Gut microbiota and bile acids changes in MASLD mice model with hepatic PLD1 knockout

Abstract

Hepatocyte phospholipase D1 (PLD1) knockout alleviates metabolic dysfunction-associated steatotic liver disease (MASLD) in mice, but the underlying mechanism is largely unknown. In this study, the mice are divided into four groups: Con (wild-type mice with normal control diet), HFHC (wild-type mice with high-fat diet), Con_KO (hepatocyte PLD1-knockout mice with normal control diet), and HFHC_KO (hepatocyte PLD1-knockout mice with high-fat diet). Intestinal contents of mice are analyzed via metagenomics and metabolomics, and the liver bile acids are assessed by mass spectrometry imaging. The results show that at the phylum level the abundance of Bacillota in the intestines of MASLD model mice is significantly increased, whereas that of Bacteroidota significantly is decreased. However, after the deletion of hepatocyte PLD1, Pseudomonadota and Candidatus Bathyarchaeota are significantly decreased in the MASLD model mice. At the species level, compared with that in the Con group, the abundance of Faecalibaculum rodentium is significantly increased in the HFHC group, whereas hepatocyte PLD1 knockout causes the abundances of Desulfovibrionaceae bacterium LT0009 and Lachnospiraceae bacterium 10-1 to be significantly decreased. In terms of intestinal bile acids, the levels of two bile acids (hyodeoxycholic acid and glycolithocholic acid) differ between the HFHC_KO group and the HFHC group. Association analysis shows that Faecalibaculum co-occurs with DCA, βMCA, ΩMCA and αMCA, while probiotic Bacteroides uniformis is significantly correlated with UDCA, 12-KetoLCA, and 7-KetoLCA. Finally, mass spectrometry imaging reveals that the TCA and TDCA contents in the liver are significantly decreased after PLD1 knockout in hepatocytes. These findings demonstrate that hepatocyte PLD1 knockout alters the gut microbiota and bile acids profiles, suggesting that PLD1 deficiency may modulate MASLD progression by changing intestinal microbiota-bile acid homeostasis.

1. Introduction

Metabolic dysfunction-associated steatotic liver disease (MASLD) has emerged as the most prevalent chronic liver disease, and with the transformation of modern lifestyles and the increasing prevalence of metabolic syndrome, such as obesity, the prevalence of MASLD is likely to continue rising. MASLD is associated with an increased risk of cardiovascular events, extrahepatic malignancy, liver failure, and hepatocellular carcinoma [1–3]. Nevertheless, to date, there remains a lack of clear and reliable therapeutic approaches for MASLD in clinical practice.

An increasing number of researchers are now exploring the interaction between the gut-liver axis and MASLD, using the gut microbiota as a key entry point to search for interventions for liver lesions in MASLD. Previous studies have demonstrated that the structure of the gut microbiota in MASLD patients often undergoes significant alterations, and the most typical feature is an imbalance in the ratio of Bacillota to Bacteroidetes, which is closely associated with the development of hepatic steatosis and obesity, strongly indicating the potential role of dysbiosis of the gut microbiota in the pathogenesis of MASLD [4]. In patients with metabolic dysfunction-associated steatohepatitis (MASH), the abundance of Bacteroides and Ruminococcus is significantly increased, whereas that of Prevotella is decreased [5]. In addition, animal studies have provided compelling evidence for the potential role of the gut microbiota in MASLD [6]. Studies have shown that the ingestion of the intestinal microbiota from MASLD mice by normal mice can cause the transmission of MASLD-related disease symptoms [7].

Phospholipase D1 (PLD1), an isoenzyme of phospholipase D (PLD), has a unique biological function and can decompose phosphatidylcholine into phosphatidic acid. As an important lipid second messenger, phosphatidic acid is widely involved in the regulation of intracellular signaling pathways [8]. PLD1 is ubiquitously expressed on the surface of mammalian cells [8] and participates in lipid metabolism by regulating the accumulation of lipid droplets [9,10]. In our prior studies, PLD1 was found to be significantly upregulated in the livers of MASLD mice, and the disease was markedly alleviated when hepatic PLD1 was absent [11]. Although previous studies have highlighted the close relationship between the gut microbiota and MASLD, there is currently no evidence regarding the impact of hepatic PLD1 on the gut microbiota.