Oral Presentation ESA-SRB-ANZBMS 2024 in conjunction with ENSA

Investigating the effects of prolonged culture on antioxidant capacity in commercially available insemination media and its impact on protecting spermatozoa against oxidative stress (#178)

Emma M Pyneandee 1 , Robert J Aitken 1 2 , Geoffry N De Iuliis 1 , Hassan W Bakos 1 2
  1. University of Newcastle, Newcastke, NSW, Australia
  2. Memphasys ltd, Sydney, NSW, Australia

Spermatozoa lack significant intracellular antioxidant protection and are particularly susceptible to oxidative stress (OS) and downstream effects of OS result in infertility. Since culture conditions can impact OS in spermatozoa, supplementation of culture media with antioxidants may provide some protection to these cells. The aim of this study was to evaluate the antioxidant protection offered by five commercially available ART media during prolonged culture. Antioxidant protection was assessed using the RoXstaTM assays, which measured lipid peroxide, hydrogen peroxide, and free radical scavenging activity. The media tested were GIVF Plus and GxIVF, supplemented with three antioxidants (Vitrolife), sperm preparation media and Sequential Fertilisation media (Origio) and Quinn’s Fertilisation media (Origio). Sperm parameters of unfractionated samples, motility and DNA integrity, were assessed following 24-hour culture. Motility and mitochondrial reactive oxidative species (ROS) production by spermatozoa were assessed under stress-conditions (arachidonic acid and hydrogen peroxide). The presence of Human Serum Albumin (HSA) was assessed using BCA and SDS-PAGE. All statistical analysis were performed in JMP, with ANOVA and Tukey–Kramer test. This study found that culture media had different antioxidant activity across the RoXstaTM assays. GxIVF, did not have a higher antioxidant activity when compared to GIVF Plus. The antioxidant activity in culture media significantly changed over a 24-hour culture period. Further investigations revealed HSA as the primary source of antioxidant activity in these media. Using normal donor samples, motility was unaffected by antioxidant activity. DNA fragmentation trended lower in media with antioxidants and higher concentrations of HSA. When comparing antioxidant supplementation in Vitrolife’s media, the presence of HSA significantly decreased mitochondrial ROS following exposure to arachidonic acid. Finally, the presence of HSA recovered motility following exposure to hydrogen peroxide. These findings underscore the need for optimizing the supplementation of IVF culture media with stable antioxidants.