Motility patterns of Trypanosoma cruzi trypomastigotes correlate with the efficiency of parasite invasion in vitro

Abstract Numerous works have demonstrated that trypanosomatid motility is relevant for parasite replication and sensitivity. Nonetheless, although some findings indirectly suggest that motility also plays an important role during infection, this has not been extensively investigated. This work is aimed at partially filling this void for the case of Trypanosoma cruzi. After recording swimming T. cruzi trypomastigotes (CL Brener strain) and recovering their individual trajectories, we statistically analyzed parasite motility patterns. We did this with parasites that swim alone or above monolayer cultures of different cell lines. Our results indicate that T. cruzi trypomastigotes change their motility patterns when they are in the presence of mammalian cells, in a cell-line dependent manner. We further performed infection experiments in which each of the mammalian cell cultures were incubated for 2 h together with trypomastigotes, and measured the corresponding invasion efficiency. Not only this parameter varied from cell line to cell line, but it resulted to be positively correlated with the corresponding intensity of the motility pattern changes. Together, these results suggest that T. cruzi trypomastigotes are capable of sensing the presence of mammalian cells and of changing their motility patterns accordingly, and that this might increase their invasion efficiency.

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The Pseudomonas aeruginosa product pyochelin interferes with Trypanosoma cruzi infection and multiplication in vitro

Transactions of the Royal Society of Tropical Medicine and Hygiene ◽

10.1093/trstmh/trz136 ◽

2020 ◽

Vol 114 (7) ◽

pp. 492-498 ◽

Cited By ~ 1

Author(s):

Gabriele Sass ◽

Laura C Miller Conrad ◽

Terrence-Thang H Nguyen ◽

David A Stevens

Keyword(s):

Pseudomonas Aeruginosa ◽

Trypanosoma Cruzi ◽

Mammalian Cells ◽

Iron Acquisition ◽

Vero Cells ◽

Dependent Manner ◽

Wild Type ◽

Current Standard ◽

Cruzi Infection

Abstract Background Bacteria are sources of numerous molecules used in treatment of infectious diseases. We investigated effects of molecules produced by 26 Pseudomonas aeruginosa strains against infection of mammalian cell cultures with Trypanosoma cruzi, the aetiological agent of Chagas disease. Methods Vero cells were infected with T. cruzi in the presence of wild-type P. aeruginosa supernatants or supernatants of mutants with defects in the production of various virulence, quorum sensing and iron acquisition factors. Quantification of T. cruzi infection (percentage of infected cells) and multiplication (number of amastigotes per infected cell) was performed and cell viability was determined. Results Wild-type P. aeruginosa products negatively affected T. cruzi infection and multiplication in a dose-dependent manner, without evident toxicity for mammalian cells. PvdD/pchE mutation (loss of the P. aeruginosa siderophores pyoverdine and pyochelin) had the greatest impact on anti–T. cruzi activity. Negative effects on T. cruzi infection by pure pyochelin, but not pyoverdine, or other P. aeruginosa exoproducts studied, were quantitatively similar to the effects of benznidazole, the current standard therapy against T. cruzi. Conclusions The P. aeruginosa product pyochelin showed promising activity against T. cruzi and might become a new lead molecule for therapy development.

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β-Chemokines Enhance Parasite Uptake and Promote Nitric Oxide-Dependent Microbiostatic Activity in Murine Inflammatory Macrophages Infected with Trypanosoma cruzi

Infection and Immunity ◽

10.1128/iai.67.9.4819-4826.1999 ◽

1999 ◽

Vol 67 (9) ◽

pp. 4819-4826 ◽

Cited By ~ 101

Author(s):

Júlio C. S. Aliberti ◽

Fabiana S. Machado ◽

Janeusa T. Souto ◽

Ana P. Campanelli ◽

Mauro M. Teixeira ◽

...

