Low-dose doxycycline inhibits hydrogen peroxide-induced oxidative stress, MMP-2 up-regulation and contractile dysfunction in human saphenous vein grafts

Abstract

Background:

Cardiopulmonary bypass (CPB) applied during coronary artery bypass grafting (CABG), promotes inflammation, generation of reactive oxygen species (ROS) and up-regulation of matrix metalloproteinases (MMPs). All these complications may lead to contractile dysfunction, restenosis and early graft failure, restricting long-term efficacy of bypass grafts. Low-dose doxycycline is a potent MMP inhibitor and ROS scavenger. In this study, we aimed to investigate the effects of doxycycline on ROS generation, MMP regulation and contractile dysfunction induced by H₂O₂ in human saphenous vein (HSV) grafts.

Methods:

HSV grafts (n=7) were divided into four groups after removing endothelial layer by mechanical scratching and incubated with 10 µM H₂O₂ and/or 10 µM doxycycline for 16 hrs. Untreated segments served as control. Concentration–response curves to noradrenaline (NA), potassium chloride (KCl), serotonin (5-HT) and papaverine were performed. Superoxide anion and other ROS levels were determined by using lucigenin- and luminol-enhanced chemiluminescence assays, respectively. Expression/activity of gelatinases (MMP-2/MMP-9) was examined by gelatin zymography. MMP-13 expression was evaluated by immunostaining/immunoscoring.

Results:

H₂O₂ incubation increased superoxide anion and other ROS levels. Doxycycline prevented these increments. H₂O₂ suppressed contractile responses to NA, KCl and 5-HT. Doxycycline ameliorated contractions to NA and KCl but not to 5-HT. H₂O₂ or doxycycline did not altered relaxation to papaverine. MMP-2 and MMP-13 expression increased with H₂O₂, but doxycycline inhibited MMP-2 up-regulation/activation.

Conclusion:

Low-dose doxycycline may have beneficial effects on increased oxidative stress, MMP up-regulation/activation and contractile dysfunction in HSV grafts.

Keywords:

• hydrogen peroxide
• doxycycline
• oxidative stress
• matrix metalloproteinase
• human saphenous vein

Acknowledgments

The authors would like to thank to Prof. Bekir Ugur Ergur for contributions to immunohistochemical analyses, to Hatice Uluer for valuable statistical consultancy and to Scientific Research Foundation of Ege University for the financial support.

Disclosure

The authors report no conflicts of interest in this work.