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MicroRNAs (miRNAs) are a class of small endogenous single-stranded noncoding RNA molecules that have important roles in several biological processes. Research in human and laboratory animals has shown that miRNAs can regulate genes associated with type 2 diabetes mellitus and metabolic syndrome, and that the levels of specific miRNAs circulating in the bloodstream can serve as potential biomarkers for the diagnosis and prognosis of these diseases. We hypothesized that insulin-resistant (IR) horses would have a different circulating miRNA profile than those that are healthy. Fifteen nonpregnant mares housed at the Virginia Tech Middelburg Agricultural Research and Extension Center were evaluated for insulin sensitivity, with the frequent sampling intravenous glucose tolerance test. Selected mares, representing the most insulin-sensitive (IS, n = 3) and IR (n = 3) states, and paired for age, weight, and body condition, underwent miRNA profiling. Serum samples were collected, miRNA extracted, and microarray analysis performed to investigate the presence and relative amount of 340 equine miRNAs. Confirmation by quantitative real-time polymerase chain reaction revealed that miRNA was present in the serum of all animals. Results demonstrated different miRNA profiles between groups: Six miRNAs were expressed only in IS mares, five miRNAs were found to have lower quantity in IR mares relative to the IS ones, and three miRNAs were higher quantity in IR mares relative to the IS ones. The novel results of this preliminary study suggest potential new tools that could be developed for the diagnosis and treatment of metabolic syndrome in horses. [ABSTRACT FROM AUTHOR]

Maternal diet has been shown to impact fetal development across a variety of species. Given the reported effects of omega-3 long-chain polyunsaturated fatty acid (n-3 LCPUFA) supplementation on health across species, it was hypothesized that supplementation of algal-based n-3 LCPUFA to mares around conception would alter the maternal uterine environment and modify gene expression in early conceptuses. Nonlactating, light horse mares were supplemented with concentrate (n = 6) or an isocaloric diet containing 0.06 g/kg body weight n-3 LCPUFA (primarily docosahexaenoic acid [DHA], n = 7) at least 60 days before first sample collection. Four consecutive ovulatory cycles were monitored. Uterine endometrial samples were obtained 12.5 days postovulation from cycles 1 (uninseminated), 3, and 4 (bred to a common stallion); embryos were flushed from cycles 3 and 4. Biopsies from supplemented mares were greater in DHA and total omega-3 fatty acid concentrations ( P < .05), indicating successful tissue incorporation. Uninseminated n-3 LCPUFA supplemented mares displayed increased PTGFS and reduced IL6 gene expression ( P < .01) than controls. Supplemented pregnant mares had increased mRNA abundance of PTGES ( P = .05), PTGFS ( P = .03), and SLCO2A1 ( P = .01), while supplemented barren mares exhibited lower PTGFS ( P = .05) and IL6 ( P < .01) expression than control mares. In the conceptus, relative transcript abundance of trophoblast and endoderm markers ( GATA3 , GATA4 , GATA6, TFAP2A , ELF3 ) was greater ( P < .05) in embryos from supplemented mares. These results indicate algae-derived n-3 LCPUFA supplementation prior to breeding alters the maternal uterine environment and modifies expression of genes in preimplantation equine conceptuses. [ABSTRACT FROM AUTHOR]

Mature, nonpregnant, Thoroughbred mares were used to determine the influence of high-starch and high-sugar diets on postprandial inflammation. Plasma samples were obtained hourly from mares (n = 12) consuming one of two treatment diets, either a diet high starch and sugar (STR) or the control (CON) diet that was low in starch and sugar. Plasma was analyzed for concentrations of lipopolysaccharide (LPS) and the inflammatory cytokines interleukin (IL)-1β and IL-6. Hour 0 was included as a covariate in the statistical model, and where interactions between the covariate and other model variables existed, simple effect means were separated at three levels of the covariate: lower 95% confidence limit (CL), mean, and upper 95% CL. For horses with low ( P = .016) and average ( P = .065) initial LPS concentrations, LPS was greater or tended to be greater in STR compared with CON at hour 2 after feeding. No other differences were detected for LPS concentrations. For horses with low ( P = .037), average ( P = .006), and high ( P = .001) initial IL-1β concentrations, plasma IL-1β was greater in STR than CON at hour 2 after feeding. For horses with high initial IL-1β concentrations, IL-1β also tended to be greater at hour 3 ( P = .077). For horses with low ( P = .022) or average ( P = .063) initial IL-6 concentrations, IL-6 was greater or tended to be greater at hour 1 than 0. No effect of diet was detected for horses that started with high initial IL-6 concentrations. High-starch and high-sugar diets increase postprandial IL-1β concentrations, and it is likely that this effect is independent of LPS. [ABSTRACT FROM AUTHOR]