Keyword(s):

Trypanosoma Cruzi ◽

Neutralizing Antibodies ◽

Specific Inhibitor ◽

No Production ◽

Dependent Manner ◽

Macrophage Infection ◽

Parasite Replication ◽

Ifn Γ

ABSTRACT In the present study, we describe the ability of Trypanosoma cruzi trypomastigotes to stimulate the synthesis of β-chemokines by macrophages. In vivo infection with T. cruzi led to MIP-1α, RANTES, and JE/MCP1 mRNA expression by cells from peritoneal inflammatory exudate. In addition, in vitro infection with T. cruzi resulted in expression of β-chemokine MIP-1α, MIP-1β, RANTES, and JE mRNA by macrophages. The expression of the β-chemokine MIP-1α, MIP-1β, RANTES, and JE proteins by murine macrophages cultured with trypomastigote forms ofT. cruzi was confirmed by immunocytochemistry. Interestingly, macrophage infection with T. cruzi also resulted in NO production, which we found to be mediated mainly by β-chemokines. Hence, treatment with anti-β-chemokine-specific neutralizing antibodies partially inhibited NO release by macrophages incubated with T. cruzi parasites. Further, the addition of the exogenous β-chemokines MIP-1α, MIP-1β, RANTES, and JE/MCP-1 induced an increased T. cruzi uptake, leading to enhanced NO production and control of parasite replication in a dose-dependent manner. l-NMMA, a specific inhibitor of thel-arginine–NO pathway, caused a decrease in NO production and parasite killing when added to cultures of macrophages stimulated with β-chemokines. Among the β-chemokines tested, JE was more potent in inhibiting parasite growth, although it was much less efficient than gamma interferon (IFN-γ). Nevertheless, JE potentiates parasite killing by macrophages incubated with low doses of IFN-γ. Together, these results suggest that in addition to their chemotactic activity, murine β-chemokines may also contribute to enhancing parasite uptake and promoting control of parasite replication in macrophages and may play a role in resistance to T. cruziinfection.

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Antifungal activity of dendritic cell lysosomal proteins against Cryptococcus neoformans

Scientific Reports ◽

10.1038/s41598-021-92991-6 ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Benjamin N. Nelson ◽

Savannah G. Beakley ◽

Sierra Posey ◽

Brittney Conn ◽

Emma Maritz ◽

...

Keyword(s):

Antifungal Activity ◽

Cryptococcus Neoformans ◽

Mammalian Cells ◽

Cysteine Proteases ◽

Dependent Manner ◽

Life Threatening ◽

Opportunistic Fungal Pathogen ◽

Dose Dependent ◽

Lysosomal Proteins

AbstractCryptococcal meningitis is a life-threatening disease among immune compromised individuals that is caused by the opportunistic fungal pathogen Cryptococcus neoformans. Previous studies have shown that the fungus is phagocytosed by dendritic cells (DCs) and trafficked to the lysosome where it is killed by both oxidative and non-oxidative mechanisms. While certain molecules from the lysosome are known to kill or inhibit the growth of C. neoformans, the lysosome is an organelle containing many different proteins and enzymes that are designed to degrade phagocytosed material. We hypothesized that multiple lysosomal components, including cysteine proteases and antimicrobial peptides, could inhibit the growth of C. neoformans. Our study identified the contents of the DC lysosome and examined the anti-cryptococcal properties of different proteins found within the lysosome. Results showed several DC lysosomal proteins affected the growth of C. neoformans in vitro. The proteins that killed or inhibited the fungus did so in a dose-dependent manner. Furthermore, the concentration of protein needed for cryptococcal inhibition was found to be non-cytotoxic to mammalian cells. These data show that many DC lysosomal proteins have antifungal activity and have potential as immune-based therapeutics.

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PapRIV, a BV-2 microglial cell activating quorum sensing peptide

Scientific Reports ◽

10.1038/s41598-021-90030-y ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Yorick Janssens ◽

Nathan Debunne ◽

Anton De Spiegeleer ◽

Evelien Wynendaele ◽

Marta Planas ◽

...

Keyword(s):

Quorum Sensing ◽

Cell Model ◽

Dependent Manner ◽

Communication Signals ◽

Gram Positive Bacteria ◽

A Cell ◽

Gastro Intestinal Tract

AbstractQuorum sensing peptides (QSPs) are bacterial peptides produced by Gram-positive bacteria to communicate with their peers in a cell-density dependent manner. These peptides do not only act as interbacterial communication signals, but can also have effects on the host. Compelling evidence demonstrates the presence of a gut-brain axis and more specifically, the role of the gut microbiota in microglial functioning. The aim of this study is to investigate microglial activating properties of a selected QSP (PapRIV) which is produced by Bacillus cereus species. PapRIV showed in vitro activating properties of BV-2 microglia cells and was able to cross the in vitro Caco-2 cell model and reach the brain. In vivo peptide presence was also demonstrated in mouse plasma. The peptide caused induction of IL-6, TNFα and ROS expression and increased the fraction of ameboid BV-2 microglia cells in an NF-κB dependent manner. Different metabolites were identified in serum, of which the main metabolite still remained active. PapRIV is thus able to cross the gastro-intestinal tract and the blood–brain barrier and shows in vitro activating properties in BV-2 microglia cells, hereby indicating a potential role of this quorum sensing peptide in gut-brain interaction.

