ArticleAnimal bioscience2026
Lysophosphatidic acid-mediated enhancement of porcine oocyte chemical enucleation for somatic cell nuclear transfer.
Article in Animal bioscience, 2026. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Not yet cited in PubMed.
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Abstract
objectiveProtocols for somatic cell nuclear transfer (SCNT) remain technically challenging and inefficient. Demecolcine and nocodazole (Noc) have been used for chemically assisted enucleation to improve cloning efficiency; however, the procedure continues to be labor-intensive. This study aimed to use lysophosphatidic acid (LPA) to enhance Noc-mediated chemical enucleation (CE), thereby simplifying the SCNT procedure.
methodsPorcine MII oocytes were subject to CE by incubation with Noc and various concentrations of LPA for 1 h. Actin signals were evaluated to determine the optimal LPA. After treatment with Noc and 25 μM LPA for 1 h, spindle reformation was assayed following subsequent washout. Noc/LPA CE-treated oocytes were used for SCNT via direct microinjection of donor cells into the ooplasm. Reconstructed embryos were recovered for 2 h in PZM-5 prior to activation with ionomycin and N,N,N',N'-tetrakis (2-pyridylmethyl) ethylenediamine. Activated reconstructed embryos were cultured in 6-DMAP for 4 h, then in chaetocin for an additional 20-24 h to promote demethylation and acetylation. Cleavage and blastocyst rates were evaluated on Day 2 and Day 6, respectively.
resultsThe culture of the matured oocytes in Noc/LPA for 1 h, the percentage of oocytes exhibiting actin signals increased significantly from 12.5% to 87.6% with the addition of 25 μM LPA. Although spindles reformed in 59.0% of oocytes 1 h after Noc/LPA washout, 83.9% of oocytes retained spindles enclosed by actin rings; enabling the direct microinjection of donor cells into the ooplasm within 30 min. Blastocyst development rates currently vary from 9.5% to 16.6% after 6 days of culture in PZM-5.
conclusionAlthough the blastocyst rate requires further improvement, we have shown that enucleation of MII pig oocytes can be successfully achieved via Noc/LPA treatment. This protocol simplifies the SCNT procedure while ensuring consistent blastocyst development in pig in vitro maturation oocytes.
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