Shikonin reduces oedema induced by phorbol ester by interfering with I[kappa]B[alpha] degradation thus inhibiting translocation of NF-[kappa]B to the nucleus

Background and purpose: In the present paper we studied the effect of shikonin on ear oedema induced by 12-O-tetradecanoylphorbol-13-acetate (TPA), and determined the mechanisms through which shikonin might exert its topical anti-inflammatory action. Experimental approach: Acute ear oedema was induced in mice by topical application of TPA. The in vitro assays used macrophages RAW 264.7 cells stimulated with lipopolysaccharide. Cyclooxygenase-2, inducible nitric oxide synthase, protein kinase C[alpha], extracellular signal-regulated protein kinase (ERK), phosphorylated ERK (pERK), c-Jun N-terminal kinase (JNK), pJNK, p38, p-p38, p65, p-p65, inhibitor protein of nuclear factor-[kappa]B (NF-[kappa]B) (I[kappa]B[alpha]) and pI[kappa]B[alpha] were measured by Western blotting, activation and binding of NF-[kappa]B to DNA was detected by reporter gene and electrophoretic mobility shift assay, respectively, and NF-[kappa]B p65 localization was detected by immunocytochemistry. Key results: Shikonin reduced the oedema (inhibitory dose 50 = 1.0 mg per ear), the expression of cyclooxygenase-2 (70%) and of inducible nitric oxide synthase (100%) in vivo. It significantly decreased TPA-induced translocation of protein kinase C[alpha], the phosphorylation and activation of ERK, the nuclear translocation of NF-[kappa]B and the TPA-induced NF-[kappa]B-DNA-binding activity in mouse skin. Moreover, in RAW 264.7 cells, shikonin significantly inhibited the binding of NF-[kappa]B to DNA in a dose-dependent manner and the nuclear translocation of p65. Conclusions and implications: Shikonin exerted its topical anti-inflammatory action by interfering with the degradation of I[kappa]B[alpha], thus inhibiting the activation of NF-[kappa]B. [PUBLICATION ABSTRACT]