Enhanced expression of glutathione peroxidase protects islet β cells from hypoxia-reoxygenation.

Lepore DA, Shinkel TA, Fisicaro N, Mysore TB, Johnson LEA, d'Apice AJF, Cowan PJ. Enhanced expression of glutathione peroxidase protects islet β cells from hypoxia-reoxygenation. Xenotransplantation 2004; 11: 53–59. © Blackwell Munksgaard, 2004 The survival of pancreatic islet β–cell xenografts and allografts may be affected by damaging reactive oxygen and nitrogen species generated during hypoxia-reoxygenation. Peroxynitrite, which is formed from superoxide and nitric oxide, appears to be an important mediator of β-cell destruction. The intracellular antioxidant enzymes glutathione peroxidase-1 (Gpx-1) and copper–zinc superoxide dismutase (CuZn SOD) detoxify peroxynitrite and superoxide, respectively. The aim of this study was to examine whether enhanced expression of Gpx-1 and/or CuZn SOD protected NIT-1 mouse insulinoma cells from hypoxia–reoxygenation injury. Stable transfectants expressing human Gpx-1 or CuZn SOD were isolated and tested for their resistance to hydrogen peroxide (H₂O₂) and menadione, which generates superoxide intracellularly. Clones expressing one or both enzymes were subjected to hypoxia in glucose-free medium for 18 h, followed by reoxygenation in complete medium for 1.5 h. Cell viability was measured using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazoliumbromide (MTT) reduction assay. Increases of up to two fold in Gpx or total SOD activity protected NIT-1 cells from H₂O₂ and menadione. Expression of Gpx-1 significantly increased NIT-1 survival following hypoxia-reoxygenation (viability 65 ± 9% vs. control 15 ± 3%, P < 0.001) but CuZn SOD expression had no effect (15 ± 1%). Expression of both enzymes was no more protective (60 ± 6%) than expression of Gpx-1 alone. Genetic manipulation of islet β cells to increase expression of Gpx-1 may protect them from oxidative injury associated with the transplantation procedure. [ABSTRACT FROM AUTHOR]