Generation of stable L. major+EGFP-LUC and simultaneous comparison between EGFP and luciferase sensitivity

Taheri, Tahereh; Nik, Hana Saberi; Seyed, Negar; Doustdari, Fatemeh; Etemadzadeh, Mohammad-Hossein; Torkashvand, Fatemeh; Rafati, Sima

doi:10.1016/j.exppara.2015.01.008

Cited by 20 publications

(27 citation statements)

References 31 publications

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“…Pixel counting and measurement of the lesions were performed using KODAK molecular image software version 5.3. Results were reported as "net intensity", a quantitative measurement defined as the number of green pixels in a given area (region of interest) multiplied by the average intensity of each pixel [22,23]. This experiment was performed 7 weeks post challenge since earlier imaging barely discriminates fluorescent signal difference among groups.…”

Section: In Situ Imaging Of Anesthetized Mice For Egfp Fluorescencementioning

confidence: 99%

Assessment of Protection Induced by DNA and Live Vaccine Encoding Leishmania MHC Class I Restricted Epitopes against L. major Challenge in Balb/c Mice Model

Zandieh¹,

Kashi²,

Taheri³

2015

J Microb Biochem Technol

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Section: In Situ Imaging Of Anesthetized Mice For Egfp Fluorescencementioning

confidence: 99%

Assessment of Protection Induced by DNA and Live Vaccine Encoding Leishmania MHC Class I Restricted Epitopes against L. major Challenge in Balb/c Mice Model

Zandieh¹,

Kashi²,

Taheri³

2015

J Microb Biochem Technol

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“…The isolated splenocytes (~3 × 10 6 cell/mL) were stimulated in vitro with F/T antigen (20 μg/mL of L. tropica or 10 μg/mL of L. major ) for 5 days at 37°C in 5% CO 2 as previously described . The supernatants were mixed with equal volume of Griess reagent in duplicate and were incubated for 10 minutes at room temperature.…”

Section: Methodsmentioning

confidence: 99%

“…for 5 days at 37°C in 5% CO 2 as previously described. 13 The supernatants were mixed with equal volume of Griess reagent in duplicate and were incubated for 10 minutes at room temperature.…”

Section: Stimulation Of Splenocytes and Nitric Oxide (No) Assaymentioning

confidence: 99%

“…To evaluate the in vivo infection in mice, six mice from each nor-NOHA-Ltrop treated (G1) and untreated PBS-Ltrop (G2) were randomly selected for bioluminescence as previously described. 13,19 All groups were given D-luciferin potassium salt (Caliper Life Science, dissolved in calcium and magnesium-free PBS at 15 mg/mL concentration) by intraperitoneal injection then were anesthetizated using intraperitoneal ketamine (10%)-xylazine (2%). Monitoring was conducted by KODAK imaging system (In Vivo Imaging system FX Pro), and the images were taken with two different modes at different exposure times, luciferase (~20 minutes), and white (1 seconds).…”

Section: In Vivo Imaging Through Bioluminescencementioning

confidence: 99%

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The outcome of arginase activity inhibition in BALB/c mice hosting Leishmania tropica

Nahidi

Gholami

Taslimi

et al. 2020

Parasite Immunology

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Two species of Leishmania (L), L. tropica and L. major, are among the main causative agents of cutaneous leishmaniasis. Arginase (ARG) is an essential enzyme for cell growth, thus an attractive drug target. In this study, we tried to survey the inhibitory impact of ARG by nor‐NOHA (N‐ω‐hydroxy‐L‐nor‐arginine) on in vivo infection caused by L. tropica. BALB/c mice were inoculated with L. tropicaEGFP‐LUC (Ltrop) or L. majorEGFP‐LUC (Lmj) and then were treated by nor‐NOHA. ARG inhibitor only indicated a delay in generation of a cutaneous lesion in inoculated footpad with nor‐NOHA‐Ltrop and nor‐NOHA‐Lmj. ARG activity has been significantly reduced in nor‐NOHA‐Ltrop group. In this group, ARG activity inhibition correlated with increased levels of nitric oxide (NO). In both inoculated mice with Ltrop or Lmj, parasite load showed a significant decrease at later steps during the CL course post‐treatment. In vivo bioluminescence intensity did not show any ARG's inhibitory effect on treated‐Ltrop. The findings verified that the ARG activity may partially control the L. tropica infection in BALB/c mice through reduction of parasite proliferation and parasite killing through NO generation. This effect is dose‐dependent.

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“…Several probes, such as the green fluorescent protein (GFP), enhanced green fluorescent protein (EGFP), mCherry red fluorescent protein, and near-infrared fluorescent proteins, as well as the firefly luciferase (LUC) reporter gene, have been stably integrated into the parasite genome and have been widely used to monitor the in vivo intracellular proliferation of Leishmania parasites (12–24). Taheri et al have reported utilizing two reporter proteins, EGFP and LUC, to quantify the parasite load and increase the in vivo experimental sensitivity (25). Published work has shown a strong correlation between the parasite load and luciferase activity or fluorescence emission, which makes the use of transgenic Leishmania parasites a useful tool to monitor disease progression and the efficacy of antileishmanial drugs in animal models (14, 18, 20, 22).…”

Section: Introductionmentioning

confidence: 99%

Use of Optical Imaging Technology in the Validation of a New, Rapid, Cost-Effective Drug Screen as Part of a TieredIn VivoScreening Paradigm for Development of Drugs To Treat Cutaneous Leishmaniasis

Caridha

Parriot

Hudson

et al. 2017

Antimicrob Agents Chemother

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In any drug discovery and development effort, a reduction in the time of the lead optimization cycle is critical to decrease the time to license and reduce costs. In addition, ethical guidelines call for the more ethical use of animals to minimize the number of animals used and decrease their suffering. Therefore, any effort to develop drugs to treat cutaneous leishmaniasis requires multiple tiers of in vivo testing that start with higher-throughput efficacy assessments and progress to lower-throughput models with the most clinical relevance. Here, we describe the validation of a high-throughput, first-tier, noninvasive model of lesion suppression that uses an in vivo optical imaging technology for the initial screening of compounds. A strong correlation between luciferase activity and the parasite load at up to 18 days postinfection was found. This correlation allows the direct assessment of the effects of drug treatment on parasite burden. We demonstrate that there is a strong correlation between drug efficacy measured on day 18 postinfection and the suppression of lesion size by day 60 postinfection, which allows us to reach an accurate conclusion on drug efficacy in only 18 days. Compounds demonstrating a significant reduction in the bioluminescence signal compared to that in control animals can be tested in lower-throughput, more definitive tests of lesion cure in BALB/c mice and Golden Syrian hamsters (GSH) using Old World and New World parasites.

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Generation of stable L. major+EGFP-LUC and simultaneous comparison between EGFP and luciferase sensitivity

Cited by 20 publications

References 31 publications

Assessment of Protection Induced by DNA and Live Vaccine Encoding Leishmania MHC Class I Restricted Epitopes against L. major Challenge in Balb/c Mice Model

Assessment of Protection Induced by DNA and Live Vaccine Encoding Leishmania MHC Class I Restricted Epitopes against L. major Challenge in Balb/c Mice Model

The outcome of arginase activity inhibition in BALB/c mice hosting Leishmania tropica

Use of Optical Imaging Technology in the Validation of a New, Rapid, Cost-Effective Drug Screen as Part of a TieredIn VivoScreening Paradigm for Development of Drugs To Treat Cutaneous Leishmaniasis

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