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Monitoring of Intracellular Nitric Oxide in Leishmaniasis: Its Applicability in Patients with Visceral Leishmaniasis

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Title Monitoring of Intracellular Nitric Oxide in
Leishmaniasis: Its Applicability in Patients
with Visceral Leishmaniasis
 
Creator Sarkar, Avijit
Saha, Piu
Mandal, Goutam
Mukhopadhyay, Debanjan
Roy, Susmita
Singh, Sunny Kumar
Das, Sukhen
Goswami, R P
Saha, Bibhuti
Kumar, Deepak
Das, Padma
Chatterjee, Mitali
 
Subject Cell Biology & Physiology
 
Description Nitric oxide (NO) has been demonstrated to be a principal effector molecule responsible
for mediating intracellular killing of Leishmania parasites, the causative organism
of leishmaniasis. As measurement of intracellular NO remains a challenge to
biologists, we have developed a flow cytometric approach to perform real time biological
detection of NO within Leishmania parasites and parasitized macrophages
using a membrane permeable derivative of diaminofluorescein [4,5-diaminofluorescein
diacetate (DAF-2DA)]. Initially, assay optimization was performed in Leishmania
donovani promastigotes, assay specificity being confirmed using both a NO donor
[S-nitroso-N-acetyl-penicillamine (SNAP)] and a NO scavenger [2-(4-carboxyphenyl)-
4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide, C-PTIO]. Using 40 lM DAF-
2DA, basal levels of intracellular NO were measured which varied in different Leishmania
species; addition of conventional anti-leishmanial drugs, antimony and miltefosine
translated into a dramatic increase in DAF-2T fluorescence. Furthermore, the
assay also measured levels of NO in macrophages, but needed a 20 fold lower concentration
of DAF-2DA, being 2 lM. Following parasitization, levels of NO decreased
which was normalized following treatment with anti-leishmanial drugs. Similarly
monocytes of patients with visceral leishmaniasis at disease presentation showed
decreased levels of NO which too reverted on completion of treatment. Taken together,
this study opens new perspectives of research regarding monocyte function
and provides a real time approach for monitoring the effect of anti-leishmanial compounds
 
Publisher John Wiley & Sons
 
Date 2011
 
Type Article
PeerReviewed
 
Format application/pdf
 
Identifier http://www.eprints.iicb.res.in/1249/1/CYTOMETRY_PART_A__Volume_79A_(1)_35%2D45;2011[147].pdf
Sarkar, Avijit and Saha, Piu and Mandal, Goutam and Mukhopadhyay, Debanjan and Roy, Susmita and Singh, Sunny Kumar and Das, Sukhen and Goswami, R P and Saha, Bibhuti and Kumar, Deepak and Das, Padma and Chatterjee, Mitali (2011) Monitoring of Intracellular Nitric Oxide in Leishmaniasis: Its Applicability in Patients with Visceral Leishmaniasis. Cytometry Part A, 79A (1). pp. 35-45. ISSN 1552-4922
 
Relation http://dx.doi.org/10.1002/cyto.a.21001
http://www.eprints.iicb.res.in/1249/