The Effect of Cilostamide on Gap Junction Communication Dynamics, Chromatin Remodeling, and Competence Acquisition in Pig Oocytes Following Parthenogenetic Activation and Nuclear Transfer

In the pig, the efficiency of in vitro embryo production and somatic cell nuclear transfer (SCNT) procedures remains limited. It has been suggested that prematuration treatments (pre-IVM) based on the prolongation of a patent bidirectional crosstalk between the oocyte and the cumulus cells through gap junction mediate communication (GJC), together with the maintenance of a proper level of cAMP, could improve the developmental capability of oocytes. The aim of this study was to assess: (i) dose dependent effects of cilostamide on nuclear maturation kinetics; (ii) the relationship between treatments on GJC functionality and large-scale chromatin configuration changes; (iii) and the impact of treatments on developmental competence acquisition after parthenogenic activation (PA) and SCNT. Accordingly, COC were collected from 3-6 mm antral follicles and cultured for 24 h in defined culture medium with or without 1 μM cilostamide. GJC functionality was assessed by Lucifer Yellow microinjection, while chromatin configuration was evaluated by fluorescence microscopy after nuclear staining. Cilostamide administration sustained functional coupling up to 24 h of culture and delayed meiotic resumption as only 25.6% of cilostamide-treated oocytes reached ProMI stage compared to the control (69.7%; P<0.05). Moreover, progressive chromatin condensation was delayed before meiotic resumption based upon G2/M biomarker phosphoprotein epitope acquisition using immunolocalization. Importantly, cilostamide treatment under these conditions, improved oocyte developmental competence as reflected in higher blastocyst quality after both parthenogenetic activation and SCNT