MAPK, AKT/FoxO3a and mTOR pathways are involved in cadmium regulating the cell cycle, proliferation and apoptosis of chicken follicular granulosa cells

Abstract

The occurrence of cadmium (Cd) in feed is a major problem in animal health and production. Studies have confirmed that Cd depresses egg production of laying hens, which is closely related to follicular atresia. This study aimed to assess the toxic impacts of Cd on the ovarian tissue, and to examine the mechanism of Cd-induced granulosa cell proliferation and apoptosis. Results from the nitric oxide (NO) and malondialdehyde (MDA) content, total superoxide dismutase (T-SOD), glutathione peroxide (GSH-Px), total nitric oxide synthase (T-NOS) and adenosine triphosphatase (ATPase) activities, terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling (TUNEL) assay, and hematoxylin-eosin (H & E) staining indicated that excess Cd induced oxidative stress, granulosa cell apoptosis and follicular atresia in the layer ovary. Low-dose Cd exposure (1 μM) induced the granulosa cell proliferation, upregulated the mRNA levels of RSK1 and RHEB, activated FoxO3a, AKT, ERK1/2, mTOR and p70S6K1 phosphorylation, and promoted cell cycle progression from phase G1 to S. However, high-dose Cd exposure (15 μM) induced reactive oxygen species (ROS) generation and cell apoptosis, upregulated the mRNA levels of the inflammatory factors, ASK1, JNK, p38 and TAK1, downregulated the expressions of RSK1 and RHEB genes, and inhibited the phosphorylation of ERK1/2, mTOR and p70S6K1 proteins, and the cell cycle progression. Rapamycin pre-treatment completely blocked the phosphorylation of mTOR and p70S6K1 proteins, and the cell cycle progression induced by 1 μM Cd, and accelerated 15 μM Cd-induced cell apoptosis and cell cycle arrest. The microRNA sequencing result showed that 15 μM Cd induced differential expression of microRNA genes, which may regulate AKT, ERK1/2 and mTOR signaling and cell cycle progression by regulating the activity of G proteins and cell cycle-related proteins. Conclusively, these results indicated that Cd can cause the ovarian damage and follicular atresia, and regulate cell cycle, cell proliferation or apoptosis of granulosa cells through MAPK, AKT/FoxO3a and mTOR pathways in laying hens.