Fluoride is abundant in the environment and is an essential trace element in living organisms. However, prolonged excessive fluoride intake can lead to fluorosis, which poses a threat to the reproductive health of animals and humans. Although previous research has mainly focused on animal models, the impact of fluoride on ovine follicular granulosa cells (GCs) has not been comprehensively elucidated. This study employed RNA-Seq technology to elucidate the toxic effects of fluoride on ovine follicular GCs and its mechanism of action. Culturing primary ovine follicular GCs in vitro and subjecting them to fluoride treatment revealed 3218 differentially expressed genes (DEGs), with 2278 upregulated and 940 downregulated. Significantly, this study unveiled fluoride’s induction of endoplasmic reticulum (ER) stress in cells, triggering a cascade involving the PERK pathway factor ATF4, leading to cell death via DDIT3/CHOP activation and the subsequent upregulation of CHAC1, ATF3, ERO1α, and TRIB3. These findings provide crucial insights into the toxicity of fluoride in ovine, offering a foundation for mitigating fluoride-related losses in the farming industry.

Zearalenone (ZEN), a non-steroidal estrogenic fungal toxin widely present in forage, food, and their ingredients, poses a serious threat to animal and human reproductive health. ZEN also threatens ovine, a major source of human food and breeding stock. However, the mechanisms underlying the impact of ZEN on the in vitro maturation (IVM) of ovine oocytes remain unclear. This study aimed to elucidate these mechanisms using the Smart-seq2 technology. A total of 146 differentially expressed genes were obtained, using Smart-seq2, from sheep oocytes cultured in vitro after ZEN treatment. ZEN treatment inhibited RUNX2 and SPP1 expression in the PI3K signaling pathway, leading to the downregulation of THBS1 and ultimately the downregulation of TNFAIP6; ZEN can also decrease TNFAIP6 by reducing PTPRC and ITGAM. Both inhibit in vitro maturation of ovine oocytes and proliferation of cumulus cells by downregulating TNFAIP6. These findings provide data and a theoretical basis for elucidating ZEN’s toxicity mechanisms, screening therapeutic drugs, and reducing ZEN-related losses in the ovine industry.

Zearalenone (ZEN) is a common fungal toxin with reproductive toxicity in various grains. It poses a serious threat to ovine and other animal husbandry industries, as well as human reproductive health. Therefore, investigating the mechanism of toxicity and screening antagonistic drugs are of great importance. In this study, based on the natural compound library and previous Smart-seq2 results, antioxidant and anti-apoptotic drugs were selected for screening as potential antagonistic drugs. Three natural plant compounds (oxysophoridine, rutin, and phellodendrine) were screened for their ability to counteract the reproductive toxicity of ZEN on ovine oocytes in vitro using quantitative polymerase chain reaction (qPCR) and reactive oxygen species detection. The compounds exhibited varying pharmacological effects, notably impacting the expression of antioxidant (GPX, SOD1, and SOD2), autophagic (ATG3, ULK2, and LC3), and apoptotic (CAS3, CAS8, and CAS9) genes. Oxysophoridine promoted GPX, SOD1, ULK2, and LC3 expression, while inhibiting CAS3 and CAS8 expression. Rutin promoted SOD2 and ATG3 expression, and inhibited CAS3 and CAS9 expression. Phellodendrine promoted SOD2 and ATG3 expression, and inhibited CAS9 expression. However, all compounds promoted the expression of genes related to cell cycle, spindle checkpoint, oocyte maturation, and cumulus expansion factors. Although the three drugs had different regulatory mechanisms in enhancing antioxidant capacity, enhancing autophagy, and inhibiting cell apoptosis, they all maintained a stable intracellular environment and a normal cell cycle, promoted oocyte maturation and release of cumulus expansion factors, and, ultimately, counteracted ZEN reproductive toxicity to promote the in vitro maturation of ovine oocytes. This study identified three drugs that antagonize the reproductive toxicity of ZEN on ovine oocytes, and compared their mechanisms of action, providing data support and a theoretical basis for their subsequent application in the ovine breeding industry, reducing losses in the breeding industry, screening of ZEN reproductive toxicity antagonists and various toxin antagonists, improving the study of ZEN reproductive toxicity mechanisms, and even protection of human reproductive health.