Integrin activation protects pulmonary endothelial cells from the genotoxic effects of bleomycin.
Integrin activation promotes the survival of endothelial cells undergoing diverse forms of stress. Here we determined the ability of integrins to inhibit DNA strand breakage by bleomycin (BLM), a DNA-cleaving antitumor antibiotic that causes acute endothelial injury and subsequent pulmonary fibrosis. We found that BLM produced DNA breakage in cultured murine lung endothelial cells (MLEC) within 45 min of treatment as measured by DNA sedimentation and in situ labeling of 3'-OH by nick translation (ISNT). Two hours after the removal of BLM, we found a marked but incomplete reduction in DNA strand breakage as measured by ISNT, indicating that the damage was reversible. DNA sedimentation and ISNT demonstrated that strand breakage due to BLM was inhibited in MLEC cultured on fibronectin, and no evidence of breakage was found 2 h after removal of the drug in ISNT experiments. Gelatin, type IV collagen, laminin, and the integrin ligand peptide Gly-Arg-Gly-Asp-Ser-Pro, but not the inactive Gly-Arg-Ala-Asp-Ser-Pro peptide, also inhibited DNA strand breakage. Activation of integrins, either by coating surfaces with antibodies to alpha 5-, beta 1-, or beta 3-integrin subunits or by receptor clustering with the soluble antibodies, inhibited BLM-induced DNA breakage. Inhibition of BLM-induced DNA strand breakage by soluble beta 1-integrin antibody increased with increasing antibody concentration and duration of receptor clustering before BLM treatment. Thus integrin activation protects pulmonary endothelial cells from the genotoxic effects of BLM.
Hoyt
D G
DG
Department of Pharmacology, University of Pittsburgh School of Medicine, Pennsylvania 15261, USA.
Rizzo
M
M
Gerritsen
M E
ME
Pitt
B R
BR
Lazo
J S
JS
D000818:Animals; D000906:Antibodies / Q000494:pharmacology; D001761:Bleomycin / Q000633:toxicity*; D002478:Cells, Cultured; D004247:DNA / Q000187:drug effects; D004249:DNA Damage; D004727:Endothelium / Q000187:drug effects* / Q000473:pathology / Q000502:physiology; D016326:Extracellular Matrix Proteins / Q000494:pharmacology; D016023:Integrins / Q000276:immunology / Q000502:physiology*; D008070:Lipopolysaccharides / Q000494:pharmacology; D008168:Lung / Q000187:drug effects* / Q000473:pathology / Q000502:physiology; D051379:Mice; D008810:Mice, Inbred C57BL; D009153:Mutagens; D009842:Oligopeptides / Q000494:pharmacology
D000906:Antibodies; D016326:Extracellular Matrix Proteins; D016023:Integrins; D008070:Lipopolysaccharides; D009153:Mutagens; D009842:Oligopeptides; C101396:glycyl-arginyl-alanyl-aspartyl-seryl-proline; D001761:Bleomycin; D004247:DNA; C058161:glycyl-arginyl-glycyl-aspartyl-seryl-proline