NH4Cl promotes apoptosis and inflammation in bovine mammary epithelial cells via the circ02771/miR-194b/TGIF1 axis

Abstract

Excess ammonia (NH3) in the circulation of dairy animals can reduce animal health and the quality of products for human consumption. To develop effective prevention and treatment methods, it is essential to examine the molecular mechanisms through which excess NH3 may affect the mammary gland. The present study used bovine mammary epithelial cells (BMECs) to evaluate the effects of exogenous NH4Cl on the abundance of circular RNAs (circRNAs) using high-throughput sequencing. Among the identified circRNAs, circ02771 was the most significantly upregulated by exogenous NH4Cl (P<0.05), with a fold change of 4.12. The results of the apoptosis and proliferation assays, transmission electron microscopy, H&E staining, and immunohistochemistry revealed that circ02771 increased apoptosis and inflammation. A double luciferase reporter assay revealed that circ02771 targeted miR-194b, and the overexpression of circ02771 (pcDNA-circ02771) reduced (P<0.05) the expression of miR-194b and led to apoptosis and inflammation. Circ02771 also enhanced the expression of transforming growth factor beta-induced factor homeobox 1 (TGIF1), which is a target gene of miR-194b. Overall, this study suggests that the circ02771/miR-194b/TGIF1 axis plays a role in mediating the effects of NH4Cl on BMECs. Therefore, this axis provides a novel target to help control hazards within the mammary gland from high circulating NH4Cl levels.