Avaliação do efeito sinérgico dos peptídeos 8WH5 e 7WH5 na inibição do crescimento de Candida albicans
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Data
2023-05-18
Autores
Mendes, Isadora Lino [UNESP]
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Editor
Universidade Estadual Paulista (Unesp)
Resumo
Candida albicans é uma espécie de fungo polimórfico, que geralmente se apresenta como um microrganismo comensal da microbiota humana. Entretanto, em muitos indivíduos, tal espécie pode se desenvolver como um patógeno oportunista, capaz de causar infecções. Pelo fato de apresentar resistência aos medicamentos mais utilizados para tratamento, a pesquisa por novas moléculas antifúngicas e por novas combinações destas se torna de extrema importância. A associação de moléculas bioativas, além de diminuir o surgimento de mecanismos de resistência, pode fazer com que ocorra a redução da concentração das substâncias analisadas, sem causar mudanças na ação antimicrobiana. Nesse contexto, os peptídeos da família das Histatinas se mostram como moléculas promissoras a terapêutica, sendo a Histatina-5 (Hst5) e seus fragmentos 7WH5 e 8WH5, alguns dos representantes da classe. Tendo em vista os elementos supracitados, o objetivo deste trabalho foi avaliar a ocorrência de efeito sinérgico decorrente da combinação dos peptídeos 7WH5 e 8WH5 e do peptídeo 8WH5 com o fármaco fluconazol (Flu) na inibição do crescimento de C. albicans. Para a obtenção dos peptídeos foi utilizada a metodologia de síntese de peptídeos em fase sólida, e a purificação e caracterização destes ocorreu, respectivamente, por cromatografia líquida de alta eficiência e espectrometria de massas. A concentração inibitória mínima (CIM) dos peptídeos e do fármaco para as cepas C. albicans ATCC 90028, C. albicans ATCC 18804 e C. albicans ATCC 10231 foi determinada pelo método de diluição em microplacas. Já a avaliação do efeito sinérgico na inibição do crescimento de C. albicans nas três cepas anteriormente mencionadas foi realizada através do ensaio checkerboard, por meio da combinação de diferentes concentrações das substâncias testadas. Por fim, foi realizado o ensaio killing-assay, para observar o perfil temporal de ação das melhores combinações e das moléculas isoladas, e posteriormente realizar uma comparação entre ambos.
Candida albicans is a polymorphic fungal species which usually presents itself as a commensal microorganism in the human microbiota. However, in many individuals, this species may develop as an opportunistic pathogen capable of causing infections. Due to the fact of presenting resistance to the most used drugs for treatment, the search for new antifungal molecules and new combinations of these becomes of extreme importance. The association of bioactive molecules, in addition to reducing the appearance of resistance mechanisms, may reduce the concentration of the analyzed substances, without causing changes in the antimicrobial action. In this context, peptides from Histatins family show to be promising molecules for therapeutics, being Histatin-5 (Hst5) and its fragments 7WH5 and 8WH5 some representatives of this class. Considering the above-mentioned elements, the aim of this work is to evaluate the occurrence of synergistic effect due to the combination of peptides 7WH5 and 8WH5, and of peptide 8WH5 with the drug fluconazole (Flu), in the inhibition of C. albicans growth. To obtain the peptides the solid phase peptide synthesis methodology was used, and the purification and characterization of these occurred, respectively, by high-performance liquid chromatography and mass spectrometry. The minimum inhibitory concentration (MIC) of peptides and the drug for C. albicans ATCC 90028, C. albicans ATCC 18804 and C. albicans ATCC 10231 strains was determined by the microplate dilution method. The evaluation of the synergistic effect on the inhibition of growth of C. albicans in the three strains mentioned above was performed by the checkerboard test, through the combination of different concentrations of the substances tested. Lastly, the killing-assay was carried out to observe the temporal profile of action of the best combinations and the isolated molecules, and then to do a comparison between both.
Candida albicans is a polymorphic fungal species which usually presents itself as a commensal microorganism in the human microbiota. However, in many individuals, this species may develop as an opportunistic pathogen capable of causing infections. Due to the fact of presenting resistance to the most used drugs for treatment, the search for new antifungal molecules and new combinations of these becomes of extreme importance. The association of bioactive molecules, in addition to reducing the appearance of resistance mechanisms, may reduce the concentration of the analyzed substances, without causing changes in the antimicrobial action. In this context, peptides from Histatins family show to be promising molecules for therapeutics, being Histatin-5 (Hst5) and its fragments 7WH5 and 8WH5 some representatives of this class. Considering the above-mentioned elements, the aim of this work is to evaluate the occurrence of synergistic effect due to the combination of peptides 7WH5 and 8WH5, and of peptide 8WH5 with the drug fluconazole (Flu), in the inhibition of C. albicans growth. To obtain the peptides the solid phase peptide synthesis methodology was used, and the purification and characterization of these occurred, respectively, by high-performance liquid chromatography and mass spectrometry. The minimum inhibitory concentration (MIC) of peptides and the drug for C. albicans ATCC 90028, C. albicans ATCC 18804 and C. albicans ATCC 10231 strains was determined by the microplate dilution method. The evaluation of the synergistic effect on the inhibition of growth of C. albicans in the three strains mentioned above was performed by the checkerboard test, through the combination of different concentrations of the substances tested. Lastly, the killing-assay was carried out to observe the temporal profile of action of the best combinations and the isolated molecules, and then to do a comparison between both.
Descrição
Palavras-chave
Peptídeos, Sinergismo, Candida albicans, Histatina-5, Fluconazol, Peptides, Synergism, Histatin-5, Fluconazole, Antifúngicos, Antifungal agents