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Previous issue date: 2007-02-27 / Ouratea cuspidata (Ochnaceae) crude extracts were added to experimental systems
containing either host macrophages, or Leishmania braziliensis promastigotes (L566). The
biological effect by the use of the hexane (OCH), methanol (OCMeH) and ethyl-acetate
(OCAcEt) extracts added at different concentrations: 2mg/mL; 4mg/mL; 8mg/mL, 16mg/mL
e 32mg/mL. The OCMeOH was the only fraction to which the anti parasitic potentiality was
achieved at doses rates that generated bio protective effects in the host cells. The parasiticide
action, due by direct exposure of promastigotes to OCMeOH, as well as the parasitic growth,
and morphological alterations intermediary metabolism alterations (mitochondrial activity)
were concomitantly measured. Biological macrophagic functions were also evaluated using
Cricetus cricetus peritoneal macrophages as a model (Mf). Bio-protective assays were carried
out in order to determine the free radical generation by the extract constituents. The results
were compared with non treated L566 and Mf. 78% of the 16 mg/106 Mf/mL treated Mf
were viable but a significant decrease of their phagocytosis capability (92%) was detected.
Such alterations were evident in 62% of the cells treated with 32mg/106 Mf/mL. At this
concentration cells were 70% viable and presented 98% phagocytosis suppression. The
macrophages enzymatic- mitochondrial activity was gradually diminished, with 87% activity
at exposures to 8mg/106 Mf/mL; 66,3% at exposures to 16mg/106 Mf/mL, and 49% at
exposures to 32mg/106 Mf/mL, respectively. The anti parasitic effects were not associated to
the promastigotes lack of viability. Instead, a significant rising on the L566 cells counting was
detected when compared to control non treated parasites, from the first 6 hours treatment until
24 hours exposure. Morphological changes were detected in 62% of the 8mg/106 L566/mL
parasite treated cells; 78% after 16mg/106 L566/mL treatment, and 98% of the cells presented
morphological changes after 32mg/106 L566/mL treatment. Cells were either round-shaped,
showing incomplete mitosis, or presenting double flagella, suggesting that the present extract
containing substances that might interfere in the parasitic topoisomerase function. The
parasitary mitochondrial activity evidenced the occurrence of a metabolic acceleration due by
the OCMeOH treatment. The mitochondrial enzymatic activity was of 220% at exposures to
8mg/106 L566/mL; 389% at exposures to 16mg/106 L566/mL and of 480% at exposures to
32mg/106 L566/mL, respectively, when compared to the non treated parasites (100%). The
anti parasitic potential (Therapeutic Index) was considered to be positive (acceptable)
(TI=4,0) when estimated in function of the morphological changes observed at the extract
concentration of 32mg/106 Mf/mL (LD62%), and at 8mg/106 L566/mL (ED62%),
respectively. Since bi-flavonoids are the main constituents present in the OCMeOH, results
were suggestive the anti parasitic effect was due to this group of secondary metabolites. As
such biologically active molecules are known COX2 selective inhibitors, its internal use
should be avoided. Otherwise, bi- flavonoids are good candidate substances to be applied as
topical medicine to treat the American Tegumentar Leishmaniasis. / Os efeitos dos extratos de Ouratea cuspidata (Ochnaceae) foram avaliadas a partir da
exposi??o de c?lulas hospedeiras e de promastigotas de Leishmania (Viannia) braziliensis
(L566) ?s fra??es oriundas da sua parti??o hex?nica (OCH), metan?lica (OCMeOH) e acetato
de et?lica (OCAcEt). Dentre estas, a fra??o OCMeOH foi a ?nica que apresentou potencial
antiparasit?rio em doses que geraram bioprote??o aos sistemas hospedeiros. Assim,
