Combined "in vitro" treatment of Amphotericin-B and Clomipramine on promastigotes of Leishmania braziliensis
Leishmaniasis, an infectious disease caused by protozoa of the genus Leishmania, is a public health problem around the world. Amphotericin-B, a drug used for its treatment, is often toxic to patients. The search for new drugs should identify molecular targets in the parasite...
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| Autores principales: | , , , , , , , |
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| Formato: | Artículo revista |
| Lenguaje: | Español |
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Universidad Nacional Córdoba. Facultad de Ciencias Médicas. Secretaria de Ciencia y Tecnología
2019
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| Materias: | |
| Acceso en línea: | https://revistas.unc.edu.ar/index.php/med/article/view/25679 |
| Aporte de: |
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I10-R327-article-25679 |
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ojs |
| institution |
Universidad Nacional de Córdoba |
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I-10 |
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R-327 |
| container_title_str |
Revista de la Facultad de Ciencias Médicas de Córdoba |
| language |
Español |
| format |
Artículo revista |
| topic |
Leishmaniasis Amphotericin-B clomipramine trypanothione reductase Leishmaniasis tripanotiona reductasa clomipramina anfotericina-B |
| spellingShingle |
Leishmaniasis Amphotericin-B clomipramine trypanothione reductase Leishmaniasis tripanotiona reductasa clomipramina anfotericina-B Amelia, E Juarez, M Vejarez, M Konigheim, B Aguilar, J Juarez, V Satrauss, M Rivarola, W Combined "in vitro" treatment of Amphotericin-B and Clomipramine on promastigotes of Leishmania braziliensis |
| topic_facet |
Leishmaniasis Amphotericin-B clomipramine trypanothione reductase Leishmaniasis tripanotiona reductasa clomipramina anfotericina-B |
| author |
Amelia, E Juarez, M Vejarez, M Konigheim, B Aguilar, J Juarez, V Satrauss, M Rivarola, W |
| author_facet |
Amelia, E Juarez, M Vejarez, M Konigheim, B Aguilar, J Juarez, V Satrauss, M Rivarola, W |
| author_sort |
Amelia, E |
| title |
Combined "in vitro" treatment of Amphotericin-B and Clomipramine on promastigotes of Leishmania braziliensis |
| title_short |
Combined "in vitro" treatment of Amphotericin-B and Clomipramine on promastigotes of Leishmania braziliensis |
| title_full |
Combined "in vitro" treatment of Amphotericin-B and Clomipramine on promastigotes of Leishmania braziliensis |
| title_fullStr |
Combined "in vitro" treatment of Amphotericin-B and Clomipramine on promastigotes of Leishmania braziliensis |
| title_full_unstemmed |
Combined "in vitro" treatment of Amphotericin-B and Clomipramine on promastigotes of Leishmania braziliensis |
| title_sort |
combined "in vitro" treatment of amphotericin-b and clomipramine on promastigotes of leishmania braziliensis |
| description |
Leishmaniasis, an infectious disease caused by protozoa of the genus Leishmania, is a public health problem around the world. Amphotericin-B, a drug used for its treatment, is often toxic to patients. The search for new drugs should identify molecular targets in the parasite and absent in the host. Trypanotione reductase is an exclusive enzyme of the order Kinetoplastide, which makes it useful as a molecular target. Tricyclic antidepressants, such as clomipramine (CLO) are good inhibitors of this enzyme. The objective of this work was to analyze the combined “in vitro” treatment of Amphotericin-B (ANF) and CLO on promastigotes of L. braziliensis.
We performed a cytotoxicity (MTT) test on Vero cells, incubating 2.5x105 cells/ml with ANF = 25-400µg/ml and CLO = 25-1000 µg/ml and a combined effect test (checkerboard) in which 3x106promastigotes/ml were incubated with ANF = 0.002-0.032µg/ml and CLO = 0.3-2µg/ml. We determined the values of the 50% inhibitory concentration (IC50) of both drugs, evaluated the effect of the combination calculating the Combination Index (CI: CI> 1 antagonistic; CI = 1 additive; CI <1 synergic) and plottedisobolograms from the IC50 values of each of the drugs and the combinations. The ultrastructural alterations in the leishmanias were evaluated with electron microscopy.
IC50 values upon promastigotes were 0.018μg/ml for ANF and 1.098μg/ml for CLO, being 25 and 108 times lower than the cytotoxic concentration, respectively. IC50 values upon Vero cells were 0.44μg/ml for ANF and 119μg/ml for CLO. The CI value of 0.18 indicates a synergistic effect. The IC50 of the combinations were located below the line of additivity in the isobolograms, which indicates a synergistic effect between ANF and CLO, coinciding with the result obtained with the CI. These results coincide with those observed in the microphotographs, since the greatest alterations corresponded to the leishmanias treated with the combination.
The observed synergistic effect may be the result of the complementary mechanisms of action of both drugs, which would potentiate the lethal effect, causing the death of the parasite at lower concentrations than the monotherapy and without obvious toxic effects to the host.