Abstract: Frozen semen provides several advantages to the breeder relative to fresh or cooled semen. However, some stallions are undesirable candidates for semen freezing due to poor post-thaw motility or longevity caused by membrane damage, osmotic stress, and oxidative stress during cryopreservation. The objective of this study was to determine the effect of post-thaw addition of pentoxifylline, caffeine, or taurine on sperm motility and longevity in equine frozen semen. Pentoxifylline, caffeine, or taurine was incorporated immediately into thawing frozen semen from nine warmblood stallions. Spermatozoa motility and longevity parameters were recorded and analyzed for each additive and for an untreated control. Of the three additives, only pentoxifylline improved total and progressive semen motility relative to that of untreated control. Pentoxifylline also increased semen curvilinear velocity, average path velocity, and straight line velocity relative to those of caffeine, taurine, or control. Semen treated with pentoxifylline also showed greater longevity relative to that of caffeine- or taurine-treated or untreated semen. Taurine improved linearity in comparison to that of semen treated with pentoxifylline, caffeine, or control but did not improve other parameters. Pentoxifylline may be useful in enhancing the quality of equine frozen semen and therefore improving its fertility. Additional studies are warranted that examine the effect of these additives on the conception rate. Pentoxifylline can be used to increase motility and longevity of equine frozen semen and theoretically increase probability of conception and overall breeding success rates. [Copyright &y& Elsevier]

Abstract: Kinematic variables of the walk in adult horses have been well described in the literature, but few studies have investigated growth-associated changes in these parameters. The objective of this study was to quantify linear and temporal walk patterns in Warmblood foals during the preweaning growth period. Nine foals were videotaped at the walk at 3, 11, and 21 weeks of age. Repeated-measures analyses were used to compare trait means between age groups. No significant effects owing to gender were found. Although stride length and stride duration increased as foals aged, neither differed across age groups when adjusted for wither height or velocity. Most kinematic variables did not differ across age groups when adjusted for Froude number. Overstride distance decreased by more than 40% in a linear manner from 3 to 21 weeks, and had an inverse relationship with distance between diagonal limbs during stance phase. Diagonal stance duration was greater than lateral stance duration for all age groups, indicating foals did not achieve an even, four-beat rhythm by the end of the study period. Changes in walk kinematics over time were independent of differences in velocity and increasing height during growth, and may indicate the need to account for body length or other morphometrics when assessing gait parameters in growing animals. Further research is needed during postweaning growth to determine when kinematic variables become consistent with those of adult horses. [Copyright &y& Elsevier]

Abstract: This study evaluated onset and incidence of palmar process fractures in foals and investigated corresponding changes in hoof conformation. Radiographs of 17 warmblood foals were taken at 1 week and at 2, 4, 6, 8, and 12 months of age and examined for evidence of palmar process fractures. Hoof parameters were measured from the radiographs, and comparisons were made between fractured and nonfractured feet. All foals suffered at least one fracture during the study period, with an average age of 3.68 ± 0.20 months at fracture occurrence. Hoof conformation and fracture occurrence were not significantly correlated. Foals consistently demonstrated higher lateral heights in the right foot and higher medial heights in the left foot. Hoof angle increased until 3 months of age and then gradually dropped. Hoof-pastern axis exhibited a broken forward conformation, but approached a more correct conformation by 12 months of age. There was a significant reduction in variation of variables for hoof and pastern angle in both feet, and hoof-pastern axis in the right foot. Hoof conformation does not appear to be a causative factor of palmar process fractures. It is possible that these fractures are a normal part of bone remodeling. [Copyright &y& Elsevier]

• Novel investigation of serum proteome associated with reproductive status. • Potential identification of systemic biomarkers of early pregnancy. • Detection of early pregnant histotroph proteins in serum from early pregnant mares. The reproductive cycle and early pregnancy represent dynamic physiological states in mammals, but mechanisms involved in early pregnancy in the domestic horse remain poorly understood. Proteins in uterine secretions have been studied, but the proteome of peripheral serum during various reproductive states has not been investigated. This study characterized and compared the serum proteome in the domestic horse during various reproductive states. Serum was collected from three mares during: (1) estrus (day [d] -1; d 0 = ovulation), (2) diestrus (d 12.5, non-mated), (3) early pregnancy (d 12.5, pregnant), and (4) nonpregnant (d 12.5, unsuccessfully mated) states. Serum proteins in each sample were analyzed by Nano LC-MS/MS, and 308 proteins were identified. Differentially-expressed proteins (DEP; > 1.5-fold or < - 0.5-fold) were identified by comparison of protein relative abundance between reproductive states: (1) diestrus compared to estrus (DEP = 71), (2) pregnant compared to diestrus (DEP = 72), and (3) non-pregnant compared to pregnant (DEP = 81). DEPs were analyzed for biological function using PANTHER (pantherdb.org). Several pregnancy-specific proteins previously identified in equine pregnant histotroph, including Apolipoprotein A-I, Complement C3, and Histone H4, were detectable in the serum. The ability to detect these biomarkers in serum provides a more readily available option for investigating and understanding early equine pregnancy. [ABSTRACT FROM AUTHOR]

