Effects of dimethyl sulfoxide on apolipoprotein A-I in the human hepatoma cell line, HepG2.

Exposure of HepG2 cells to 1% (v/v) dimethyl sulfoxide (DMSO), an effective free radical scavenger, for 24 h resulted in a 2-fold increase in the levels of apolipoprotein (apo) A-I mRNA and secreted protein, with no significant change in apoA-II, apoB, and apoE mRNA and protein levels. The induction of apoA-I was accompanied by a 50% increase in secreted HDL. Nuclear run-off assays indicated that the transcription rate of the apoA-I gene was also increased 2-fold in DMSO-treated cells. Consistent with nuclear run-off assays, transient transfection experiments, using a series of pGL2-derived luciferase reporter constructs containing the human apoA-I proximal promoter, demonstrated that DMSO treatment increased apoA-I promoter activity 2-fold. We have identified a potential 'antioxidant response element' (ARE) in the apoA-I promoter that may be responsible for the increase in apoA-I transcriptional activity by DMSO. Gel mobility shift assays with an apoA-I-ARE revealed increased levels of a specific protein-DNA complex that formed with nuclear extracts from DMSO-treated cells. The formation of this complex is sequence specific as determined by DNA competition studies. When a copy of the ARE was inserted upstream of the SV40 promoter in a luciferase reporter plasmid, a significant 2-fold induction in luciferase activity was observed in HepG2 cells in the presence of DMSO. In contrast, a plasmid containing a mutated apoA-I-ARE did not confer responsiveness to DMSO treatment. Furthermore, pGL2 (apoA-I-250 mutant ARE), in which point mutations eliminated the ARE in the apoA-I promoter, showed no increase in luciferase activity in response to DMSO. These results implicate protein-DNA interactions at the antioxidant response element region in the transcriptional induction of human apoA-I gene expression by DMSO.