Glycosaminoglycans Modify Elastase Action In Vitro and Enhance Elastase-Induced Cell Death in Cultured Fibroblasts

Figure 3

HNE-like peptidase activity in NCM. Supernatants from incubation of neutrophils with DMEM were kept in 100 mM HEPES buffer pH 7.5, containing 500 mM NaCl and 10% DMSO for 10 min at 37°C. After that, the substrate MeOSuc-Ala-Ala-Pro-Val-AMC (4.0 μM) was added. Substrate hydrolysis was followed by the increasing in the fluorescence (𝜆_exc=360nm and 𝜆_em=460nm) in the spectrofluorimeter. HNE concentration in NCM was determined using a standard curve of MeOSuc-Ala-Ala-Pro-Val-AMC hydrolysis by commercial HNE.