High incidence of multiple aster formation in vitrified-warmed bovine oocytes after in vitro fertilization

In vitro-matured bovine oocytes do not tolerate vitrification as well as mature murine or human oocytes. Delayed first cleavage in vitrified and in vitro-fertilized bovine oocytes may be responsible for the decreased yield of blastocysts in vitro. Because formation of sperm-aster and the subsequent assembly of microtubule network play an important role for migration and fusion of both pronuclei, aster formation in vitrified-warmed oocytes was analyzed by confocal laser-scanning microscopy. At 10 h post-insemination (hpi), proportions of oocytes fertilized normally were comparable between the vitrified and fresh control groups (67 and 70%, respectively). Proportions of oocytes that exhibited microtubule assembly were similar between the two groups (95% each), but the proportion of oocytes with multiple asters was higher in the vitrified group when compared with the fresh control group (68 vs 29%, P < 0.05). Both migration and development of two pronuclei were adversely affected by multiple aster formation. In the next experiment, multiple asters observed in 5.5 vs 8 hpi pronuclear zygotes were located near the male pronucleus, suggesting that those multiple asters were not the cytoplasmic asters of maternal origin. In conclusion, multiple aster formation frequently observed in vitrified-warmed bovine oocytes may be related to loss of ooplasmic function responsible for normal microtubule assembly from the sperm-aster.ArticleTHERIOGENOLOGY. 77(5):908-915 (2012)journal articl

Stimulatory effect of Rho-associated coiled-coil kinase (ROCK) inhibitor on revivability of in vitro-produced bovine blastocysts after vitrification

Inhibition of Rho-associated kinase (ROCK) activity promoted recovery and growth of frozen-thawed human embryonic stem cells. The primary objective was to determine if a ROCK inhibitor (Y-27632) in post-thaw culture medium unproved revivability of vitrified IVP bovine blastocysts. Expanding or expanded blastocysts (7 d after IVF) were vitrified (minimum volume cooling procedure, using a Cryotop) in 15% ethylene glycol. 15% DMSO and 0 5 M sucrose When post-warm blastocysts were cultured in inSOF medium, survival rate (re-expansion of blastocoel at 24 h of culture) was improved (P < 0 05) by the addition of 10 mu M Y-27632 (94 9 +/- 2 4%, mean +/- SEM) compared to a control (78 0 +/- 6 0%) Conversely. after 48 h of culture, there were no significant differences in hatching late (62.8 +/- 11 1 vs 59 6 +/- 9.4%) and mean total cell number (135 2 +/- 13 1 vs. 146 7 +/- 13 3) In non-vitrified IVP bovine blastocysts, the hatching rate on Day 9 was improved by Y-27632 (91 7 +/- 3 8 vs 54 7 +/- 8 9%. P < 0 05). with no difference in mean total cell number of blastocysts (230 0 +/- 23 0 vs 191 2 +/- 22 2, P = 0 23). In an additional experiment, Y-27632 was added to culture medium on either Day 0, Day 2. or Day 4 (and remained present until Day 8). resulting in no improvement in blastocyst yield compared to a control group (7.5 +/- 2 1, 31 4 +/- 2 3, 36 2 +/- 3.2. and 28.6 +/- 6.9%. respectively) In conclusion, adding a ROCK inhibitor to post-thaw culture medium improved revivability of IVP bovine blastocysts after vitrification and warmingArticleTHERIOGENOLOGY. 73(8):1139-1145 (2010)journal articl

Microtubule assembly and in vitro development of bovine oocytes with increased intracellular glutathione level prior to vitrification and in vitro fertilization

Although vitrification is a useful technique for preservation of bovine oocytes, the yield of blastocysts derived from the vitrified oocytes is still low. We have recently reported a new type of cryoinjury, multiple aster formation, by which pronuclear migration and development of vitrified–warmed and in vitro-fertilized bovine oocytes are impaired. The aim of the present study was to investigate the effect of glutathione (GSH) content of vitrified bovine oocytes on multiple aster formation and subsequent in vitro development. Treatment of bovine cumulus–oocyte complexes with β-mercaptoethanol (βME) and l-cysteine (Cys) during in vitro maturation resulted in 2.5-fold higher GSH content not only in fresh control but also in vitrified–warmed oocytes. The percentage of normally fertilized zygotes exhibiting sperm aster(s) was >95% in all four groups (with or without βME/Cys × fresh control or vitrified). The frequency of multiple aster formation in vitrified oocytes (three-fold higher than that in fresh control oocytes) was not affected by the increased level of intracellular GSH with βME/Cys. Consequently, the migration and development of pronuclei as well as the yield of blastocysts from vitrified–warmed oocytes (17 versus 41%) were not improved. In addition, there was no effect of increased GSH level on the yield of blastocysts in fresh control groups.ArticleZYGOTE. 22(4):476-482 (2014)journal articl

