# FOXA3 Alleviates Lipid Deposition in Primary Bovine Hepatocytes by Inhibiting SREBP1 and Cell Proliferation

## TL;DR

FOXA3 helps reduce fat buildup in cow liver cells by suppressing fat production genes and cell growth, offering a new way to prevent liver disease in dairy cows.

## Contribution

This study reveals FOXA3's novel role in mitigating lipid accumulation in bovine hepatocytes by inhibiting SREBP1 and cell proliferation.

## Key findings

- FOXA3 expression is significantly reduced in the livers of cows with fatty liver.
- FOXA3 overexpression reduces lipid accumulation by inhibiting SREBP1 and affecting cell cycle pathways.
- NEFA treatment downregulates FOXA3 and promotes triacylglycerol accumulation in hepatocytes.

## Abstract

During the periparturient period, dairy cows often experience negative energy balance, leading to elevated levels of non-esterified fatty acids (NEFA) in their blood. This condition predisposes them to fatty liver disease and ketosis. FOXA3, a crucial transcription factor involved in regulating liver metabolism, has an unclear role in the pathogenesis of fatty liver in dairy cows. This study aimed to elucidate the mechanism by which FOXA3 regulates hepatic lipid metabolism. We analyzed liver samples from dairy cows diagnosed with fatty liver, developed a lipid accumulation model using primary hepatocytes treated with NEFA, and employed FOXA3 overexpression technology. Our findings revealed a significant reduction in FOXA3 expression in the livers of dairy cows with fatty liver. Furthermore, NEFA were found to downregulate FOXA3 expression and promote lipid accumulation. In contrast, FOXA3 overexpression mitigated lipid accumulation by inhibiting lipogenesis-related genes, such as SREBP1, and influencing cell cycle pathways. This study elucidates the protective role of FOXA3 in the development of fatty liver in dairy cows, indicating that enhancing FOXA3 expression could be a novel strategy for the prevention and treatment of fatty liver and ketosis in dairy cows, with significant economic value.

During the periparturient period, negative energy balance (NEB) in dairy cows leads to increased concentrations of non-esterified fatty acids (NEFA) in the blood, which can induce fatty liver disease and ketosis. Forkhead box protein A3 (FOXA3) is a key transcription factor that regulates liver metabolism; however, its specific role in the pathogenesis of fatty liver in dairy cows remains unclear. This study aimed to investigate the mechanism by which FOXA3 regulates hepatic lipid metabolism. We collected liver samples from dairy cows with fatty liver (n = 10) and from healthy cows (n = 10). Bovine primary hepatocytes were isolated from the liver tissue of calves (n = 5), followed by NEFA treatment, and we utilized FOXA3 overexpression, immunofluorescence, and RNA sequencing (RNA-seq) to conduct our analysis. Our results demonstrated that FOXA3 expression in the livers of cows with fatty liver was significantly lower than in healthy cows. NEFA treatment resulted in the downregulation of FOXA3 protein levels in hepatocytes, promoting triacylglycerol (TAG) accumulation and the expression of lipogenesis-related genes. Conversely, FOXA3 overexpression mitigated NEFA-induced lipid accumulation, inhibited the expression of lipogenesis-related genes and proteins—particularly SREBP1—and affected cell proliferation, and the intracellular localization of FOXA3 and SREBP1. RNA-seq analysis suggested that FOXA3 may influence hepatic lipogenesis through pathways such as PI3K-Akt and the cell cycle. In summary, FOXA3 mitigates NEFA-induced hepatic lipid accumulation through a dual mechanism: regulating SREBP1 expression and inhibiting cellular proliferation. These findings highlight FOXA3′s potential as a novel target for the prevention and treatment of fatty liver disease in dairy cows.