Growth arrest and morphological changes triggered by emodin on Trypanosoma cruzi epimastigotes cultivated in axenic medium

Summary Cholestasis is a condition wherein bile flow is interrupted and lithocholic acid is known to play a key role in causing severe liver injury. In this study, we performed in-depth analysis of the morphological changes in bile canaliculi and the biological role of villin in cholestasis using lithocholic acid-stimulated HepG2 human hepatocarcinoma cells. We confirmed disruption of the bile canaliculi in liver sections from a liver allograft patient with cholestasis. Lithocholic acid caused strong cytotoxicity in HepG2 cells, which was associated with abnormal morphology. Lithocholic acid reduced villin expression, which recovered in the presence of nuclear receptor agonists. Furthermore, villin mRNA expression increased following small interfering RNA (siRNA)-mediated knockdown of the nuclear farnesoid X receptor and pregnane X receptor. Villin knockdown using siRNA caused cell growth arrest in HepG2 cells. The effect of villin-knockdown on whole-genome expression in HepG2 cells was analyzed by DNA microarray. Our data suggest that lithocholic acid caused cell growth arrest by suppressing villin expression via farnesoid X receptor and pregnane X receptor in HepG2 cells.

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Dual Role for Transforming Growth Factor β-Dependent Signaling in Trypanosoma cruzi Infection of Mammalian Cells

Infection and Immunity ◽

10.1128/iai.68.4.2077-2081.2000 ◽

2000 ◽

Vol 68 (4) ◽

pp. 2077-2081 ◽

Cited By ~ 37

Author(s):

Belinda S. Hall ◽

Miercio A. Pereira

Keyword(s):

Gene Expression ◽

Growth Factor ◽

Trypanosoma Cruzi ◽

Host Cell ◽

Cell Lines ◽

Mammalian Cells ◽

Transforming Growth Factor ◽

Growth Arrest ◽

Transforming Growth Factor Β

ABSTRACT Expression of functional transforming growth factor β (TGF-β) receptors (TβR) is required for the invasion of mammalian cells by the protozoan parasite Trypanosoma cruzi. However, the precise role of this host cell signaling complex in T. cruzi infection is unknown. To investigate the role of the TGF-β signaling pathway, infection levels were studied in the mink lung epithelial cell lines JD1, JM2, and JM3. These cells express inducible mutant TβR1 proteins that cannot induce growth arrest in response to TGF-β but still transmit the signal for TGF-β-dependent gene expression. In the absence of mutant receptor expression, trypomastigotes invaded the cells at a low level. Induction of the mutant receptors caused an increase in infection in all three cell lines, showing that the requirement for TGF-β signaling at invasion can be divorced from TGF-β-induced growth arrest. TGF-β pretreatment of mink lung cells expressing wild-type TβR1 caused a marked enhancement of infection, but no enhancement was seen in JD1, JM2, and JM3 cells, showing that the ability of TGF-β to stimulate infection is associated with growth arrest. Likewise, expression of SMAD7 or SMAD2SA, inhibitors of TGF-β signaling, did not block infection by T. cruzi but did block the enhancement of infection by TGF-β. Taken together, these results show that there is a dual role for TGF-β signaling in T. cruzi infection. The initial invasion of the host cell is independent of both TGF-β-dependent gene expression and growth arrest, but TGF-β stimulation of infection requires a fully functional TGF-β signaling pathway.

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Morphological changes during the growth cycle of axenic and monoxenic Tetrahymena pyriformis

Canadian Journal of Zoology ◽

10.1139/z76-232 ◽

1976 ◽

Vol 54 (11) ◽

pp. 2011-2018 ◽

Cited By ~ 9

Author(s):

William D. Taylor ◽

Michael A. Gates ◽

Jacques Berger

Keyword(s):