Vitrification of ICSI- and IVF-derived bovine blastocysts by minimum volume cooling procedure: effect of developmental stage and age

The objective was to investigate the effects of developmental stage (fully-expanded or expanding blastocysts) and/or age (harvested on Days 7 or 8) on post-vitrification in vitro survival of bovine blastocysts derived from intracytoplasmic sperm injection (ICSI) or in vitro fertilization (IVF). Post-warming survival (re-expansion of blastocoele within 24 h) of ICSI-derived fully-expanded blastocysts (80%) was similar to that of their IVF-derived counterparts (88%). However, the ability of ICSI-derived expanding blastocysts to survive vitrification procedures (61%) was lower than that of IVF-derived blastocysts (85%; P < 0.05), although the ICSI- and IVF-derived fresh blastocysts were of similar quality. The age of the blastocysts before vitrification did not affect cryotolerance for either ICSI-derived (73 and 59% for Days 7 and 8 embryos, respectively) or IVF-derived blastocysts (86% for both Days 7 and 8 embryos). At 24 h of post-warming culture, ICSI-derived blastocysts surviving vitrification contained a higher proportion of dead cells than their IVF-derived counterparts (5-13% vs. 2-4%; P < 0.05), but these proportions were not different from those of fresh control embryos. There was an adverse effect of vitrification on the ability of blastocysts to hatch within 72 h of culture only in IVF-derived Day 8 blastocysts (41 and 70% in vitrified and fresh control groups, respectively). In conclusion, the proportion of blastocysts that survived vitrification procedures was similar for ICSI- and IVF-derived bovine blastocysts if the former were cultured to the fully-expanded stage prior to vitrification, with no significant difference between embryos harvested on Day 7 versus Day 8.ArticleTHERIOGENOLOGY. 74(6):1028-1035 (2010)journal articl

Pressure-induced reconstitution of Fermi surfaces and spin fluctuations in S-substituted FeSe

FeSe is a unique high-Tc iron-based superconductor in which nematicity, superconductivity, and magnetism are entangled with each other in the P-T phase diagram. We performed ⁷⁷Se-nuclear magnetic resonance measurements under pressures of up to 3.9 GPa on 12% S-substituted FeSe, in which the complex overlap between the nematicity and magnetism are resolved. A pressure-induced Lifshitz transition was observed at 1.0 GPa as an anomaly of the density of states and as double superconducting (SC) domes accompanied by different types of antiferromagnetic (AF) fluctuations. The low-Tc SC dome below 1 GPa is accompanied by strong AF fluctuations, whereas the high-Tc SC dome develops above 1 GPa, where AF fluctuations are fairly weak. These results suggest the importance of the dxy orbital and its intra-orbital coupling for the high-Tc superconductivity

The melting curve of iron at extreme pressures: implications for planetary cores

Exoplanets with masses similar to that of Earth have recently been discovered in extrasolar systems. A first order question for understanding their dynamics is to know whether they possess Earth like liquid metallic cores. However, the iron melting curve is unknown at conditions corresponding to planets of several times the Earth's mass (over 1500 GPa for planets with 10 times the Earth's mass (ME)). In the density-temperature region of the cores of those super-Earths, we calculate the iron melting curve using first principle molecular dynamics simulations based on density functional theory. By comparing this melting curve with the calculated thermal structure of Super Earths, we show that planets heavier than 2ME, have solid cores, thus precluding the existence of an internal metallic-core driven magnetic field. The iron melting curve obtained in this study exhibits a steeper slope than any calculated planetary adiabatic temperature profile rendering the presence of molten metallic cores less likely as sizes of terrestrial planets increase