Quantification of Leishmania Parasites in Murine Models of Visceral Infection

Tavares, Joana; Cordeiro-da-Silva, Anabela

doi:10.1007/978-1-4939-9210-2_16

Cited by 6 publications

(7 citation statements)

References 35 publications

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“…After a ten minute incubation time to allow the distribution of the substrate in the body of the anesthetised animals, a ten-minute signal acquisition controlled by the Living Image software (Perkin Elmer) was initiated. At the end of this period, animals returned to their cage and recovered from the anaesthesia 33 .…”

Section: Methodsmentioning

confidence: 99%

“…Parasite loads were also determined by limiting dilution 26,33,34 . Briefly, organ homogenates (1 mg of spleen; 4 mg of liver) were subjected to two-fold serial dilutions in quadruplicate with Schneider’s Insect medium in 96-well microtitration plates.…”

Section: Methodsmentioning

confidence: 99%

“…The plates were incubated at 27 °C for 15 days and then each well was inspected for the presence or absence of promastigotes. The number of parasites per gram of organ was calculated as previously 26,33,34 .…”

Section: Methodsmentioning

confidence: 99%

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Murine infection with bioluminescent Leishmania infantum axenic amastigotes applied to drug discovery

Costa

Cecílio

Santarém

et al. 2019

Sci Rep

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Leishmaniasis is an important vector-borne neglected tropical disease caused by Leishmania parasites. Current anti-Leishmania chemotherapy is unsatisfactory, justifying the continued search for alternative treatment options. Herein, we demonstrate that luciferase-expressing Leishmania infantum axenic amastigotes, unlike promastigotes, are highly infectious to BALB/c mice and thus generate a robust bioluminescent signal in target organs, such as the liver and the spleen, as early as two weeks after infection. Treatment with the reference drugs amphotericin B and miltefosine was effective at reducing parasite burdens. This model allows the assessment of treatment efficacy using whole-mouse bioluminescence imaging without the need to wait several weeks for spleen infections to be detectable by this non-invasive method. In conclusion, we propose the use of this model in an initial approach to evaluate the treatment efficacy of promising chemical entities without having to sacrifice large numbers of animals or to wait several days for a readout.

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Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

See 1 more Smart Citation

Murine infection with bioluminescent Leishmania infantum axenic amastigotes applied to drug discovery

Costa

Cecílio

Santarém

et al. 2019

Sci Rep

Self Cite

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“…Results are indicated per mg of spleen and liver, or parasites per 1 × 10 6 BM cells. 65 Statistical Analysis. Statistical analyses between multiple groups were performed by using Kruskal−Wallis (nonparametric) followed by Dunn's multiple comparisons tests.…”

Section: ■ Methodsmentioning

confidence: 99%

Oral Efficacy of a Diselenide Compound Loaded in Nanostructured Lipid Carriers in a Murine Model of Visceral Leishmaniasis

Etxebeste-Mitxeltorena

Moreno

Carvalheiro

et al. 2021

ACS Infect. Dis.

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Leishmaniasis urgently needs new oral treatments, as it is one of the most important neglected tropical diseases that affects people with poor resources. The drug discovery pipeline for oral administration currently discards entities with poor aqueous solubility and permeability (class IV compounds in the Biopharmaceutical Classification System, BCS) such as the diselenide 2m , a trypanothione reductase (TR) inhibitor. This work was assisted by glyceryl palmitostearate and diethylene glycol monoethyl ether-based nanostructured lipid carriers (NLC) to render 2m bioavailable and effective after its oral administration. The loading of 2m in NLC drastically enhanced its intestinal permeability and provided plasmatic levels higher than its effective concentration (IC 50 ). In L. infantum- infected BALB/c mice, 2m -NLC reduced the parasite burden in the spleen, liver, and bone marrow by at least 95% after 5 doses, demonstrating similar efficacy as intravenous Fungizone. Overall, compound 2m and its formulation merit further investigation as an oral treatment for visceral leishmaniasis.

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“…Hence, an exact region of interest (ROI) with heterogenous morphologies can be identified which is not based on the assumption that nuclei are located in the center of the cells, to avoid inaccurate results working with primary cells and not with homogenous macrophage cell lines. In most HCS assays, the detection of Leishmania parasites is based on DNA staining [8][9][10] or transgenic parasites expressing fluorescent proteins [6,7,37,38]. The use of DNA staining can lead to inaccurate results as not only the parasitic nuclear DNA is stained, but also the mitochondrial kinetoplast DNA.…”

Section: Development Of a High Content Screening Assay For Cutaneous And Visceral Leishmania Speciesmentioning

confidence: 99%

High Content Analysis of Macrophage-Targeting EhPIb-Compounds against Cutaneous and Visceral Leishmania Species

et al. 2021

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An immunostimulatory glycolipid molecule from the intestinal protozoan parasite Entamoeba histolytica (Eh) and its synthetic analogs derived from its phosphatidylinositol-b-anchor (EhPIb) previously showed considerable immunotherapeutic effects against Leishmania major infection in vitro and in vivo. Here, we describe a high content screening assay, based on primary murine macrophages. Parasites detection is based on a 90 kDA heat shock protein-specific staining, enabling the detection of several Leishmania species. We validated the assay using L. major, L. braziliensis, L. donovani, and L. infantum as well as investigated the anti-leishmanial activity of six immunostimulatory EhPIb-compounds (Eh-1 to Eh-6). Macrophages infected with dermotropic species were more sensitive towards treatment with the compounds as their viability showed a stronger reduction compared to macrophages infected with viscerotropic species. Most compounds caused a significant reduction of the infection rates and the parasite burdens depending on the infecting species. Only compound Eh-6 was found to have activity against all Leishmania species. Considering the challenges in anti-leishmanial drug discovery, we developed a multi-species screening assay capable of utilizing non-recombinant parasite strains, and demonstrated its usefulness by screening macrophage-targeting EhPIb-compounds showing their potential for the treatment of cutaneous and visceral leishmaniasis.

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Quantification of Leishmania Parasites in Murine Models of Visceral Infection

Cited by 6 publications

References 35 publications

Murine infection with bioluminescent Leishmania infantum axenic amastigotes applied to drug discovery

Murine infection with bioluminescent Leishmania infantum axenic amastigotes applied to drug discovery

Oral Efficacy of a Diselenide Compound Loaded in Nanostructured Lipid Carriers in a Murine Model of Visceral Leishmaniasis

High Content Analysis of Macrophage-Targeting EhPIb-Compounds against Cutaneous and Visceral Leishmania Species

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