Bufalin, a
cardiotonic steroid extracted from
toad venom, has generally been known to possess a range of
biological activities; however, only a few studies have reported the anti-metastatic activity of
bufalin. In the present study, we investigated the inhibitory effects of
bufalin on cell migration and invasion, two critical cellular processes that are often deregulated during
metastasis, using the human
bladder cancer cell line, T24. Within the concentration range that was not cytotoxic,
bufalin markedly inhibited the cell motility and invasiveness of T24 cells. The inhibitory effects of
bufalin on cell invasiveness were associated with the tightening of tight junctions (TJs), which was demonstrated by an increase in transepithelial electrical resistance (TER).
Bufalin treatment also repressed the levels of
claudin proteins (claudin-2, -3 and -4) and the major components of TJs that play key roles in the control and selectivity of paracellular transport. Furthermore, the activities of
matrix metalloproteinase (
MMP)‑2 and -9 in T24 cells were dose‑dependently inhibited by treatment with
bufalin and this also correlated with a decrease in their
mRNA and
protein expression levels; however, the
mRNA and
protein levels of the
tissue inhibitor of metalloproteinase (TIMP)‑1 and -2 were increased. In addition, these effects were related to the increased phosphorylation of the extracellular signal-regulated
protein kinase (ERK) pathway. The inhibition of ERK (
PD98059) significantly prevented the bufalin‑induced suppression of T24 cell migration. These findings suggest that
bufalin inhibits the migration and invasion of T24 cells by modulating the activity of TJs and
MMPs, possibly in association with the activation of ERK.