Creating ex situ protected bioreservoirs as a powerful strategy for the reproductive biotechnology-mediated rescue of threatened Polish livestock breeds

The current article presents the state of the art of the creation of bioreservoirs of cryopreserved somatic and stem cell lines and cryopreserved or lyophilized germplasm-based resources of selected farm animal species. It also presents the agricultural, biological and biotechnological determinants, and showcases the importance of the National Research Institute of Animal Production (NRIAP) in Poland in this process. The aforementioned bioreservoirs serve as an innovative research tool used for cryogenically or lyophilizogenically assisted and species-specific ex situ conservation. The latter ensures, in the long term, not only restitution, but also perpetuation of sustainable biodiversity that underlies genotypic and phenotypic, intra- and inter-population variability within pure-breeding herds of the national conserved livestock breeds

Anabolic steroids activate the NF-KB pathway in porcine ovarian putative stem cells independently of the ZIP-9 receptor

Boldenone (Bdn) and nandrolone (Ndn) are anabolic androgenic steroids (AASs) that, as our previous studies have shown, may increase the risk of neoplastic transformation of porcine ovarian putative stem cells (poPSCs). The NF-κB pathway may be important in the processes of carcinogenesis and tumour progression. Therefore, in this work, we decided to test the hypothesis of whether Bdn and Ndn can activate the NF-κB pathway by acting through the membrane androgen receptor ZIP-9. For this purpose, the expression profiles of both genes involved in the NF-κB pathway and the gene coding for the ZIP-9 receptor were checked. The expression and localization of proteins of this pathway in poPSCs were also examined. Additionally, the expression of the ZIP-9 receptor and the concentration of the NF-κB1 and 2 protein complex were determined. Activation of the NF-κB pathway was primarily confirmed by an increase in the relative abundances of phosphorylated forms of RelA protein and IκBα inhibitor. Reduced quantitative profiles pinpointed not only for genes representing this pathway but also for unphosphorylated proteins, and, simultaneously, decreased concentration of the NF-κB1 and 2 complex may indicate post-activation silencing by negative feedback. However, the remarkably and sustainably diminished expression levels noticed for the SLC39A9 gene and ZIP-9 protein suggest that this receptor does not play an important role in the regulation of the NF-κB pathway

Molecular Mechanisms of Somatic Cell Cloning and Other Assisted Reproductive Technologies in Mammals: Which Determinants Have Been Unraveled Thus Far?—Current Status, Further Progress and Future Challenges

Taking into consideration recent reports on the successful creation of cloned rhesus monkeys [...

Molecular Mechanism and Application of Somatic Cell Cloning in Mammals—Past, Present and Future

Thus far, nearly 25 mammalian species have been cloned by intra- or interspecies somatic cell nuclear transfer (SCNT) [...

Generating Cloned Goats by Somatic Cell Nuclear Transfer—Molecular Determinants and Application to Transgenics and Biomedicine

The domestic goat (Capra aegagrus hircus), a mammalian species with high genetic merit for production of milk and meat, can be a tremendously valuable tool for transgenic research. This research is focused on the production and multiplication of genetically engineered or genome-edited cloned specimens by applying somatic cell nuclear transfer (SCNT), which is a dynamically developing assisted reproductive technology (ART). The efficiency of generating the SCNT-derived embryos, conceptuses, and progeny in goats was found to be determined by a variety of factors controlling the biological, molecular, and epigenetic events. On the one hand, the pivotal objective of our paper was to demonstrate the progress and the state-of-the-art achievements related to the innovative and highly efficient solutions used for the creation of transgenic cloned does and bucks. On the other hand, this review seeks to highlight not only current goals and obstacles but also future challenges to be faced by the approaches applied to propagate genetically modified SCNT-derived goats for the purposes of pharmacology, biomedicine, nutritional biotechnology, the agri-food industry, and modern livestock breeding.</jats:p

Extranuclear Inheritance of Mitochondrial Genome and Epigenetic Reprogrammability of Chromosomal Telomeres in Somatic Cell Cloning of Mammals

