Each year at the American Association of Equine Practitioners convention, three researchers present what they believe to be the top studies with clinical implications from the past year in the fields of surgery, medicine, and reproduction. At the 2016 meeting, held Dec. 3-6 in Orlando, Florida, Texas A&M University theriogenology professor Terry Blanchard, DVM, Dipl. ACT, shared his seven favorite breeding-related papers on topics ranging from stem cell therapy in mares to semen extender in stallions.

Post-Mating Cervical Occlusion Puts Mares at Risk of Endometritis

Mares with a cervix that fails to relax and allow contaminants to drain post-mating are at risk of developing endometritis (inflammation of the inner lining of the uterus, called the endometrium).

So European researchers evaluated whether cervical occlusion (closing or blocking the cervix, which mimics the tight cervix of an older maiden mare, for instance) after artificial insemination increased uterine fluid accumulation and inflammation. In the study, the researchers gathered endometrial swabs, biopsies, and fluid from 29 normal mares over five estrous cycles. They artificially inseminated the mares during the second and fourth estrus; immediately after one of these inseminations they inserted a clamped catheter (to simulate cervical occlusion) into the uterus. The researchers found that the clamped catheter mares had more fluid accumulation and neutrophils (a type of white blood cell used for fighting infections) present than did the mares without catheters, resulting in declining fertility and development of periglandular fibrosis (scarring).

“Closure of the cervix after artificial insemination results in pronounced inflammation of the endometrium and may result in permanent damage,” Blanchard explained. “With those mares that have a tight or fibrotic cervix, expect problems and plan on aggressive treatment (perhaps two to three times) after insemination to get inflammation under control.”

Reilas T, del Alamo MMR, Liepina E, et al. Effects on the equine endometrium of cervical occlusion after insemination. Therio 2016;85:617-624

Try Hysteroscopic Hydrotubation of the Oviducts in Mares With Fertility Issues

Veterinarians believe proteinaceous plugs can accumulate in some mares’ oviducts, which might block sperm access to the oocyte or embryo migration through the oviduct, leading to infertility.

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Researchers from Rood & Riddle Equine Hospital, in Lexington, Kentucky, recently evaluated the effect of hysteroscopic flushing of the oviducts (in which the veterinarian passes an endoscopic catheter through the cervix and uterus into the oviduct and flushes it with saline) on subsequent mare fertility, with positive results: Post-hydrotubation, 79% of study mares became pregnant or obtained an embryo within one or two estrous cycles.

“Consider hysteroscopic flushing of the oviducts in mares with unexplained fertility,” the authors concluded.

Blanchard said that while this procedure is significantly less expensive than the previously relied-upon laparoscopic surgery, it also comes with a learning curve.

“You may still have to use laparoscopy if unsuccessful in flushing the oviducts in order to inspect ovaries and oviducts for adhesions or to apply prostaglandin E2 to the surface of the oviducts in an attempt to stimulate emptying of their contents,” he said.

Bradecamp EA, Schnobrich MR. Hysteroscopic hydrotubation of the oviducts as a treatment for idiopathic infertility in the mare – a retrospective study. J Clin Therio 2016;8:337.

Rosiglitazone Might be Promising New Semen Extender

Australian researchers recently looked at a new media for storing stallion semen that would preserve sperm function longer and potentially at room temperature (rather than cooled, which some sperm don’t tolerate well). Because they previously demonstrated that stallion sperm use oxidative phosphorylation (the metabolic pathway by which cells use enzymes to oxidize nutrients) to provide energy for rapid movement, rather than anaerobic glycolysis (a metabolic process in which glucose is transformed to lactate), the team evaluated the effects of rosiglitazone (which is used to improve glucose use during type 2 diabetes treatment in humans) on stallion sperm function.

“Samples incubated with rosiglitazone displayed significantly higher motility, percentage of cells with normal mitochondrial membrane potential, adenosine triphosphate (energy) content, and glucose uptake capacity, while sperm viability was unaffected,” wrote the authors.

Stored at room temperature, sperm motility remained above 60% for six days. “Room temperature extender might be on the horizon in a year or two,” said Blanchard.

