From embryonic stem cells to blastema and MRL mice.

New scientific knowledge offers fresh opportunities for regenerative medicine and tissue repair. Among various clinical options, multipotent embryonic stem cells (ESC) prepared from inner cell masses of rabbit blastocysts have been tested over many years. More recently, stem cells have been isolated from individual tissues and from umbilical cord blood. These methods seemingly offer similar rates of repair and avoid ethical complexities arising from the need for human embryos to prepare ESC. Different methods of regenerating tissues have now emerged, based on the well-known forms of organ regeneration in urodeles such as salamanders. These methods depend on the formation of a blastema, and recent studies on MRL mice have revealed that they possess similar methods of repair as in salamanders. There is also some evidence showing that this form of repair is also active in human fetuses but not in adults. Detailed knowledge of these various forms of tissue repair is now urgently needed in order to assess the benefits of each form of treatment. These matters are discussed at the end of this review where various investigations clarify the benefits and drawbacks of these varied approaches to tissue repair.

A burgeoning science of embryological genetics demands a modern ethics.

This brief article discusses the nature of recent scientific advances in reproductive biomedicine and genetics, their moral implications and their effects on society. The pace of research has amplified exponentially, leading society into situations incomprehended by our ancestors. Early studies on reproductive biology in animals, and clinical methods such as artificial insemination by donor spermatozoa, were introduced several centuries ago and led to prolonged ethical disagreements. The 20th century witnessed the introduction of controlled ovulation in laboratory animals, the fertilization of the oocyte and preimplantation embryology in mammalian species. The second half of this century produced an avalanche of knowledge on genetics, developmental biology, the fertilization of the human oocyte in vitro, test-tube babies, preimplantation genetic diagnosis, designer babies, stem cells and a deeper understanding of molecular differentiation in the human embryo. The ethical and legal aspects of these items have led to intense debates on their rights and wrongs. The future may have even more bizarre possibilities such as producing medicines in cow's milk or trees and delaying death for many years.

The significance of parthenogenetic virgin mothers in bonnethead sharks and mice.

Two astonishing virgin births in quick succession have raised interest in parthenogenesis in cartilaginous sharks and mammals. These were believed to be exceptions until numerous female bonnethead (hammerhead) sharks were found be giving birth at the Henry Doorly Zoo in Nebraska, despite the prolonged absence of male sharks. The birth of a shark pup led to suggestions that spermatozoa from a previous coitus had persisted in the female tract of its mother and fertilized one of her eggs some months later. These proved to be incorrect because the female had been isolated for several years whereas spermatozoa persisted in the female tract for approximately 6 months. Molecular investigations into the pup's DNA failed to find any paternal contribution and proved the pup to be descended from its mother only. Just before this discovery, a study in mice had revealed that parthenogenesis could be induced by overcoming damage to embryonic development that is normally caused by gene imprinting. This was done by fusing two mouse oocytes and then inserting Igf2 into the parthenogenotes, which led to the birth of several parthenogenetic offspring. Modifying epigenesis had thus opened pathways to full-term parthenogenetic development. The birth of these parthenogenotes fulfils the attempts of earlier scientists to invoke parthenogenesis in experimental animals.

IVF, IVM, natural cycle IVF, minimal stimulation IVF - time for a rethink.

Considerable changes are afoot in the practice of assisted human conception. Doubts about its methods, especially over endocrinology, concern its complexity and its expense. IVF has spread worldwide since its beginnings in the UK, but its current practice, termed routine IVF, is being challenged by simpler routines. These include natural cycle IVF, which has been in the background for many years, minimal stimulation IVF, where doses of hormones are reduced, and the in-vitro maturation of human oocytes ready for fertilization in vitro (IVM). These three approaches are now practised in increasing numbers of IVF clinics, and may well replace routine IVF. The events leading to current interest in these methods will be discussed briefly in this review.

Genetics, epigenetics and gene silencing in differentiating mammalian embryos.

A highly complex pattern of differentiation involving maternal and embryonic factors characterizes the early development of mammalian embryos. These complex genetic and proteonomic patterns of early growth also involve various forms of gene silencing and tissue reprogramming. Understanding the nature of fundamental developmental events is hence essential to appreciate the significance of natural and induced forms of remodelling, damaged forms of gene expression and gene silencing during the initial stages of growth. Natural forms of remodelling include subtle genetic events involved in, for example, the changing nature of imprinting from before fertilization or the inactivation of one X chromosome in female blastocysts. Induced forms include the consequences of nuclear transfer and embryo cloning or the immediate effects of placing embryos in culture media. Animal and human studies are described in this paper, relating reprogramming to detailed embryological and clinical knowledge gained through the use of IVF, preimplantation genetic diagnosis and the establishment in vitro of stem cells. Attention concentrates on the consequences of variations in all growth stages from the formation of oocytes, through fertilization, the differentiation of blastocysts and early haemopoietic stages in mammalian species. Unique features of gene expression or gene modification are described for each developmental stage.