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Regulation of the tumour suppressor Fbw7α by PKC-dependent phosphorylation and cancer-associated mutations

Biochemical Journal ◽

10.1042/bj20100799 ◽

2010 ◽

Vol 432 (1) ◽

pp. 77-87 ◽

Cited By ~ 12

Author(s):

Joanne Durgan ◽

Peter J. Parker

Keyword(s):

Mammalian Cells ◽

Tumour Suppressor ◽

Dependent Manner ◽

Serine Threonine Kinase ◽

Threonine Kinase ◽

Pkc Protein Kinase C ◽

Specific Regulation ◽

The Relationship ◽

Substrate Binding Domain

Fbw7 (F-box WD40 protein 7) is a major tumour suppressor, which mediates the degradation of several potent oncogenes. PKC (protein kinase C) comprises a serine/threonine kinase family that can promote transformation when dysregulated. In the present study, we investigated the relationship between Fbw7 and PKC. Multiple members of the PKC superfamily interact with the substrate-binding domain of Fbw7. However, we find no evidence for Fbw7-mediated degradation of PKC. Instead, we demonstrate that Fbw7 is a novel substrate for PKC. Two residues within the isoform-specific N-terminus of Fbw7α are phosphorylated in a PKC-dependent manner, both in vitro and in mammalian cells (Ser10 and Ser18). Mutational analyses reveal that phosphorylation of Fbw7α at Ser10 can regulate its nuclear localization. Cancer-associated mutations in nearby residues (K11R and the addition of a proline residue at position 16) influence Fbw7α localization in a comparable manner, suggesting that mislocalization of this protein may be of pathological significance. Together these results provide evidence for both physical and functional interactions between the PKC and Fbw7 families, and yield insights into the isoform-specific regulation of Fbw7α.

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Neurotrophin Receptor TrkC Is an Entry Receptor for Trypanosoma cruzi in Neural, Glial, and Epithelial Cells

Infection and Immunity ◽

10.1128/iai.05403-11 ◽

2011 ◽

Vol 79 (10) ◽

pp. 4081-4087 ◽

Cited By ~ 16

Author(s):

Craig Weinkauf ◽

Ryan Salvador ◽

Mercio PereiraPerrin

Keyword(s):

Trypanosoma Cruzi ◽

Chagas Disease ◽

Mammalian Cells ◽

Chinese Hamster ◽

Neuronal Cell ◽

Host Cells ◽

Neurotrophin Receptor ◽

Content Type

ABSTRACTTrypanosoma cruzi, the agent of Chagas' disease, infects a variety of mammalian cells in a process that includes multiple cycles of intracellular division and differentiation starting with host receptor recognition by a parasite ligand(s). Earlier work in our laboratory showed that the neurotrophin-3 (NT-3) receptor TrkC is activated byT. cruzisurfacetrans-sialidase, also known as parasite-derived neurotrophic factor (PDNF). However, it has remained unclear whether TrkC is used byT. cruzito enter host cells. Here, we show that a neuronal cell line (PC12-NNR5) relatively resistant toT. cruzibecame highly susceptible to infection when overexpressing human TrkC but not human TrkB. Furthermore,trkCtransfection conferred an ∼3.0-fold intracellular growth advantage. Sialylation-deficient Chinese hamster ovarian (CHO) epithelial cell lines Lec1 and Lec2 also became much more permissive toT. cruziafter transfection with thetrkCgene. Additionally, NT-3 specifically blockedT. cruziinfection of the TrkC-NNR5 transfectants and of naturally permissive TrkC-bearing Schwann cells and astrocytes, as did recombinant PDNF. Two specific inhibitors of Trk autophosphorylation (K252a and AG879) and inhibitors of Trk-induced MAPK/Erk (U0126) and Akt kinase (LY294002) signaling, but not an inhibitor of insulin-like growth factor 1 receptor, abrogated TrkC-mediated cell invasion. Antibody to TrkC blockedT. cruziinfection of the TrkC-NNR5 transfectants and of cells that naturally express TrkC. The TrkC antibody also significantly and specifically reduced cutaneous infection in a mouse model of acute Chagas' disease. TrkC is ubiquitously expressed in the peripheral and central nervous systems, and in nonneural cells infected byT. cruzi, including cardiac and gastrointestinal muscle cells. Thus, TrkC is implicated as a functional PDNF receptor in cell entry, independently of sialic acid recognition, mediating broadT. cruziinfection bothin vitroandin vivo.

