Affiliation:
1. College of Veterinary Medicine, Yangzhou University, Yangzhou 225009, China
2. Joint International Research Laboratory of Agriculture and Agri-Product Safety of Ministry of Education of China, Yangzhou University, Yangzhou 225009, China
3. International Joint Research Laboratory in Universities of Jiangsu Province of China for Domestic Animal Germplasm Resources and Genetic Improvement, Yangzhou University, Yangzhou 225009, China
4. College of Animal Science and Technology, Yangzhou University, Yangzhou 225009, China
Abstract
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.
Funder
National Natural Science Foundation of China-CGIAR
Major Project of the Natural Science Foundation of Xinjiang Uyghur Autonomous Region
National Natural Science Foundation of China
Major New Varieties of Agricultural Projects in Jiangsu province
Jiangshu 333 Distinguished Talents Project Foundation
Natural Science Foundation of the Jiangsu Higher Education Institutions of China
Jiangsu Agricultural Science and Technology Innovation Fund
the Project of Jiangsu Innovation Support Programme for International Science and Technology Cooperation—“Belt and Road” Innovation Cooperation
Yangzhou Meat Sheep Industry Public Service Platform
Distinguished Talents Project Foundation of Yangzhou University, and High-end Foreign Expert Introduction Project