Nucleon axial charge in full lattice QCD.

The nucleon axial charge is calculated as a function of the pion mass in full QCD. Using domain wall valence quarks and improved staggered sea quarks, we present the first calculation with pion masses as light as 354 MeV and volumes as large as (3.5 fm)3. We show that finite volume effects are small for our volumes and that a constrained fit based on finite volume chiral perturbation theory agrees with experiment within 7% statistical errors.

Ethics and moral philosophy in the initiation of IVF, preimplantation diagnosis and stem cells.

Details of the work leading to the introduction of human IVF, animal and human stem cells, and the preimplantation diagnosis of inherited characteristics in blastocysts are outlined briefly in this paper. The progress of these studies is related to ethical issues emerging during these years. The current status of these studies is outlined, together with a brief moral philosophy as practised by the original investigators.

Changing genetic world of IVF, stem cells and PGD. A. Early methods in research.

Genetics proved essential to introduce IVF, preimplantation diagnosis (PGD) and embryo stem cells in the 1960s. Its small input in early years was confined to aspects such as timing follicle growth and ovulation. Modest understanding in the mid- to late 1980s, mostly on studies in mice, involved the actions of single genes and the balance between maternal and zygotic transcripts in preimplantation stages. Human IVF began after human oocytes were matured in vitro, and their meiotic chromosomes analysed. Their fertilization in vitro led to PGD and embryo stem cells. Unlike mouse embryos, most human embryos failed to implant, so the best had to be selected to improve IVF pregnancy rates. Initially, faster-growing embryos proved superior. Later, patterns of polarized nucleoli in pronuclei, the degree of blastomere fragmentation and growth of embryos in vitro to blastocysts provided excellent markers. Single cells could be isolated from embryos using micromanipulation. Stem cells from inner cell mass, a branch of IVF, differentiated into immortal stem cell lines in vitro if disaggregated. They formed virtually all body tissues in blastocysts cultured intact or when injected singly into recipient blastocysts. Later, the genetic controls of ES cell differentiation were assessed, together with factors switching them along specific differentiation pathways. Marker genes identified ES cells differentiating into various tissues.

Changing genetic world of IVF, stem cells and PGD. B. Polarities and gene expression in differentiating embryo cells and stem cells.

Novel genetic techniques in the later twentieth century led to new analytical methods for assessing the growth of embryos and stem cells and improve preimplantation diagnosis. Increasing attention to the nature of polarities in mouse and human embryos revealed the existence of an animal-vegetal axis in human oocytes and embryos. Combinations of meridional and transverse cleavage divisions, the latter due to spindle rotation, determined the unequal division of ooplasm to embryonic blastomeres. Blastomeres with differing functions were accordingly formed in 4-cell embryos, including founders of inner cell mass and trophectoderm. New forms of gene analysis led to the polymerase chain reaction, while fluorescence in-situ hybridization revealed astonishingly high degrees of heteroploidy in human embryos. Developmental genetics gained immense analytical power as cDNA libraries, microarrays, transcriptomes RNAi and other methods clarified the roles of hundreds of genes in pre- and early post-implantation embryos and stem cells.

Changing genetic world of IVF, stem cells and PGD. C. Embryogenesis and the differentiation of the haemopoietic system.

In this final review, attention is focused on the formation of several haemopoietic systems and their genetic markers. Very early haemopoietic precursors have been identified in mesoderm and yolk sac, as interactions arise between haemopoietic stem cells (HSC) and mesenchymal stem cells (MSC). The foundation cell for the haemopoietic system has not been identified, although several candidate cells carrying specific markers have been recognized and are highly pluripotent. Haemangioblasts were proposed as the founder haemopoietic stem cell. They may be the source of pluripotent haemopoietic cells formed in blastocyst injection chimaeras, a characteristic typical of ES cells. Their role as the founder cell of haemopoietic and mesenchymal tissues is discussed.

Time to revolutionize the triggering of ovulation.

Methods used for ovarian stimulation constantly change with advances in gonadotrophin therapy. In this Commentary, an appeal is made for more attention to the use of LH for the induction of ovulation. Its typical characteristics during the LH surge are finely balanced to induce normal ovulation and luteinization. It does not induce ovarian hyperstimulation, for example. The recent commercial availability of recombinant LH (LHr) offers a chance of escaping from the use of urinary human chorionic gonadotrophin (HCG) and its varied forms such as those with a shorter half-life. It should also avoid the weakly effective bursts of FSH and LH and weak luteal phases released associated with the use of gonadotrophin-releasing hormone agonists. Currently, large dosages of LHr are needed to match the endocrine events typical of inducing ovulation by the endogenous LH surge. In the interests of patients' safety and improved forms of luteal phase endocrinology, research should be devoted to improving the properties of rLH to make it induce surges similar to endogenous discharges. This would replace the current use of HCG to induce ovulation, with its attendant risks of ovarian hyperstimulation and luteal phase anomalies.