scholarly journals A Novel Strategy for Enhance Potentiation of Meglumine antimo-niate against Leishmania major In Vitro

2019 ◽  
Author(s):  
Farzaneh MIRZAEI ◽  
Hossein KHANAHMAD ◽  
Fatemeh NAMDAR ◽  
Shahrokh IZADI ◽  
Seyed Hossein HEJAZI
Keyword(s):  
Leishmania Major ◽  
Host Cells ◽  
Leishmania Infection ◽  
Listeriolysin O ◽  
Meglumine Antimoniate ◽  
Dose Concentration ◽  
Vitro Model ◽  
Novel Strategy ◽  
Hlya Gene

Background: We aimed to design a different method of drug delivery for increased transfer of the choice drug (meglumine antimoniate) within the host cells. Therefore, listeriolysin O (LLO), a bacterial product which is a member of pore-forming peptides was used as an enhancer factor with meglumine antimoniate in order to facilitate the transition of the drug across macrophage membrane. Methods: LLO was produced in Isfahan University of Medical Sciences in 2016, by expressing the hlyA gene in Escherichia coli and purified using affinity chromatography. Cytotoxicity of the purified protein was investigated in an in vitro model of macrophage Leishmania infection. Results: LLO was cytotoxic against murine macrophage cells (J774-A1) and amastigote forms of L. major (MRHO/IR/75/ER). It was less toxic to macrophages (CC50=2.56 μg ml-1 ±0.09) than to the parasites (IC50=1.72 μg ml-1 ±0.07). Moreover, non-cytotoxic concentration of LLO (0.006 ug ml-1) potentiated the cytotoxicity induced by low dose concentration of meglumine antimoniate. Very little dose of meglumine antimoniate was needed when combined with the LLO (IC50=12.63 μg ml-1 ±0.13) in comparison with the cytotoxicity induced when the drug is used alone (IC50=46.17 μg ml-1 ±0.28). Conclusion: The combination of pore-forming proteins with anti-leishmanial agents could increase the advantage of anti-leishmanial drugs. Since lower concentrations of anti-leishmanial drugs can reduce undesirable side effects of chemotherapy trials carried out in animal models and then in humans with this system.

scholarly journals Exposure of ichnovirus particles to digitonin leads to enhanced infectivity and induces fusion from without in an in vitro model system

2007 ◽  
Vol 88 (11) ◽  
pp. 2977-2984 ◽  
Author(s):  
Don Stoltz ◽  
Renée Lapointe ◽  
Andrea Makkay ◽  
Michel Cusson
Keyword(s):  
Outer Membrane ◽  
In Vitro Model ◽  
Model System ◽  
Host Cells ◽  
Fusion Event ◽  
Susceptible Host ◽  
In Vitro Model System ◽  
Vitro Model

Unlike most viruses, the mature ichnovirus particle possesses two unit membrane envelopes. Following loss of the outer membrane in vivo, nucleocapsids are believed to gain entry into the cytosol via a membrane fusion event involving the inner membrane and the plasma membrane of susceptible host cells; accordingly, experimentally induced damage to the outer membrane might be expected to increase infectivity. Here, in an attempt to develop an in vitro model system for studying ichnovirus infection, we show that digitonin-induced disruption of the virion outer membrane not only increases infectivity, but also uncovers an activity not previously associated with any polydnavirus: fusion from without.

scholarly journals Hydrogen Peroxide-Preconditioned Human Adipose-Derived Stem Cells Enhance the Recovery of Oligodendrocyte-Like Cells after Oxidative Stress-Induced Damage

2020 ◽  
Vol 21 (24) ◽  
pp. 9513
Author(s):  
Patricia Garrido-Pascual ◽  
Ana Alonso-Varona ◽  
Begoña Castro ◽  
María Burón ◽  
Teodoro Palomares
Keyword(s):  
Oxidative Stress ◽  
Stem Cells ◽  
Therapeutic Potential ◽  
Mild Stress ◽  
Adipose Derived Stem Cells ◽  
Vitro Model ◽  
Novel Strategy ◽  
The One ◽  
Cell Conditioned Medium