Culture Media ◽

Morphological Changes ◽

Food Limitation ◽

Food Resource ◽

Tetrahymena Pyriformis ◽

Growth Cycle ◽

Exponential Phase ◽

Type Ii ◽

Axenic Medium ◽

Morphological Differences

Morphometric data were collected from Tetrahymena pyriformis (WH-14, syngen I, mating type II) grown in two different media, one axenic and one monoxenic (with Serratia marcescens), at various times during the growth cycle. Stationary phase cells differed morphometrically from exponential phase cells in different ways in the two media. It is proposed that, in the monoxenic medium, division stopped because the food resource was depleted, while in the axenic medium division was blocked before food limitation occurred.Cells in the two culture media showed consistent morphological differences during exponential phase, the axenic cells being larger (length and width), but having smaller (in length) mouths. Significant differences between cells in replicate experiments were frequently observed.

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Mode of fibroblast growth enhancement by human interleukin-1.

The Journal of Cell Biology ◽

10.1083/jcb.106.3.813 ◽

1988 ◽

Vol 106 (3) ◽

pp. 813-819 ◽

Cited By ~ 24

Author(s):

J P Singh ◽

L D Adams ◽

P D Bonin

Keyword(s):

Dna Synthesis ◽

Morphological Changes ◽

Interleukin 1 ◽

Growth Arrest ◽

Extended Period ◽

Growth Stimulation ◽

Growth Enhancement ◽

Fibroblast Growth ◽

Wide Range ◽

Competence Induction

Previous studies have demonstrated that interleukin-1 (IL-1) stimulates fibroblast growth (Schmidt, J. A., S. B. Mizel, D. Cohn, and I. Green. 1982. J. Immunol. 128:2177-2182) and binds to specific, high affinity receptors of BALB/c3T3 cells (Bird, T. A., and J. Saklatval. 1986. Nature (Lond.). 324:263-265, 266-268). We have investigated the mechanism of fibroblast growth stimulation by IL-1. Addition of fibroblast growth factor derived from platelets (PDGF) to a quiescent culture of BALB/c3T3 cells produced 8-10-fold increase in DNA synthesis during 24-h incubation. The cellular action of PDGF was mediated through competence induction and required synergistic action of plasma-derived factors for full mitogenic activity. When tested at a wide range of concentrations (0.1-100 pM), natural IL-1 or recombinant IL-1 produced only a maximum of 5-10% of DNA synthesis elicited in response to PDGF or serum. Induction of DNA synthesis required continuous presence of IL-1 and did not exhibit synergism with plasma. Competence induction and mitogenic stimulation by PDGF was associated with early induction of proteins P32, P38, P46-48, P75, and changes in cytoskeletal organization. Examination of these early cellular changes showed that IL-1 did not produce similar induction of cellular proteins and the morphological changes associated with growth stimulation. These results suggest that the mode of IL-1 action on BALB/c3T3 was not through competence induction. When IL-1 was added to cells rendered competent by brief exposure to PDGF, 10-15% additional DNA synthesis occurred during the first 24 h. Extended incubation of PDGF-treated cells in the presence of IL-1 revealed that the stimulation by IL-1 occurred predominantly during the subsequent cycle of DNA replication, wherein DNA synthesis reached three- to fivefold higher than the untreated cultures. We conclude (a) IL-1 alone is not a potent mitogen for BALB/c3T3 cells, and does not bring cells out of the growth arrest Go phase, (b) treatment with PDGF renders the cells more responsive to IL-1, (c) part of the IL-1 action on competent cells may be characterized as progression inducing activity, further, (d) our results indicate that action of IL-1 on PDGF-treated cells produces sustained DNA synthesis for an extended period, perhaps by preventing the entry of cells into growth arrest Go phase.

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Proteasome activity is required for the stage-specific transformation of a protozoan parasite.

Journal of Experimental Medicine ◽

10.1084/jem.184.5.1909 ◽

1996 ◽

Vol 184 (5) ◽

pp. 1909-1918 ◽

Cited By ~ 68

Author(s):

J González ◽

F J Ramalho-Pinto ◽

U Frevert ◽

J Ghiso ◽

S Tomlinson ◽

...

