[In vitro experimental study on the upregulation of cellular lactylation modification caused by HiAlc <i>Kpn</i> metabolites via the initiation of cell lipid peroxidation in liver cells].

<b>Objective:</b> To investigate the impact of High Alcohol-Producing <i>Klebsiella pneumoniae</i> (HiAlc <i>Kpn</i>) on hepatocyte function and explore its regulatory mechanism from the perspective of epigenetic modifications. <b>Methods:</b> Using the HepG2 cell line as the research model, the study involved exposing the cells to alcohol and three different HiAlc <i>Kpn</i> strains <i>in vitro</i>, dividing them into a control group, alcohol-treated group, W8 group, 3-24 group, and 4-26 group. The effect of HiAlc <i>Kpn</i> on liver cell proliferation was investigated using the Incucyte live cell imaging system, and the apoptotic level of liver cells was determined using flow cytometry. The fluorescence confocal microscopy combined with live cell probes was used to detect lipid accumulation and intracellular ROS levels in liver cells. The amount of mitochondrial damage was determined using flow cytometry combined with the seahorse cell metabolism analyzer, and changes in protein levels undergoing global lactylation modification were investigated using Western blotting. <b>Results:</b> Compared with the control group, HiAlc <i>Kpn</i> strains W8, 3-24 and 4-26 could decrease the proliferation rate and increase the ratio of apoptosis of hepatocyte HepG2 cells. The results of high-content cell imaging showed that the fluorescence points of ROS enrichment in HepG2 cells were increased after HiAlc <i>Kpn</i> treatment. The lipid accumulation was significantly increased by oil red O and BODIPY staining. The number of oil droplets and fluorescence points was higher than those in the control group and alcohol treatment group. The results of flow cytometry showed that the ratio of JC-1 monomer/polymer was significantly increased after alcohol and three kinds of HiAlc <i>Kpn</i> were treated and the W8 treatment group was about six times higher than the control group (<i>P</i><0.05). Seahorse Energy Metabolism System's mitochondrial pressure test results showed that the extracellular acidification degree and oxygen consumption rate were significantly reduced by the HiAlc <i>Kpn</i> 4-26 strain. Western blot analysis showed that the pan-lactylation modification level increased after high-concentration alcohol treatment and the increased rate of pan-lactylation modification in the 1 000 mmol/L alcohol group was about three times that of the control group. HiAlc <i>Kpn</i> W8 and 3-24 strains resulted in four or two-times pan-lactylation modification increases compared with the control group, and the differences were statistically significant (<i>P</i><0.05). <b>Conclusion:</b> HiAlc <i>Kpn</i> can induce lipid peroxidation in hepatic cells by regulating the increase in histone pan-lactylation modification levels, leading to mitochondrial damage, impaired cell proliferation capacity and increased apoptosis levels.