Oral Presentation ESA-SRB 2023 in conjunction with ENSA

GM-CSF treatment of frozen/thawed bovine sperm improves sperm function and quality. (#160)

Annie Whitty 1 2 , Kylie Dunning 1 3 , Karen Kind 2 4 , Nicole Mcpherson 1 5 , Mark Nottle 1
  1. The University of Adelaide, School of Biomedicine, & Robinson Research Institute, Adelaide Medical School, The University of Adelaide, Adelaide, SOUTH AUSTRALIA, Australia
  2. The Davies Livestock Research Centre, The University of Adelaide, Adelaide, SA, Australia
  3. Australian Research Council Centre of Excellence for Nanoscale BioPhotonics, Institute for Photonics and Advanced Sensing, Adelaide, South Australia, Australia
  4. School of Animal and Veterinary Sciences, The University of Adelaide, Adelaide, South Australia, Australia
  5. Freemason Centre for Male Health and Wellbeing, Adelaide, South Australia, Australia

Sperm cryopreservation is essential for cattle in vitro embryo production (IVP). However, many sperm are rendered unviable post-thaw [1]. Granulocyte macrophage colony stimulating factor (GM-CSF) is present in seminal plasma and the female reproductive tract and its receptors are found on bovine sperm [2-4]. Whether GM-CSF can improve the health of post-thaw bovine sperm is unknown. The present study aimed to determine if in vitro addition of GM-CSF to frozen/thawed bovine sperm could improve sperm function, fertilisation and embryo development outcomes following IVP.

 

Thawed bovine sperm (N=4 bulls/4reps) were treated with 0, 0.1, 1, 2 or 10 ng/ml of recombinant bovine GM-CSF at 38.5C for 45 min and assessed for motility on CASA and quality parameters: glucose uptake, mitochondrial ROS, intracellular Ca+, and mitochondrial membrane potential (MMP) by flow cytometry. Capacitation as measured by chlortetracycline staining, and DNA integrity (HALO sperm) were examined following 5 h. IVP was performed on in vitro matured oocytes (N=200/5rep/group), fertilised with post-thaw bovine sperm treated with 2 ng/ml GM-CSF. Presumptive zygotes were cultured in 6% CO2; 7% O2; N2 balance, and day 8 blastocyst development examined. Cell numbers were determined by differential staining.

 

GM-CSF (2 and 10 ng/ml) increased progressive and rapid motility (10%), and glucose uptake (13%), while 1, 2, and 10 ng increased capacitation (17%) of post-thaw treated sperm (P<0.05), but had no effect on mitochondrial ROS, intracellular Ca+, MMP or DNA integrity. Oocytes fertilised with treated sperm had increased fertilisation rates (69.6 vs 81.7, P=0.006), hatching blastocyst rates (5.6% vs 7.8%, P= 0.048), and blastocyst inner cell mass numbers (35.3 vs 41.8, P=0.026) compared with control.

 

Our data suggest that GM-CSF treatment of frozen/thawed bovine sperm increases sperm function and improves IVP embryo quality and could be a useful addition to cattle IVP protocols to increase embryo yield and quality.   

 

 

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