OxLDL induces macrophage gamma-GCS-HS protein expression: a role for oxLDL-associated lipid hydroperoxide in GSH synthesis.

Oxidized LDL (oxLDL) produced a rapid depletion of intracellular glutathione (GSH) followed by an adaptive increase in J774 A.1 macrophages. OxLDL also induced a transient increase in the levels of gamma-glutamylcysteine synthetase heavy subunit (gamma-GCS-HS), representing the catalytic subunit of the rate-limiting enzyme for de novo GSH synthesis. The induction took place within 3 h, with maximum levels observed by 10 h of treatment. Pretreatment of oxLDL with ebselen inhibited GSH depletion and attenuated the gamma-GCS-HS induction. OxLDL-associated lipid hydroperoxides and their decomposition product aldehydes are two major components thought to account for GSH depletion in macrophages. Ebselen pretreatment had only a minor effect on malondialdehyde levels, whereas peroxide content was essentially abolished, suggesting that oxLDL-associated hydroperoxides may mediate both GSH depletion and gamma-GCS-HS induction. Acetylated LDL (AcLDL) also caused a moderate induction of gamma-GCS-HS protein along with a 30% transient increase in GSH by 3;-6 h, suggesting a minor involvement of scavenger receptor-mediated signaling of GSH synthesis. However, the level of gamma-GCS induction by AcLDL was insufficient to cause a sustained increase in GSH. Macrophages with higher glutathione peroxidase (GPx) activity experienced a more rapid and extensive depletion of GSH when treated with oxLDL under similar conditions, along with greater resistance to oxLDL- or peroxide-induced cytotoxicity. We conclude that oxLDL-associated peroxides are primarily responsible for GSH depletion, creating an oxidizing environment required for gamma-GCS induction and compensatory GSH synthesis. This is facilitated in cells expressing high GPx activity through a rapid depletion of GSH in the face of a peroxide challenge.