Effect of hypoxia on stimulatory effect of TGF-beta 1 on MMP-2 and MMP-9 activities in mouse fibroblasts

OBJECTIVE: Because chronic low-grade hypoxia has been implicated in the pat hogenesis of fibrosis and postoperative adhesion formation we hypothesized that hypoxia may modulate the effect of transforming growth factor-beta1 (T GF-beta1) on matrix metalloproteinase (MMP)-2 and MMP-9 at both transcripti onal and translational levels. METHODS: Mouse fibroblasts were placed in a hypoxic environment with or wit hout 1 ng/mL TGF-beta1 for varying periods of time. Zymography was performe d on cell supernatants collected after each treatment. Gelatinolytic bands corresponding to MMP-2 and MMP-9 were quantiated by densitometry, Multiplex reverse transcriptase-polymerase chain reaction (RT-PCR) was also performe d for MMP-2 and MMP-9 on total RNA extracted from cells after each treatmen t. Analysis of PCR-amplified products was performed by 2% agarose gel follo wed by ethidium bromide staining of DNA bands, Scanning densometry was used to determine the ratio of intensity of each band relative to beta -actin. RESULTS: Hypoxia resulted in a 64% decrease in MMP-9 activity and 80% decre ase in MMP-9 mRNA level but did not affect MMP-2 mRNA level or activity. TG F-beta1 treatment resulted in 180% and 50% increases in MMP-2 and MMP-9 act ivities, respectively, Increases of 37.5% and 40% in MMP-2 and MMP-9 mRNA l evels, respectively, were seen. However, under hypoxic conditions, TGF-beta 1 resulted in a 160% increase and 45% decrease in MMP-2 and MMP-9 activitie s and a 37.5% increase and 71% decrease in MMP-2 and MMP-9 mRNA levels, res pectively. CONCLUSION: Hypoxia suppresses the stimulatory effect of TGF-beta1 on the a ctivity, of MMP-9, but not MMP-2. This may suggest an important role for MM P-9 under hypoxic conditions in the pathogenesis of tissue fibrosis and pos toperative adhesion formation. (J Soc Gynecol Investig 2000;7:348-54) Copyr ight (C) 2000 by the Society for Gynecologic Investigation.