Oxidative stress associated with neuroinflammation is a key process involved in the pathophysiology of neurodegenerative diseases, and therefore, has been proposed as a crucial target for new therapies. Recently, the therapeutic potential of human adipose-derived stem cells (hASCs) has been investigated as a novel strategy for neuroprotection. These cells can be preconditioned by exposing them to mild stress in order to improve their response to oxidative stress. In this study, we evaluate the therapeutic potential of hASCs preconditioned with low doses of H2O2 (called HC016 cells) to overcome the deleterious effect of oxidative stress in an in vitro model of oligodendrocyte-like cells (HOGd), through two strategies: i, the culture of oxidized HOGd with HC016 cell-conditioned medium (CM), and ii, the indirect co-culture of oxidized HOGd with HC016 cells, which had or had not been exposed to oxidative stress. The results demonstrated that both strategies had reparative effects, oxidized HC016 cell co-culture being the one associated with the greatest recovery of the damaged HOGd, increasing their viability, reducing their intracellular reactive oxygen species levels and promoting their antioxidant capacity. Taken together, these findings support the view that HC016 cells, given their reparative capacity, might be considered an important breakthrough in cell-based therapies.

scholarly journals Modelling Bovine Granuloma Formation In Vitro upon Infection with Mycobacterium Avium Subspecies Paratuberculosis

2019 ◽  
Vol 6 (4) ◽  
pp. 80 ◽  
Author(s):  
J. Hunter Rice ◽  
Margaret M. McDaniel ◽  
Alyson Holland ◽  
Shigetoshi Eda
Keyword(s):  
Mycobacterium Avium ◽  
Mycobacterium Avium Subspecies Paratuberculosis ◽  
Clinical Signs ◽  
Host Cells ◽  
Economic Losses ◽  
Multiple Parameters ◽  
Cell Mediated Immune Response ◽  
A Cell ◽  
Vitro Model

Mycobacterium avium subspecies paratuberculosis (Map) causes chronic granulomatous disease in cattle and ruminant livestock, causing substantial economic losses. Current vaccines delay clinical signs but cannot train the immune system to fully eradicate latent Map. During latency, Map uses host defenses, cage-like macrophage clusters called granuloma, as incubators for months or years. We used an in vitro model to investigate the early coordination of macrophages into granuloma upon Map infection over ten days. We found that at multiplicities of infection (MOI; Map:macrophages) of 1:2 and below, the macrophages readily form clusters and evolve pro-inflammatory cytokines in keeping with a cell-mediated immune response. At higher MOIs, viability of host macrophages is negatively impacted. At 1:4 MOI, we quantified viable Map in our model and confirmed that intracellular Map reproduced over the first five days of infection. Host cells expressed Type 1-specific cytokines, and Map-infected macrophages displayed reduced motility compared to Map-exposed, uninfected macrophages, suggesting an important role for uninfected macrophages in the early aggregative response. Reported is the first in vitro JD granuloma model capturing Map and macrophage viability, size distribution of resulting clusters, motility of monocyte-derived macrophages, and cytokine response during clustering, allowing quantitative analysis of multiple parameters of the Map-specific granulomatous response.

scholarly journals Fibroblasts as Host Cells in Latent Leishmaniosis

2000 ◽  
Vol 191 (12) ◽  
pp. 2121-2130 ◽  
Author(s):  
Christian Bogdan ◽  
Norbert Donhauser ◽  
Reinhard Döring ◽  
Martin Röllinghoff ◽  
Andreas Diefenbach ◽  
...  
Keyword(s):  
Lymph Nodes ◽  
Leishmania Major ◽  
Chronic Phase ◽  
Host Cells ◽  
Confocal Laser ◽  
Intracellular Parasites ◽  
Important Host ◽  
Mechanisms Of Persistence ◽  
Activated Fibroblasts

Intracellular parasites are known to persist lifelong in mammalian hosts after the clinical cure of the disease, but the mechanisms of persistence are poorly understood. Here, we show by confocal laser microscopy that in the draining lymph nodes of mice that had healed a cutaneous infection with Leishmania major, 40% of the persisting parasites were associated with fibroblasts forming the reticular meshwork of the lymph nodes. In vitro, both promastigotes and amastigotes of L. major infected primary skin or lymph node fibroblasts. Compared with macrophages, cytokine-activated fibroblasts had a reduced ability to express type 2 nitric oxide synthase and to kill intracellular L. major. These data identify fibroblasts as an important host cell for Leishmania during the chronic phase of infection and suggest that they might serve as safe targets for the parasites in clinically latent disease.

scholarly journals Intracellular Survival of Leishmania major in Neutrophil Granulocytes after Uptake in the Absence of Heat-Labile Serum Factors

