Fig 3. Representative zymograms showing gelatinolytic activity in serum from patients with ascending aortic/aortic root aneurysms and healthy controls.

Increasing amounts of total protein were analyzed to detect possible signals close to detection limit. Lane 1: Human full length MMP-2 as delivered. Lane 2: Human full length MMP-2 activated with 2 mM APMA for 2 hours. Asterisk indicates 67 kDa intermediate MMP-2 isoform present in MMP-2 standard with and without APMA incubation and in serum incubated with APMA. A. Serum from patients with ascending aortic aneurysms was analyzed in total protein amounts from 15 μg to 80 μg. Whereas signals corresponding to pro-MMP-2 increased, no signals relatable to active MMP-2 became visible. B. Serum from patients with ascending aortic aneurysms was incubated with 2mM APMA for 2 hours at 37°C and analyzed in increasing amounts of total protein from 15 μg to 40 μg. Signals related to pro-MMP-2 and intermediate MMP-2 became stronger with increasing amount of total protein. But no signal relatable to the 65kDa active MMP-2 appeared. C. Serum from healthy controls was analyzed in total protein amounts of 15 μg until 80 μg. signals increased in the same way as described for the serum samples from patients with ascending aortic aneurysms. No signals corresponding to active MMP-2 appeared. D. Serum from healthy controls was analyzed in total protein amounts of 15 μg until 40 μg after incubation with APMA for 2 hours at 37°C. Signals for pro-MMP-2 and intermediate MMP-2 increased with protein amount but no signals relatable to active MMP-2 appeared.