By Dr. Tom Arjun E. Stout EBVS
The first reports of in vitro embryo production (IVEP) by conventional in vitro fertilization and intracytoplasmic sperm injection in horses date respectively from approximately 30 and 25 years ago. However, IVEP has only become established in clinical practice during the last decade.
The initial slow uptake of IVEP was largely because the likelihood of success was too low to make it an economically viable means of breeding horses. During the last decade, the balance has shifted, primarily because of significant improvements in the efficiency of recovering immature oocytes from live donor mares (historically <25%; now >50%) and in the successful culture of zygotes to the blastocyst stage in vitro (historically <10%; now >20%). It has also been established that immature oocytes can be ‘held’ at room temperature for at least 24 hours, allowing overnight transport to a laboratory with expertise in IVEP. Moreover, because in vitro-produced embryos can be cryopreserved with no appreciable reduction in viability, they can be shipped and stored until a suitable recipient mare is available for transfer. Most importantly, in an established equine ovum pick-up intracytoplasmic sperm injection (OPU-ICSI) program, blastocyst production rates now exceed 1 per procedure, and posttransfer foaling rates exceed 50%, such that overall efficiency betters that of either embryo flushing or oocyte transfer. Moreover, OPU-ICSI can be performed year round and allows embryo production from mares with severe acquired subfertility and extremely efficient use of scarce or expensive frozen semen. Cumulatively, these factors have stimulated rapid growth in demand for IVEP among sport horse breeders.
1. Introduction
The first foal produced via in vitro fertilization (IVF) was born in 1990 [1]. Subsequent demand for in vitro embryo production (IVEP) in commercial horse breeding was however, until the last decade, very limited. This was in part because of the failure to develop reliable protocols for conventional IVF (i.e., fertilization of oocytes by coincubation with spermatozoa), which in turn appears to be primarily a function of an inability to adequately trigger stallion spermatozoa to capacitate and penetrate equine oocytes ex vivo (for review, refer to the article by Leemans et al [2]).
However, even though the necessity to induce sperm activation was circumvented by the demonstration that equine oocytes could be successfully fertilized by direct injection of a spermatozoon into the ooplasm (i.e., by intracytoplasmic sperm injection [ICSI]: for review, refer to the article by Squires et al [3]), there were still a number of significant obstacles to the establishment of equine IVEP as a practically and economically viable means of producing foals; these included the relative inefficiency of immature oocyte recovery from live donor mares (for review, refer to the article by Hinrichs [4]) and the relatively low success of culturing ICSI-derived zygotes to the blastocyststage where they can be transferred nonsurgically into the uterus, rather than surgically into the oviduct, of a recipient mare[5].
Although most improvements in the various steps required to produce embryos in vitro, namely ovum pick-up, in vitro oocyte maturation (IVM), ICSI, and in vitro embryo culture, have been incremental, there have been one or two major breakthroughs that have markedly improved the likelihood of success and, thereby, stimulated interest in ovum pick-up and intracytoplasmic sperm injection (OPU-ICSI) among horse breeders.
The first indication that OPU-ICSI could compete with embryo flushing or oocyte transfer in a clinical setting came when Galli et al [6] reported recovering a mean of 8.4 oocytes per OPU (with an efficiency of 50% per aspirated follicle) and generating 0.6 blastocysts per procedure. This compares favorably with conventional embryo transfer (ET) where embryo recovery in commercial programs typically ranges between 60 and 65% when mares are inseminated exclusively with fresh semen [7,8] but only between 30% and 50% when chilled-transported and frozen-thawed semen are used predominantly for insemination [9], where success rates are compromised by the high proportion of aged, subfertile donor mares in many ET programs... To read the complete article you need to be a subscriber
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