(R)-α-Lipoic Acid Reverses the Age-Associated Increase in Susceptibility of Hepatocytes to tert-Butylhydroperoxide both In Vitro and In Vivo

Abstract

Hepatocytes were isolated from young (3-5 months) and old (24-28 months) rats and incubated with various concentrations of tert-butylhydroperoxide (t-BuOOH). The t-BuOOH concentration that killed 50% of cells (LC50) in 2 hr declined nearly two-fold from 721 +/- 32 microM in cells from young rats to 391 +/- 31 microM in cells from old rats. This increased sensitivity of hepatocytes from old rats may be due, in part, to changes in glutathione (GSH) levels, because total cellular and mitochondrial GSH were 37.7% and 58.3% lower, respectively, compared to cells from young rats. Cells from old animals were incubated with either (R)- or (S)-lipoic acid (100 microM) for 30 min prior to the addition of 300 microM t-BuOOH. The physiologically relevant (R)-form, a coenzyme in mitochondria, as opposed to the (S)-form significantly protected hepatocytes against t-BuOOH toxicity. Dietary supplementation of (R)-lipoic acid [0.5% (wt/wt)] for 2 weeks also completely reversed the age-related decline in hepatocellular GSH levels and the increased vulnerability to t-BuOOH as well. An identical supplemental diet fed to young rats did not enhance the resistance to t-BuOOH, indicating that antioxidant protection was already optimal in young rats. Thus, this study shows that cells from old animals are more susceptible to oxidant insult and (R)-lipoic acid, after reduction to an antioxidant in the mitochondria, effectively reverses this age-related increase in oxidant vulnerability.

Is α‐lipoic acid a scavenger of reactive oxygen species in vivo? Evidence for its initiation of stress signaling pathways that promote endogenous antioxidant capacity

“…LA ultimately increased intracellular GSH by inducing the transcription of both the catalytic and regulatory subunits of c-glutamylcysteine ligase, which is the rate-controlling enzyme for GSH synthesis. The improved GSH status afforded by LA also reversed the age-related increased susceptibility of hepatocytes to tert-butyl hydroperoxide, a model alkylhydroperoxide that is detoxified in a GSH-dependent manner (14). Thus, LA appears to be an inducer of Nrf2-mediated antioxidant gene expression, which significantly increases the cellular capacity to synthesize GSH.…”

“…Despite these caveats, it is clear that dietary LA provides a remarkable range of positive therapeutic benefits, which includes limiting the neurological complications from diabetes, lowering the age-associated increases in oxidative damage, and reducing the toxicity from free radical generating redox-active transition metals (14)(15)(16)(17). All these outcomes implicate an antioxidant role for LA in vivo.…”

Decline in transcriptional activity of Nrf2 causes age-related loss of glutathione synthesis, which is reversible with lipoic acid

“…Nowhere is this loss more pronounced than in the age-related decline in hepatic glutathione (GSH) levels. GSH is the principal low molecular weight thiol antioxidant and the cosubstrate for a variety of antioxidant and anti-xenobiotic (Phase II) enzymes (4)(5)(6)(7). Decline in constitutive GSH levels adversely affects cellular thiol redox balance and potentially leaves the cell susceptible to a number of internal and environmental stresses.…”

“…Due to the central role of GSH in cellular protective mechanisms, the induction of enzymes required for its synthesis represents a key adaptive response to oxidative injury. In aging, however, when basal levels of oxidative stress become elevated, GSH and the enzymes from which it is synthesized do not concomitantly increase but actually decline in many tissues (4,7,10,11). This lack of a cellular compensatory response to loss in GSH and the existence of a prooxidant state in aging cells suggest that the coordination of cellular antioxidant defenses may be altered with age.…”

“…Moreover, previous studies show that thiol-reactive substances such as 3H-1,2-dithiole-3-thione (D3T), pyrrolidine dithiocarbamate (PDTC) (24), sulforaphane, and (R)-␣-lipoic acid (LA) (7,25,26) act as potent chemopreventive agents that increase cellular GSH and Phase II response. Thus, we also sought to understand whether potential lesions in basal regulation of Nrf2 prevent its activation by the disulfide chemoprotectant, lipoic acid.…”