Keyword(s):

Essential Role ◽

Morphological Changes ◽

Protozoan Parasite ◽

Eukaryotic Cells ◽

Cysteine Proteinases ◽

Axenic Medium ◽

Cytoplasmic Proteins ◽

Intracellular Parasites

A prominent feature of the life cycle of intracellular parasites is the profound morphological changes they undergo during development in the vertebrate and invertebrate hosts. In eukaryotic cells, most cytoplasmic proteins are degraded in proteasomes. Here, we show that the transformation in axenic medium of trypomastigotes of Trypanosoma cruzi into amastigote-like organisms, and the intracellular development of the parasite from amastigotes into trypomastigotes, are prevented by lactacystin, or by a peptide aldehyde that inhibits proteasome function. Clasto-lactacystin, an inactive analogue of lactacystin, and cell-permeant peptide aldehyde inhibitors of T. cruzi cysteine proteinases have no effect. We have also identified the 20S proteasomes from T. cruzi as a target of lactacystin in vivo. Our results document the essential role of proteasomes in the stage-specific transformation of a protozoan.

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Inhibition of NAD+-dependent histone deacetylases (sirtuins) causes growth arrest and activates both apoptosis and autophagy in the pathogenic protozoan Trypanosoma cruzi

Parasitology ◽

10.1017/s0031182013001704 ◽

2014 ◽

Vol 141 (6) ◽

pp. 814-825 ◽

Cited By ~ 17

Author(s):

PHERCYLES VEIGA-SANTOS ◽

LISSA CATHERINE REIGNAULT ◽

KILIAN HUBER ◽

FRANZ BRACHER ◽

WANDERLEY DE SOUZA ◽

...

Keyword(s):

Trypanosoma Cruzi ◽

Chagas Disease ◽

Histone Deacetylases ◽

Morphological Changes ◽

Light And Electron Microscopy ◽

Chronic Phase ◽

Physiological Processes ◽

Low Concentrations ◽

Disease Therapy ◽

New Compounds

SUMMARYChagas disease, which is caused by the parasite Trypanosoma cruzi, affects approximately 7–8 million people in Latin America. The drugs available to treat this disease are ineffective against chronic phase disease and are associated with toxic side effects. Therefore, the development of new compounds that can kill T. cruzi at low concentrations is critically important. Herein, we report the effects of a novel 3-arylideneindolin-2-one that inhibits sirtuins, which are highly conserved proteins that are involved in a variety of physiological processes. The compound KH-TFMDI was tested against the epimastigote, trypomastigote and amastigote forms of T. cruzi, and its effects were evaluated using flow cytometry, light and electron microscopy. KH-TFMDI inhibited the replication of T. cruzi intracellular amastigotes with an IC50 of 0·5±0·2 μm, which is significantly lower than the IC50 of benznidazole. The compound also lysed the highly infectious bloodstream trypomastigotes (BST) with LC50 values of 0·8±0·3 μm at 4 °C and 2·5±1·1 μm at 37 °C. KH-TFMDI inhibited cytokinesis and induced several morphological changes in the parasite, leading to its death by apoptosis and autophagy. This study highlights sirtuins as a potential new target for Chagas disease therapy.

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Effect of Crotalus viridis viridis snake venom on the ultrastructure and intracellular survival of Trypanosoma cruzi

Parasitology ◽

10.1017/s0031182010000958 ◽

2010 ◽

Vol 138 (1) ◽

pp. 46-58 ◽

Cited By ~ 19

Author(s):

CAMILA M. ADADE ◽

BRUNO LEMOS CONS ◽

PAULO A. MELO ◽

THAÏS SOUTO-PADRÓN

Keyword(s):