2002 ◽  
Vol 70 (2) ◽  
pp. 826-835 ◽  
Author(s):  
Helmut Laufs ◽  
Kerstin Müller ◽  
Jens Fleischer ◽  
Norbert Reiling ◽  
Nicole Jahnke ◽  
...  
Keyword(s):  
Leishmania Major ◽  
Intracellular Survival ◽  
Host Cells ◽  
Polymorphonuclear Neutrophil ◽  
Heat Inactivation ◽  
Neutrophil Granulocytes ◽  
Complement Components ◽  
Serum Factors

ABSTRACT The role of polymorphonuclear neutrophil granulocytes (PMN) in defense against the intracellular parasite Leishmania is poorly understood. In the present study, the interaction of human PMN with Leishmania major promastigotes was investigated in vitro. In the presence of fresh human serum, about 50% of PMN phagocytosed the parasites within 10 min and the parasite uptake led to PMN activation, resulting in the killing of most ingested parasites. Heat inactivation of the serum markedly reduced the rate of early parasite phagocytosis, suggesting a role of complement components in the early uptake of Leishmania. However, over 50% of PMN were able to ingest parasites in the presence of heat-inactivated serum if the coincubation was extended to 3 h. After 3 h, 10% of the PMN were found to internalize Leishmania even under serum-free conditions. These findings indicate that PMN possess mechanisms for both opsonin/complement-dependent and -independent uptake of Leishmania. Both pathways of uptake could be partially blocked by anti-CR3 antibody. Mannan-binding lectin was found not to be involved in this process. When phagocytosed in the absence of opsonin, the majority of Leishmania parasites survived intracellularly in PMN for at least 1 day. These data suggest a dual role of PMN in the early response to L. major infection. On the one hand, PMN can rapidly eliminate the intracellular parasites, and on the other hand, Leishmania can survive intracellularly in PMN. These data, together with the finding that intact parasites were seen in PMN isolated from the skin of infected mice, suggest that PMN can serve as host cells for the intracellular survival of Leishmania within the first hours or days after infection.

scholarly journals Coiling Phagocytosis of Trypanosomatids and Fungal Cells

1998 ◽  
Vol 66 (9) ◽  
pp. 4331-4339 ◽  
Author(s):  
M. G. Rittig ◽  
K. Schröppel ◽  
K.-H. Seack ◽  
U. Sander ◽  
E.-N. N’Diaye ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Saccharomyces Cerevisiae ◽  
Legionella Pneumophila ◽  
Leishmania Major ◽  
Posterior Pole ◽  
Host Cells ◽  
Human Monocytes ◽  
Eukaryotic Microorganisms ◽  
Microscopic Studies

ABSTRACT Coiling phagocytosis has previously been studied only with the bacteria Legionella pneumophila and Borrelia burgdorferi, and the results were inconsistent. To learn more about this unconventional phagocytic mechanism, the uptake of various eukaryotic microorganisms by human monocytes, murine macrophages, and murine dendritic cells was investigated in vitro by video and electron microscopy. Unconventional phagocytosis of Leishmania spp. promastigotes, Trypanosoma cruzi trypomastigotes,Candida albicans hyphae, and zymosan particles fromSaccharomyces cerevisiae differed in (i) morphology (rotating unilateral pseudopods with the trypanosomatids, overlapping bilateral pseudopods with the fungi), (ii) frequency (high withLeishmania; occasional with the fungi; rare with T. cruzi), (iii) duration (rapid with zymosan; moderate with the trypanosomatids; slow with C. albicans), (iv) localization along the promastigotes (flagellum of Leishmania major andL. aethiopica; flagellum or posterior pole of L. donovani), and (v) dependence on complement (strong with L. major and L. donovani; moderate with the fungi; none with L. aethiopica). All of these various types of unconventional phagocytosis gave rise to similar pseudopod stacks which eventually transformed to a regular phagosome. Further video microscopic studies with L. major provided evidence for a cytosolic localization, synchronized replication, and exocytic release of the parasites, extending traditional concepts about leishmanial infection of host cells. It is concluded that coiling phagocytosis comprises phenotypically similar consequences of various disturbances in conventional phagocytosis rather than representing a single separate mechanism.

scholarly journals Inhibition of ABC Transporters Abolishes Antimony Resistance in Leishmania Infection

