Journal article
Oxidant Generation by Single Infected Monocytes after Short-Term Fluorescence Labeling of a Protozoan Parasite
Infection and immunity, Vol.75(2), pp.1017-1024
02/2007
DOI: 10.1128/IAI.00914-06
PMCID: PMC1828521
PMID: 17118986
Abstract
Leishmania
spp. are intracellular protozoa residing in mononuclear phagocytes.
Leishmania
organisms are susceptible to microbicidal responses generated in response to phagocytosis. Assuming that both phagocyte and parasite populations are heterogeneous, it is advantageous to examine the response of individual cells phagocytosing living parasites. Because
Leishmania
spp. lose virulence during the raising of transfectants, we developed a method to label live
Leishmania chagasi
short-term with fluorescent dyes. Up to six parasite divisions were detected by flow cytometry after labeling with carboxyfluorescein diacetate succinimidyl ester (CFSE), dioctadecyl-tetramethylindo carbocyanine perchlorate, or chloromethyl tetramethylrhodamine. Labeled parasites entered mononuclear phagocytes as determined by confocal and time-lapse microscopy. Dihydroethidium (DHE) was used to detect macrophage-derived oxidants generated during phagocytosis. Presumably
Leishmania
organisms are opsonized with host serum/tissue components such as complement prior to phagocytosis. Therefore, we investigated the effects of opsonization and found that this increased the efficiency of CFSE-labeled parasite entry into monocytes (84.6% ± 8.8% versus 20.2% ± 3.8% monocytes infected;
P
< 0.001). Opsonization also increased the percentage of phagocytes undergoing a respiratory burst (66.0% ± 6.3% versus 41.0% ± 8.3% of monocytes containing CFSE-labeled parasites;
P
< 0.001) and the magnitude of oxidant generation by each infected monocyte. Inhibitor data indicated that DHE was oxidized by products of the NADPH oxidase. These data suggest that opsonized serum components such as complement lead to more efficient entry of
Leishmania
into their target cells but at the same time activate the phagocyte oxidase to generate microbicidal products in infected cells. The parasite must balance these positive and negative survival effects in order to initiate a viable infection.
Details
- Title: Subtitle
- Oxidant Generation by Single Infected Monocytes after Short-Term Fluorescence Labeling of a Protozoan Parasite
- Creators
- Haeok K Chang - Universidade Federal da Bahia, Salvador, BA, BrazilColin Thalhofer - Universidade Federal da Bahia, Salvador, BA, BrazilBreck A Duerkop - Universidade Federal da Bahia, Salvador, BA, BrazilJoanna S Mehling - Universidade Federal da Bahia, Salvador, BA, BrazilShilpi Verma - Universidade Federal da Bahia, Salvador, BA, BrazilKenneth J Gollob - Universidade Federal da Bahia, Salvador, BA, BrazilRoque Almeida - Universidade Federal da Bahia, Salvador, BA, BrazilMary E Wilson - Universidade Federal da Bahia, Salvador, BA, Brazil
- Resource Type
- Journal article
- Publication Details
- Infection and immunity, Vol.75(2), pp.1017-1024
- DOI
- 10.1128/IAI.00914-06
- PMID
- 17118986
- PMCID
- PMC1828521
- NLM abbreviation
- Infect Immun
- ISSN
- 0019-9567
- eISSN
- 1098-5522
- Publisher
- American Society for Microbiology
- Language
- English
- Date published
- 02/2007
- Academic Unit
- Microbiology and Immunology; International Programs; Epidemiology; Internal Medicine
- Record Identifier
- 9984001230902771
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