Reduction-oxidation (redox) state regulation of matrix metalloproteinase activity in human fetal membranes

OBJECTIVE: The mechanisms underlying membrane rupture at term and preterm a re obscure. Collagenolytic activity of matrix metalloproteinases in amnioch orionic membranes increases during spontaneous term and preterm labor assoc iated with intra-amniotic infection. We sought to test the hypothesis that reduction-oxidation homeostasis, which is altered in inflammatory states, d irectly regulates amniochorionic matrix metalloproteinases. STUDY DESIGN: Membranes were collected from 7 patients undergoing elective cesarean delivery at term, rinsed thoroughly, and immediately incubated in phosphate-buffered sodium chloride solution at 37 degrees C for 24 hours. M atrix metalloproteinase activity in the culture medium was assayed by subst rate-gel electrophoresis and normalized against the dry weight of the tissu e incubated. Superoxide anions were generated in the presence of membranes by a xanthine (2 mmol/L) and xanthine oxidase (20 mU/mL) mixture and monito red by reduction of ferri-cytochrome c to ferro-cytochrome c. Incubations w ere performed in the presence of xanthine alone, a xanthine-xanthine oxidas e mixture, superoxide dismutase (500 U/ml), a xanthine-xanthine oxidase-sup eroxide dismutase mixture, nitro-c-arginine (a nitric oxide synthase inhibi tor, 1 mmol/L), xanthine-xanthine oxidase-nitro-L-arginine, S-nitroso-N-ace tylpenicillamine (a nitric oxide donor, 10 mmol/L), xanthine-xanthine oxida se-S-nitroso-N-acetylpenicillamine, N-acetylcysteine (a thiol-containing an tioxidant, 0.1, 1, or 10 mmol/L), lipopolysaccharide (100 ng/mL), or lipopo lysaccharide-N-acetylcysteine. Intracellular generation of superoxide anion s was monitored by the reduction of nitroblue tetrazolium to formazan. RESULTS: Basal matrix metalloproteinase 9 and matrix metalloproteinase 2 le vels were detected in all samples. Superoxide anions significantly increase d matrix metalloproteinase 9 activity but did not increase matrix metallopr oteinase 2 activity, which effect was reversed by the addition of superoxid e dismutase. N-acetylcysteine reduced basal activity of both matrix metallo proteinase 9 and matrix metalloproteinase 2 to 20%. importantly N-acetylcys teine completely inhibited intracellular formazan formation in cultured mem branes both in the absence and in the presence of lipopolysaccharide. Neith er nitric oxide synthase inhibition nor the nitric oxide donor S-nitroso-N- acetylpenicillamine had any effect on fetal membrane matrix metalloproteina se activity. CONCLUSION: Matrix metalloproteinase activity in human fetal membranes is r eduction-oxidation (redox)-regulated. Matrix metalloproteinase 9 activity i n human fetal membranes is directly increased by superoxide anion, a byprod uct of macrophages and neutrophils. Neither nitric oxide donors nor nitric oxide synthase inhibitors significantly affect matrix metalloproteinase act ivity in human fetal membranes. The glutathione precursor N-acetylcysteine dramatically inhibits amniochorionic matrix metalloproteinase activity in a ddition to inhibiting intrinsic superoxide generation within the tissue. Th us thiol-reducing agents, such as N-acetylcysteine, may be beneficial in pr eventing preterm premature rupture of the membranes.