High copper levels induce inflammation and oxidative stress in a cell culture model of Wilson's disease

The trans-Golgi network compartment acts as one of the major hubs controlling intracellular copper flux by hosting the copper transporter ATP7B. Dysfunctions in hepatocytes leads to cellular copper accumulation, culminating in autophagy, disruption of mitochondrial activity and cell death. Our aim was to compare the cellular responses to copper in wild type (WT) and ATP7B knockout (KO) hepatoma cells (HepG2) and investigating the effect of high copper levels on oxidative stress and inflammatory processes.

The response of HepG2-WT and HepG2-ATP7B-KO cells to CuCl2 was investigated through cell viability (CCK8 Assay) and transcriptional changes. Oxidative stress induction was assessed by GSH/GSSG ratio after CuCl2 treatment. NFĸB and AP1 promoter activity was determined in vitro after CuCl2 treatment using the luciferase reporter and LEGENDPlexTM assay.

Multiple component analysis of GEO data (GSE107323) identified gene expression patterns in ATP7B-KO cells that are associated with autophagy, ion transport and cell intrinsic immunological signals (NFκB and AP1). Quantitative PCR partially validated the expression of selected genes. CuCl2 treatment resulted in upregulation of luciferase reporter activity in NFκB and AP1 promoter assays leading to secretion of TNF, IL1B and IL8. Oxidative stress, determined by decreased GSH/GSSG ratio and increased H2O2 levels was observed 24 h after CuCl2 treatment.

Inflammation is a self-defensive reaction that aims to eliminate or neutralise harmful stimuli and restore tissue integrity. The present data showed that in addition to metal ion transport and autophagy, processes involved in inflammation and oxidative stress were induced by high copper in an ATP7B-KO cell culture model.

Publication History

Article published online:
20 January 2025