Sex-dependent effect of sublethal copper concentrations on de novo cholesterol synthesis in astrocytes and their possible links to variations in cholesterol and amyloid precursor protein levels in neuronal membranes.

BACKGROUND: Cholesterol (Cho) is an essential lipophilic molecule in cells; however, both its decrease and its increase may favor the development of neurological diseases such as Alzheimer's disease (AD). Although copper (Cu) is an essential trace metal for cells, the increased plasma concentration of its free form has been linked with AD development and severity. AD affects aged people, but its prevalence and severity are higher in women than in men. We have previously shown that Cu promotes Cho de novo synthesis in immature neurons as well as increased Cho in membrane rafts and Aß levels in culture medium, but there are no results yet regarding sex differences in the effects of sublethal Cu exposure on Cho de novo synthesis. METHODS: We examined the potential sex-specific impact of sublethal Cu concentrations on de novo Cho synthesis in primary cultures of male and female astrocytes. We also explored whether this had any correlation with variations in Cho and APP levels within neuronal membrane rafts. RESULTS: Flow cytometry analysis demonstrated that Cu treatment leads to a greater increase in ROS levels in female astrocytes than in males. Furthermore, through RT-PCR analysis, we observed an upregulation of SREBP-2 and HMGCR. Consistently, we observed an increase in de novo Cho synthesis. Finally, western blot analysis indicated that the levels of ABCA1 increase after Cu treatment, accompanied by a higher release of radiolabeled Cho and an elevation in Cho and APP levels in neuronal membrane rafts. Importantly, all these results were significantly more pronounced in female astrocytes than in males. CONCLUSIONS: Our findings confirm that Cu stimulates Cho synthesis in astrocytes, both in a ROS-dependent and -independent manner. Moreover, female astrocytes displayed elevated levels of HMGCR, and de novo Cho synthesis compared to males following TBH and Cu treatments. This corresponds with higher levels of Cho released into the culture medium and a more significant Cho and APP rise within neuronal rafts. We consider that the increased risk of AD in females partly arises from sex-specific responses to metals and/or exogenous substances, impacting key enzyme regulation in various biochemical pathways, including HMGCR.

Alzheimer's disease (AD) primarily affects the elderly and is linked to excess cholesterol (Cho) and copper (Cu). It is more prevalent and severe in women. Previous research suggested that Cu may enhance Cho synthesis in developing neurons, raising Cho levels in specialized membrane structures (rafts) and Aß protein in the culture medium. However, the specific effects of Cu exposure on Cho synthesis in males and females are not entirely understood. We conducted experiments using astrocytes, the primary cells in the brain that produce Cho in adults, and neurons, both from male and female rats. We exposed them to non-lethal levels of Cu to explore its potential sex-related effects on (1) Cho metabolism in astrocytes, and (2) The relationship between the Cho released by astrocytes and the levels of Cho and amyloid precursor protein (APP) in neuronal membrane rafts. Our findings suggested that reactive oxygen species (ROS)-responsive sensitivity is higher in females than in male astrocytes. Cu, alongside ROS, promoted Cho synthesis, with female astrocytes being more susceptible. These released more Cho into the medium after Cu exposure, and Cho and APP levels were also higher in female neuronal rafts exposed to Cu-treated astrocyte-conditioned medium. Our results thus imply that the higher risk of AD in females may arise partly from sex-related disparities in cellular responses to external substances, impacting such crucial biochemical pathways as Cho synthesis.

Effect of Sublethal Copper Overload on Cholesterol De Novo Synthesis in Undifferentiated Neuronal Cells.

Although copper (Cu) is an essential trace metal for cells, it can induce harmful effects as it participates in the Fenton reaction. Involuntary exposure to Cu overload is much more common than expected and has been linked with neurodegeneration, particularly with Alzheimer's disease (AD) evidenced by a positive correlation between free Cu in plasma and the severity of the disease. It has been suggested that Cu imbalance alters cholesterol (Chol) homeostasis and that high membrane Chol promotes the amyloidogenic processing of the amyloid precursor protein (APP) secreting the ß-amyloid (Aß) peptide. Despite the wide knowledge on the effects of Cu in mature brain metabolism, the consequence of its overload on immature neurons remains unknown. Therefore, we used an undifferentiated human neuroblastoma cell line (SH-SY5Y) to analyze the effect of sublethal concentrations of Cu on 1- de novo Chol synthesis and membrane distribution; 2-APP levels in cells and its distribution in membrane rafts; 3-the levels of Aß in the culture medium. Our results demonstrated that Cu increases reactive oxygen species (ROS) and favors Chol de novo synthesis in both ROS-dependent and independent manners. Also, at least part of these effects was due to the activation of 3-hydroxy-3-methyl glutaryl CoA reductase (HMGCR). In addition, Cu increases the Chol/PL ratio in the cellular membranes, specifically Chol content in membrane rafts. We found no changes in total APP cell levels; however, its presence in membrane rafts increases with the consequent increase of Aß in the culture medium. We conclude that Cu overload favors Chol de novo synthesis in both ROS-dependent and independent manners, being at least in part, responsible for the high Chol levels found in the cell membrane and membrane rafts. These may promote the redistribution of APP into the rafts, favoring the amyloidogenic processing of this protein and increasing the levels of Aß.