A investigação fitoquímica do preparado mais ativo foi conduzida segundo Matos (1997). Os teores de sódio e de potássio presentes na fração mais potente, foram determinados por fotometria de chama. A força de contração atrial foi captada isometricamente e, depois amplificada, foi armazenada em computador. O transiente de cálcio intracelular foi avaliado com microscopia confocal de varredura a laser. As correntes de cálcio sarcolemais foram medidas em cardiomiócitos submetidos à técnica do patch clamp ( whole cell ). A fração aquosa (FAq) foi a que apresentou maior rendimento (69,40%) e a que exerceu maior efeito inotrópico negativo (CE50 = 305 ± 41,00 mg/L, Hill = 1,46 ± 0,19). Na sua constituição foram detectadas as seguintes classes de metabólitos secundários: taninos e saponinas, com reação fortemente positiva, e os polifenóis, com reação positiva (flavonóis, flavononóis, flavonas, xantonas, fenóis e flavonóides). Em 1 g/L de FAq foram encontrados 1,91 mM de potássio e 0,15 mM de sódio. A adição de FAq ao Tyrode não modificou significativamente a concentração desses íons. Os tempos de contração e de relaxamento, bem como o tempo de acoplamento eletromecânico não foram alterados pela FAq. Contudo, ela reduziu os Índices de Eficiência da contração e do relaxamento. A FAq aboliu completamente o fenômeno de Bowditch induzido por alta frequência de estimulação, indicando que ela reduz a entrada desse íon nas células. Com base nessa evidência, foram realizados protocolos para aprofundar o conhecimento sobre a participação das correntes de cálcio no mecanismo cardiodepressor da FAq. Esta fração produziu os seguintes resultados: 1) aboliu completamente o efeito inotrópico positivo do isoproterenol (10-1 a 103 pM); 2) deslocou para a direita a curva concentração-efeito para o CaCl2 (0,5 a 7,0 mM), aumentando a CE50 de 1,12 ± 0,07 (Hill = 1,5) para 7,23 ± 0,47 mM (Hill = 7,4) (n = 3; p < 0,05); 3) aboliu completamente o efeito inotrópico positivo do (-) BAY K8644 (5 a 2000 nM); 4) reduziu em cerca de 20% o pico da fluorescência intracelular correspondente ao transiente de cálcio citoplasmático (controle: n = 30 células; teste: n = 27 células; 4 animais); 5) não modificou a velocidade de decaimento do sinal de fluorescência, o que significa que ela não interfere com o funcionamento da bomba de cálcio do retículo sarcoplasmático; 6) reduziu em 23% a densidade de corrente de cálcio tipo-L que variou de -6,29 ± 0,34 para -4,9 ± 0,2 A/F (n = 5 animais, p < 0,05). 1) a FAq foi a fração com maior potência inotrópica; 2) os principais metabólitos secundários presentes na FAq foram taninos, saponinas e polifenóis; 3) a FAq reduz a força de contração do átrio; 4) o mecanismo da ação cardiodepressora da FAq sobre a contratilidade miocárdica se deve à diminuição da disponibilização do cálcio durante a contração.

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