Characterization of valproic acid-initiated homologous recombination.
BACKGROUND
Valproic acid (VPA) is a frequently used antiepileptic agent and known teratogen. Previous research suggests that inhibition of histone deacetylases (HDACs) may play a role in VPA-induced teratogenicity. We have also shown that VPA exposure leads to both an increase in reactive oxygen species (ROS) production and increased frequency of homologous recombination (HR).


METHODS
In the present study, we evaluated the role of HDAC inhibition in VPA-initiated HR to determine if HDAC inhibition could alter repair activity and/or cause DNA double-strand breaks (DSBs), which would then initiate repair. Histone acetylation status was assessed to determine if VPA exposure led to HDAC inhibition in CHO 33 cells.


RESULTS
Our results demonstrate that VPA (5 mM) exposure leads to increased acetylated histone H3 and H4 protein levels after 10 to 24 hr. Secondly, in our recombination assay where an artificial DNA DSB was induced in CHO 33 cells to assess repair activity, VPA exposure did not affect the repair activity of VPA-initiated HR. Subsequently, to determine if VPA could increase susceptibility to DNA DSBs, the number of gamma-H2AX foci was assessed using immunocytochemistry and results revealed an increase in gamma-H2AX foci after 10- to 24-hr exposure to VPA.


CONCLUSIONS
Although we demonstrated the protective effect of polyethylene glycol-catalase against VPA-induced HR and the generation of intracellular ROS within 24 hr, we did not observed an increase in DNA oxidation. These studies suggest that HDAC inhibition and ROS signaling may play roles in DNA maintenance and cell-cycle arrest in initiating DNA damage and repair.
Sha
Kevin
K
Department of Pharmacology and Toxicology, Queen's University, Kingston, Ontario, Canada.
Winn
Louise M
LM
D000107:Acetylation; D000818:Animals; D000927:Anticonvulsants / Q000494:pharmacology*; D016466:CHO Cells / Q000187:drug effects* / Q000378:metabolism; D002374:Catalase / Q000494:pharmacology; D006224:Cricetinae; D003412:Cricetulus; D004247:DNA / Q000187:drug effects; D053903:DNA Breaks, Double-Stranded / Q000187:drug effects; D004249:DNA Damage; D004260:DNA Repair / Q000187:drug effects; D056572:Histone Deacetylase Inhibitors / Q000494:pharmacology*; D006657:Histones / Q000378:metabolism; D018384:Oxidative Stress; D011092:Polyethylene Glycols / Q000494:pharmacology; D017382:Reactive Oxygen Species / Q000378:metabolism; D011995:Recombination, Genetic / Q000187:drug effects*; D015398:Signal Transduction; D013723:Teratogens / Q000494:pharmacology*; D014635:Valproic Acid / Q000494:pharmacology
D000927:Anticonvulsants; D056572:Histone Deacetylase Inhibitors; D006657:Histones; D017382:Reactive Oxygen Species; D013723:Teratogens; C070295:catalase-polyethylene glycol; D011092:Polyethylene Glycols; D014635:Valproic Acid; D004247:DNA; D002374:Catalase