Cyanotoxin cylindrospermopsin disrupts lipid homeostasis and metabolism in a 3D in vitro model of the human liver.

Cylindrospermopsin, a potent hepatotoxin produced by harmful cyanobacterial blooms, poses environmental and human health concerns. We used a 3D human liver in vitro model based on spheroids of HepG2 cells, in combination with molecular and biochemical assays, automated imaging, targeted LC-MS-based proteomics, and lipidomics, to explore cylindrospermopsin effects on lipid metabolism and the processes implicated in hepatic steatosis. Cylindrospermopsin (1 µM, 48 h) did not significantly affect cell viability but partially reduced albumin secretion. However, it increased neutral lipid accumulation in HepG2 spheroids while decreasing phospholipid levels. Simultaneously, cylindrospermopsin upregulated genes for lipogenesis regulation (SREBF1) and triacylglycerol synthesis (DGAT1/2) and downregulated genes for fatty acid synthesis (ACLY, ACCA, FASN, SCD1). Fatty acid uptake, oxidation, and lipid efflux genes were not significantly affected. Targeted proteomics revealed increased levels of perilipin 2 (adipophilin), a major hepatocyte lipid droplet-associated protein. Lipid profiling quantified 246 lipid species in the spheroids, with 28 significantly enriched and 15 downregulated by cylindrospermopsin. Upregulated species included neutral lipids, sphingolipids (e.g., ceramides and dihexosylceramides), and some glycerophospholipids (phosphatidylethanolamines, phosphatidylserines), while phosphatidylcholines and phosphatidylinositols were mostly reduced. It suggests that cylindrospermopsin exposures might contribute to developing and progressing towards hepatic steatosis or metabolic dysfunction-associated steatotic liver disease (MASLD).

Domino-like effect of C112R mutation on ApoE4 aggregation and its reduction by Alzheimer's Disease drug candidate.

BACKGROUND: Apolipoprotein E (ApoE) ε4 genotype is the most prevalent risk factor for late-onset Alzheimer's Disease (AD). Although ApoE4 differs from its non-pathological ApoE3 isoform only by the C112R mutation, the molecular mechanism of its proteinopathy is unknown. METHODS: Here, we reveal the molecular mechanism of ApoE4 aggregation using a combination of experimental and computational techniques, including X-ray crystallography, site-directed mutagenesis, hydrogen-deuterium mass spectrometry (HDX-MS), static light scattering and molecular dynamics simulations. Treatment of ApoE ε3/ε3 and ε4/ε4 cerebral organoids with tramiprosate was used to compare the effect of tramiprosate on ApoE4 aggregation at the cellular level. RESULTS: We found that C112R substitution in ApoE4 induces long-distance (> 15 Å) conformational changes leading to the formation of a V-shaped dimeric unit that is geometrically different and more aggregation-prone than the ApoE3 structure. AD drug candidate tramiprosate and its metabolite 3-sulfopropanoic acid induce ApoE3-like conformational behavior in ApoE4 and reduce its aggregation propensity. Analysis of ApoE ε4/ε4 cerebral organoids treated with tramiprosate revealed its effect on cholesteryl esters, the storage products of excess cholesterol. CONCLUSIONS: Our results connect the ApoE4 structure with its aggregation propensity, providing a new druggable target for neurodegeneration and ageing.