α-lipoic acid ameliorates consequences of copper overload by up-regulating selenoproteins and decreasing redox misbalance

Abstract

α-lipoic acid (LA) is an essential cofactor for mitochondrial dehydrogenases and is required for cell growth, metabolic fuel production, and antioxidant defense. In vitro, LA binds copper (Cu) with high affinity and as an endogenous membrane permeable metabolite could be advantageous in mitigating the consequences of Cu overload in human diseases. We tested this hypothesis in 3T3-L1 preadipocytes with inactivated Cu transporter Atp7a; these cells accumulate Cu and show morphologic changes and mitochondria impairment. Treatment with LA corrected the morphology of Atp7a −/− cells similar to the Cu chelator bathocuproinedisulfonate (BCS) and improved mitochondria function; however, the mechanisms of LA and BCS action were different. Unlike BCS, LA did not decrease intracellular Cu but instead increased selenium levels that were low in Atp7a −/− cells. Proteome analysis confirmed distinct cell responses to these compounds and identified upregulation of selenoproteins as the major effect of LA on preadipocytes. Upregulation of selenoproteins was associated with an improved GSH:GSSG ratio in cellular compartments, which was lowered by elevated Cu, and reversal of protein oxidation. Thus, LA diminishes toxic effects of elevated Cu by improving cellular redox environment. We also show that selenium levels are decreased in tissues of a Wilson disease animal model, especially in the liver, making LA an attractive candidate for supplemental treatment of this disease.

ATP7A-dependent copper sequestration contributes to termination of β-CATENIN signaling during early adipogenesis

“…We propose that Atp7a-mediated transient Cu deficit is beneficial for initiation of lipogenesis and lipid accumulation, and serves as an integral component of the entire differentiation program. The effects of Cu on cytoskeleton and cell morphology were previously observed in 3T3-L1 cells [ 23 ], as well as in other cell types [ 32 , 33 ]. Our studies suggest that β-catenin, which coordinates gene transcription and cell adhesion, could be an important factor in the observed morphological changes; however, the role of Cu in adipocyte morphology need to be studied further.…”

“…To further examine significance of Atp7a function in adipocyte differentiation, we used 3T3-L1- Atp7a −/− cells, previously generated using CRISPR/Cas9. These cells lack the Atp7a-dependent Cu export and accumulate Cu ([ 22 , 23 ], Figure 1 D and Suppl. Figure 3A ).…”

“…The elevation of Cu in either Atp7a −/− skin fibroblasts or adipocytes was previously shown to induce oxidative stress [ 22 , 23 ]. Consequently, we tested whether normal 3T3-L1 cell differentiation is accompanied by changes in redox balance, i.e.…”

“…Two molecular mechanisms may explain β-catenin stabilization. First, excess Cu is known to cause oxidative stress, and 3T3-L1- Atp7a −/− cells were previously shown to have redox misbalance [ 22 , 23 ]. Oxidative stress can cause changes in β-catenin plasma-membrane retention/nuclear distribution and increase expression of Wnt target genes [ 35 ].…”

Bioorthogonal Cu Single‐Atom Nanozyme for Synergistic Nanocatalytic Therapy, Photothermal Therapy, Cuproptosis and Immunotherapy

“…Intriguingly, upon irradiation with a 1064 nm laser, cell viability was further reduced, and the IC 50 value decreased to 7.66 μM (Figure 4d), which was consistent with the results of calcein-AM/propidium iodide (PI) staining assay (Figure S24). Surprisingly, we found that IC 50 values were significantly enhanced by 2.58-fold, 4.81-fold and > 18-fold when we employed cuproptosis inhibitors, including UK 5099 (an inhibitor of mitochondrial pyruvate uptake), [17] antimycin A (an inhibitor of the electron transport chain) [18] and bathocuproinedisulfonic acid (BCS) (a copper chelator) [19] to incubate CuSACO plus Ac 4 ManNAz-treated cells than that of CuSACO plus Ac 4 ManNAz group, which displayed that CuSACO induced cell death via cuproptosis. To further validate the CuSACO-mediated cuproptosis, 4T1 cells treated with CuSACO plus Ac 4 ManNAz, CuSA plus Ac 4 ManNAz or PBS (control group) were collected and subjected to western blotting analysis.…”