Although human in vitro fertilization (IVF) has been successful since the 1970s, similar reproductive technology in horses has lagged behind. Success rates stubbornly hover between zero and 30%, and only two live foals have been born using IVF.

The main problem seems to be with the ability of sperm to penetrate the egg. Ongoing research, much of it at Texas A&M University (TAMU), has led to the ability to successfully mature horse oocytes in vitro (in the laboratory). However, achieving in vitro sperm capacitation—which involves a series of changes that sperm must undergo in order to be able to fertilize an egg—has proved more complicated.

A solution could soon be at hand, though: Leticia Vivani, DVM, MS, a PhD student working jointly in the labs of Katrin Hinrichs, DVM, PhD, Dipl. ACT, and Dickson Varner, DVM, PhD, Dipl. ACT, at the Texas A&M College of Veterinary Medicine & Biomedical Sciences (CVM), is working to understand the reasons behind this problem.

“We are trying to understand the factors that regulate the process of sperm capacitation,” Vivani said. “Of course, this happens perfectly well when we breed a mare, but not when we try to mimic this in vitro. We do not know the reason, but I am almost sure there is something ‘special’ within the mare reproductive tract, something that might not be needed in other species, but that makes stallion sperm undergo all these changes.”

Hinrichs’ lab has determined that for some reason, equine sperm are different from those of other species. For example, incubation conditions that successfully induce hyperactivation (the whip-like tail motion needed to penetrate the egg) and subsequent fertilization in other species fail to do so with equine sperm.

“There can be so many things that can be affecting capacitation, and there is very little research done on equine sperm physiology,” Vivani said. “This is good, in a way, because anything you do is new, is innovative, but at the same time, the research can be very challenging and frustrating.”

Some progress is being made in understanding equine sperm capacitation. In 2009, by inducing hyperactivated motility with a substance called procaine, researchers at Cornell University achieved the highest fertilization rate to date (61%). Unfortunately, this is not a practicable solution for embryo production because procaine is toxic to the embryos.

“This was nevertheless an important finding,” Vivani said, “because it showed us that the failure of IVF was likely due to a sperm-related problem. We now know that there is something difficult about inducing appropriate equine sperm motility in vitro, and that may be why IVF rates have been so low.”

Vivani, who earned her DVM degree in her native Argentina in 2001 and her Master of Science degree at the University of Massachusetts in 2010, has long wanted to study at TAMU with this team of researchers.

“I have been fascinated with equine reproduction and the reasons behind the failure of IVF in the horse in particular, since my DVM graduation,” Vivani said. “Therefore, I’ve been dreaming of working with Dr. Hinrichs and with Dr. Varner since I was in Argentina. I had been writing to them and calling them for years before I was eventually able to make it work to come to Texas A&M, first as a visiting researcher and then as a PhD student.” She began her program in May 2012 and plans to graduate in 2016.

“Horses are an excellent model for human comparative studies,” Vivani said. “Mice, for example, do not age as women do, but the changes that mares undergo are very similar to women’s changes with aging. When mares reach a certain age, their reproductive efficiency decreases as a result; changes in hormone levels, follicular development, and oocyte quality are very similar in older mares and older women. So it’s a great model for human medicine.”

Just as in human medicine, owners of horses for whom natural reproduction has failed turn to assisted reproduction technologies. In horses, this often means using intracytoplasmic sperm injection (ICSI), a technique of “bypassing” standard IVF by injecting a sperm directly into the egg. The ICSI technique is also sometimes used in humans, especially when more traditional IVF has not worked. At the moment, the only way to successfully produce a horse embryo in vitro is through the use of ICSI, which many breeders are increasingly using.

“However, is not the most physiological way,” Vivani said, “and is not always practical, as it requires time, sophisticated equipment, and trained personnel. That is why I focus on how to make IVF work.”

Ironically, the One Health approach, which usually means translating findings from animals into human medicine, works backward in this case: IVF works well in humans, and has for more than 30 years, however, it is not yet successful in horses, and perhaps going back to the basics of reproduction can help explain why that is the case.

“In the beginning basic research was hard for mebecause I was trained as a veterinarian and I just wanted to see results,” Vivani said, “My advisor for my master’s degree, who is a basic researcher, made me stop and ask why things work or not, and I’m very grateful that he did.”

In fact, the IVF technique was pioneered through basic research that led to the discovery of capacitation.

“Dr. Hinrichs and other researchers like her have focused on really understanding physiological processes related to reproduction,” Vivani said. “For many years we, working in equine reproduction, tried one thing and if it didn’t work, we tried a completely different thing without trying to figure out why it didn’t work, and this process explains why there is so little information in this area.”

Despite all of the challenges, Vivani says that she finds her work extremely satisfying, partly due to her mentors: “Dr. Hinrichs and Dr. Varner are so encouraging with their students and really value their work,” Vivani said. “They always encourage me to learn as much as I can. Studying with them has been a wonderful experience—truly a dream come true.

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