Prostaglandin H synthase (PHS)‐1/2‐dependent oxidative DNA damage and cytotoxicity caused by neurotransmitters, their precursors and metabolites

Abstract

PHS-1 and PHS-2 are implicated in neurodegeneration, but the mechanism is unclear. Neurotransmitters, their precursors and metabolites may serve as substrates for PHS-1/2-dependent bioactivation in the brain to free radicals that generate potentially neurotoxic reactive oxygen species (ROS). To assess this hypothesis, ovine PHS-1 was incubated in vitro with DNA and dopamine (DA) or its precursor or metabolites. DA (p<0.05), L-dihydroxyphenylalanine (L-DOPA) (p<0.05) and dihydroxyphenylacetic acid (DOPAC) (p<0.05) were oPHS-1 substrates, resulting in PHS-dependent ROS formation that initiated DNA oxidation, quantified as 8-oxo-2′-deoxyguanosine. CHO-K1 cell lines expressing human PHS-2 (hPHS-2) and untransformed CHO-K1 cells were used to investigate hPHS-2-dependent cytotoxicity. PHS activity, quantified by the conversion of arachidonic acid (AA) to prostaglandin E2, was up to 40-fold higher in hPHS-2 cells compared to untransformed CHO-K1 cells and activities were reduced by withholding AA (p<0.01). Cytotoxicity, measured by lactate dehydrogenase release, was higher in hPHS-2 cells treated with DA, L-DOPA, DOPAC or homovanillic acid versus vehicle control after a 6 hour incubation (p<0.001). AA-activation increased hPHS-2 cytotoxicity (p<0.05), while cytotoxicity was lower in untransformed CHO-K1 cells (p<0.05). These results show that DA, its precursor and metabolites are substrates for PHS-1/2-dependent bioactivation, causing ROS-mediated oxidative DNA damage and cytotoxicity. This mechanism may be involved in neurodegenerative changes associated with aging and drugs like amphetamines, which initiate neurotransmitter release. [Support: CIHR, CIHR/Rx&D HRF]