Association between embryonic loss and damage to the zona pellucida by invasive micromanipulation during oviductal transfer of early-stage embryos in pigs.

The aim of this study was to examine the impact of zona pellucida damage, which might arise during somatic cell nuclear transfer (SCNT), on the development and survival of transferred embryos. The zonae pellucidae of in vitro matured oocytes were either punctured with 8- to 10-microm square-ended nuclear injection pipettes and piezo pulses or slit with 35- to 40-microm enucleation pipettes. Intact oocytes were used as controls. These oocytes were electroactivated to induce parthenogenesis and transferred to the oviducts of estrus-synchronized recipient gilts. After 5 to 7 days, the recipient uteri were flushed to collect embryos, and embryonic development (morula-blastocyst stage embryos/collected embryos) and survival (viable embryos/collected embryos) were determined. In total, 221 zona-punctured, 129 zona-slitted and 57 intact embryos were transplanted into four, two and two gilts, respectively. The efficiency of embryo recovery was similar in all groups (64.3 to 79.1%). However, the zona-penetrated and incised embryos exhibited unstable development and survival compared with the controls; development and survival of the control embryos were 94.7 and 87.7%, whereas those of the zona-punctured embryos were 69.0 and 47.9% (P<0.01) and those of the zona-slit embryos were 64.7 and 50.0% (P<0.01). Cells with large foci that appeared to be macrophage giant cells were observed at the surface or inside the degenerated zona-damaged embryos. These results indicate that the recipient's immune response to damage to the zona pellucida may impair embryonic development after transplantation to the oviduct. This may be one of the factors causing the reduced efficiency of live progeny production by SCNT.

Is intracytoplasmic sperm injection (ICSI) a safe procedure? What do we learn from early pregnancy data about ICSI?

The aim of this study was to evaluate the safety of the intracytoplasmic sperm injection (ICSI) procedure by analysing early pregnancy data from ICSI and in-vitro fertilization (IVF) patients. In all, 50 ICSI pregnancies were compared with 226 IVF pregnancies. Comparisons were made during the first 9 weeks after the theoretical last menstrual period (7 weeks after oocyte retrieval) with regard to epidemiological data, plasma hormonal concentrations and transvaginal ultrasonographical findings. Although patients were significantly (P < 0.001) younger in ICSI (31 years) than in IVF pregnancies (33 years), their duration of infertility was similar. Miscarriage and multiple gestation rates were not significantly different in ICSI pregnancies (respectively 24 and 24%) from those found after IVF (32 and 29%). The probability of developmental arrest of the intrauterine sac (miscarriages and vanishing twins) was similar in both ICSI (16%) and IVF (25%) cases. The mean plasma hormonal concentrations starting from day 11 after oocyte retrieval were similar in both groups. Every ICSI and IVF pregnancy showed an embryo with cardiac activity at 7 weeks. Early pregnancy data did not show any abnormal findings for pregnancies achieved using ICSI compared to those achieved by IVF.

[The effect of the micromanipulations used for transgenosis on mouse development]. / Vliianie mikromanipuliatsii, ispol'zuemykh dlia transgeneza, na razvitie myshei.

Non-specific effects of micromanipulation techniques used for producing transgenic mice on processes of embryonic development were studied. Zygotes obtained from C57BL and BALBxDD mice were treated as follows: (1) incubated in culture medium; (2) the male pronucleus punctured with a glass microneedle; (3) microinjected with a buffer solution; and (4) DNA (mouse P-35 oncogene with human insulin gene promoter) injected into the male pronucleus. Then zygotes were transferred into oviducts of syngeneic or allogeneic pseudopregnant females. Such treatment resulted in the intrauterine death of embryos, as well as in birth of the dead or non-viable offspring with numerous defects of development. Zygote pronucleus puncturing is the most damaging manipulation, since its effect exceeds that of the zygote incubation and is comparable with the effect of buffer of DNA injections.

Micromanipulation of embryos and germ cells: an approach to gene therapy?

Recent advances in mammalian gamete and embryo micromanipulation have stimulated the scientific and medical communities, and to some degree the public at large, to become aware that treatment of genetic disease by direct alteration of the genetic code may soon be possible. Because these micromanipulation techniques result in modification of the genotype at the earliest stages of development, such "gene therapy" affects not only the conceptus itself but also its germ cells. Thus such genetic modifications are heritable and can be transmitted indefinitely to succeeding generations of progeny. In the presentation, both narrow and broad definitions of gene therapy will be considered with respect to the techniques upon which they are based, their potential for treatment of genetic disease, and their current feasibility.