Cell-associated IL-8 in human blood monocytes: analysis by flow cytometry

Area of Science:

• Immunology
• Cell Biology

Background:

• Cell-associated cytokines, like IL-1 and TNF, are biologically active on cell surfaces.
• Extracellular IL-8 is a key neutrophil chemoattractant, but cell-associated IL-8 remains understudied.

Purpose of the Study:

• To investigate the presence and localization of intracellular and cell-surface IL-8 in human blood monocytes.
• To predict the in vivo biological activity of cell-associated IL-8.

Main Methods:

• Flow cytometry was used to analyze intracellular and cell-surface IL-8 in monocytes.
• Mononuclear cells were fixed and either left untreated or permeabilized with saponin.
• Immunohistochemical analysis confirmed differential cytokine localization.

Main Results:

• Lipopolysaccharide (LPS) stimulation increased intracellular IL-8 in monocytes but did not result in detectable cell-surface IL-8.
• IL-1 beta was detected both intracellularly and on the cell surface of LPS-stimulated monocytes.
• ELISA and flow cytometry confirmed quantitative changes in intracellular IL-8 in an LPS-stimulated monocytic cell line.

Conclusions:

• LPS stimulation leads to a time-dependent increase in intracellular IL-8, but not cell-surface IL-8, in monocytes.
• Cell-associated IL-8 is unlikely to have a significant physiological function.
• Flow cytometry is a valuable tool for simultaneously examining cell-surface and intracellular cytokines.