Pig oocyte activation using a Zn²⁺ chelator, TPEN

Area of Science:

• Reproductive Biology
• Developmental Biology
• Biochemistry

Background:

• Somatic cell nuclear transfer (SCNT) requires efficient artificial oocyte activation.
• Current methods often rely on increasing intracellular calcium (Ca2+).
• The role of zinc (Zn2+) in oocyte activation is less understood.

Purpose of the Study:

• To investigate the potential of TPEN, a zinc chelator, to enhance oocyte activation during SCNT.
• To determine if TPEN-mediated oocyte activation is independent of intracellular Ca2+ increase.
• To assess the developmental efficiency of cloned embryos using TPEN-assisted activation.

Main Methods:

• Treatment of mature pig oocytes with TPEN at various concentrations and durations.
• Assessment of oocyte activation efficiency compared to conventional methods.
• Evaluation of embryo development to the blastocyst stage after TPEN treatment.
• Production of cloned piglets via SCNT using TPEN-assisted activation.

Main Results:

• TPEN treatment reduced available Zn2+ in oocytes but did not affect cytosolic Ca2+ levels.
• TPEN alone did not improve activation efficiency compared to conventional methods.
• Co-incubation with low TPEN concentrations (5-10 μM) significantly increased blastocyst formation.
• TPEN (100-200 μM) combined with a small Ca2+ stimulus significantly enhanced blastocyst rates.
• Cloned piglets were produced without observable abnormalities, indicating TPEN's efficacy.

Conclusions:

• TPEN can serve as a key reagent for oocyte activation in SCNT.
• TPEN-assisted activation enhances cloned embryo developmental potential independently of Ca2+.
• This novel approach offers an efficient method for improving SCNT outcomes.