High molecular mass kininogen inhibits cathepsin G-induced platelet activation by forming a complex with cathepsin G

Abstract

Introduction: Preliminary studies have shown that high molecular mass kininogen (HK) inhibits cathepsin G-induced platelet activation. However, the potential mechanism underlying this inhibitory effect remains to be elucidated.

Materials and methods: Suspensions of washed and gel-filtered platelets were used in radioligand binding and aggregation studies. The amidolytic activity of cathepsin G was measured using specific chromogenic substrate. Western blot technique was utilised to explore the potential complex formation between cathepsin G and HK. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis was used to analyse the cleavage products of HK.

Results: At a concentration of 1 microM, HK completely blocked cathepsin G-induced platelet shape change and secretion of ATP. HK inhibited cathepsin G-induced platelet aggregation in a concentration-dependent manner with an IC(50) of 0.48 microM. Moreover, HK was found to inhibit binding of (125)I-cathepsin G to gel-filtered platelets. (125)I-cathepsin G forms a complex with HK. The complex formation did not affect the amidolytic activity of cathepsin G. HK was proteolysed upon interaction with cathepsin G.

Conclusion: Our results show that high molecular mass kininogen down-regulates cathepsin G-induced platelet activation by forming a complex with cathepsin G and thus prevents binding of cathepsin G to platelets. These kininogen-cathepsin G interactions may be potential targets for pharmacological intervention.