Elsevier

Life Sciences

Volume 81, Issue 2, 20 June 2007, Pages 109-114
Life Sciences

Cryptotanshinone inhibits macrophage migration by impeding F-actin polymerization and filopodia extension

https://doi.org/10.1016/j.lfs.2007.04.028Get rights and content

Abstract

We evaluated the anti-inflammatory effects of cryptotanshinone and tanshinone IIA, two major tanshinones isolated from Salvia miltiorrhiza, on chemoattractant-induced cell migration in RAW264.7 macrophages. Results showed that cryptotanshinone inhibited cell migration toward complement 5a (C5a) and macrophage inflammatory protein-1α (MIP-1α) in a concentration-dependent manner. In contrast, tanshinone IIA displayed less or even no effect on cell migration evoked by these chemoattractants. Both C5a- and MIP-1α-induced migration were clearly inhibited by cytochalasin B (an inhibitor of actin polymerization), but not by colchicine (an inhibitor of microtubule polymerization). Fluorescence staining demonstrated that cryptotanshinone as well as cytochalasin B, effectively reversed cell polarization and filopodia extension induced by both chemoattractants. Furthermore, C5a-evoked increase in F-actin fluorescence intensity was significantly suppressed by cryptotanshinone. Based on these observations, we suggest that cryptotanshinone exerts anti-migrating activity possibly by impeding F-actin polymerization and filopodia formation.

Introduction

Dried roots of Salvia miltiorrhiza Bunge (Danshen) have been used in traditional Chinese medicine for the treatment of several pathologies including coronary heart disease, hepatitis, and chronic renal failure (Lu and Foo, 2002). Cryptotanshinone and tanshinone IIA (Fig. 1) are two major tanshinones in this plant. Tanshinone showed a variety of biological activities including anti-inflammation (Kim et al., 2002) and cytotoxicity against human tumor cell lines (Lin and Chang, 2000, Ryu et al., 1997, Yuan et al., 2003). One of the therapeutic objectives in inflammation is to reduce inflammatory cell infiltration. The inappropriate recruitment of leukocytes from blood to the site of infection is an essential process of inflammation and can result in tissue destruction. A variety of products present at the site of inflammation can act as chemotactic agents, including formylmethionyl peptides, platelet activating factor, leukotriene B4, and complement products. Tanshinone IIA was reported to show inhibitory actions on leukocyte chemotactic migration (Gao, 1985, Zhou et al., 1997). Cryptotanshinone was previously observed to possess the most powerful antibacterial activity among tanshinones (Lee et al., 1999) and counteract inflammation through inhibiting cyclooxygenase II activity (Jin et al., 2006) and endothelin-1 expression (Zhou et al., 2006). Nevertheless, there is no report of the effects of cryptotanshinone on inflammatory cell infiltration. In this study, complement 5a (C5a) and macrophage inflammatory protein (MIP-1α) were used to induce cell migration, a crucial determinant of leukocyte trafficking, to evaluate the anti-inflammatory properties of cryptotanshinone.

Redistribution of F-actin fibers and the formation of pseudopodia are important events in cell locomotion (Watts, 1996). Leukocyte migration is mediated by the coordinated activation of the cytoskeleton and associated chemoattractant receptors (Ma et al., 1998, Postma et al., 2004). The major component of the cytoskeleton is actin which, in response to chemotactic stimuli, is rapidly and transiently converted from a monomeric, globular form, G-actin, to a needle-like filamentous form, F-actin (Gupta and Campenot, 1996). Although cell migration cannot be attributed to F-actin polymerization alone, the redistribution of F-actin fibers and the formation of pseudopodia are important events in cell locomotion (Watts, 1996). Thus, we also attempted to characterize the effect of cryptotanshinone on chemoattractant-induced morphological change, F-actin reorganization and pseudopodia formation.

Section snippets

Cell culture conditions

RAW264.7 (American Type Culture Collection, TIB 71, Rockville, MD) macrophage-like cells were cultured in Dulbecco's modified Eagle's medium (DMEM, Gibco BRL, India, USA) supplemented with 10% heat-inactivated fetal calf serum, penicillin, and streptomycin (Biological Industries, Israel) at 37 °C in a humidified atmosphere in the presence of 5% CO2 (Chiou et al., 2003a, Chiou et al., 2003b).

Natural products

Cryptotanshinone and tanshinone IIA were isolated by our laboratory (Sun et al., 2006). The dried roots

Effects of cryptotanshinone and tanshinone IIA on C5a- and MIP-1α-induced chemotactic migration

The stimulation dose of 1 μg/ml of C5a was selected according to our previous findings (Chiou et al., 2003a, Chiou et al., 2004). Non-stimulated control macrophages displayed spontaneous migration with a total cell number of 72 ± 16. The concentration gradient generated by 1 μg/ml of C5a induced an 8-fold increase (cell number: 615 ± 49) in cell migration as compared with non-stimulated control and was represented as 100% (Fig. 2). Cryptotanshinone (1–30 μM) alone did not influence the spontaneous

Discussion

We report here that cryptotanshinone displayed inhibitory effect on chemotactic migration and was more potent than tanshinone IIA, suggesting that cryptotanshinone might be one of the active component existing in S. miltiorrhiza Bunge (Danshen). Indeed, our results indicated that cryptotanshinone not only inhibited C5a-induced migration, but also inhibited cell migration in response to MIP-1α. These observations indicated that cryptotanshinone may act as an inhibitor to block a variety of

Acknowledgment

This work was supported by National Research Institute of Chinese Medicine (NRICM95-DBCMR-01), Taipei, Taiwan, ROC.

References (24)

  • W.F. Chiou et al.

    Antiinflammatory properties of piperlactam S: modulates complement 5a induced chemotaxis and inflammatory cytokines production in macrophages

    Planta Medica

    (2003)
  • A.K. Cross et al.

    Chemokines induced migration and changes in actin polymerization in adult rat brain microglia and a human fetal microglial cell line in vitro

    Journal of Neuroscience Research

    (1999)
  • Cited by (12)

    • Cryptotanshinone downregulates the profibrotic activities of hypertrophic scar fibroblasts and accelerates wound healing: A potential therapy for the reduction of skin scarring

      2016, Biomedicine and Pharmacotherapy
      Citation Excerpt :

      Fibrosis is not only accompanied by excessive collagen accumulation, it also associates with increased fibroblast migration. CT has been reported previously to inhibit TNF-α-treated human aortic smooth muscle cells [19] and macrophage migration toward complement 5a and macrophage inflammatory protein-1α [5]. Our study showed CT could suppress HSFs migration.

    View all citing articles on Scopus
    View full text