The current research was conducted to explore the in vitro developmental outcome and cytological/molecular quality of porcine nuclear-transferred (NT) embryos reconstituted with adult bone marrow-derived mesenchymal stem cells (ABM-MSCs) that were epigenetically transformed by treatment with nonspecific inhibitor of histone deacetylases, known as trichostatin A (TSA). The cytological quality of cloned blastocysts was assessed by estimation of the total cells number (TCN) and apoptotic index. Their molecular quality was evaluated by real-time PCR-mediated quantification of gene transcripts for pluripotency- and multipotent stemness-related markers (Oct4, Nanog, and Nestin). The morula and blastocyst formation rates of NT embryos derived from ABM-MSCs undergoing TSA treatment were significantly higher than in the TSA-unexposed group. Moreover, the NT blastocysts generated using TSA-treated ABM-MSCs exhibited significantly higher TCN and increased pluripotency extent measured with relative abundance of Oct4 and Nanog mRNAs as compared to the TSA-untreated group. Altogether, the improvements in morula/blastocyst yields and quality of cloned pig embryos seem to arise from enhanced abilities for promotion of correct epigenetic reprogramming of TSA-exposed ABM-MSC nuclei in a cytoplasm of reconstructed oocytes. To our knowledge, we are the first to report the successful production of mammalian high-quality NT blastocysts using TSA-dependent epigenomic modulation of ABM-MSCs.

The present study sought to examine whether trichostatin A (TSA)‐assisted epigenetic transformation of porcine bone marrow (BM)‐derived mesenchymal stem cells (BM‐MSCs) affects the transcriptional activities of pluripotency‐related genes (Oct4, Nanog, c‐Myc, Sox2 and Rex1), multipotent stemness‐related gene (Nestin) and anti‐apoptotic/anti‐senescence‐related gene (Survivin). Epigenetically transformed or non‐transformed BM‐MSCs that had been transcriptionally profiled by qRT‐PCR and had been analysed for different stages of apoptosis progression provided a source of nuclear donor cells for the in vitro production of cloned pig embryos. TSA‐mediated epigenomic modulation has been found to enhance the multipotency extent, stemness and intracellular anti‐ageing properties of porcine BM‐MSCs. This has been confirmed by the relative abundances for Nanog, c‐Myc Rex1, Sox2 and Survivin mRNAs in TSA‐exposed BM‐MSCs that turned out to be significantly higher than those of TSA‐unexposed BM‐MSCs. Additionally, TSA‐assisted epigenomic modulation of BM‐MSCs did not impact the caspase‐8 activity, Bax protein expression and the incidence of TUNEL‐positive cells. In conclusion, the considerably elevated quantitative profiles of Sox2, Rex1, c‐Myc, Nanog and Survivin mRNA transcripts seem to trigger improved reprogrammability of TSA‐treated BM‐MSC nuclei in cloned pig embryos that thereby displayed remarkably increased blastocyst formation rates as compared to those noticed for embryos derived from TSA‐untreated BM‐MSCs.

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Growth and development traits are economically important in animal production, especially in pig breeding. Therefore, the porcine GHRL gene is considered as a candidate gene responsible for growth rate and body weight. The aim of our study was to identify new polymorphisms in the GHRL gene in pig. Ten novel single nucleotide polymorphisms (SNP's) were detected: four substitutions in exons, four in introns and two mutations in promoter region. We evaluated the GHRL mRNA abundance in porcine stomachs (fundus ventriculis) and ghrelin protein concentration in plasma in three breeds: Polish Landrace, Polish Large White and Pietrain. The results showed that transcript abundance of GHRL gene was significantly higher in Polish Landrace than in other two breeds (P<0.05). The mutation c.-93A>G located in the promoter region affected expression of the GHRL gene. The AA genotype animals showed a significantly (P<0.05) higher expression level when compared to AG genotype animals.

The objective of the present study was to evaluate the effect of hyaluronan (HA) during IVM on meiotic maturation, embryonic development, and the quality of oocytes, granulosa cells (GC), and obtained blastocysts. COCs were matured in vitro in control medium and medium with additional 0.035% or 0.07% of exogenous HA. The meiotic maturity did not differ between the analysed groups. The best rate and the highest quality of obtained blastocysts were observed when 0.07% HA was used. A highly significant difference (P < 0.001) was noted in the mean number of apoptotic nuclei per blastocyst and in the DCI between the 0.07% HA and the control blastocysts (P < 0.01). Our results suggest that addition of 0.035% HA and 0.07% HA to oocyte maturation media does not affect oocyte nuclear maturation and DNA fragmentation. However, the addition of 0.07% HA during IVM decreases the level of blastocysts DNA fragmentation. Finally, our results suggest that it may be risky to increase the HA concentration during IVM above 0.07% as we found significantly higher Bax mRNA expression levels in GC cultured with 0.07% HA. The final concentration of HA being supplemented to oocyte maturation media is critical for the success of the IVP procedure.