Sonmez_2026_Turk.J.Biol_50_245

BACKGROUND/AIM: Polyunsaturated fatty acids (PUFAs) and the hepatic cholinergic axis are critical regulators of lipid metabolism and inflammatory signaling; however, their time-dependent interplay in hepatocytes remains incompletely understood. Clarifying how n-6 and n-3 PUFAs interact with cholinergic pathways may provide mechanistic insight into metabolic dysfunction-associated steatotic liver disease (MASLD). MATERIALS AND METHODS: HepG2 cells were used to examine the combined effects of butyrylcholinesterase (BChE) overexpression and PUFA exposure. Cells were treated with linoleic acid (LA; n-6) or alpha-linolenic acid (alpha-LA; n-3) for 24 or 48 h. Gene expression analyses were performed to assess pathways involved in de novo lipogenesis, beta-oxidation, cholinergic signaling, and inflammatory responses. Cholinesterase activities were measured enzymatically, secreted cytokines were profiled, and molecular docking simulations were performed to evaluate potential alpha-LA interactions with the alpha7 nicotinic acetylcholine receptor (alpha7 nAChR) and M3 muscarinic acetylcholine receptor (M3 mAChR). RESULTS: An early antilipogenic profile and changes suggestive of increased fatty-acid oxidation (SREBP-1c, ACC1, and CPT1A) were induced by alpha-LA at 24 h and were partially attenuated by 48 h. LA exhibited a biphasic pattern characterized by modest early suppression followed by a pronounced proinflammatory profile at 48 h (TNF, IL6, and COX-2). BChE overexpression was associated with increased expression of lipogenic and triglyceride-related genes, whereas cotreatment with PUFAs resulted in time- and class-specific modulation. Cholinergic markers exhibited divergent expression patterns: BCHE + alpha-LA increased CHAT and CHRNA7 expression at 48 h, whereas ACHE mRNA expression increased under both BCHE + LA and BCHE + alpha-LA conditions. Enzymatic analyses demonstrated increased total cholinesterase activity in the BChE and alpha-LA groups, while molecular docking supported the potential accommodation of alpha-LA within the binding pockets of alpha7 nAChR and M3 mAChR. CONCLUSION: PUFA class and exposure duration jointly shape hepatocellular metabolic and inflammatory states, with cholinergic signaling acting as a modulatory node relevant to MASLD.