Trypanosoma cruzi Cell Death Induced by the Morita-Baylis-Hillman Adduct 3-Hydroxy-2-Methylene-3-(4-Nitrophenylpropanenitrile)

Abstract

Chagas disease, caused by the protozoan Trypanosoma cruzi, remains a serious health concern due to the lack of effective vaccines or satisfactory treatment. In the search for new compounds against this neglected disease, we have previously demonstrated that the compound 3-Hydroxy-2-methylene-3-(4-nitrophenylpropanenitrile) (MBHA3), derived from the Morita-Baylis-Hillman reaction, effectively caused a loss of viability in both the epimastigote and trypomastigote forms. However, the mechanisms of parasite death elicited by MBHA3 remain unknown. The aim of this study was to better understand the morphophysiological changes and the mechanism of cell death induced by MBHA3 treatment on T. cruzi. To perform this analysis, we used confocal microscopy and flow cytometry to monitor the fluorescent probes such as annexin-V/propidium iodide (AV/PI), calcein-AM/ethidium homodimer (CA/EH), acridine orange (AO) and rhodamine 123 (Rho 123). Lower concentrations of MBHA3 led to alterations in the mitochondrial membrane potential and AO labeling, but did not decrease the viability of the epimastiogote forms, as determined by the CA/EH and AV/PI assays. Conversely, treatment with higher concentrations of MBHA3 led to extensive plasma membrane damage, loss of mitochondrion membrane potential, DNA fragmentation and acidification of the cytoplasm. Our findings suggest that at higher concentrations, MBHA3 induces T. cruzi epimastigote death by necrosis in a mitochondrion-dependent manner.

Synthesis and Biological in vitro and in vivo Evaluation of 2‐(5‐Nitroindazol‐1‐yl)ethylamines and Related Compounds as Potential Therapeutic Alternatives for Chagas Disease

“…Subsequently,t he parasites were resuspended in 0.5 mL PBS with 10 mgmL À1 Rho 123 (Sigma-Aldrich) or 10 mgmL À1 AO (Sigma-Aldrich) for 20 min. [66] Finally,t he samples were washed twice with ice-cold PBS, dispersed in 1mLo fP BS and immediately analyzed by flow cytometry (Becton Dickinson FACSAria III). The data were captured and analyzed using BD FACSDiva v8.01 software (Becton Dickinson Biosciences, 2350 Qume Drive, San Jose, CA, USA).…”

A coating composed of chitosan andCymbopogon citratus(Dc. Ex Nees) essential oil to control Rhizopus soft rot and quality in tomato fruit stored at room temperature

“…If the PI penetrates the cell, membrane integrity has been lost and a red staining occurs (Sandes et al, 2014). After the staining step, the spores were collected via centrifugation (14,000 × g for 10 min) and washed twice with PBS to remove residual dye.…”

Trypanosoma cruzi biochemical changes and cell death induced by an organometallic platinum‐based compound

“…The mechanism of cell death by necrosis involves an increase in the cell volume, which leads to the rupture of the plasma membrane and release of cell material. Apart from the damage to the plasma membrane, no specific biochemical markers have been attributed to necrosis . At last, autophagy involves the sequestration of excessive, old, and unneeded cytoplasmic organelles into large double‐membrane vesicles, called autophagosomes, which are loaded into lysosomes for degradation, with no accompanying inflammatory response …”