28 November 2016
Faculty of Biology Medicine and Health, University of ManchesterAppeared in BioNews 879
A major factor limiting the effectiveness of current assisted reproduction technologies (ARTs) is the availability of high quality oocytes (egg cells) from a patient. In a recent paper, Shoukhrat Mitalipov and colleagues investigate the use of polar body nuclear transfer (PBNT) as a means of generating healthy, high-quality oocytes from a polar body of a patient oocyte and a donor oocyte that had its own nuclear material removed.
Oocytes are produced through meiosis – the process of chromosome replication and cell division which produces sex cells. They begin as primary oocytes – early forms of egg cells that contain two copies of the body's chromosomes. During fetal development, the meiosis of primary oocytes is paused. Meiosis of individual oocytes only continues years later, during a woman's monthly menstrual cycle. Primary oocytes then progress through the rest of meiosis, dividing into two cells, each of which contains one copy of the body's chromosomes. One of these becomes the secondary oocyte, and the other a much smaller cell-like structure called the polar body (PB1). As a result, PB1 is a waste product of the developmental process of producing oocytes, acting only as a repository for the excess, second copies of chromosomes that the egg cell needs to get rid of in order to combine with a sperm cell. The meiosis of the secondary oocytes is then paused and only resumes at the onset of fertilisation, whereupon a second polar body (PB2) is produced.
Mitalipov and colleagues have previously demonstrated that when the nucleus of a secondary oocyte is removed and replaced with the nucleus of an adult cell, this can give rise to a functional oocyte. In this new paper, this idea is moved forward by transplanting the nuclear content of PB1, which is theoretically identical to that of a secondary oocyte, to create a healthy oocyte capable of being fertilised, and completing meiosis. Indeed, this has been achieved before in mice and has produced viable, fertile offspring; however, this is the first instance of PBNT being successful in humans. Of the 31 (of 32) PBNT oocytes that were successfully fertilised through intracytoplasmic sperm injection (ICSI), 19 progressed through meiosis, representing a success rate of 61 percent. This is impressive, albeit significantly lower than the 79 percent of control embryos. The researchers then cultured the embryos to the blastocyst stage, to assess their developmental capacity and their genetic integrity. While 75 percent of control embryos reached blastocyst stage, only nine of the remaining PBNT embryos reached this point. This comprises 42 percent of the embryos that were successfully fertilised – 30 percent of the original 32 PBNT oocytes in the experiment.
So, what have we learned from this paper and what might the applications of this new knowledge be? The first and most interesting point is that the genetic integrity of PB1 appears to be intact, despite that fact that it plays no further part in normal development. This tells us that defective material is not partitioned into PB1 for discarding, and therefore one wonders what the evolutionary advantage of this is. In practical terms, it is probably an issue of timing, as PB1 in humans does tend to degenerate and visibly fragment with time, such that in human clinical ART the presence of both PBs is not always seen post-fertilisation. So, for this method to work, PB1 has to be captured before it degenerates.
In terms of practical applications, using PB1 in PBMT with a donor oocyte effectively generates two genetically similar (not identical) oocytes for the donor patient, so a 'two for the price of one' secondary oocyte. These 'extra' oocytes could have also uses in the generation of human embryonic stem cell (hESC) lines, in avoidance of mitochondrial disease transmission or in routine clinical IVF treatment.
With this in mind, the authors conducted a rigorous evaluation of the genetic and epigenetic status of the PBNT embryos and in hESCs derived from them. They found no significant changes in methylation (an epigenetic modification), or in mRNA expression profiles (a measure of gene expression). However, 186 genes were detected to be differentially regulated in PBNT embryos compared to controls. Much further analysis is required on whether differential regulation of these genes would affect the health of any offspring who might result from this methodology. It is also possible that there may yet be differences that were not detected by the limited replicate number and scope of this first-in-human study.
Due to the obvious limitation of relying upon donor oocytes, this methodology is unlikely to ever 'double' the number of eggs available to IVF patients, and will certainly not double IVF success rates. However, uses in future applications as a means to transplant healthy genetic material from a patient into a healthy donor oocyte are promising.