Choline attenuates NEFA-induced hepatic steatosis via GNMT regulation in hepatocytes

TL;DR

Choline reduces liver fat buildup in calf and human liver cells by regulating GNMT and related metabolic pathways.

Contribution

Identifies GNMT as a key target of choline in mitigating hepatic steatosis and reveals the AMPK/Myc/GNMT signaling axis as a novel regulatory mechanism.

Findings

Choline reduces NEFA-induced triglyceride accumulation and cytotoxicity in hepatocytes.

GNMT expression is upregulated by choline, and its knockdown reverses choline's protective effects on lipid and bile acid metabolism.

AMPK inhibition reduces GNMT expression and increases Myc, suggesting AMPK/Myc axis mediates choline's effects.

Abstract

To elucidate the molecular mechanisms by which choline regulates hepatic lipid metabolism under negative energy balance conditions, we established non-esterified fatty acid (NEFA)-induced hepatic steatosis models in both calf primary hepatocytes and human LO2 hepatocytes. Choline supplementation significantly reduced intracellular triglyceride accumulation and cytotoxicity induced by NEFA exposure. Transcriptomic profiling identified glycine N-methyltransferase (GNMT) as a key differentially expressed gene. Subsequent experiments confirmed that choline upregulated GNMT expression at both the mRNA and protein levels in a concentration-dependent manner. Knockdown of GNMT reversed the beneficial effects of choline on genes related to lipid synthesis (FAS, ACC), fatty acid oxidation (CPT1), lipoprotein assembly (ApoB100, MTTP), and bile acid metabolism (CYP7A1, CYP27A1, BSEP). Furthermore, inhibition of AMP-activated protein kinase (AMPK) reduced GNMT protein expression and elevated Myc, a negative transcriptional regulator of GNMT, suggesting that choline may regulate GNMT through the AMPK/Myc axis. Collectively, our findings demonstrate that choline alleviates NEFA-induced lipid accumulation and hepatocellular damage by modulating lipid and bile acid metabolism through GNMT, with the AMPK/Myc/GNMT signaling axis playing a pivotal regulatory role. These results provide mechanistic insights into the hepatic protective effects of choline and suggest GNMT as a potential therapeutic target for metabolic disorders in dairy cows and beyond.

The online version contains supplementary material available at 10.1007/s44154-025-00264-3.

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