Metastasis plays an important role in mortality of cancer patients. Migration and invasion are the major characteristics of tumor metastasis. The induction of matrix metalloproteinases (MMPs) such as MMP-2 and -9 are particularly important for the invasiveness of various cancer cells. Bufalin, a class of toxic steroids, was purified from the skin glands of Bufo gargarizans or Bufo melanostictus; it is known to inhibit proliferation of human cancer cells. In this study, we investigated the antiinvasive mechanisms of bufalin in the human hepatocellular cancer cell line SK-Hep1. Bufalin significantly reduced serum-induced cell invasion and migration. Furthermore, bufalin markedly inhibited MMP-2 and -9 activity, mRNA expression and protein levels in SK-Hep1 cells. Bufalin attenuated phosphoinisitide-3-kinase (PI3K) and phosphorylation of AKT which was associated with reduced levels of nuclear factor kappa B (NF-κB). Bufalin also suppressed protein levels of FAK and Rho A, VEGF, MEKK3, MKK7, and uPA and it diminished NF-κB translocation. Based on these observations, we propose that bufalin is acts as an antiinvasive agent by inhibiting MMP-2 and -9 and involving PI3K/AKT and NF-κB pathways. Bufalin is a potential therapeutic agent that may have efficacy in preventing the invasion and metastasis of malignant liver tumors.

Triptolide, a traditional Chinese medicine, obtained from Tripterygium wilfordii Hook F, has anti-inflammatory, antiproliferative, and proapoptotic properties. We investigated the potential efficacy of triptolide on murine leukemia by measuring the triptolide-induced cytotoxicity in murine leukemia WEHI-3 cells in vitro. Results indicated that triptolide induced cell morphological changes and induced cytotoxic effects through G0/G1 phase arrest, induction of apoptosis. Flow cytometric assays showed that triptolide increased the production of reactive oxygen species, Ca release and mitochondrial membrane potential (ΔΨ ), and activations of caspase-8, -9, and -3. Triptolide increased protein levels of Fas, Fas-L, Bax, cytochrome c, caspase-9, Endo G, Apaf-1, PARP, caspase-3 but reduced levels of AIF, ATF6α, ATF6β, and GRP78 in WEHI-3 cells. Triptolide stimulated autophagy based on an increase in acidic vacuoles, monodansylcadaverine staining for LC-3 expression and increased protein levels of ATG 5, ATG 7, and ATG 12. The in vitro data suggest that the cytotoxic effects of triptolide may involve cross-talk between cross-interaction of apoptosis and autophagy. Normal BALB/c mice were i.p. injected with WEHI-3 cells to generate leukemia and were oral treatment with triptolide at 0, 0.02, and 0.2 mg/kg for 3 weeks then animals were weighted and blood, liver, spleen samples were collected. Results indicated that triptolide did not significantly affect the weights of animal body, spleen and liver of leukemia mice, however, triptolide significant increased the cell populations of T cells (CD3), B cells (CD19), monocytes (CD11b), and macrophage (Mac-3). Furthermore, triptolide increased the phagocytosis of macrophage from peripheral blood mononuclear cells (PBMC) but not effects from peritoneum. Triptolide promoted T and B cell proliferation at 0.02 and 0.2 mg/kg treatment when cells were pretreated with Con A and LPS stimulation, respectively; however, triptolide did not significant affect NK cell activities in vivo. © 2016 Wiley Periodicals, Inc. Environ Toxicol 32: 550-568, 2017.

Bufalin has been shown to exhibit multiple pharmacological activities, including induction of apoptosis in many types of cancer cell lines. Osteosarcoma is a type of cancer which is difficult to treat and the purpose of this study was to investigate the effects of bufalin on the migration and invasion of human osteosarcoma U-2 OS cells. The wound healing assay and Boyden chamber transwell assay were used for examining the migration of U-2 OS cells. Western blotting and gelatin zymography assays were used for theexpression and activities of metalloproteinase (MMP)-2, MMP-7 or MMP-9 levels. Western blotting analysis also was used for measuring the levels of growth factor receptor-bound protein 2 (GRB2), son of sevenless homolog 1 (SOS1), c-Jun N-terminal kinases 1/2 (JNK1/2), extracellular signal-regulated kinase 1/2 (ERK1/2), and p38 in bufalin-treated U-2 OS cells. Bufalin inhibited the cell migration and invasion of U-2 OS cells in vitro. Moreover, bufalin reduced MMP-2 and MMP-9 enzyme activities of U-2 OS cells. Bufalin also suppressed the protein level of MMP-2 and reduced the levels of mitogen-activated protein kinases (MAPKs) such as JNK1/2 and ERK1/2 signals in U-2 OS cells. Our results suggest that signaling pathways for bufalin-inhibited migration and invasion of U-2 OS cells might be mediated through blocking MAPK signaling and resulting in the inhibition of MMP-2. Bufalin could be a useful agent to develop as a novel antitumor agent by virtue of its ability to inhibit tumor cell migration and invasion.