Evaluation of drugs and stationary growth on the cell cycle of Giardia intestinalis

Reaume, Chase; Moore, Bradley J.; Hernández, Paula C.; Ruzzini, Antonio C.; Chlebus, Matthew; Wasserman, Moisés; Yee, Janet

doi:10.1016/j.molbiopara.2012.11.005

Cited by 10 publications

(10 citation statements)

References 17 publications

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“…It is noteworthy that the cell cycle arrest is not complete and this allows a portion of the cell population to continue through the cell cycle. The effect of different drugs on the Giardia cell cycle has been reported to show different phenomena, including partial arrest in different cell cycle stages depending on the drug under study ( Reaume et al, 2013 ). In our work, ABZ-treated trophozoites showed a pattern consistent with Giardia trophozoites displaying incomplete cytokinesis upon exposure to microtubule-acting compounds ( Mariante et al, 2005 ).…”

Section: Discussionmentioning

confidence: 99%

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Albendazole induces oxidative stress and DNA damage in the parasitic protozoan Giardia duodenalis

et al. 2015

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The control of Giardia duodenalis infections is carried out mainly by drugs, among these albendazole (ABZ) is commonly used. Although the cytotoxic effect of ABZ usually involves binding to β-tubulin, it has been suggested that oxidative stress may also play a role in its parasiticidal mechanism. In this work the effect of ABZ in Giardia clones that are susceptible or resistant to different concentrations (1.35, 8, and 250 μM) of this drug was analyzed. Reactive oxygen species (ROS) were induced by ABZ in susceptible clones and this was associated with a decrease in growth that was alleviated by cysteine supplementation. Remarkably, ABZ-resistant clones exhibited partial cross-resistance to H2O2, whereas a Giardia H2O2-resistant strain can grow in the presence of ABZ. Lipid oxidation and protein carbonylation in ABZ-treated parasites did not show significant differences as compared to untreated parasites; however, ABZ induced the formation of 8OHdG adducts and DNA degradation, indicating nucleic acid oxidative damage. This was supported by observations of histone H2AX phosphorylation in ABZ-susceptible trophozoites treated with 250 μM ABZ. Flow cytometry analysis showed that ABZ partially arrested cell cycle in drug-susceptible clones at G2/M phase at the expense of cells in G1 phase. Also, ABZ treatment resulted in phosphatidylserine exposure on the parasite surface, an event related to apoptosis. All together these data suggest that ROS induced by ABZ affect Giardia genetic material through oxidative stress mechanisms and subsequent induction of apoptotic-like events.

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Section: Discussionmentioning

confidence: 99%

“…Finally cell pellets were washed, stained with PI (1 μM in PBS), washed and resuspended in small volume (200–300 μL) for analysis in a FACS Calibur Flow Cytometer in at least 10,000 events per sample. The histogram areas were identified as reported by Reaume et al (2013) .…”

Section: Methodsmentioning

confidence: 99%

Albendazole induces oxidative stress and DNA damage in the parasitic protozoan Giardia duodenalis

et al. 2015

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“…In brief, parasites were washed with PBS, fixed in 70% ethanol for 30 min, washed again with PBS and incubated overnight at 4 o C in RNAse A (0.1 mg/mL). Finally, PI was added at a final concentration of 50 µg/mL and after 30 min at room temperature samples were quantified in a FACSCalibur flow cytometer with filter at 585 nm and cell cycle stages in histogram regions were identified as reported by Reaume et al 13 In these assays 25,000 cells per sample (three independent experiments) were recorded.…”

Section: Methodsmentioning

confidence: 99%

Ultrastructural and proapoptotic-like effects of kaempferol in Giardia duodenalis trophozoites and bioinformatics prediction of its potential protein target