2007 ◽  
Vol 52 (3) ◽  
pp. 1080-1093 ◽  
Author(s):  
Jayati Mookerjee Basu ◽  
Ananda Mookerjee ◽  
Rajdeep Banerjee ◽  
Manik Saha ◽  
Subhankar Singh ◽  
...  
Keyword(s):  
Abc Transporters ◽  
Leishmania Donovani ◽  
Host Cells ◽  
Leishmania Infection ◽  
Blood Monocytes ◽  
Glycoprotein P ◽  
Kala Azar ◽  
Antimony Resistance ◽  
Parasite Replication

ABSTRACT The emergence of antimony (Sb) resistance has jeopardized the treatment of visceral leishmaniasis in various countries. Previous studies have considered the part played by leishmanial parasites in antimony resistance, but the involvement of host factors in the clinical scenario remained to be investigated. Here we show that unlike infection with Sb-sensitive (Sbs) Leishmania donovani, infection with Sb-resistant (Sbr) L. donovani induces the upregulation of multidrug resistance-associated protein 1 (MRP1) and permeability glycoprotein (P-gp) in host cells, resulting in a nonaccumulation of intracellular Sb following treatment with sodium antimony gluconate (SAG) favoring parasite replication. The inhibition of MRP1 and P-gp with resistance-modifying agents such as lovastatin allows Sb accumulation and parasite killing within macrophages and offers protection in an animal model in which infection with Sbr L. donovani is otherwise lethal. The occurrence of a similar scenario in clinical cases is supported by the findings that unlike monocytes from SAG-sensitive kala-azar (KA) patients, monocytes from SAG-unresponsive KA patients overexpress P-gp and MRP1 and fail to accumulate Sb following in vitro SAG treatment unless pretreated with inhibitors of ABC transporters. Thus, the expression status of MRP1 and P-gp in blood monocytes may be used as a diagnostic marker for Sb resistance and the treatment strategy can be designed accordingly. Our results also indicate that lovastatin, which can inhibit both P-gp and MRP1, might be beneficial for reverting Sb resistance in leishmaniasis as well as drug resistance in other clinical situations, including cancer.

scholarly journals Development of a New Antileishmanial Aziridine-2,3-Dicarboxylate-Based Inhibitor with High Selectivity for Parasite Cysteine Proteases

2015 ◽  
Vol 60 (2) ◽  
pp. 797-805 ◽  
Author(s):  
Caroline Schad ◽  
Ulrike Baum ◽  
Benjamin Frank ◽  
Uwe Dietzel ◽  
Felix Mattern ◽  
...  
Keyword(s):  
Leishmania Major ◽  
Neglected Tropical Diseases ◽  
Cathepsin L ◽  
Selective Inhibition ◽  
Cysteine Proteases ◽  
Host Cells ◽  
Th2 Response ◽  
Content Type

ABSTRACTLeishmaniasis is one of the major neglected tropical diseases of the world. Druggable targets are the parasite cysteine proteases (CPs) of clan CA, family C1 (CAC1). In previous studies, we identified two peptidomimetic compounds, the aziridine-2,3-dicarboxylate compounds 13b and 13e, in a series of inhibitors of the cathepsin L (CL) subfamily of the papain clan CAC1. Both displayed antileishmanial activityin vitrowhile not showing cytotoxicity against host cells. In further investigations, the mode of action was characterized inLeishmania major. It was demonstrated that aziridines 13b and 13e mainly inhibited the parasitic cathepsin B (CB)-like CPC enzyme and, additionally, mammalian CL. Although these compounds induced cell death ofLeishmaniapromastigotes and amastigotesin vitro, the induction of a proleishmanial T helper type 2 (Th2) response caused by host CL inhibition was observedin vivo. Therefore, we describe here the synthesis of a new library of more selective peptidomimetic aziridine-2,3-dicarboxylates discriminating between host and parasite CPs. The new compounds are based on 13b and 13e as lead structures. One of the most promising compounds of this series is compound s9, showing selective inhibition of the parasite CPsLmaCatB (a CB-like enzyme ofL. major; also namedL. majorCPC) andLmCPB2.8 (a CL-like enzyme ofLeishmania mexicana) while not affecting mammalian CL and CB. It displayed excellent leishmanicidal activities againstL. majorpromastigotes (50% inhibitory concentration [IC50] = 37.4 μM) and amastigotes (IC50= 2.3 μM). In summary, we demonstrate a new selective aziridine-2,3-dicarboxylate, compound s9, which might be a good candidate for futurein vivostudies.

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