Trypanosoma Cruzi ◽

Chagas Disease ◽

Morphological Changes ◽

Chemotherapeutic Agents ◽

Patient Treatment ◽

Medical Treatments ◽

Crotalus Viridis ◽

Cell Staining ◽

Host Cell Cytoplasm ◽

Potential Use

SUMMARYChagas' disease, caused by Trypanosoma cruzi, affects 16–18 million people in Central and South America. Patient treatment is based on drugs that have toxic effects and limited efficacy. Therefore, new chemotherapeutic agents need to be developed. Snake venoms are sources of natural compounds used in various medical treatments. We observed that Crotalus viridis viridis venom was effective against all developmental forms of T. cruzi. Ultrastructural analysis revealed swelling of mitochondria, blebbing and disruption of the plasma membrane, loss of cytoplasm components and morphological changes of the cell. Staining with propidium iodide and rhodamine 123 confirmed the observed alterations in the plasma and mitochondrial membranes, respectively. The effects of the venom on the parasite intracellular cycle were also analysed. Pre-infected LLC-MK2 cells incubated with Cvv venom showed a 76–93% reduction in the number of parasites per infected cell and a 94–97·4% reduction in the number of parasites per 100 cells after 96 h of infection. Free trypomastigotes harvested from the supernatants of Cvv venom-treated cells were incapable of initiating a new infection cycle. Our data demonstrate that Cvv venom can access the host cell cytoplasm at concentrations that cause toxicity only to the amastigote forms of T. cruzi, and yields altered parasites with limited infective capacity, suggesting the potential use of Cvv venom in Chagas' disease chemotherapy.

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Single cell monitoring of growth arrest and morphological changes induced by transfer of wild-type p53 alleles to glioblastoma cells.

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.92.4.1008 ◽

1995 ◽

Vol 92 (4) ◽

pp. 1008-1012 ◽

Cited By ~ 31

Author(s):

E. G. Van Meir ◽

K. Roemer ◽

A. C. Diserens ◽

T. Kikuchi ◽

S. A. Rempel ◽

...

Keyword(s):

Single Cell ◽

Morphological Changes ◽

Growth Arrest ◽

Wild Type ◽

Glioblastoma Cells ◽

Cell Monitoring ◽

Wild Type P53

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In vitro studies and preclinical evaluation of benznidazole microparticles in the acute Trypanosoma cruzi murine model

Parasitology ◽

10.1017/s0031182020002310 ◽

2020 ◽

pp. 1-10

Author(s):

Marcela S. Rial ◽

Katia P. Seremeta ◽

Mónica I. Esteva ◽

Jacqueline Búa ◽

Claudio J. Salomon ◽

...

Keyword(s):

Trypanosoma Cruzi ◽

Chagas Disease ◽

Murine Model ◽

Morphological Changes ◽

Quantitative Polymerase Chain Reaction ◽

Parasitic Infection ◽

Parasite Load ◽

Enzyme Linked Immunosorbent Assay ◽

Significant Difference

Abstract Chagas disease is a serious parasitic infection caused by Trypanosoma cruzi. Unfortunately, the current chemotherapeutic tools are not enough to combat the infection. The aim of this study was to evaluate the trypanocidal activity of benznidazole-loaded microparticles during the acute phase of Chagas infection in an experimental murine model. Microparticles were prepared by spray-drying using copolymers derived from esters of acrylic and methacrylic acids as carriers. Dissolution efficiency of the formulations was up to 3.80-fold greater than that of raw benznidazole. Stability assay showed no significant difference (P > 0.05) in the loading capacity of microparticles for 3 years. Cell cultures showed no visible morphological changes or destabilization of the cell membrane nor haemolysis was observed in defibrinated human blood after microparticles treatment. Mice with acute lethal infection survived 100% after 30 days of treatment with benznidazole microparticles (50 mg kg−1 day−1). Furthermore, no detectable parasite load measured by quantitative polymerase chain reaction and lower levels of T. cruzi-specific antibodies by enzyme-linked immunosorbent assay were found in those mice. A significant decrease in the inflammation of heart tissue after treatment with these microparticles was observed, in comparison with the inflammatory damage observed in both infected mice treated with raw benznidazole and untreated infected mice. Therefore, these polymeric formulations are an attractive approach to treat Chagas disease.

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