Kallistatin is a
plasma protein with anti-inflammatory properties. In this study, we investigated the role and mechanisms of
kallistatin in inhibiting endothelial
inflammation through its
heparin-binding domain. We showed that recombinant wild-type
kallistatin dose-dependently competed with
tumor necrosis factor (
TNF)-alpha binding to
TNF-alpha receptor in endothelial cells, whereas
kallistatin mutant at the
heparin-binding domain had no effect.
Kallistatin, but not
kallistatin mutant at the
heparin-binding domain, abrogated
TNF-alpha-induced endothelial cell activation, as evidenced by inhibition of
TNF receptor 1-associated death domain
protein activation, inhibitor of
nuclear factor kappaB-alpha degradation,
nuclear factor kappaB translocation, and
p38 mitogen-activated protein kinase phosphorylation, as well as
cell adhesion molecule and
cytokine expression. Moreover,
kallistatin, but not
kallistatin mutant at the
heparin-binding domain, inhibited
TNF-alpha-induced human monocytic THP-1 cell adhesion to endothelial cells and prevented
vascular endothelial growth factor-induced endothelial permeability. In mice,
kallistatin gene delivery prevented vascular leakage provoked by
complement factor C5a, whereas delivery of
kallistatin heparin mutant gene had no effect. Similarly, gene transfer of
kallistatin, but not the
kallistatin heparin mutant, inhibited
collagen/adjuvant-induced
arthritis in rats. These results indicate that
kallistatin's
heparin-binding site plays an essential role in preventing
TNF-alpha-mediated endothelial activation and reducing
vascular endothelial growth factor-induced vascular permeability, resulting in attenuation of vascular
inflammation in cultured endothelial cells and animal models. This study identifies a protective role of
kallistatin in
vascular injury, thereby implicating the therapeutic potential of
kallistatin for vascular and inflammatory diseases.