Argüello-García

Calzada

García-Hernández

et al. 2020

Mem. Inst. Oswaldo Cruz

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BACKGROUND Kaempferol (KPF) is a flavonoid with antiparasitic activity including experimental giardiasis which mechanism of action is unknown. OBJECTIVE To analyse the cytotoxic effects of KPF on Giardia duodenalis trophozoites and to identify a likely parasite target of this compound. METHODS We used inhibitory concentrations of KPF (IC 25 , IC 50 and IC 100 ) and albendazole (ABZ) as reference drug. The ultrastructure of the trophozoites was analysed by transmission electron microscopy (TEM) whilst apoptosis/necrosis, production of reactive oxygen species (ROS) and cell cycle progression were assessed by flow cytometry (FCM) and confocal laser microscopy (CLM). Ligand-protein docking analyses were carried out using KPF structure from a drug library and crystal structure of a G. duodenalis aldose reductase ( GdAldRed ) homolog. RESULTS KPF provoked appearance of perinuclear and periplasmic spaces devoid of cytosolic content and multilamellar structures. KPF induced proapoptotic death associated with partial arrest in the S phase without ROS production. Bioinformatics approaches predicted that GdAldRed is a viable KPF target (ΔG = -7.09 kCal/mol), exhibiting 92% structural identity and a similar coupling pattern as its human homolog. CONCLUSIONS KPF exerted a proapoptotic effect on G. duodenalis trophozoites involving partial interruption of DNA synthesis without oxidative stress or structure damage to chromatin and cytoskeletal structures. GdAldRed is a likely target underlying its antigiardial activity.

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“…Furthermore, the giardia cell cycle has several unusual and unexpected features. For example, the giardia is able to continue to grow in size when DNA synthesis is blocked ( 13 , 14 ) and is able to enter mitosis with double-stranded DNA breaks ( 15 ) or with defective spindles ( 16 ). These observations suggest that the giardia is able to override or has defective cell cycle checkpoints that are essential in other eukaryotic cells.…”

Section: Introductionmentioning

confidence: 99%

“…Another giardia cell cycle synchronization protocol involves an initial incubation with 100 nM nocodazole for 2 h followed by 6 μM aphidicolin for 6 h, which results in the arrest of cells in G 1 /S ( 18 ). However, aphidicolin induces double-stranded DNA breaks ( 15 ) and nocodazole induces endoreplication ( 13 , 14 ) as side effects in the treated giardia cells. Fluorouracil, hydroxyurea, colchicine, and demecolcine treatments and nutrient deprivation were also tested as agents for giardia synchronization but resulted in no effect on the giardia cell cycle, nonspecific blocks, or irreversible effects ( 14 ).…”

Section: Introductionmentioning

confidence: 99%

Drug-Free Approach To Study the Unusual Cell Cycle of Giardia intestinalis

Horlock-Roberts

Reaume

Dayer

et al. 2017

mSphere

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Giardias are among the most commonly reported intestinal protozoa in the world, with infections seen in humans and over 40 species of animals. The life cycle of giardia alternates between the motile trophozoite and the infectious cyst. The regulation of the cell cycle controls the proliferation of giardia trophozoites during an active infection and contains the restriction point for the differentiation of trophozoite to cyst. Here, we developed counterflow centrifugal elutriation as a drug-free method to obtain fractions of giardia cultures enriched in cells from the G1, S, and G2 stages of the cell cycle. Analysis of these fractions showed that the cells do not show side effects associated with the drugs used for synchronization of giardia cultures. Therefore, counterflow centrifugal elutriation would advance studies on key regulatory events during the giardia cell cycle and identify potential drug targets to block giardia proliferation and transmission.

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Evaluation of drugs and stationary growth on the cell cycle of Giardia intestinalis

Cited by 10 publications

References 17 publications

Albendazole induces oxidative stress and DNA damage in the parasitic protozoan Giardia duodenalis

Albendazole induces oxidative stress and DNA damage in the parasitic protozoan Giardia duodenalis

Ultrastructural and proapoptotic-like effects of kaempferol in Giardia duodenalis trophozoites and bioinformatics prediction of its potential protein target

Drug-Free Approach To Study the Unusual Cell Cycle of Giardia intestinalis

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