No evidence for paternal mtDNA transmission to offspring or extra-embryonic tissues after ICSI.

There is a risk that ICSI may increase the transmission of mtDNA diseases to children born after this technique. Knowledge of the fate and transmission of paternal mitochondrial DNA is important since mutations in mitochondrial DNA have been described in oligozoospermic males. We have used an adaptation of solid phase mini-sequencing to exclude the presence of levels of paternal mtDNA >0.001% in ICSI families. This method is more sensitive than those used in previous studies and is sufficient to detect the likely paternal contribution (approximately 0.1-0.5% from simple calculations of expected dilution during fertilization). Using this method, we were able to detect concentrations as low as 0.001% paternal mtDNA in a maternal mtDNA background. No paternal mtDNA was detected in the embryonic (blood or buccal swabs) tissue of children born after ICSI nor in extra-embryonic tissue (placenta or umbilical cord). In conclusion, we did not detect paternal mtDNA in blood, buccal swabs, placenta or umbilical cord of children born after ICSI. We have found no evidence that ICSI increases the risk of paternal transmission of mtDNA and hence of mtDNA disorders.

A postal survey of embryo transfer practice in the UK.

In this postal survey a questionnaire was sent to all unit directors in the UK to determine their attitudes to the factors influencing embryo transfer practice. They were requested to rate each step on a scale of 1-10, where 1 was irrelevant and 10 very important. A total of 80 practitioners from 40 units replied. Over 50% of the corresponding practitioners were consultants, 33% were middle-grade clinicians, and 12% were infertility nurse specialists. The factor that got the highest rating was the need for a standardized protocol for all unit staff regarding embryo transfer technique. The second critical factor voted by the respondents was the presence of blood on the embryo transfer catheter. Not touching the uterine fundus was deemed to be the third most important factor while the type of embryo transfer catheter used was a very close fourth. Prolonged bed rest following embryo transfer was voted the least important factor to influence the outcome. The wide variations in practice and choice of catheters encountered in this survey are indications of the divided opinion and lack of concrete evidence on which to base any firm decisions. The need for large clinical studies to assess clearly whether higher pregnancy rates will result from modifications in embryo transfer practice is highlighted.

Uptake of a mixture of amino acids by mouse blastocysts.

Preimplantation mouse embryos exhibit growth at the blastocyst stage when there is a dramatic increase in protein content between days 4 and 5 after hCG treatment. This increse in protein synthesis requires the uptake of amino acids from the surrounding medium, but the consumption of individual amino acids present as a mixture has not been examined. We therefore measured the simultaneous depletion of 18 amino acids by mouse blastocysts in culture on days 4 and 5 after hCG treatment. Two culture media were used: one with amino acids present at concentrations largely based on those reported to give optimal rates of embryo development (M16/mix AA), and a second with the amino acids each at 0.1 mmol l-1 (M16/0.1 AA). After derivatization with o-phthaldialdehyde to yield highly fluorescent products, amino acid concentrations in the media were measured by HPLC. On day 4, seven amino acids were depleted from M16/mix AA at rates significantly greater than zero: aspartate, arginine, glycine, alanine, isoleucine, leucine and lysine; on day 5, nine amino acids were depleted at significant rates: aspartate, glutamate, asparagine, tyrosine, methionine, valine, phenylalanine, isoleucine and leucine. Average amino acid depletion from M16/mix AA was 32% greater on day 5 than on day 4. Aspartate, glutamate, arginine, isoleucine and leucine appeared to be the key amino acids sought by embryos on both days tested. Embryos grown in amino-acid-containing media appeared morphologically normal and their volume increased compared with embryos cultured in the absence of amino acids.