Unraveling the role of glucosylated sterols in modulating glucocorticoid receptor activity: insights from zebrafish models and cellular studies

Glucosylated sterols are lipid molecules consisting of a sterol backbone and a glucose unit. Although their role in the cellular metabolism remains largely unclear, disruption in glucosylated sterols homeostasis seems to be associated with an increased risk of developing neurodegenerative disorders. This suggests that GlcSt may play a critical role in neuronal health and proper functioning of the nervous system. Among GlcSt, β-glucosyl-cholesterol is synthesized by the lysosomal enzyme glucocerebrosidase (GBA1) in humans, while certain pathogens, including Helicobacter pylori, produce α-glucosyl-cholesterol. Interestingly, mutations in the GBA1 gene, encoding GBA1, are the major known genetic risk factor for Parkinson’s disease (PD) and infections caused by Helicobacter pylori have been associated with a higher prevalence of PD in affected patients. GlcSt levels can also be influenced by dietary intake. β-glucosyl-sitosterol (β-GlcSito), a plant-derived GlcSt abundantly present in the seeds of Cycas micronesica, a native plant of south-east Asia, has gained attention due to its link with the development of the neurodegenerative disease Amyotrophic Lateral Sclerosis-Parkinsonism Dementia Complex (ALS-PDC). ALS-PDC can exhibit features of both PD and ALS, including neuronal loss, protein inclusions, motor deficits, dementia, and muscle atrophy. This neurodegenerative disease affected the Chamorro people of Guam, who traditionally consumed Cycas seed products, fostering the connection between the occurrence of the disorder and the dietary intake of β-GlcSito. Among the most prevalent neurodegenerative diseases, PD and ALS are characterized by the presence of non-motor symptoms that may emerge long before the appearance of the canonical motor symptoms associated with the loss of dopaminergic and motor neurons, respectively. Gastrointestinal dysfunctions such as constipation and delayed food transit together with the presence of intestinal inflammation are commonly observed in PD and ALS patients even years before the definition of a formal diagnosis. This fosters the urgence to explore the mechanisms underlying the deep connection existing between the gut and the brain, and how one could influence the other in the emergence and progression of neurodegenerative diseases. In this context, my thesis project aims at investigating the role of GlcSt, and in particular of β-GlcSito, in modulating detrimental effect in the intestine, thus promoting the gut inflammation that may be prodromal to neurodegeneration. Data previously produced in the lab evidenced that β-GlcSito can interact with the glucocorticoid receptor (GR) and modulate its activity. Since β-GlcSito interacts with the GR penetrating the nucleus, it was hypothesized that it could pass through a cholesterol channel (as β-GlcSito is a derivative of cholesterol). Supporting this option was the work conducted by some colleagues, who demonstrated that the affinity of the molecule for ten cholesterol transporter channels was very low. Consequently, an alternative pathway was considered, specifically a glucose transporter channel. Therefore, this thesis project is focused on the characterization of the impact of β-GlcSito treatment on GR nuclear translocation by overexpressing GR-EGFP in cellular models, and to assess the expression of GR-regulated genes through qPCR analysis in intestinal cells. Moreover, the study of the effects of β-GlcSito administration to mutant zebrafish lines, lacking a functional response to sterol intake, specifically, cyp11c1 -/-, deficient in corticosteroid biosynthesis, gr -/-, devoid of GR-mediated signaling and NF-κB line that allows the visualization of NF-κB pathway. These in vivo models will help elucidate the implications of sterol signaling disruption in the context of gut–brain axis impairment and neurodegeneration.