Study of apoptosis-related markers in ram spermatozoa

Viability Tests, Active Caspase‐3 and −7, and Chromatin Structure in Ram Sperm Selected Using the Swim‐Up Procedure

“…Some studies have reported the presence of active caspases in sperm of different species (Paasch et al, 2004;Martí et al, 2008), and distribution along the sperm appears to be specific in human (Paasch et al, 2004) and ram (Martí et al, 2008). We do not know whether the patterns of caspase activity observed in this study correspond to caspase-3 or -7 because the FLICA reagent interacts with the enzymatic reactive center of an active caspase through a specific recognition sequence (aspartic acid-glutamic acid-valine-aspartic acid, for caspase-3 and -7) and subsequently attaches covalently to a cysteine through the fluoromethyl ketone moiety (Ekert et al, 1999).…”

“…Because sperm motility is an indicator of cell viability, it is assumed that SU-processed sperm are of good quality. Some studies have demonstrated that membrane integrity and mitochondrial function are high, whereas DNA fragmentation and caspase expression decrease after SU processing (Sakkas et al, 2000;Martí et al, 2006Martí et al, , 2008Kotwicka et al, 2008). We evaluated in neat and SU samples cytoplasmic esterases and mitochondrial dehydrogenases as indicators of cytoplasmic membrane integrity and mitochondrial function by using calcein AM and resazurin, respectively.…”

“…However, DNA fragmentation may originate from processes other than apoptosis (Sakkas et al, 1999), and because intrinsic and extrinsic paths of apoptosis converge in caspase activation, it is a more reliable marker of cell death. Caspase activity has been detected in the sperm of humans (Weng et al, 2002;Kotwicka et al, 2008) and rams (Martí et al, 2006(Martí et al, , 2008, and both procaspases and caspases have been immunolocated in the ram (Martí et al, 2008); however, the specific location of active caspases is unknown (Martí et al, 2006(Martí et al, , 2008. Previous studies have reported that ejaculates with a high number of sperm positive for caspases are negatively correlated with fertilization rate after in vitro fertilization (Marchetti et al, 2004), but this relationship has not been described following intracytoplasmic sperm injection (Said et al, 2006).…”

“…The extrinsic and intrinsic pathways to apoptosis converge with the activation of caspases (Chang and Yang, 2000), which induces cell death (Johnson et al, 2000). Some ejaculated sperm express markers of apoptosis (Kotwicka et al, 2008;Martí et al, 2008), which are negatively correlated with semen quality (Weng et al, 2002;Almeida et al, 2005) and fertility (Fatehi et al, 2006). Using SU to sort sperm reduces the number of sperm expressing caspases (Martí et al, 2006(Martí et al, , 2008.…”

“…Early and late markers of apoptosis, such as phosphatidylserine translocation (Paasch et al, 2003;Kotwicka et al, 2008;Martí et al, 2008) and DNA fragmentation (Younglai et al, 2001;Sakkas et al, 2002;Martí et al, 2008), have been used to evaluate sperm quality under different conditions (Sakkas et al, 2000;Paasch et al, 2003;Ricci et al, in press). Phosphatidylserine translocation has been associated with maturation (Barroso et al, 2000;de Vries et al, 2003) and capacitation of sperm (Kotwicka et al, 2002).…”