Journal: Scientific Reports
Article Title: Human osteoarthritis knee joint synovial fluids cleave and activate Proteinase-Activated Receptor (PAR) mediated signaling
doi: 10.1038/s41598-023-28068-3
Figure Lengend Snippet: Cleavage of enzyme class preferred fluorogenic substrates by healthy and OA knee joint synovial fluids. Fluorogenic substrates were incubated with the healthy and OA synovial fluids (10%) with and without enzyme inhibitors (PPACK, STI, BB-94 or ONO-4817) and cleavage of ( A ) Bz-FVR-AMC (thrombin-like enzymes), ( B ) Boc-QAR-AMC (trypsin-like enzymes), and ( C ) MCA-KPLGL-Dpa(DNP)-AR-NH 2 (MMPs) substrates was monitored. Representative kinetic traces of four healthy and four patient samples obtained with ( D ) Bz-FVR-AMC (10 min) and ( E ) Boc-QAR-AMC (10 min), and ( F ) four patient samples with MCA-KPLGL-Dpa(DNP)-AR-NH 2 (60 min) substrates. The data represents the mean ± SEM ( N = 1–3). One-Way ANOVA Kruskal–Wallis test was utilized to assess differences between groups. p < 0.05 was considered statistically significant.
Article Snippet: The fluorogenic substrates, Bz-Phe-Val-Arg-AMC.HCl (Thrombin substrate III) and Boc-Gln-Ala-Arg-AMC.HCl (Trypsin substrate) were from Bachem, Suc-Ala-Ala-Pro-Phe-AMC (Chymotrypsin substrate II) and Z-Gly-Gly-Arg-AMC.HCl (Urokinase substrate III) were from Calbiochem, and MCA-Lys-Pro-Leu-Gly-Leu-Dpa(DNP)-Ala-Arg-NH 2 (MMP substrate FS-6) was from Sigma-Aldrich.
Techniques: Incubation