procederam-se an?lises de a??o parasiticida direta do extrato OCMeOH sobre formas
promastigotas, onde, tamb?m, foram avaliados o crescimento, altera??es morfol?gicas e
fun??es do metabolismo mitocondriais. Do mesmo modo, foram realizadas observa??es
semelhantes em macr?fagos peritoneais de hamsters Cricetus cricetus (Mf) afim de se
determinar a toxicidade relativa para o hospedeiro. Finalmente, foram realizados ensaios
complementares de bioprote??o e gera??o de radicais livres, com a finalidade de confirmar a
poss?vel aplica??o terap?utica dos constituintes presentes no extrato. Em todas as etapas
experimentais, foram utilizadas cinco concentra??es de OCMeOH (2mg/mL; 4mg/mL;
8mg/mL; 16mg/mL e 32mg/mL), sendo que os resultados obtidos foram comparativamente
avaliados em rela??o a sistemas de controle (L566 e Mf n?o tratados). Os (Mf) tratados com
16 mg/106 Mf/mL do extrato sofreram uma diminui??o significativa (92%) de sua capacidade
fagocit?ria, mas permaneceram vi?veis, em sua maioria (78%). As altera??es morfol?gicas
mostraram-se mais evidentes em 62% das c?lulas tratadas com 32mg/106 Mf/mL, com 70%
de viabilidade e 98% de inibi??o da fagocitose. A atividade enzim?tica-mitocondrial
macrof?gica apresentou diminui??o gradativa, com a preserva??o de 87% das fun??es
enzim?ticas nas c?lulas tratadas com 8mg/106 Mf/mL; 66,3% daquelas tratadas com
16mg/106 Mf/mL e 49% das tratadas com 32mg/106 Mf/mL. As L566 n?o apresentaram a
perda da viabilidade ap?s exposi??o de 24horas ?s diferentes concentra??es de OCMeOH. Ao
contr?rio, houve um aumento significativo no n?mero de promastigotas que, a partir de seis
horas de cultivo, mostraram-se sempre superiores ? quantidade de parasitos n?o tratados.
Foram detectadas altera??es morfol?gicas em 62% das L566 tratadas com 8mg/106 L566/mL;
78% daquelas tratadas com 16mg/106 L566/mL e 98% das tratadas com 32mg/106 L566/mL.
As promastigotas tratadas apresentaram-se arredondadas, com mitose incompleta, ou
apresentando dois flagelos, sugerindo que as subst?ncias presentes no extrato podem interferir
nas topoisomerases parasit?rias. A atividade mitocond rial parasit?ria evidenciou a ocorr?ncia
de acelera??o metab?lica induzida pelo tratamento com OCMeOH. A atividade enzim?ticamitocondrial
foi de 220% no tratamento com 8mg/106 L566/mL; de 389% com 16mg/106
L566/mL e 480% com 32mg/106 L566/mL quando comparadas ao controle (100%). O
potencial antiparasit?rio (Indice Terap?utico) foi considerado positivo (IT=4,0) em fun??o das
altera??es morfol?gicas observadas nas concentra??es de 32mg/106 Mf/mL (LD62%), e
8mg/106 L566/mL (ED62%). Sendo biflavon?ides os constituintes presentes em maior
propor??o, atribuiu-se a este grupo de subst?ncias a a??o antiparasit?ria e imunot?xica
observadas para OCMeOH. Os bi-flavon?ides presentes no extrato metan?lico de Ouratea
cuspidata est?o entre os inibidores seletivos de COX2, sugerindo o desenvolvimento de
pesquisas que visem sua utiliza??o t?pica em animais naturalmente infectados.
| Identifer | oai:union.ndltd.org:IBICT/oai:localhost:tede/746 |
| Date | 27 February 2007 |
| Creators | Ribeiro, Renata da Silva |
| Contributors | Freire, Ronald Bastos |
| Publisher | Universidade Federal Rural do Rio de Janeiro, Curso de P?s-Gradua??o em Ci?ncias Veterin?rias, UFRRJ, Brasil, Parasitologia Veterin?ria |
| Source Sets | IBICT Brazilian ETDs |
| Language | Portuguese |
| Detected Language | English |
| Type | info:eu-repo/semantics/publishedVersion, info:eu-repo/semantics/masterThesis |
| Format | application/pdf |
| Source | reponame:Biblioteca Digital de Teses e Dissertações da UFRRJ, instname:Universidade Federal Rural do Rio de Janeiro, instacron:UFRRJ |
| Rights | info:eu-repo/semantics/openAccess |
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