HT-2 toxin, a type A trichothecene mycotoxins produced by Fusarium, is a widespread contaminant of animal feed and food products (Ok et al., 2013). It has become a serious problem in Europe and China due to its contamination of grains and drinking water (Langseth and Rundberget, 1999, Pleadin et al., 2017). HT-2 toxin has been shown to cause male reproductive toxicity in rabbits by suppressing spermatogenesis and testosterone secretion (Kolesarova et al., 2017). Due to its high activity of oxygen ring and carbon-oxygen double bond, HT-2 toxin can cause intracellular reactive oxygen species (ROS) accumulation, thereby damaging DNA and leading to cell death (Zhang et al., 2017a, Zhang et al., 2017b).

Programmed cell death, including apoptosis and ferroptosis, can been induced by ROS accumulation (Dixon et al., 2012). Apoptosis is regulated by several signal pathways, including extrinsic and mitochondrial pathways. ROS can stimulate the activation of both pathways. Apoptosis is regulated by several signal pathways, including extrinsic (Romero et al., 2015) and mitochondrial pathway (Green and Llambi, 2015). Given evidence has demonstrated that ROS can stimulate the activation of both pathways (Chaudhary et al., 2010, Elkin et al., 2018). Moreover, attributable to higher levels of polyunsaturated fatty acids, cellular and organelle membranes are vulnerable to ROS damage which named “lipid peroxidation” (Que et al., 2018). Ferroptosis, a newly discovered modality of cell death, is mainly caused by lipid peroxidation and intracellular iron catalytic activity. characterized by intracellular metabolic process glutaminolysis and acculturation of ROS, as well as mitochondria membrane potential decrease (Dixon et al., 2012, Yang et al., 2014).

Melatonin, as potent antioxidant, has long been known to effectively eliminate ROS to prevent cells from oxidative stress (Zhang et al., 2020). In addition, melatonin can promote the secretion of testosterone in estrus animals (Deng et al., 2018, Li and Zhou, 2015). LCs are one of the principal cells that produce testosterone and are important for spermatogenesis and body maturation (Zhao et al., 2020). However, the in vitro function of melatonin on apoptosis and ferroptosis of HT-2 toxin-treated LCs has yet to be determined, and the mechanisms of its regulation remain elusive.

In current study, we investigated that HT-2 toxin resulted in apoptosis and ferroptosis on sheep LCs in vitro. Moreover, adding melatonin ameliorated the damage of HT-2 toxin and reduced intracellular ROS by maintaining G6PD expression in HT-2 toxin-treated LCs in vitro. Our founding will provide fundamental evidence for the regulatory mechanisms of melatonin in anti-ferroptosis and anti-apoptosis and the effects of melatonin on male reproduction.