Method for analysis of intracellular production of free radicals in vivo
Publish date: 2001-01-01
Report number: FOI-R--0074--SE
Written in: English
Inhaltion of toxic substances may give rise to an acute inflammatory response leading to recruitment of neutrophils into the lung. Upon activation, these cells start to produce reactive oxygen metabolites that are finally released into the surrounding tissue. Because of their microbicidal properties, these oxygen radicals play an important role in the host defence to microbes, but since their action is non-specific they could also be harmful to the host. Research on treatment of acute pulmonary inflammation is on-going at the Swedish Defence Research Agency, Division of NBC Defence. By using a method of intracellular staining with a fluorescent probe (dichlorodihydrofluorescensindiacetate) in vitro, the neutrophil production of reactive radicals can be detected. In order to improve the analysis of cell activation during inflammation, the aim of this study was to develop a method for intracellular staining of oxygen radicals in vivo. Inflammation in peritoneum or the lunf was triggered in mice through intraperitoneal injection or aerosol inhalation of the bacterial toxin lipopolysaccharide (LPS). To avoid dilution effects, the probe was administered directly into the target organ of inflammation either by intraperitoneal injection of by tracheal instillation. Cells from either locality was isolated and labelled with a specific antibody against neutrophils followed by analysis of oxidative activity by flow cytometry or fluorescence microscopy.