FLOW CYTOMETRIC DETECTION OF CELL-ASSOCIATED CYTOKINES IN ALVEOLAR MACROPHAGES

To elucidate the cytokine-producing capacity of alveolar macrophages ( AMs), we have introduced a method of flow cytometry combined with sapo nin treatment to detect the cell-associated cytokines. We studied bron choalveolar lavage fluid cells from six patients with sarcoidosis (SAR ) and six control (CTL) subjects, Cells were either left uncultured, o r cultured with and without lipopolysaccharide (LPS), then treated wit h paraformaldehyde and saponin and analysed for cell-associated interl eukin-1 beta (IL-1 beta) and tumour necrosis factor-alpha (TNF-alpha) by flow cytometry, Cell-associated IL-1 beta and TNF-alpha were also a nalysed by immunoassays. The flow cytometric cytokine values were corr elated with the immunoreactive cell-associated cytokines (IL-1 beta: r =0.45, p<0.05; TNF-alpha: r=0.82, p<0.001), The histograms of cell-ass ociated IL-1 beta yielded a single peak both in the patients and contr ols, whereas the histograms of cell-associated TNF-alpha exhibited two peaks in SAR patients, but just a single peak in the CTL subjects, Th e mean value of the cell-associated TNF-alpha in LPS(+) AMs was higher in the SAR patients than in the CTL subjects (p<0.001). In conclusion , the flow cytometric method can be applied to the semiquantitative de tection of cell-associated cytokines in alveolar macrophages at the si ngle cell level.