The Molecular Quality and Mitochondrial Activity of Porcine Cumulus–Oocyte Complexes Are Affected by Their Exposure to Three Endocrine-Active Compounds under 3D In Vitro Maturation Conditions

Abstract

Thus far, the potential short- and long-term detrimental effects of a variety of environmental chemicals designated as endocrine-active compounds (EACs) have been found to interfere with histo- and anatomo-physiological functions of the reproductive system in humans and wildlife species. For those reasons, this study sought to examine whether selected EACs, which encompass the fungicide vinclozolin (Vnz), the androgenic anabolic steroid nandrolone (Ndn) and the immunosuppressant cyclosporin A (CsA), affect the developmental competence and molecular quality (MQ) of porcine cumulus–oocyte complexes (COCs) subjected to in vitro maturation (IVM) under 3D culture conditions. The COCs underwent 3D-IVM in the presence of Vnz, Ndn or CsA for 48 h. To explore whether the selected EACs induce internucleosomal DNA fragmentation in cumulus cells (CCs), TUNEL-assisted detection of late apoptotic cells was performed. Additionally, for the detailed evaluation of pro- and antiapoptotic pathways in COCs, apoptosis proteome profiler arrays were used. To determine changes in intracellular metabolism in COCs, comprehensive assessments of mitochondrial ultrastructure and activity were carried out. Moreover, the relative abundances (RAs) of mRNAs transcribed from genes that are involved in scavenging reactive oxygen species (ROS), such as SIRT3 and FOXO3, and intramitochondrial bioenergetic balance, such as ATP synthase subunit (ATP5A1), were ascertained. Finally, to investigate the extent of progression of oocyte maturation, the intraooplasmic levels of cAMP and the RAs of mRNA transcripts encoding regulatory and biocatalytic subunits of a heterodimeric meiosis-promoting factor, termed cyclin B1 (CCNB1) and cyclin-dependent kinase 1 (CDC2), were also estimated. The obtained results provide, for the first time, strong evidence that both Vnz and Ndn decrease the developmental competence of oocytes and stimulate apoptosis processes in CCs. The present study is also the first to highlight that Vnz accelerates the maturation process in immature oocytes due to both increased ROS production and the augmented RA of the CCNB1 gene. Furthermore, Vnz was proven to trigger proapoptotic events in CCs by prompting the activity of the FOXO3 transcription factor, which regulates the mitochondrial apoptosis pathway. In turn, Ndn was shown to inhibit oocyte maturation by inducing molecular events that ultimately lead to an increase in the intraooplasmic cAMP concentration. However, due to the simultaneous enhancement of the expression of TNF-β and HSP27 proteins in CCs, Ndn might be responsible for the onset of their neoplastic transformation. Finally, our current investigation is the first to clearly demonstrate that although CsA did not interfere with the nuclear and cytoplasmic maturation of oocytes, by inducing mitophagy in CCs, it disrupted oocyte metabolism, consequently attenuating the parameters related to the MQ of COCs. Summing up, Vnz, Ndn and CsA reduced not only the processes of growth and IVM but also the MQ of porcine COCs, which might make them unsuitable for assisted reproductive technologies (ARTs) such as in vitro fertilization by either gamete co-incubation or intracytoplasmic sperm injection (ICSI) and cloning by somatic cell nuclear transfer (SCNT).

Combination of CNP, MT and FLI during IVM Significantly Improved the Quality and Development Abilities of Bovine Oocytes and IVF-Derived Embryos

“…Elucidating the importance of CNP, MT, and FLI for cytobiochemical and molecular balance of intracellular anti-and pro-oxidative processes in IVM-derived heifer/cow oocytes might be helpful for elaborating a variety of biotechnological and embryological parameters. The combination of these parameters might result in the efficient generation of bovine and other mammalian high-quality embryos not only by standard IVF [77,78] but also by intracytoplasmic sperm injection (ICSI) [79,80] or by somatic cell nuclear transfer (SCNT) [81,82]. Informed Consent Statement: Not applicable.…”

Effects of Heat Stress on Bovine Oocytes and Early Embryonic Development—An Update

“…Such a situation leads to disturbances in the energy metabolism in oocytes and embryos, which fundamentally impacts their developmental competence. Finally, precisely recognizing the biological factors responsible for the heat stress-mediated induction of impairments in reproductive physiology and endocrinology in cattle and other mammalian species might enable the multifaceted estimation of genomic, epigenomic, transcriptomic and proteomic profiles in the embryos, fetuses and offspring created and multiplied by such novel assisted reproductive technologies and in vitro embryo production strategies as the ex vivo fertilization of oocytes, with the aid of either classic penetration by capacitated spermatozoa [ 91 , 92 ] or intra-cytoplasmic sperm injection [ 93 , 94 ] and somatic cell cloning [ 95 , 96 ]. The harmful effects of heat stress on cow reproduction are of key importance to the economic status of the farm, producing financial losses every year.…”

Effects of IGF-1 on the Three-Dimensional Culture of Ovarian Preantral Follicles and Superovulation Rates in Mice

“…The results of the current research have proven that designing modern and powerful approaches to the murine ex vivo 3D model of IGF-1-assisted growth and maturation of ovarian secondary preantral follicles in order to obtain satisfactory outcomes of IVM might be reliable and feasible for the purposes of achieving high efficiency rates in such novel ARTs as IVF/ICSI in humans and other mammalian species [ 65 , 66 , 67 , 68 ] or somatic cell cloning in other mammalian species [ 69 , 70 , 71 , 72 ].…”