Zinc Regulates Meiotic Resumption in Porcine Oocytes via a Protein Kinase C-Related Pathway

Zhao, Minghui; Kwon, Jin-Sook; Liang, Shuang; Kim, Seon-Hyang; Li, Yinghua; Oh, Jeill; Kim, Nam‐Hyung; Cui, Xiang‐Shun

doi:10.1371/journal.pone.0102097

Cited by 20 publications

(17 citation statements)

References 39 publications

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“…The amount of Zn 2+ in the oocyte increases during maturation and can regulate the timing of meiotic exit in mammalian oocytes [18]. This increase in the level of Zn 2+ during oocyte maturation has also been reported in pigs [30]. It was suggested that the release of Zn 2+ from FBXO43, a cytostatic factor with a Zn 2+ binding domain, relieves APC/C from FBXO43-mediated inhibition, thus initiating the degradation of MPF [20].…”

Section: Discussionmentioning

confidence: 95%

Pig oocyte activation using a Zn2+ chelator, TPEN

et al. 2015

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Artificial oocyte activation is a critical step during somatic cell nuclear transfer (SCNT). Most of current activation protocols focus on inducing an increase in the intracellular free Ca2+ concentration of the oocyte. Here, we have utilized a zinc chelator, TPEN, to enhance the efficiency of oocyte activation during SCNT. TPEN treatment of matured pig oocytes resulted in the reduction of available Zn2+ in pig oocytes; however, the cytosolic Ca2+ concentration in the oocytes was not affected by the TPEN treatment. When various concentrations (100 – 250 μM) and incubation durations (45 min – 2.5 hours) of TPEN were used to activate oocytes, the efficiency of oocyte activation was not different from conventional activation methods. When oocytes that were activated by conventional activation methods were incubated with a lower concentration of TPEN (5 – 10 μM), a significant increase in embryos developing to the blastocyst stage was observed. In addition, when oocytes receiving a small Ca2+ stimulus were further activated by higher concentration of TPEN (100 – 200 μM), a significant increase in the frequency of blastocyst formation was observed, compared to a conventional activation method. This result indicated that TPEN can be a main reagent in oocyte activation. No increase in the cytosolic Ca2+ level was detected when oocytes were exposed to various concentrations of TPEN, indicating the ability of TPEN to induce oocyte activation is independent of an intracellular Ca2+ increase. We were able to produce clones through SCNT using the TPEN-assisted activation procedure, and the piglets produced through the process did not show any signs of abnormality. In this study, we have developed an efficient way to utilize TPEN to increase the developmental potential of cloned embryos.

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Section: Discussionmentioning

confidence: 95%

Pig oocyte activation using a Zn2+ chelator, TPEN

et al. 2015

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“…Follicles that were 3–6 mm in diameter were aspirated. Cumulus-oocyte complexes (COCs) that were surrounded by a minimum of three cumulus cells were selected for culture [ 27 ]. In brief, the COCs were washed three times in TL-HEPES supplemented with 0.1% polyvinyl alcohol (PVA, w/v) and 0.05 g/L gentamycin.…”

Section: Methodsmentioning

confidence: 99%

Scriptaid Treatment Decreases DNA Methyltransferase 1 Expression by Induction of MicroRNA-152 Expression in Porcine Somatic Cell Nuclear Transfer Embryos

