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), with the maintenance of a proper level of cAMP, could improve the developmental capability of oocytes. The aim of this study was to assess: 1) dose-dependent effects of cilostamide on nuclear maturation kinetics, 2) the relationship between treatments on GJC functionality and large-scale chromatin configuration changes, and 3) the impact of treatments on developmental competence acquisition after parthenogenetic activation (PA) and SCNT. Accordingly, cumulus-oocyte complexes were collected from 3- to 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 for up to 24 h of culture and delayed meiotic resumption, as only 25.6% of cilostamide-treated oocytes reached the pro-metaphase I 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.
Avantea, Laboratory of Reproductive Technologies, Cremona, Italy.
Cecilia
C
Dieci
Valentina
V
Lodde
Federica
F
Franciosi
Irina
I
Lagutina
Irene
I
Tessaro
Silvia C
SC
Modina
David F
DF
Albertini
Giovanna
G
Lazzari
Cesare
C
Galli
Alberto M
AM
Luciano
D000818:Animals; D002450:Cell Communication / Q000187:drug effects*; D002478:Cells, Cultured; D042002:Chromatin Assembly and Disassembly / Q000187:drug effects*; D054885:Cumulus Cells / Q000187:drug effects / Q000502:physiology; D004305:Dose-Response Relationship, Drug; D005260:Female; D017629:Gap Junctions / Q000187:drug effects*; D008540:Meiosis / Q000187:drug effects; D053652:Nuclear Transfer Techniques / Q000662:veterinary*; D009865:Oocytes / Q000187:drug effects* / Q000502:physiology; D009866:Oogenesis / Q000187:drug effects; D010312:Parthenogenesis / Q000187:drug effects* / Q000502:physiology; D015363:Quinolones / Q000494:pharmacology*; D013552:Swine / Q000502:physiology*
D015363:Quinolones; C021294:cilostamide
PDE3; SCNT; chromatin; cumulus cells; cyclic adenosine monophosphate (cAMP); embryonic development; gamete biology; gap junctions; meiosis; oocyte; parthenogenetic activation; phosphodiesterases; porcine/pig