Analysis of apoptosis and DNA damage in bovine cumulus cells after exposurein vitroto different zinc concentrations

Abstract

The purpose of this study was to investigate the effect of Zn (zinc) concentration on CCs (cumulus cells) during in vitro maturation. For this purpose, DNA integrity of CCs by addition of different Zn concentrations [0 (control); 0.7 μg/ml (Zn1); 1.1 μg/ml (Zn2) and 1.5 μg/ml (Zn3)] to the culture medium was evaluated by comet assay. In addition, early apoptosis was analysed by annexin staining assay. CCs treated with Zn showed a significant decrease in the DNA damage in a dose-dependent manner. Comet assay analysed for TM (tail moment) was significantly higher in cells cultured without Zn (control, P<0.01) with respect to cells treated with Zn (control: 5.24±16.05; Zn1: 1.13±5.31; Zn2: 0.10±0.36; Zn3: 0.017±0.06). All treatments were statistically different from the control (P = 0.014 for Zn1; P<0.01 for Zn2 and Zn3). The frequency of apoptotic cells was higher in the control group (control: 0.142±0.07; Zn1: 0.109±0.0328; Zn2:0.102±0.013; Zn3: 0.0577±0.019). Statistical differences were found between control and Zn1 (P = 0.0308), control and Zn2 (P = 0.0077), control and Zn3 (P<0.0001), Zn1 and Zn3 (P<0.001) and Zn2 and Zn3 (P = 0.0004). No differences were found between Zn1 and Zn2. In conclusion, low Zn concentrations increase DNA damage and apoptosis in CCs cultured in vitro. However, adequate Zn concentrations 'protect' the integrity of DNA molecule and diminish the percentage of apoptotic CC.

Mapping the zinc‐transporting system in mammary cells: Molecular analysis reveals a phenotype‐dependent zinc‐transporting network during lactation

“…The precise contribution of Zn reuptake in mammary gland Zn metabolism is not understood but it clearly plays a critical role in mammary gland Zn metabolism Zip3‐null mice (Kelleher et al, 2009) (as well as Zip1‐null mice; SLK, unpublished observations) have serious defects in mammary gland morphology. Mammary gland function is dependent upon the maintenance of a highly differentiated secretory cell‐type and we speculate that Zn signaling from the previously secreted milk pool, plays a novel and key role in regulating processes essential to maintaining differentiation such as modulating ATP biogenesis (Lemire et al, 2008), regulating apoptosis (Anchordoquy et al, 2011) and regulating Zn‐finger transcription factors such as Egr1 and Egr2 which are critical for cellular differentiation (Boyle et al, 2009). Further studies are needed to explore the relevance of Zn reuptake on mammary cell function.…”

Maternal control of oocyte quality in cattle “a review”

“…Among these trace elements, zinc, copper, and manganese play an essential role in reproduction. It has been demonstrated that low concentrations of zinc and copper during in vitro maturation (IVM) induce apoptosis and affect the DNA integrity of cumulus cells, which negatively influence the developmental competence of bovine oocytes (Anchordoquy et al, 2011;Picco et al, 2010Picco et al, , 2012. However, adequate zinc (1.5 g/mL) and manganese (6 ng/mL) concentrations during IVM increased the intracellular glutathione concentrations and protect the bovine COCs from apoptosis (Anchordoquy et al, 2014;Picco et al, 2010).…”

Effect of complexed trace minerals on cumulus-oocyte complex recovery and in vitro embryo production in beef cattle1,2

“…More specifically, copper (Cu), iodine (I), manganese (Mn), selenium (Se), and Zinc (Zn) all participate in reproductive processes and are crucial for early embryonic development (Hostetler et al, 2003). Supplementation with Zn, Mn, and Cu improves cumulus-oocyte complex (COC) maturation, reduces apoptosis of cumulus cells, and increases the success of early embryonic development through the blastocyst stage in multiple species including the mouse, cow, pig, and human, when incorporated into in vitro embryo production systems (Gao et al, 2007; Anchordoquy et al, 2011, 2014a, 2014b; Ménézo et al, 2011; Picco et al, 2012; Jeon et al, 2014; Geravandi et al, 2017). Trace minerals have been shown to play a role in the formation of metalloenzymes, which are important in several metabolic processes, including lipid metabolism, glucose utilization, DNA synthesis and transport, and free radical metabolism (Cunnane et al, 1993; De Haan et al, 1994; Townsend et al, 1994; Jovanovic-Peterson and Peterson, 1996; Hostetler et al, 2003).…”