A novel anti-inflammatory peptide inhibits endothelial cell cytoskeletal rearrangement, nitric oxide synthase translocation, and paracellular permeability increases.

The endothelial cell (EC) membrane-cytoskeletal interface in part maintains plasma membrane integrity and promotes cell-cell apposition. Nonmuscle filamin (ABP-280), an actin crosslinking protein, promotes orthogonal branching of F-actin and is the major protein that links the peripheral actin network to the plasma membrane through its C-terminal glycoprotein binding site. In response to bradykinin, filamin translocates from the cell periphery to the cytosol within 1 min. A synthetic peptide, corresponding to filamin's C-terminal calcium/calmodulin-dependent protein kinase II phosphorylation site (CaM peptide), prevents calcium-activated filamin translocation in permeabilized bovine pulmonary artery EC. The myristoylated permeable form of this peptide inhibits bradykinin-induced filamin translocation and F-actin rearrangement in cultured intact ECs. In addition, bradykinin-induced paracellular gap formation is significantly attenuated by CaM peptide, which suggests that the presence of a filamin-based peripheral F-actin network is essential for maintaining EC barrier function. Moreover, CaM peptide reduces wound-induced EC migration rate by 40%, which indicates that F-actin rearrangement is required for efficient cell motility. The CaM peptide affects other bradykinin-induced inflammatory responses. EC nitric oxide synthase (eNOS) translocates from the cell membrane to the nuclear fraction within 1-2 min of bradykinin treatment. Pretreatment with CaM peptide inhibits eNOS translocation. However, the peptide has no effect on bradykinin-induced von Willebrand Factor release. In summary, the CaM peptide exhibits several anti-inflammatory properties that include maintaining EC junctional stability and inhibiting eNOS translocation.