Antioxidative treatment inhibits the release of thrombogenic tissue factor from irradiation- and cytokine-induced endothelial cells.
OBJECTIVE
The aim of this study was to investigate the effect of the antioxidants pyrrolidine dithiocarbamate (PDTC) and N-acetylcysteine (NAC) on the ionizing radiation (IR)- and tumor necrosis factor-alpha (TNF-alpha) induced tissue factor (TF) expression and its release from human umbilical vein endothelial cells (HUVECs).


METHODS
HUVECs were irradiated with a single dose of either 5 Gy or 10 Gy and stimulated with TNF-alpha (10 ng/mL) in the presence or absence of PDTC and NAC, respectively. Quantitative real-time PCR, ELISA, and TF activity measurements were performed, including TF activity in the supernatant. Apoptosis was detected by flow cytometric active caspase-3 measurement and formation of reactive oxygen species (ROS) by chemiluminescence.


RESULTS
We demonstrated a thus far uninvestigated persistent induction of TF expression in HUVECs after treatment with IR and TNF-alpha. Combined stimulation with IR and TNF-alpha led to an immense shedding of microparticle-associated TF which was positively correlated with apoptosis and ROS formation. Antioxidative pre-treatment reduced not only apoptosis and ROS formation, but also the release of thrombogenic microparticles.


CONCLUSIONS
Antioxidative treatment inhibited apoptosis and shedding of microparticles, thereby reducing thrombogenicity. Thus, antioxidants may help to prevent late thrombosis after antiproliferative treatment when used in combination with anticoagulants.
Szotowski
Björn
B
Department of Cardiology and Pneumology, Charité-Universitätsmedizin Berlin, Campus Benjamin Franklin, Berlin, Germany.
Antoniak
Silvio
S
Goldin-Lang
Petra
P
Tran
Quoc-Viet
QV
Pels
Klaus
K
Rosenthal
Peter
P
Bogdanov
Vladimir Y
VY
Borchert
Hans-Hubert
HH
Schultheiss
Heinz-Peter
HP
Rauch
Ursula
U
D000111:Acetylcysteine / Q000494:pharmacology*; D000704:Analysis of Variance; D000975:Antioxidants / Q000494:pharmacology*; D017209:Apoptosis; D053148:Caspase 3 / Q000032:analysis; D002478:Cells, Cultured; D042783:Endothelial Cells / Q000187:drug effects / Q000378:metabolism* / Q000528:radiation effects; D004797:Enzyme-Linked Immunosorbent Assay; D005434:Flow Cytometry; D006801:Humans; D008163:Luminescent Measurements; D011759:Pyrrolidines / Q000494:pharmacology*; D011839:Radiation, Ionizing; D017382:Reactive Oxygen Species / Q000032:analysis / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013859:Thiocarbamates / Q000494:pharmacology*; D013925:Thromboplastin / Q000032:analysis / Q000378:metabolism*; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology; D014471:Umbilical Veins
D000975:Antioxidants; D011759:Pyrrolidines; D017382:Reactive Oxygen Species; D013859:Thiocarbamates; D014409:Tumor Necrosis Factor-alpha; C020972:pyrrolidine dithiocarbamic acid; D013925:Thromboplastin; D053148:Caspase 3; D000111:Acetylcysteine