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Nature and dynamics of nucleosome release from neoplastic and non-neoplastic cells.

BACKGROUND
Circulating nucleosomes are elevated in the blood of patients with malignant and non-malignant diseases. Here, we investigated the nature and the dynamics of their release in functional cell studies.


MATERIALS AND METHODS
Leukemia blasts were exposed to the intrinsic inducers of apoptotic cell death, cytosine arabinoside (AraC; 10 μg/ml) and etoposide (50 μg/ml), and cell death markers lactate dehydrogenase (LDH) and the nucleosomes were measured in the supernatants at 0, 24, 48, 72, and 96 hours after drug application. In addition, HepG2 cells were exposed to extrinsic apoptosis-inducing tumor necrosis factor-related apoptosis-inducing ligand (TRAIL; 0.5 and 1.0 ng/ml) and the nucleosomes were measured in the supernatants after 0, 24, 48, and 72 hours. Finally, neutrophils preactivated by phorbol myristate acetate (PMA) were co-incubated with platelet-rich plasma (PRP) in the presence of collagen (type I; 8 μg/ml) for 15 or 30 minutes at 37°C, and the nucleosome release into the supernatant was quantified.


RESULTS
During treatment with AraC, cell viability constantly decreased. LDH and nucleosome levels increased at 24 h and peaked at 48 h after exposure to AraC and etoposide. While LDH declined after 96 h, the nucleosomes' levels were still elevated. Similarly, nucleosomes increased dose-dependently 24 h after exposure to TRAIL and reached a peak at 48 h. After 72 h, the nucleosomes' levels decreased again. While there was only a minor release of nucleosomes from PMA-stimulated neutrophils, co-incubation with PRP resulted in a strongly increased nucleosome release after 30 minutes.


CONCLUSION
Nucleosomes are released from cells stimulated intrinsically or extrinsically to undergo apoptotic cell death in a time- and dose-dependent manner. Further mechanisms of release may be their active secretion from stimulated neutrophils when co-incubated with PRP, as may be observed during bacterial inflammation and thrombosis.