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Effects of thiol compounds and antioxidants on the activity of MMP-2 and -9 after neonatal hypoxia-reoxygenation Matrix metalloproteinases (MMPs) have been implicated in the degradation of the extracellular matrix in tissue remodeling (Stetler-Stevenson, 1996), where MMP-2 and -9 are believed to be involved in cancer and cardiovascular diseases (Woessner, 1998). Although thiol compounds and antioxidants have been evaluated for their influence on MMP-2 and -9 activities, their effect on the activity of MMP-2 and -9 found in clinical conditions is not available. Therefore the aim of this study was to evaluate the effect of thiol compounds and antioxidants on the activity of MMP-2 and -9 found after neonatal hypoxia-reoxygenation. The tissue activity of MMP-2 and -9 was first determined by gelatin zymography in the brain, heart, liver, kidney and small intestine of 6 piglets (1-4 day old, 1.4-2.2 kg) that underwent alveolar hypoxia (10-12% O2) for 2 hours then resuscitated with 100% O2 for 1 hour followed by room air for 3 hours. We then used human recombinant MMP-2 and -9 ( Calbiochem, USA) at the highest tissue activity identified after neonatal hypoxia-reoxygenation to study the inhibitory effect of amino acids and antioxidants with or without thiol groups. These compounds included: amino acids containing thiol L-cysteine (Cys), DL-homocysteine (Hcy), L-methionine (Met) and not containing thiol L-histidine (His), antioxidants containing thiol L-glutathione (GSH) and N-acetyl-L-cysteine (NAC) and not containing thiol ascorbic acid (Asc), and oxidized GSH (GSSG)(0.03-10 mM; n=8 in each concentration). IC50 values of the gelatinolytic action were determined and compared by one-way ANOVA. Liver had the highest activity of MMP-9, whereas no significant differences were found in MMP-2 activities among the tissues studied. The compounds used in this study showed differential inhibitory effects on the activity of MMP-2 and -9 (6.0 and 0.6 ng/lane, respectively). The potencies of inhibition (IC50 ) for these compounds were widely different (Table 1). The order of the potency of inhibition of these compounds for MMP-2 was Cys ≥ His = Asc = GSH ≥ GSSG ≥ Hcy ≥ NAC > Met, whereas for MMP-9, it was Cys ≥ Asc ≥ His > GSH > Hcy > NAC > GSSG >> Met. Table 1: Differential inhibition of MMP-2 and -9 by thiol compounds and antioxidants
The IC50 s (mM, mean ± s.e. mean) were determined using human recombinant MMP-2 and -9 at the activity after hypoxia-reoxygenation in newborn piglets. At the activity of MMP-2 and -9 measured after neonatal hypoxia-reoxygenation, cysteine showed the highest potency of inhibition. The other compounds showed different potencies of inhibition, regardless of the presence or absence of the thiol group or the antioxidant property of the compound.
Stetler-Stevenson, W.G. (1996). Am J Pathol, 148, 1345-1350. |
