D-propranolol attenuates lysosomal iron accumulation and oxidative injury in endothelial cells.
The influence of selected beta-receptor blockers on iron overload and oxidative stress in endothelial cells (ECs) was assessed. Confluent bovine ECs were loaded with iron dextran (15 muM) for 24 h and then exposed to dihydroxyfumarate (DHF), a source of reactive oxygen species, for up to 2 h. Intracellular oxidant formation, monitored by fluorescence of 2',7'-dichlorofluorescin (DCF; 30 microM), increased and peaked at 30 min; total glutathione decreased by 52 +/- 5% (p < 0.01) at 60 min. When the ECs were pretreated 30 min before iron loading with 1.25 to 10 microM d-propranolol, glutathione losses were attenuated 15 to 80%, with EC(50) = 3.1 microM. d-Propranolol partially inhibited the DCF intensity increase, but atenolol up to 10 microM was ineffective. At 2 h, caspase 3 activity was elevated 3.2 +/- 0.3-fold (p < 0.01) in the iron-loaded and DHF-treated ECs, and cell survival, determined 24 h later, decreased 47 +/- 6% (p < 0.01). Ten micromoles of d-propranolol suppressed the caspase 3 activation by 63% (p < 0.05) and preserved cell survival back to 88% of control (p < 0.01). In separate experiments, 24-h iron loading resulted in a 3.6 +/- 0.8-fold increase in total EC iron determined by atomic absorption spectroscopy; d-propranolol at 5 microM reduced this increase to 1.5 +/- 0.4-fold (p < 0.01) of controls. Microscopic observation by Perls' staining revealed that the excessive iron accumulated in vesicular endosomal/lysosomal structures, which were substantially diminished by d-propranolol. We previously showed that propranolol could readily concentrate into the lysosomes and raise the intralysosomal pH; it is suggested that the lysosomotropic properties of d-propranolol retarded the EC iron accumulation and thereby conferred the protective effects against iron load-mediated cytotoxicity.
Mak
I Tong
IT
Department of Biochemistry and Molecular Biology, Division of Experimental Medicine, George Washington University Medical Center, 2300 Eye Street, N.W. Ross Hall, Rm 443, Washington DC 20037, USA. itmak@gwu.edu
Chmielinska
Joanna J
JJ
Nedelec
Lucie
L
Torres
Armida
A
Weglicki
William B
WB
D000319:Adrenergic beta-Antagonists / Q000494:pharmacology*; D000818:Animals; D053148:Caspase 3; D020169:Caspases / Q000378:metabolism; D002417:Cattle; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects; D042783:Endothelial Cells / Q000187:drug effects* / Q000201:enzymology / Q000378:metabolism; D005978:Glutathione / Q000378:metabolism; D007501:Iron / Q000378:metabolism*; D008247:Lysosomes / Q000187:drug effects* / Q000201:enzymology / Q000378:metabolism; D018384:Oxidative Stress; D011433:Propranolol / Q000494:pharmacology*; D017382:Reactive Oxygen Species / Q000378:metabolism
D000319:Adrenergic beta-Antagonists; D017382:Reactive Oxygen Species; D011433:Propranolol; D007501:Iron; D053148:Caspase 3; D020169:Caspases; D005978:Glutathione