The effectiveness of somatic cell nuclear transfer (SCNT) in mammals seems to be still characterized by the disappointingly low rates of cloned embryos, fetuses, and progeny generated. These rates are measured in relation to the numbers of nuclear-transferred oocytes and can vary depending on the technique applied to the reconstruction of enucleated oocytes. The SCNT efficiency is also largely affected by the capability of donor nuclei to be epigenetically reprogrammed in a cytoplasm of reconstructed oocytes. The epigenetic reprogrammability of donor nuclei in SCNT-derived embryos appears to be biased, to a great extent, by the extranuclear (cytoplasmic) inheritance of mitochondrial DNA (mtDNA) fractions originating from donor cells. A high frequency of mtDNA heteroplasmy occurrence can lead to disturbances in the intergenomic crosstalk between mitochondrial and nuclear compartments during the early embryogenesis of SCNT-derived embryos. These disturbances can give rise to incorrect and incomplete epigenetic reprogramming of donor nuclei in mammalian cloned embryos. The dwindling reprogrammability of donor nuclei in the blastomeres of SCNT-derived embryos can also be impacted by impaired epigenetic rearrangements within terminal ends of donor cell-descended chromosomes (i.e., telomeres). Therefore, dysfunctions in epigenetic reprogramming of donor nuclei can contribute to the enhanced attrition of telomeres. This accelerates the processes of epigenomic aging and replicative senescence in the cells forming various tissues and organs of cloned fetuses and progeny. For all the above-mentioned reasons, the current paper aims to overview the state of the art in not only molecular mechanisms underlying intergenomic communication between nuclear and mtDNA molecules in cloned embryos but also intrinsic determinants affecting unfaithful epigenetic reprogrammability of telomeres. The latter is related to their abrasion within somatic cell-inherited chromosomes

Extranuclear Inheritance of Mitochondrial Genome and Epigenetic Reprogrammability of Chromosomal Telomeres in Somatic Cell Cloning of Mammals

The effectiveness of somatic cell nuclear transfer (SCNT) in mammals seems to be still characterized by the disappointingly low rates of cloned embryos, fetuses, and progeny generated. These rates are measured in relation to the numbers of nuclear-transferred oocytes and can vary depending on the technique applied to the reconstruction of enucleated oocytes. The SCNT efficiency is also largely affected by the capability of donor nuclei to be epigenetically reprogrammed in a cytoplasm of reconstructed oocytes. The epigenetic reprogrammability of donor nuclei in SCNT-derived embryos appears to be biased, to a great extent, by the extranuclear (cytoplasmic) inheritance of mitochondrial DNA (mtDNA) fractions originating from donor cells. A high frequency of mtDNA heteroplasmy occurrence can lead to disturbances in the intergenomic crosstalk between mitochondrial and nuclear compartments during the early embryogenesis of SCNT-derived embryos. These disturbances can give rise to incorrect and incomplete epigenetic reprogramming of donor nuclei in mammalian cloned embryos. The dwindling reprogrammability of donor nuclei in the blastomeres of SCNT-derived embryos can also be impacted by impaired epigenetic rearrangements within terminal ends of donor cell-descended chromosomes (i.e., telomeres). Therefore, dysfunctions in epigenetic reprogramming of donor nuclei can contribute to the enhanced attrition of telomeres. This accelerates the processes of epigenomic aging and replicative senescence in the cells forming various tissues and organs of cloned fetuses and progeny. For all the above-mentioned reasons, the current paper aims to overview the state of the art in not only molecular mechanisms underlying intergenomic communication between nuclear and mtDNA molecules in cloned embryos but also intrinsic determinants affecting unfaithful epigenetic reprogrammability of telomeres. The latter is related to their abrasion within somatic cell-inherited chromosomes.</jats:p

Assessment of in Vitro Developmental Capacity of Porcine Nuclear-Transferred Embryos Reconstituted with Cumulus Oophorus Cells Undergoing Vital Diagnostics for Apoptosis Detection / Ocena zdolności rozwojowych in vitro klonalnych zarodków świni rekonstytuowanych z jąder komórek wzgórka jajonośnego poddawanych przyżyciowej diagnostyce w kierunku wykrywania apoptozy

Abstract The objective of the current investigation was to extensively compare the in vitro developmental capabilities between cloned pig embryos reconstructed with the cell nuclei of either cumulus oophorus cells or adult dermal fibroblast cells that were both evaluated as non-apoptotic on the basis of YO-PRO-1- and Annexin V-eGFP-mediated vital analysis for programmed cell suicide. In Group I, the competences of nuclear-transferred (NT) embryos that were derived from non-apoptotic/ non-necrotic (i.e., YO-PRO-1- and Annexin V-eGFP-negative) cumulus cells to complete their development to the morula and blastocyst stages were maintained at the proportions of 155/364 (42.6%) and 54/364 (14.8%), respectively. In Group II, NT embryos that were reconstituted with non-apoptotic and/or non-necrotic adult cutaneous fibroblast cells developed to the morula and blastocyst stages at the rates of 207/358 (57.8%) and 110/358 (30.7%), respectively. Although the in vitro developmental potential of porcine NT embryos derived from non-apoptotic/non-necrotic cumulus cells was significantly lower (P&lt;0.001) than that of NT embryos reconstructed with adult dermal fibroblast cells, the obtained morula/blastocyst formation rates turned out to be considerably higher as compared to the rates reported by other investigators. Altogether, to our knowledge, the comprehensive research aimed at the determination of preimplantation developmental outcomes of cloned pig embryos produced using nuclear donor somatic cells of different provenance (cumulus oophorus cells or adult cutaneous fibroblast cells) that were vitally diagnosed for the lack of proapoptotic transformations in their plasma membranes has not yet been accomplished.</jats:p