Andrographolide interferes with binding of nuclear factor-kappaB to DNA in HL-60-derived neutrophilic cells

1. Andrographolide, the major active component from Andrographis paniculata, has shown to possess anti-inflammatory activity. Andrographolide inhibits the expression of several proinflammatory proteins that exhibit a nuclear factor kappa B (NF-kappaB) binding site in their gene. 2. In the present study, we analyzed the effect of andrographolide on the activation of NF-kappaB induced by platelet-activating factor (PAF) and N-formyl-methionyl-leucyl-phenylalanine (fMLP) in HL-60 cells differentiated to neutrophils. 3. PAF (100 nM) and fMLP (100 nM) induced activation of NF-kappaB as determined by degradation of inhibitory factor B alpha (IkappaB alpha) using Western blotting in cytosolic extracts and by binding to DNA using electrophoretic mobility shift assay (EMSA) in nuclear extracts. 4. Andrographolide (5 and 50 microM) inhibited the NF-kappaB-luciferase activity induced by PAF. However, andrographolide did not reduce phosphorylation of p38 MAPK or ERK1/2 and did not change IkappaB alpha degradation induced by PAF and fMLP. 5. Andrographolide reduced the DNA binding of NF-kappaB in whole cells and in nuclear extracts induced by PAF and fMLP. 6. Andrographolide reduced cyclooxygenase-2 (COX-2) expression induced by PAF and fMLP in HL-60/neutrophils. 7. It is concluded that andrographolide exerts its anti-inflammatory effects by inhibiting NF-kappaB binding to DNA, and thus reducing the expression of proinflammatory proteins, such as COX-2.