Effect of METTL3 Gene on Lipopolysaccharide Induced Damage to Primary Small Intestinal Epithelial Cells in Sheep.

Newborn lambs are susceptible to pathogenic bacterial infections leading to enteritis, which affects their growth and development and causes losses in sheep production. It has been reported that N6-methyladenosine (m6A) is closely related to innate immunity, but the effect of m6A on sheep small intestinal epithelial cells (IECs) and the mechanism involved have not been elucidated. Here, we investigated the effects of m6A on lipopolysaccharide (LPS)-induced inflammatory responses, apoptosis and oxidative stress in primary sheep IECs. First, the extracted IECs were identified by immunofluorescence using the epithelial cell signature protein cytokeratin 18 ( CK18 ), and the cellular activity of IECs induced by different concentrations of LPS was determined by the CCK8 assay. Meanwhile, LPS could induce the upregulation of mRNA and protein levels of IECs cytokines IL1β , IL6 and TNFα and the apoptosis marker genes caspase-3 , caspase-9 , Bax , and apoptosis rate, reactive oxygen species (ROS) levels and mRNA levels of CAT , Mn-SOD and CuZn-SOD , and METTL3 were found to be upregulated during induction. It was hypothesized that METTL3 may have a potential effect on the induction of IECs by LPS. Overexpression and knockdown of METTL3 in IECs revealed that a low-level expression of METTL3 could reduce the inflammatory response, apoptosis and ROS levels in LPS-induced IECs to some extent. The results suggest that METTL3 may be a genetic marker for potential resistance to cellular damage.