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We examined growing oocytes collected from follicles remaining in superovulated rabbit ovaries, that were grown (in vitro growth, IVG) and matured (in vitro maturation, IVM) in vitro. We produced somatic cell nuclear transfer (SCNT) embryos using the mature oocytes and examined whether these embryos have the ability to develop to the blastocyst stage. In addition, we examined the effects of trichostatin A (TSA), a histone deacetylase inhibitor (HDACi), on the developmental competence of SCNT embryos derived from IVG–IVM oocytes. After growth for 7 days and maturation for 14–16 h in vitro, the growing oocytes reached the metaphase II stage (51.4%). After SCNT, these reconstructed embryos reached the blastocyst stage (20%). Furthermore, the rate of development to the blastocyst stage and the number of cells in the blastocysts in SCNT embryos derived from IVG–IVM oocytes were significantly higher for TSA-treated embryos compared with TSA-untreated embryos (40.6 versus 21.4% and 353.1 ± 59.1 versus 202.5 ± 54.6, P < 0.05). These results indicate that rabbit SCNT embryos using IVG–IVM oocytes have the developmental competence to reach the blastocyst stage.
Histone acetylation is one of the major mechanisms of epigenetic reprogramming of gamete genomes after fertilization to establish a totipotent state for normal development. In the present study, the effects of trichostatin A (TSA), an inhibitor of histone deacetylase, during in vitro fertilization (IVF) of bovine oocytes on subsequent embryonic development were investigated. Cumulus-enclosed oocytes obtained from slaughterhouse bovine ovaries were matured in vitro and subjected to IVF in a defined medium supplemented with 0 (control), 5, 50, and 500 nM TSA for 18 h. After IVF, presumptive zygotes were cultured in modified synthetic oviductal fluid (mSOF) medium until 168 h postinsemination (hpi). Some oocytes were immunostained using antibody specific for histone H4-acetylated lysine 5 at 10 hpi. Cleavage, blastocyst development and cell number of inner cell mass (ICM) and trophectoderm (TE) of blastocysts were assessed. TSA treatment enhanced histone acetylation that was prominent in decondensed sperm nuclei. TSA did not affect the postfertilization cleavage, blastocyst rates, and TE cell number. However, it significantly enhanced ICM cell number (p < 0.05). These results indicate that TSA treatment during IVF of bovine oocytes does not affect blastocyst development but alters the cell number of ICM, suggesting that overriding epigenetic modification of the genome during fertilization has a carryover effect on cell proliferation and differentiation in preimplantation embryos. Thus, further environmental quality controls in assisted reproductive technologies are needed in terms of factors which affect chromatin remodelling.
A 10-year-old girl required replacement of the aortic valve because of massive regurgitation caused by Kawasaki disease at the age of 3 months. Postoperatively, left ventricular function and mass improved remarkably, with reduction of both hypertrophy and interstitial fibrosis. During a follow-up period of 10 years, she has been free from any cardiac event.
A 32 Mbit chain FeRAM™ stack with 0.20μm minimum feature size was etched with two subsequent lithography/RIE steps: in mask step 1 the platinum/SRO (strontium ruthenium oxide) top electrode and the PZT (lead zirconate titanate) layer, in mask step 2 the bottom electrode together with the Ir/IrO2 diffusion barrier were etched. The stack was etched with various chlorine based chemistries. High temperature etching processes were applied to suppress residues by the formation of volatile etching byproducts resulting in a highly anisotropic etching profile and low redeposition.
Profile angles of 75° for step 1 and 80° for step 2 could be achieved. For the thin SRO-layer a separate etching recipe was developed to avoid surface roughening caused by micromasking. The influence of etching temperature and different gas chemistries on the etching behavior was evaluated. Reliable end point detection and good uniformity of the individual etching processes were obtained, both being crucial for the application of a multi-step recipe. The ferroelectric properties of the capacitor were confirmed by hysteresis measurements. This demonstrates that the ferroelectric properties were conserved during RIE etch processes at high temperature.
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