Glucose-induced oxidative stress in vascular contractile cells: comparison of aortic smooth muscle cells and retinal pericytes.
Free radical-mediated damage to vascular cells may be involved in the pathogenesis of diabetic vasculopathy. The aim of this study was to compare the extent of glucose-induced oxidative stress in both vascular smooth muscle cells (VSMCs) and pericytes and the effect on antioxidant enzyme gene expression and activities. Porcine aortic VSMC and retinal pericytes were cultured in either 5 or 25 mmol/l glucose for 10 days. Intracellular malondialdehyde (MDA) was measured as a marker of peroxidative damage, and mRNA expression of CuZn-SOD, MnSOD, catalase, and glutathione peroxidase (GPX) were measured by Northern analysis. Glutathione (GSH) was also measured. There was a significant increase in MDA in VSMCs in 25 mmol/l glucose (1.34 +/- 0.11 vs. 1.88 +/- 0.24 nmol/mg protein, 5 vs. 25 mmol/l D-glucose, mean +/- SE, n = 15, P < 0.01), but not in pericytes (0.38 +/- 0.05 vs. 0.37 +/- 0.05 nmol/mg protein, n = 11). There was a significant decrease in GSH in both cell types (VSMC, 1.40 +/- 0.13 vs. 0.69 +/- 0.12 nmol/mg protein, n = 15, P < 0.001; pericytes, 1.97 +/- 0.17 vs. 0.94 +/- 0.16 nmol/mg protein, n = 11, P < 0.001). mRNA expression of CuZnSOD and MnSOD was increased only in VSMCs (by 58.5 +/- 8.1 and 41.0 +/- 6.9%, respectively, n = 8, P < 0.01). CuZnSOD protein was increased by approximately 120% (P < 0.00001). None of the antioxidant enzyme activities was altered between 5 and 25 mmol/l glucose in either cell type. Both MnSOD activities and GSH concentrations were higher in pericytes compared with VSMC under basal (5 mmol/l) conditions (P < 0.05 and P < 0.02, respectively). These results demonstrate glucose-induced reduction of GSH in both cells, but only in VSMC is there evidence of oxidant damage in the form of lipid peroxidation, implying significant differences in intracellular responses to glucose between contractile cells in the macro- and microvasculature.
Sharpe
P C
PC
Department of Clinical Biochemistry, The Queen's University of Belfast, Northern Ireland, UK. p.sharpe@qub.ac.uk
Liu
W H
WH
Yue
K K
KK
McMaster
D
D
Catherwood
M A
MA
McGinty
A M
AM
Trimble
E R
ER
D000818:Animals; D000975:Antioxidants / Q000378:metabolism; D001011:Aorta / Q000166:cytology / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D005947:Glucose / Q000494:pharmacology*; D005978:Glutathione / Q000378:metabolism; D015227:Lipid Peroxidation / Q000187:drug effects; D008315:Malondialdehyde / Q000378:metabolism; D008833:Microcirculation / Q000187:drug effects; D009131:Muscle, Smooth, Vascular / Q000166:cytology / Q000187:drug effects*; D018384:Oxidative Stress / Q000187:drug effects*; D012160:Retina / Q000166:cytology / Q000187:drug effects*; D013552:Swine; D014661:Vasoconstriction / Q000187:drug effects*
D000975:Antioxidants; D008315:Malondialdehyde; D005978:Glutathione; D005947:Glucose