Swegen A, Lambourne SR, Aitken RJ, et al. Rosiglitazone improves stallion sperm motility, ATP content, and mitochondrial function. Biol of Reprod 2016, in press. DOI:10.1095/biolreprod.116.142687

Cushion Centrifugation Can Salvage Some Urine-Contaminated Semen

Urine can negatively affect a stallion’s sperm quality if it contaminates the semen during breeding. A group of researchers evaluated the effect of the level of urine contamination on sperm quality and whether cushion centrifugation to remove urine would improve sperm survival rates after cooling. The team looked at 10%, 20%, 30%, and 40% urine concentration.

“In general, most sperm quality measures declined with increasing urine concentration starting immediately after semen collection,” the authors wrote. However, they found that cushion centrifugation and resuspension of the sperm in semen extender preserved the quality of several urine-contaminated samples, even after 24 hours in cooled storage.

“Cushion centrifugation with resuspension can salvage some urospermic (urine in the ejaculate) samples for cooling,” Blanchard concluded, adding that repeat offender stallions should be managed intensively (e.g., evacuate urine prior to collection; perform drug therapy, etc.) to minimize urine contamination of ejaculates.

Voge J, Varner DD, Blanchard TL, et al. The effects of urine concentration, and cushion centrifugation to remove urine, on the quality of cool-shipped stallion sperm. Therio 2016;86:1294-1298.

Vascular Elastosis Compromises Mares’ Uterine Blood Flow

In 2015, University of California, Davis, researchers identified a link between uterine vascular elastosis (a condition in which the vessels of the uterine wall have thickened, often disruptive, changes in surrounding elastic fibers) and infertility in mares. In this follow-up study, they evaluated whether mares with vascular elastosis also have impaired uterine vasodilation (widening of the blood vessels to increase blood flow), which could similarly impair fertility. They administered the estrogen estradiol (which has vasodilatory effects) to both normal mares and mares with severe elastosis and found that uterine vessel function is, indeed, compromised in mares with elastosis. It is possible, therefore, that these vascular changes are permanent and might prevent any treatment from improving uterine blood flow, said Blanchard.

Esteller-Vico A, Liu IKM, Vaughan B, et al. Effects of estradiol on uterine perfusion in anesthetized cyclic mares affected with uterine vascular elastosis. Anim Reprod Sci 2016;164:57-63.

Stem Cell Therapy Might Help Heal Injuries to the Endometrium

There are few effective treatments for many of the pathologic changes that can occur in a mare’s endometrium. But based on findings from studies in mice, human uterine-derived stem cells (MenSCs) might offer some effective options. The goal of this study from Chinese researchers was to determine if MenSCs promote endometrial repair in mice with injured endometriums. Upon being bred, mice treated with MenSCs had increased endometrial thickness, microvessel density, pregnancy rates, and embryo development compared with untreated controls.

“Human uterine-derived stem cells could restore the endometrium and improve fertility,” said Blanchard, noting, of course, the obvious limitation of this study being conducted in a different species.

Zhang Y, Lin X, Dai Y, et al. Endometrial stem cells repair injured endometrium and induce angiogenesis via AKT and ERK pathways. Reprod 2016;152:389-402.

Stem Cell Therapy is Safe and Simple Endometrial Treatment

Chronic degenerative endometriosis is a common cause of infertility, particularly in older mares. To potentially combat this condition, researchers from Brazil evaluated the safety of bone marrow-derived stem cell (BMSC) injections into the endometrium of 16 subfertile mares. In the study, they injected 12 million BMSCs at 12 locations in each mare’s endometrium. They detected no intrauterine fluid or endometrial edema (fluid swelling) in any mare after therapy and no changes in endometrial fibrosis. The only change noted was a transient acute endometritis two weeks post-injection that resolved spontaneously within one month.

“Endometrial injections of bone marrow-derived stem cells was safe and simple using this procedure,” said Blanchard.

Alvarenga MA, Carmo MT, Segabinazzi GL, et al. Feasibility and safety of endometrial injection of autologous bone marrow mesenchymal stem cells in mares. J Equine Vet Sci 2016;42:12-18.