Figure 3.

The results of aortic valve interstitial cell culture studies in which expression of matrix metalloproteinase-2 (MMP-2) is up-regulated by tenascin-C (TN-C). A: relative amounts of mRNA in sheep (SAVIC) and human aortic valve interstitial cells (HAVIC), evaluated by competitive RT-PCR. Cells were grown on a substrate of either bovine type I collagen or collagen with TN-C (15 μg/ml). A representative agarose gel (see Materials and Methods) is shown for SAVIC demonstrating greater amounts of MMP-2 RNA in the TN-C cultures. The top bands are derived from MMP-2 reverse transcriptase (RT) products, while the bottom bands are due to the MMP-2M13 competitors. The lanes from left to right represent PCRs with a series of dilutions of the MMP-2M13 competitor. The reduction in MMP-2M13 band intensity (bottom bands) occurs in conjunction with an increase in the upper bands (MMP-2 RT). The last lane is a 100 bp DNA ladder. Bars represent the mean ± SE of 3 independent measurements. TN-C significantly increased MMP-2 expression (P < 0.05). B: gel zymograms, demonstrating the results of SAVIC cultures, with greater MMP-2 activity, both pro-, and active-form, in the TN-C cell culture lysates (lane 5) compared to control (lane 4). MMP-2 levels in the conditioned media (detected only as the pro form) were comparable in the TN-C group (lane 3) versus control group (lane 2). No MMP-9 activity was detected in these cell culture studies. Lanes 1 and 6 are molecular marker and MMP gelatinase zymography standards, respectively.