Inhibition of Matrix Metalloproteinase Activity Attenuates Tenascin-C Production and Calcification of Implanted Purified Elastin in Rats

摘要

Elastin, a major extracellular matrix protein present in arterial walls provides elastic recoil and resilience to arteries. Elastin is prone to calcification in a number of cardiovascular diseases including atherosclerosis and bioprosthetic heart valve mineralization. We have recently shown that purified elastin when implanted subdermally in rats undergoes severe calcification. In the present study, we used this elastin implant model to investigate the molecular mechanisms underlying elastin calcification. Intense matrix metalloproteinase (MMP-2) and tenascin-C (TN-C) expression were seen in the proximity of the initial cal-cific deposits at 7 days. Gelatin zymography studies showed both MMP-2 (latent and active form) and MMP-9 expression within the implants. To investigate the role of MMPs in calcification, rats were administered a MMP inhibitor, (2S-allyl-N-hydroxy-3R-isobutyl-N-(1S-methylcarbamoyl-2-phenylethyl)-succinamide (BB-1101) by daily injection, either systemically or at the implant site. The site-specific BB-1101 administration almost completely suppressed TN-C expression, as shown by immunohistochemical staining, within the implants. The systemic BB-1101 injections also significantly reduced TN-C expression within the elastin implants. Moreover, calcification of elastin implants was significantly reduced in the site-specific administration group (5.43 ± 1.03 µg/mg Ca for BB-1101 group versus 21.71 ± 1.19 for control group, P < 0.001). Alizarin Red staining clearly showed that the elastin fibers were heavily calcified in the control group, whereas in BB-1101 group the calcification was scarce with few fibers showing initial calcification deposits. The systemic administration of BB-1101 also significantly reduced elastin calcification (28.07 ± 5.81 control versus 16.92 ± 2.56 in the BB-1101 group, P < 0.05), although less than the site-specific administration. Thus, the present studies indicate that MMPs and TN-C play a role in elastin-oriented calcification.