et al. 2015

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Abnormal epigenetic reprogramming of donor nuclei after somatic cell nuclear transfer (SCNT) is thought to be the main cause of low cloning efficiencies. A growing body of evidence has demonstrated a positive role of Scriptaid, a histone deacetylase inhibitor (HDACi) that belongs to an existing class of hydroxamic acid-containing HDACis, on the development competence of cloned embryos in many species. The present study investigated the effects of Scriptaid on the development of porcine SCNT embryos in vitro and its mechanism. Treatment with 300 or 500 nM Scriptaid for 20 h after activation significantly increased the percentage of SCNT embryos that developed to the blastocyst stage and the total number of cells per blastocyst and significantly decreased the percentage of apoptotic cells in blastocysts. Scriptaid treatment significantly increased the level of histone H3 acetylated at K9 and the conversion of 5-methylcytosine into 5-hydroxymethylcytosine and significantly decreased the level of histone H3 trimethylated at K9 at the pronuclear stage. As a potential mechanism for the DNA methylation changes, our results showed that the expression of DNA methyltransferase 1 was frequently down-regulated in Scriptaid-treated embryos in comparison with untreated embryos and was inversely correlated to endogenous microRNA-152 (miR-152). Taken together, these findings illustrated a crucial functional crosstalk between miR-152 and DNMT1. Meanwhile, mRNA and protein levels of POU5F1 and CDX2 were increased in Scriptaid-treated embryos. mRNA levels of Caspase3, and Bax were significantly decreased and that of Bcl-xL was significantly increased in Scriptaid-treated embryos. In conclusion, these observations would contribute to uncover the nuclear reprogramming mechanisms underlying the effects of Scriptaid on the improvement of porcine SCNT embryos.

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“…mRNA extraction and cDNA synthesis were performed as previously described [ 28 ]. Briefly, mRNA was extracted from 10 blastocysts using Dynabeads mRNA Direct Kit (Dynal Biotech ASA, Oslo, Norway) followed by routine cDNA synthesis by reverse transcription (RT) of RNA using an oligo(dT) 12–18 primer and SuperScript Reverse Transcriptase (Invitrogen) following the manufacturer’s instructions.…”

Section: Methodsmentioning

confidence: 99%

Fe(III) Is Essential for Porcine Embryonic Development via Mitochondrial Function Maintenance

et al. 2015

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Iron is an important trace element involved in several biological processes. The role of iron in porcine early embryonic development remains unknown. In the present study, we depleted iron (III, Fe3+) with deferoxamine (DFM), a specific Fe3+ chelator, in cultured porcine parthenotes and monitored embryonic development, apoptosis, mitochondrial membrane potential, and ATP production. Results showed biphasic function of Fe3+ in porcine embryo development. 0.5 μM DFM obviously increased blastocyst formation (57.49 ± 2.18% vs. control, 43.99 ± 1.72%, P < 0.05) via reduced (P < 0.05) production of reactive oxygen species (ROS), further increased mitochondrial membrane potential and ATP production in blastocysts (P < 0.05). 0.5 μM DFM decreased mRNA expression of Caspase 3 (Casp3) and increased Bcl-xL. However, results showed a significant reduction in blastocyst formation in the presence of 5.0 μM DFM compared with the control group (DFM, 21.62 ± 3.92% vs. control, 43.99 ± 1.73%, P < 0.05). Fe3+ depletion reduced the total (DFM, 21.10 ± 8.78 vs. control, 44.09 ± 13.65, P < 0.05) and increased apoptotic cell number (DFM, 11.10 ± 5.24 vs. control, 2.64 ± 1.43, P < 0.05) in the blastocyst. An obvious reduction in mitochondrial membrane potential and ATP level after 5.0 μM DFM treatment was observed. Co-localization between mitochondria and cytochrome c was reduced after high concentration of DFM treatment. In conclusion, Fe3+ is essential for porcine embryonic development via mitochondrial function maintenance, but redundant Fe3+ impairs the function of mitochondria.

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Zinc Regulates Meiotic Resumption in Porcine Oocytes via a Protein Kinase C-Related Pathway

Cited by 20 publications

References 39 publications

Pig oocyte activation using a Zn2+ chelator, TPEN

Pig oocyte activation using a Zn2+ chelator, TPEN

Scriptaid Treatment Decreases DNA Methyltransferase 1 Expression by Induction of MicroRNA-152 Expression in Porcine Somatic Cell Nuclear Transfer Embryos

Fe(III) Is Essential for Porcine Embryonic Development via Mitochondrial Function Maintenance

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