RESULTS
To
determine if Echinacea stimulates macrophages through MAP kinase activity, the
pathway was blocked using the PD98059
inhibitor. PD98059 specifically blocks the phosphorylation of ERK and prevents
it from becoming activated. When stimulated by Echinacea, macrophages produce
the cytokine, IL-6. The amount of IL-6 produced with and without the presence of
the inhibitor was measured using ELISA.
When
stimulated with Echinacea, macrophages produce increased levels of IL-6. The
positive control for cell stimulation was LPS + IFN, a well documented
macrophage activator, which produced the highest levels of IL-6. The negative
control, media alone, produced no IL-6. When cells had been pre-treated with the
inhibitor and subsequently stimulated with Echinacea IL-6 levels dropped in
relation to the concentration of inhibitor used. Increasing concentration of ERK
inhibitor resulted in decreased IL-6 production. This is evidence that the MAP kinase pathway is involved in
the activation of macrophages by Echinacea because when blocked there is a
significant decrease in cytokine production.
The IFN
control showed that IFN alone was not responsible for macrophage activation but
acts as a second signal to either LPS or Echinacea to enhance activation because
alone it did not stimulate IL-6 production. The DMSO control showed that the
DMSO concentrations used to resuspend the inhibitor was not toxic to cells as it
did not hinder IL-6 production. Therefore, showing DMSO alone does not inhibit
IL-6, which means that any decrease in IL-6 seen with the inhibitor is a result
of ERK inhibition versus cell death.