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1.
Int. j. morphol ; 38(6): 1639-1644, Dec. 2020. graf
Article in English | LILACS | ID: biblio-1134491

ABSTRACT

SUMMARY: Previous studies from our group described the consequences of using ethanol on penile erection. Nevertheless, the molecular mechanisms surrounding microRNAs, apoptosis process and their relationship with erectile dysfunction associated with alcohol consumption are still poorly understood. The objective of this analysis was to evaluate the mechanism of apoptosis by the expression of AIF and PARP, as well as their regulatory microRNAs: miR-145, miR-210 and miR-486, in the corpus cavernosum of rats submitted to a semivoluntary alcoholism model. For this study 24 Wistar rats were divided into two groups: control (C) and treated with 20 % ethanol (A) for seven weeks. The corpus cavernosum samples were prepared for immunohistochemical analysis of AIF and PARP protein expression, and microRNAs miR-145, miR-210, miR-486 gene expression in cavernous tissue was performed by real time PCR. The immunohistochemical analysis showed little nuclear positive labeling for the protein PARP and AIF in the corpus cavernosum of control and ethanol treated animals. After analysis of miR-145, -210 and -486 microRNA expression in the 12 animals studied, no results were found with significant statistical difference between the control and alcoholized groups. The expression of AIF and PARP and their regulatory microRNAs involved in apoptotic process (miR-145, miR-210 and miR-486) were not altered in the corpus cavernosum of rats submitted to semivoluntary alcoholism.


RESUMEN: Estudios previos de nuestro grupo describieron las consecuencias del uso de etanol en la erección del pene. Sin embargo, los mecanismos moleculares que rodean a los microARN, el proceso de apoptosis y su relación con la disfunción eréctil asociada con el consumo de alcohol aún no se conocen bien. El objetivo de este análisis fue evaluar el mecanismo de apoptosis mediante la expresión de AIF y PARP, así como sus microARN reguladores: miR-145, miR-210 y miR-486, en el cuerpo cavernoso de ratas sometidas a un modelo de alcoholismo semivoluntario. Se dividieron 24 ratas Wistar en dos grupos: control (C) grupo de ratas tratadas con etanol al 20 % (A) durante siete semanas. Las muestras del cuerpo cavernoso se prepararon para el análisis inmunohistoquímico de la expresión de la proteína AIF y PARP, y la expresión del gen microRNAs miR-145, miR-210, miR-486 en tejido cavernoso se realizó por PCR en tiempo real. El análisis inmunohistoquímico mostró escaso etiquetado nuclear positivo para la proteína PARP y AIF en el cuerpo cavernoso de los animales de control y tratados con etanol. Después del análisis de la expresión de microARN miR-145, -210 y -486 no se encontraron resultados con diferencias estadísticas significativas entre los grupos control y alcoholizados. La expresión de AIF y PARP y sus microARN reguladores involucrados en el proceso apoptótico (miR-145, miR-210 y miR-486) no se alteraron en el cuerpo cavernoso de las ratas sometidas a alcoholismo semivoluntario.


Subject(s)
Animals , Rats , Apoptosis , Alcoholism/metabolism , Erectile Dysfunction/metabolism , Penis/physiopathology , Penis/chemistry , Immunohistochemistry , Rats, Wistar , MicroRNAs/analysis , MicroRNAs/genetics , MicroRNAs/metabolism , Disease Models, Animal , Alcoholism/physiopathology , Apoptosis Inducing Factor/analysis , Apoptosis Inducing Factor/genetics , Apoptosis Inducing Factor/metabolism , Real-Time Polymerase Chain Reaction , Erectile Dysfunction/physiopathology
2.
Braz. J. Pharm. Sci. (Online) ; 53(1): e15210, 2017. graf
Article in English | LILACS | ID: biblio-839446

ABSTRACT

Abstract In this study, the potential antileukemic activity of grandisin, a lignan extracted from Piper solmsianum, was evaluated against the leukemic line K562. The cytotoxicity of grandisin (0.018 to 2.365 µM) was evaluated in K562 and normal peripheral blood lymphocytes by Trypan Blue Exclusion and MTT methods after 48h exposure to the drug. In both methods, cellular viability was concentration-dependent and the IC50 values were lower than 0.85µM. Analysis of K562 cells after treatment with grandisin showed that the cell cycle was arrested in the G1 phase with a 12.31% increase, while both S and G2 phases decreased. Morphological studies conducted after the exposure of K562 to grandisin revealed changes consistent with the apoptosis process, which was confirmed by anexin V stain and caspase activation. Thus, lignan grandisin showed antileukemic activities against the K562 cell line and the cell death process occurred via apoptosis.


