Impact of Circadian Clock Gene Overexpression on Rumen Epithelial Cell Dynamics and VFA Transport Protein Expression

The circadian gene is recognized for its regulatory effects on cell proliferation and lipid metabolism across various non-ruminant cells. This study investigates the influence of gene overexpression on goat rumen epithelial cells using a constructed pcDNA3.1- plasmid, assessing its impact on circadian gene expression, cell proliferation, and mRNA levels of short-chain fatty acid (SCFA) transporters, alongside genes related to lipid metabolism, cell proliferation, and apoptosis. Rumen epithelial cells were obtained every four hours from healthy dairy goats (n = 3; aged 1.5 years; average weight 45.34 ± 4.28 kg), cultured for 48 h in vitro, and segregated into control (pcDNA3.1) and overexpressed (pcDNA3.1-) groups, each with four biological replicates. The study examined the potential connection between circadian rhythms and nutrient assimilation in ruminant, including cell proliferation, apoptosis, cell cycle dynamics, and antioxidant activity and the expression of circadian-related genes, VFA transporter genes and regulatory factors. The introduction of the pcDNA3.1- plasmid drastically elevated expression levels by 3471.48-fold compared to controls ( < 0.01), confirming effective overexpression. overexpression resulted in a significant increase in apoptosis rates ( < 0.05) and a notable reduction in cell proliferation at 24 and 48 h post-transfection ( < 0.05), illustrating an inhibitory effect on rumen epithelial cell growth. elevation significantly boosted the expression of , , , and ( < 0.05) while diminishing expression ( < 0.05). While the general expression of intracellular inflammation genes remained stable, expression notably increased. Antioxidant marker levels (SOD, MDA, GSH-Px, CAT, and T-AOC) exhibited no significant change, suggesting no oxidative damage due to overexpression. Furthermore, overexpression significantly downregulated , , , and mRNA expressions while upregulating and . These results suggest that overexpression impairs cell proliferation, enhances apoptosis, and modulates VFA transporter-related factors in the rumen epithelium. This study implies that the gene may regulate VFA absorption through modulation of VFA transporters in rumen epithelial cells, necessitating further research into its specific regulatory mechanisms.