Birth of piglets after transferring of in vitro-produced embryos pre-matured with R-roscovitine

Coy, Pilar; Romar, Raquel; Ruiz, Salvador; Cánovas, Sebastián; Gadea, J.; Vázquez, Francisco García; Matás, Carmen

doi:10.1530/rep.1.00691

Cited by 48 publications

(47 citation statements)

References 54 publications

(56 reference statements)

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“…At 18-20 hours post-insemination, putative zygotes were washed 10 times by aspiration through an automatic pipette and evaluated after fixation and staining (22). Penetration, monospermy, number of spermatozoa penetrating each oocyte, number of pronuclei and sperm decondensed heads inside each oocyte, and number of spermatozoa bound to ZP were assessed in each oocyte.…”

Section: Ivfmentioning

confidence: 99%

Timing of oviductal fluid collection, steroid concentrations, and sperm preservation method affect porcine in vitro fertilization efficiency

Ballester

Romero-Aguirregomezcorta

Soriano‐Úbeda

et al. 2014

Fertility and Sterility

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Section: Ivfmentioning

confidence: 99%

Timing of oviductal fluid collection, steroid concentrations, and sperm preservation method affect porcine in vitro fertilization efficiency

Ballester

Romero-Aguirregomezcorta

Soriano‐Úbeda

et al. 2014

Fertility and Sterility

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“…The ability of porcine oocytes to adequately develop after in vitro maturation (IVM) and fertilization (IVF), leading to live piglets, has been largely demonstrated by several laboratories (Kikuchi et al 2002;Yoshioka et al 2002Yoshioka et al , 2003Suzuki et al 2004Suzuki et al , 2006Coy et al 2005;Garcia-Rosello et al 2006a;Yong et al 2006;Maehara et al 2012;Kaneko et al 2013). Despite this fact, the efficiency of IVP of blastocysts after intracytoplasmic sperm injection (ICSI) still remains low ).…”

Section: Introductionmentioning

confidence: 99%

Male pronucleus formation after ICSI: effect of oocyte cysteine or sperm Triton X-100 treatments

García-Mengual¹,

Silvestre²,

Salvador³

et al. 2015

CZECH J ANIM SCI

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ABSTRACT:In pigs, intracytoplasmic sperm injection (ICSI) efficiency is still poor. The inadequate decondensation of the sperm chromatin, its transformation into the male pronucleus (MPN) together with the subsequent inability to activate the oocyte, seem to be the main causes of the low ICSI efficiency. In order to improve the MPN formation we took two different approaches. On the one hand, the in vitro culture (IVC) medium post-ICSI was supplemented with 1.71mM cysteine (CYS). Alternatively, the sperm membrane was digested with Triton X-100 (TX) before ICSI, to improve the exposure of the sperm chromatin to the oocyte cytoplasm. After 6 h post-ICSI, the activation rate was significantly higher in TX group (70.0%) compared with CYS and control groups (42.2% and 48.9%, respectively; P < 0.05). However, no significant differences between the three groups were observed in terms of the number of pronuclei, 2PN (oocytes with 2 pronuclei and no visible sperm), and 1PN + sperm (oocytes with 1 pronucleus and one sperm head). At 22 h post-ICSI, the activation rates were similar in TX, CYS, and control groups (73.1, 78.9, and 75.7%, respectively). In addition, we did not observe significant differences between TX, CYS, and control groups for the number of pronuclei, 2PN (52.6, 56.7, and 50%, respectively) or 1PN + sperm (21.1, 33.3, and 32.1%, respectively). While no cleavage was observed in the CYS group, no significant differences in the cleavage rate were observed between control (21.3%) and TX (10.5%) groups. In summary, and under our conditions, neither CYS supplement, nor sperm TX pre-treatment were able to improve MPN formation at 6 and 22 h post-ICSI. However, the sperm TX pre-treatment improved oocyte activation at 6 h post-ICSI, although 22 h post-ICSI such a beneficial effect did not persist.Keywords: porcine; assisted reproductive technology; pronuclear formation List of abbreviations: ICSI = intracytoplasmic sperm injection, PN = pronucleus, MPN = male pronucleus, IVC = in vitro culture, CYS = cysteine, TX = Triton X-100, IVP = in vitro production, IVM = in vitro maturation, IVF = in vitro fertilization, EGF = epidermal growth factor, GSH = glutathione, COC = cumulus oocyte complex, PZM-3 = porcine zygote medium-3, MM199 = maturation medium, HM199 = handling medium 242

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“…Most recently, Coy et al (2004) reported the birth of live piglets derived from roscovitine-treated oocytes.…”

Section: Introductionmentioning

confidence: 99%

Maintenance of meiotic arrest in bovine oocytes using the S-enantiomer of roscovitine: effects on maturation, fertilization and subsequent embryo development in vitro

Coy¹,

Romar²,

Payton³

et al. 2005

Reproduction

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The overall objective was to evaluate the effectiveness of the S-enantiomer of roscovitine (inhibitor of p34 cdc2 /cyclin B kinase)to maintain bovine cumulus -oocyte complexes at the germinal vesicle (GV) stage for extended times after removal from antral follicles without compromising subsequent maturation, fertilization and embryo development. Oocytes were cultured in 0, 12.5, 25 or 50 mmol/l S-roscovitine for 24 h. Hoechst staining showed that 50 mmol/l S-roscovitine maintained >90% of oocytes at the GV stage and inhibited gonadotropin-induced cumulus expansion. Fewer oocytes underwent nuclear maturation after in vitro maturation (Hoechst staining) when cultured in 50 mmol/l S-roscovitine for 66 versus 21 or 42 h. Zona pellucida (ZP) hardening (pronase resistance), cortical granule types (lens culinaris agglutinin -fluorescein isothiocyanate), nuclear maturation and fertilization with frozen-thawed spermatozoa (Hoechst staining) were assessed after culture of oocytes in 50 mmol/l S-roscovitine for 0, 24 or 48 h. Neither ZP hardening, nor nuclear maturation nor fertilization were altered by roscovitine culture for 48 h. A higher proportion of oocytes had a type III cortical granule pattern (premature translocation to the oolemma) after roscovitine culture for 48 h. However, embryo development was not compromised as cleavage, development to 8-16 cell and blastocyst stages were at least comparable in control and roscovitine-treated oocytes. In conclusion, the studies have shown that S-roscovitine reversibly maintained bovine oocytes at the GV stage for 48 h. However, maintenance of oocytes in static culture for 48 h was not sufficient to improve development above non-treated controls.

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Birth of piglets after transferring of in vitro-produced embryos pre-matured with R-roscovitine

Cited by 48 publications

References 54 publications

Timing of oviductal fluid collection, steroid concentrations, and sperm preservation method affect porcine in vitro fertilization efficiency

Timing of oviductal fluid collection, steroid concentrations, and sperm preservation method affect porcine in vitro fertilization efficiency

Male pronucleus formation after ICSI: effect of oocyte cysteine or sperm Triton X-100 treatments

Maintenance of meiotic arrest in bovine oocytes using the S-enantiomer of roscovitine: effects on maturation, fertilization and subsequent embryo development in vitro

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