Subject(s)
Gene Expression Regulation, Leukemic/genetics , Lignans/pharmacokinetics , K562 Cells/classification , Apoptosis Inducing Factor/analysis , Leukemia, Myelogenous, Chronic, BCR-ABL Positive/drug therapy , Piperaceae/classification
3.
Braz. j. pharm. sci ; 48(3): 497-505, July-Sept. 2012. ilus, graf
Article in English | LILACS | ID: lil-653464

ABSTRACT

Chronic myeloid leukemia (CML) is a clonal myeloproliferative disease that shows apoptosis resistance. The introduction of imatinib mesylate has revolutionized the treatment of CML, but imatinib resistance may develop at any time and inevitably leads to disease progression. Synadenium umbellatum Pax. belongs to the Euphorbiaceae family and is popularly used in Brazil for the treatment of cancer. The cytotoxicity of Euphorbiaceae is associated with the ability of these plants and their bioactive compounds to induce apoptotic tumor cell death. Therefore, we aimed to investigate the cytotoxicity and the mechanisms of death induced by S. umbellatum extract in leukemic cells. S. umbellatum cytotoxicity was evaluated by trypan blue exclusion assay and flow cytometric analysis of the cell cycle; the mechanisms involved in K-562 cell death were investigated by light microscopy and flow cytometry. The results demonstrate that S. umbellatum is cytotoxic to leukemic cells in a concentration-dependent manner. Morphological analysis revealed that S. umbellatum treatment induced K-562 cell death by an apoptotic pathway. Furthermore, data indicate ROS overproduction, alterations in mitochondrial membrane potential, phosphatidylserine externalization and activation of caspase 9. Taken together, the results demonstrate that S. umbellatum extract arrested the cell cycle and triggered apoptosis at several levels in K-562 cells.


A leucemia mielóide crônica (LMC) é uma doença mieloproliferativa clonal, que apresenta resistência à apoptose. A introdução do mesilato de imatinibe revolucionou o tratamento da LMC, porém a resistência ao imatinibe pode ser desenvolvida em qualquer tempo e, inevitavelmente, leva à progressão da doença. Synadenium umbellatum Pax. pertence à família Euphorbiaceae e é usado popularmente no Brasil para o tratamento do câncer. A citotoxicidade das Euphorbiaceae está associada com a capacidade dessas plantas e seus compostos bioativos em induzir apoptose em células tumorais. Portanto, este trabalho teve como objetivo investigar a citotoxicidade e os mecanismos de morte induzidos por S. umbellatum em células leucêmicas. A citotoxicidade de S. umbellatum foi avaliada pelo ensaio de exclusão do azul de tripano e a análise do ciclo celular foi feita por citometria de fluxo. Os mecanismos envolvidos na morte celular das células K-562 foram investigados por microscopia óptica e por citometria de fluxo. Os resultados demonstraram que S. umbellatum é citotóxico para células leucêmicas de uma maneira dependente da concentração. A análise morfológica revelou que o tratamento com S. umbellatum induziu as célula K-562 à morte por via apoptótica. Além disso, os dados indicam aumento de ERO S, alterações no potencial de membrana mitocondrial, externalização da fosfatidilserina e ativação de caspase 9. Em conjunto, os resultados demonstram que S. umbellatum promoveu retenção do ciclo celular das células K-562 e induziu estas células à morte por apoptose.


Subject(s)
Apoptosis Inducing Factor/analysis , Euphorbiaceae/classification , K562 Cells/immunology , Cell Cycle
4.
Arq. bras. med. vet. zootec ; 62(2): 258-264, abr. 2010. ilus
Article in Portuguese | LILACS | ID: lil-551825

ABSTRACT

Apoptose e seus mecanismos reguladores são eventos fisiológicos cruciais para a manutenção da homeostase placentária, e o desequilíbrio desses processos pode comprometer a função placentária e, consequentemente, o sucesso da gravidez. Neste estudo, investigou-se a apoptose utilizando-se histomorfometria em lâminas coradas em HE e submetidas à reação de TUNEL. Além disso, avaliou-se a expressão de Bcl-2 e das caspases 8 e 3, pela reação de polimerase em cadeia em tempo real, em placentas saudáveis em diferentes estádios de gestação. Amostras de placentônios de vacas com quatro, seis e nove meses de gestação foram colhidas e processadas. O índice apoptótico aumentou progressivamente com o avanço da gestação. Tanto o Bcl-2 quanto as caspases 3 e 8 foram expressas nos três períodos estudados, sendo a expressão de Bcl-2 menor que a de caspase 8, que é menor que a de caspase 3. Estes resultados indicam que essas moléculas estão envolvidas na via apoptótica ativada na maturação placentária, exibindo um padrão de expressão ao longo da gestação e contribuindo para o equilíbrio fisiológico da celularidade e renovação celular na placenta bovina.


Apoptosis and its regulating mechanisms are crucial physiological events for the maintenance of the placental homostasis; and disequilibrium of these processes may compromise placental function and the success of the pregnancy. In this study, apoptosis was investigated by histomorphometry using slides stained with HE and TUNEL reaction. Besides that, Bcl-2 and caspases 8 and 3 expression were evaluated by real time polymerase chain reaction in healthy placentas under different gestacional ages. Samples of placentones of cows at 4th, 6th, and 9th months of gestation were harvested and processed. The apoptotic index gradually increased with the advance of the gestation. Bcl-2 and caspases 3 and 8 were expressed in all the studied periods, being the expression of Bcl-2 lower than that of caspase 8, which was lower than caspase 3. These results indicate that these molecules are involved in the activated apoptotic way in the placental maturation, showing a standard expression throughout the gestation and contributing for the physiological balance of the cellularity and cellular turn over in bovine placenta.


Subject(s)
Animals , Cattle , Caspases/analysis , Caspases/adverse effects , Caspases/isolation & purification , Apoptosis Inducing Factor/analysis , Apoptosis Inducing Factor/deficiency , Placenta/anatomy & histology , Placenta/embryology , Cattle/anatomy & histology , Cattle/abnormalities , Cattle/surgery , Homeostasis , In Situ Nick-End Labeling/methods , In Situ Nick-End Labeling/veterinary , Pregnancy, Animal
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