Treatment of clinical laminitis usually fails to prevent some degree of persistent disability; thus, intervention should aim at avoiding risk factors and preventing the disease. Efficiency of intervention would be improved by identifying predisposed horses and ponies. A herd of 160 healthy ponies included 54 previously laminitic (PL) and 106 never laminitic (NL). Pedigree analysis was consistent with dominant inheritance partially suppressed in males. Blood analysis revealed higher plasma concentrations of insulin and triglycerides but not cortisol, glucose, or free fatty acids in the PL group. Proxies for insulin sensitivity and β-cell responsiveness, which were calculated from plasma insulin and glucose, indicated compensated insulin resistance in the PL group. A prelaminitic metabolic syndrome (PLMS) was derived statistically to have cut-off points for the 2 proxies, hypertriglyceridemia, and body condition score. It had a total predictive power of 78%. It identified 62 ponies with PLMS, and 98 as PLMS-free. Two months later, pasture starch concentration doubled, and 13 clinical cases of laminitis developed, 11 in the PLMS group and 2 in the PLMS-free group, giving an odds ratio of 10.4 (P = 0.0006). The PLMS can be used to identify predisposed ponies in need of special care; the efficiency of intervention would increase nearly 3-fold in the present case. It enables the design of new interventions suitable for testing. The PLMS also might influence market values. [ABSTRACT FROM AUTHOR]

Abstract: The objective of this study was to examine the effect of different insemination techniques and extenders on the volume of liquid dispensed from insemination equipment. The method of insemination has a significant effect on the volume of semen deposited into the mare''s uterus when low volumes are used. Insemination pipettes that allow for direct deposit of straw contents into the uterus are preferred. Aspiration of semen into a pipette is preferred over aspiration into a syringe with deposition through a pipette when direct deposit is not possible. Use of a pipette with a smaller lumen and less length of contact with liquid provides better results. Contact of semen with equipment may allow for residual liquid accumulation on the luminal surfaces and a decrease in overall semen dose. Extenders with differing amounts of egg yolk did not influence volume of liquid dispensed. [Copyright &y& Elsevier]

With increasing public scrutiny on animal welfare, it behooves those involved in the equine industry to revisit best management practices to ensure these support healthy horses. There is little published research on how horses are used in the equine industry, particularly in therapeutic horseback riding (THR) programs. Although there is a large amount of information on the benefits of THR programs to the participants, there is little published information available about the horses. Therefore, the objective of this survey was to gather data regarding horse use and care in Professional Association of Therapeutic Horsemanship International (PATH Intl.)–affiliated THR programs in the United States to help establish a foundation for a standard of care. A 20-question survey sent to 659 PATH Intl.–affiliated THR programs returned a 40% response rate. Demographics demonstrated that the median number of horses in each program was 10; geldings outnumbered mares; most horses were between 16 and 20 years of age; and Quarter Horse or stock-type breeds predominated. Median session length was 8 weeks and median lesson length was 45 minutes. Horses were typically ridden by clients 4 days/week and 2 hours/day. Most horses were donated to the programs, participated for approximately 7 years, and left because of aging. Limb lameness and back soreness were the top health issues noted, with only a small percentage of colic and ulcers reported. More horses received nonsteroidal anti-inflammatory drugs for a lameness issue, chiropractic adjustment, and massage than any other supplemental care or complementary therapy. Based on data gathered in this survey, THR horses were not worked excessively. Horses were ridden less than PATH Intl.'s maximum recommendation of 6 hours/day and 6 days/week and less than those used in university programs. Horses in THR programs also appeared to have fewer reported health issues as compared with data in other national reports. • Most therapy horses were geldings, aged 16–20 years, and a stock-type breed. • Horses were ridden an average of 4 days/week and 2 hours/day. • Limb lameness and back soreness were the top health issues reported. • Chiropractic adjustment, massage, and NSAIDs were the most common complementary therapies. • Horses were typically ridden less than the PATH Intl. maximum recommendation. [ABSTRACT FROM AUTHOR]