Leishmaniasis; Amphotericin-B; clomipramine; trypanothione reductase
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| publisher |
Universidad Nacional Córdoba. Facultad de Ciencias Médicas. Secretaria de Ciencia y Tecnología |
| publishDate |
2019 |
| url |
https://revistas.unc.edu.ar/index.php/med/article/view/25679 |
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I10-R327-article-256792024-08-27T18:26:04Z Combined "in vitro" treatment of Amphotericin-B and Clomipramine on promastigotes of Leishmania braziliensis Tratamiento combinado “in vitro” de Anfotericina-B y Clomipramina sobre promastigotes de Leishmania braziliensis Amelia, E Juarez, M Vejarez, M Konigheim, B Aguilar, J Juarez, V Satrauss, M Rivarola, W Leishmaniasis Amphotericin-B clomipramine trypanothione reductase Leishmaniasis tripanotiona reductasa clomipramina anfotericina-B Leishmaniasis, an infectious disease caused by protozoa of the genus Leishmania, is a public health problem around the world. Amphotericin-B, a drug used for its treatment, is often toxic to patients. The search for new drugs should identify molecular targets in the parasite and absent in the host. Trypanotione reductase is an exclusive enzyme of the order Kinetoplastide, which makes it useful as a molecular target. Tricyclic antidepressants, such as clomipramine (CLO) are good inhibitors of this enzyme. The objective of this work was to analyze the combined “in vitro” treatment of Amphotericin-B (ANF) and CLO on promastigotes of L. braziliensis. We performed a cytotoxicity (MTT) test on Vero cells, incubating 2.5x105 cells/ml with ANF = 25-400µg/ml and CLO = 25-1000 µg/ml and a combined effect test (checkerboard) in which 3x106promastigotes/ml were incubated with ANF = 0.002-0.032µg/ml and CLO = 0.3-2µg/ml. We determined the values of the 50% inhibitory concentration (IC50) of both drugs, evaluated the effect of the combination calculating the Combination Index (CI: CI> 1 antagonistic; CI = 1 additive; CI <1 synergic) and plottedisobolograms from the IC50 values of each of the drugs and the combinations. The ultrastructural alterations in the leishmanias were evaluated with electron microscopy. IC50 values upon promastigotes were 0.018μg/ml for ANF and 1.098μg/ml for CLO, being 25 and 108 times lower than the cytotoxic concentration, respectively. IC50 values upon Vero cells were 0.44μg/ml for ANF and 119μg/ml for CLO. The CI value of 0.18 indicates a synergistic effect. The IC50 of the combinations were located below the line of additivity in the isobolograms, which indicates a synergistic effect between ANF and CLO, coinciding with the result obtained with the CI. These results coincide with those observed in the microphotographs, since the greatest alterations corresponded to the leishmanias treated with the combination. The observed synergistic effect may be the result of the complementary mechanisms of action of both drugs, which would potentiate the lethal effect, causing the death of the parasite at lower concentrations than the monotherapy and without obvious toxic effects to the host. Leishmaniasis; Amphotericin-B; clomipramine; trypanothione reductase La leishmaniasis, una enfermedad infecciosa causada por protozoarios del género Leishmania, es un problema de salud pública alrededor del mundo. Anfotericina-B, fármaco utilizado en el tratamiento, es a menudo tóxico para los pacientes. La búsqueda de nuevos fármacos debe identificar blancos moleculares en el parásito y ausentes en el huésped. La tripanotiona reductasa es una enzima exclusiva del orden Kinetoplastide, por lo que le confiere utilidad como blanco molecular. Antidepresivos tricíclicos, como clomipramina (CLO) son buenos inhibidores de esta enzima. El objetivo del presente trabajo fue analizar el tratamiento combinado “in vitro” de Anfotericina-B (ANF) y CLO sobre promastigotes de L. braziliensis. Realizamos un ensayo de citotoxicidad (MTT) sobre celulas Vero, incubando 2,5x105 cel/mL con ANF=25-400µg/mL y CLO=25-1000µg/mL y un ensayo de efecto combinado (checkerboard) en el que se incubaron 3x106 promastigotes/mL con ANF=0,002-0,032µg/mL y CLO=0,3-2µg/mL. Determinamos los valores de la concentración inhibitoria del 50% (IC50) de ambas drogas y evaluamos el efecto de la combinación con el Índice de Combinación (CI: CI>1 antagónico; CI=1 aditivo; CI<1 sinérgico) y graficamos isobologramas a partir de los valores de IC50 de cada una de las drogas y de las combinaciones. Las alteraciones ultraestructurales de las leishmanias se evaluaron con microscopia electrónica. El IC50 de ANF fue de 0,018μg/ml y de CLO 1,098μg/ml sobre promastigotes, siendo 25 y 108 veces menor que la concentración citotóxica. Los valores de IC50 sobre la línea celular Vero fueron 0,44μg/ml para ANF y 119μg/ml para CLO. El valor del CI de 0,18 indica efecto sinérgico. Los isobologramas muestran que los IC50 de las combinaciones se ubicaron por debajo de la línea de aditividad, lo que indica un efecto sinérgico entre ANF y CLO, coincidiendo con el resultado obtenido con el índice de combinación. Estos resultados coinciden con lo observado en las microfotografías, ya que las mayores alteraciones corresponden a las leishmanias tratadas con la combinación. El efecto sinérgico observado puede ser resultado de los mecanismos de acción complementarios de ambas drogas, que estarían potenciando el efecto letal, produciendo la muerte del parásito a menores concentraciones que la monoterapia y sin efectos tóxicos evidentes sobre el huésped. Universidad Nacional Córdoba. Facultad de Ciencias Médicas. Secretaria de Ciencia y Tecnología 2019-10-10 info:eu-repo/semantics/article info:eu-repo/semantics/publishedVersion application/pdf https://revistas.unc.edu.ar/index.php/med/article/view/25679 Revista de la Facultad de Ciencias Médicas de Córdoba.; 2019: Suplemento JIC XX Revista de la Facultad de Ciencias Médicas de Córdoba; 2019: Suplemento JIC XX Revista da Faculdade de Ciências Médicas de Córdoba; 2019: Suplemento JIC XX 1853-0605 0014-6722 10.31053/1853.0605.v76.nSuplemento spa https://revistas.unc.edu.ar/index.php/med/article/view/25679/27391 Derechos de autor 2019 Universidad Nacional de Córdoba https://creativecommons.org/licenses/by-nc/4.0 |