Cryptotanshinone inhibits chemotactic migration in macrophages through negative regulation of the PI3K signaling pathway

Background and purpose: Cryptotanshinone, the major tanshinone isolated from Salvia miltiorrhiza Bunge, exhibits antiinflammatory activity. However, there is no report on the effect of cryptotanshinone on recruitment of leukocytes to inflammatory sites. We therefore assessed the effects of cryptotanshinone on macrophage chemotaxis. Experimental approach: Macrophage migration induced by complement 5a (C5a) or macrophage inflammatory protein-1 alpha (MIP-1 alpha) was measured in vitro. Intracellular kinase translocation and phosphorylation was assessed by Western blotting. Key results: RAW264.7 cell migration towards C5a (1 mu gml-1 alpha) was significantly inhibited by cryptotanshinone (1, 3, 10 and 30 mu M) in a concentration- dependent manner. Primary human macrophages stimulated by C5a were similarly inhibited. C5a-evoked migration in RAW264.7 cells was significantly suppressed by wortmannin (phosphatidylinositol 3-kinase (PI3K) inhibitor), PD98059 (MEK1/2 inhibitor) and SB203580 (p38 mitogen- activated protein kinase (MAPK) inhibitor), but not by SP600125 (c-Jun N-terminal kinase (JNK) inhibitor), suggesting that activation of PI3K, ERK1/2 and p38 MAPK signal pathways was involved in responses to C5a. Western blotting revealed that cryptotanshinone significantly inhibited PI3K-p110 gamma membrane translocation and phosphorylation of Akt (PI3K downstream effector protein) and ERK1/2 induced by C5a. However, neither p38 MAPK nor JNK phosphorylation was affected by cryptotanshinone. Wortmannin significantly attenuated C5a-induced PI3K-p110 gamma translocation, Akt and ERK1/2 phosphorylation. PD98059 suppressed ERK1/2 phosphorylation but failed to modify PI3K-p110 gamma translocation by C5a stimulation. Furthermore, MIP-1 alpha-induced cell migration and PI3K-p110 gamma translocation were also inhibited by cryptotanshinone in a concentration-dependent manner. Conclusions and implications: Inhibition of macrophage migration by cryptotanshinone involved inhibition of PI3K activation with consequent reduction of phosphorylation of Akt and ERK1/2.