Insulin resistance (IR) is characterized by an increase in biomarkers of systemic inflammation and susceptibility to laminitis in horses. Impacts on reproduction include a lengthened interovulatory period in horses. Dietary omega-3 (docosahexaenoic acid [DHA]) promotes anti-inflammatory processes, has been implicated in health benefits, and can reduce cytokine secretion. This preliminary study investigated the impact of IR as well as the influence of dietary supplementation (DHA) on the uterine fluid proteome in early pregnant horses. Mares were artificially inseminated; uterine fluid and embryos were collected on d 12.5 after ovulation. Uterine fluid was pooled for metabolic and diet categories (n = 8; n = 2 per metabolic and dietary status) and concentrated, and the proteome was analyzed using tandem mass spectrometry (iTRAQ). Five proteins met differential abundance criteria (±1.5-fold change, P <.05) in all comparisons (Control C, IS vs. C, IR; C, IS vs. DHA, IS; C, IR vs. DHA, IR). Serum amyloid A, afamin, and serotransferrin were upregulated in C, IR mares but downregulated in DHA, IR mares when compared to C, IS and C, IR, respectively. Quantitative PCR supported mass spectrometry results. The presence of serum amyloid A and serotransferrin in histotroph of IR mares potentially indicates an inflammatory response not seen in IS counterparts. These preliminary findings provide novel evidence on the potential impact of insulin resistance and DHA supplementation on the secreted equine uterine proteome during early pregnancy. • Uterine secretions were examined via iTRAQ-labeled tandem mass spectrometry. • Insulin resistance appears to increase expression of inflammatory markers, without clinical symptoms during early pregnancy. • Serum amyloid A and afamin increase in uterine secretions from insulin-resistant mares. • Dietary docosahexaenoic acid provided to insulin-resistant mares may reduce inflammatory markers. [ABSTRACT FROM AUTHOR]

When fed to horses, high-starch diets elevate plasma concentrations of interleukin-1β (IL-1β) as soon as 1 hour posteating. This increase in IL-β is possibly because of changes in intestinal pH that result from rapid bacterial fermentation of starches and sugars in the digestive tract. The purpose of this research was to investigate the effect of feeding 0.9 kg of grass hay 30 minutes before feeding a concentrate meal on the postprandial rise in IL-1β, compared with control horses receiving the same concentrate without hay first. Six mature geldings were used in a switchback design. Horses were fasted overnight before being offered a concentrate feed that provided 1.2 g/kg bodyweight of nonstructural carbohydrates. Plasma was harvested 30 minutes before hay feeding and 1, 2, 4, 6, and 8 hours postfeeding. Concentrations of IL-1β and d-lactate were analyzed by repeated measures analysis of variance. The hay-first treatment reduced (P =.034) postprandial concentrations of IL-1β at all time points compared with the control horses. An interaction between hour and treatment was detected for mean d-lactate concentrations (P =.037), with lower concentrations in hay-first fed horses at postfeeding hours 1, 2, and 4, compared with control horses. Given these findings, we believe that feeding a small amount of hay before feeding a meal of moderate starch and sugar content reduced the negative effects of rapid starch and sugar fermentation in the equine digestive tract, evidenced by reduced postprandial d-lactate and IL-1β concentrations. • Consuming 1.2 g nonstructural carbohydrates per kilogram bodyweight after an overnight fast results in increased plasma concentrations of interleukin 1β. • Feeding a small amount of hay before feeding a meal of starch and sugar inhibited the rise in postprandial interleukin 1β concentrations. • Plasma d-lactate concentrations were lower in hay-first fed horses than concentrate-first fed horses. [ABSTRACT FROM AUTHOR]

Abstract: Yearling morphometric measurements and variables related to career racing performance were collected on 260 Thoroughbreds to determine correlations between and among these traits. Morphometric measurements were adjusted to 365 days based on interpolation from adjacent age points and included wither and hip height, body length, distal limb length, cannon circumference, heartgirth circumference, and chest width. Measures of career racing ability included total earnings, win percentage, and Standard Starts Index (SSI). Wither height and hip height were favorably correlated with lifetime earnings, SSI, and win percentage. Body length and heartgirth were positively correlated with SSI and win percentage. Leg traits were not correlated with any performance parameters. Horses that had placed in or won a stakes race tended to be taller at the wither and hip as yearlings. Horses categorized as preferring turf tended to be taller at the wither and hip at 365 days than those that preferred to run on dirt. No significant difference in yearling morphometric measurements between sprinters and routers was detected. In addition to phenotype and pedigree information, morphometric measurements taken in the yearling year may allow for more accurate estimation of future performance in the Thoroughbred racehorse. [Copyright &y& Elsevier]