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Monocyte Infiltration and Differentiation in 3D Multicellular Spheroid Cancer Models

Pathogens ◽

10.3390/pathogens10080969 ◽

2021 ◽

Vol 10 (8) ◽

pp. 969

Author(s):

Natasha Helleberg Madsen ◽

Boye Schnack Nielsen ◽

Son Ly Nhat ◽

Søren Skov ◽

Monika Gad ◽

...

Keyword(s):

Cell Line ◽

Cancer Cell ◽

Cancer Cell Line ◽

Dependent Manner ◽

Multicellular Spheroid ◽

Tumor Associated Macrophages ◽

Cancer Models ◽

Ht 29 ◽

Mcf 7

Tumor-associated macrophages often correlate with tumor progression, and therapies targeting immune cells in tumors have emerged as promising treatments. To select effective therapies, we established an in vitro 3D multicellular spheroid model including cancer cells, fibroblasts, and monocytes. We analyzed monocyte infiltration and differentiation in spheroids generated from fibroblasts and either of the cancer cell lines MCF-7, HT-29, PANC-1, or MIA PaCa-2. Monocytes rapidly infiltrated spheroids and differentiated into mature macrophages with diverse phenotypes in a cancer cell line-dependent manner. MIA PaCa-2 spheroids polarized infiltrating monocytes to M2-like macrophages with high CD206 and CD14 expression, whereas monocytes polarized by MCF-7 spheroids displayed an M1-like phenotype. Monocytes in HT-29 and PANC-1 primarily obtained an M2-like phenotype but also showed upregulation of M1 markers. Analysis of the secretion of 43 soluble factors demonstrated that the cytokine profile between spheroid cultures differed considerably depending on the cancer cell line. Secretion of most of the cytokines increased upon the addition of monocytes resulting in a more inflammatory and pro-tumorigenic environment. These multicellular spheroids can be used to recapitulate the tumor microenvironment and the phenotype of tumor-associated macrophages in vitro and provide more realistic 3D cancer models allowing the in vitro screening of immunotherapeutic compounds.

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Epinephrine Stimulates Cell Proliferation and Induces Chemoresistance in Myeloma Cells through the β-Adrenoreceptor in vitro

Acta Haematologica ◽

10.1159/000478517 ◽

2017 ◽

Vol 138 (2) ◽

pp. 103-110 ◽

Cited By ~ 5

Author(s):

Yang Liu ◽

Xiaochen Yu ◽

Junling Zhuang

Keyword(s):

Cell Proliferation ◽

Cell Growth ◽

Cell Line ◽

Receptor Subtype ◽

Dependent Manner ◽

Myeloma Cells ◽

U266 Cell ◽

Β Receptor ◽

U266 Cell Line

Objectives: To explore the effect of the β-adrenoreceptor signaling pathway on myeloma cells. Methods: The myeloma U266 cell line was treated with epinephrine and propranolol. Cell proliferation was analyzed by MTS assay. Apoptosis was detected by flow cytometry. The β-receptor subtype and the key enzyme of epinephrine were identified by reverse transcription polymerase chain reaction (RT-PCR). Results: Epinephrine (5-50 μM) promoted U266 cell growth in a dose-dependent manner and neutralized the inhibition effect of bortezomib (25 and 50 ng/mL) in vitro. Cell proliferation was inhibited by a β-receptor antagonist, propranolol, at a concentration of 50-200 μM. The proportions of early and late apoptotic cells were enhanced after treatment with propranolol. The expression of caspase 3/7, 8, and 9 was elevated in propranolol-treated myeloma cells. Both β1- and β2-adrenoceptor mRNAs were expressed in the U266 cell line. Key enzymes dopamine hydroxylase and tyrosinehydroxylase were identified in myeloma cells. Conclusions: Our results reveal that epinephrine stimulates myeloma cell growth in vitro while the β-blocker propranolol has an antiproliferative effect, indicating that stress hormones may trigger the progression of myeloma.

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