Abstracts / Journal of Equine Veterinary Science 31 (2011) 230-356 295

a large group of horses and submitted the results to statisticalanalysis, providing a comprehensive groundwork for research onthe relationship between hoof shape and exercise.

References

[1] Bailey C, Rose R, Reid S, Hodgson D. Wastage in the AustralianThoroughbred racing industry: a survey of Sydney trainers. Aust Vet J1997;75:64-6.

[2] Arabian A, Lanovaz J, Clayton H. Determination of hoof mass andcentre of mass from morphological measurements. Equine Vet J2001;33:46-9.

[3] Moleman M, van Heel M, van Weeren P, Back W. Hoof growthbetween two shoeing sessions leads to a substantial increase of themoment about the distal, but not the proximal, interphalangeal joint.Equine Vet J 2006;38:170-4.

[4] Decurnex V, Anderson G, Davies H. Influence of different exerciseregimes on the proximal hoof circumference in young Thoroughbredhorses. Equine Vet J 2009;41:233-6.

[5] Back W, Clayton H. Equine Locomotion. 1st ed. U.K: W.B. Saunders;2001.

[6] Kummer M, Gygax D, Lischer C, Auer J. Comparison of the trimmingprocedure of six different farriers by quantitative evaluation of hoofradiographs. Vet J 2009;179:401-6.

[7] Kummer M, Geyer H, Imboden I, Auer J, Lischer C. The effect of hooftrimming on radiographic measurements of the front feet of normalWarmblood horses. Vet J 2006;172:58-66.

[8] van Heel M, Moleman M, Barneveld A, van Weeren P, Back W.Changes in location of centre of pressure and hoof-unrollmentpattern in relation to an 8-week shoeing interval in the horse. EquineVet J 2005;37:536-40.

The Effects of Non-Steroidal Anti-Inflammatory DrugAdministration on Systemic and Muscle Inflammation inMature and Aged Horses

A.L. Wagner, R.B. Ennis, A. Betancourt, A.A. Adams, D.W.Horohov, and K.L. UrschelUniversity of Kentucky, Lexington, KY, USA

Introduction: Aging has been associated with chronic inflam-mation in humans, rodents [1], and horses [2]. Chronicinflammation, characterized by increased levels of inflammatorycytokines, has been linked to decreased physical performanceand increased incidence of injury[1,3] and has been associatedwith the development of sarcopenia in the elderly [1]. Non-steroidal anti-inflammatory drug (NSAID) administration isa common treatment for localized and systemic inflammation inhorses; however, little is known about the effects at the musclelevel. In rodents, NSAID administration has shown potential toblunt the effects of inflammation on sarcopenia, by increasingmuscle protein synthesis [4,5]. To date, little is known aboutmuscle levels of inflammatory cytokines in horses, especiallyaged horses. The objectives of this study were to characterizeboth systemic and muscle levels of inflammatory cytokines inmature and aged horses, and to determine the effects of 4 wk ofNSAID administration on both systemic and muscle levels ofinflammatory cytokines in mature and aged horses.Materials and Methods: Six aged (mean age 23.5; range 21-26 yold) and six mature (mean age 11; range 7-14 y old) horses ofmixed-breed were obtained from the University of KentuckyDepartment of Veterinary Sciences0 Maine Chance Farm. Withineach age group there were three geldings and three mares. Allanimals were group housed on dry lots, and individually fed installs two meals daily (at 0800 and 1500). Diets were designed tomeet the NRC [6] requirements for mature horses.Muscle biopsies were collected from the gluteus medius prior tothe initiation of NSAID administration (0 wk) and after 2 wk and 4wk of NSAID administration (2g/day phenylbutazone). Biopsies

were collected at a depth of 8 cm [7] following a 6-h period ofintermittent steady state feeding. Tissue samples were frozen inRNALater and stored at -20�C until RNAwas extracted. In order todetermine systemic levels of inflammatory cytokines, venipunc-ture blood samples were collected into PAXgene� Blood RNAtubes (Quiagen, Inc., Santa Clarita, CA) at 0 and 4wks. Tubes weregradually brought to �80�C and stored until assayed.Muscle samples were homogenized using the bead beatingtechnique into the RNA-Stat60 solution (Tel- Test, Friendswood,TX). Total RNA was then isolated using phenol-chloroformextraction (BioPhotometer, Eppendorf, Hamburg, Germany). TotalRNAwas isolated from PAXgene� tubes using the PAXgene� RNABlood Kit. The mRNA expression of inflammatory cytokines(IFN-g, IL-1b, IL-6, IL-10, and TNFa) and an internal housekeepinggene (b-GUS) was determined using real time-PCR [8].Changes in mRNA expression were calculated using the DDCTmethod [2], where DDCT ¼ (Avg. gene of interest CT – Avg. b-GUSCT)horse – (Avg. gene of interest CT – Avg. b-GUS CT)calibrator. Themean DCTaveraged for mature horses at the 0 wk sampling periodset as the calibrator for each individual cytokine. Results areexpressed as relative quantity (RQ) calculated as 2–DDCT. Theeffects of age, time on NSAID (time), and the interaction ofage*time on RQ for both muscle and systemic cytokines wereanalyzed using repeated measures in the MIXED models proce-dure of SAS (SAS Institute, Cary, NC). Sex was initially tested asa main effect, but was ultimately removed from the modelbecause it was not significant for muscle or systemic cytokines.Results and Discussion: There was no effect of age, time, orage*time interaction (P > .05) for muscle or systemic IL-6 andTNFa, and systemic IFN-g. However, there was an age*timeinteraction trend (P¼ .07) for muscle IFN-g; where mature horsesshowed a 11.3% decrease as opposed to a 33.8% increase withNSAID treatment in the aged horses. IL-1b decreased with time inboth the muscle (over 450%) and blood (over 30%; P ¼ .04) withNSAID treatment, regardless of age. IL-10 was the only inflam-matory cytokine to be affected by age; where IL-10 was 53.3%higher in the muscle of the aged horses (P ¼ .04), but showeda trend to be 73% lower in the blood (P ¼ .09) compared to themature horses. However, there was no effect of time (P > .05) onIL-10 in the muscle or blood, regardless of age.In agreement with a previous study [2], there were no agedifferences in blood IFN-g. Blood TNFa and IL-10 have beenshown to be higher in older horses [2]. However, this was not seenin the current study, which may be attributed to the mature horseage used in each study (11 compared to 4.52 y old). These resultsimply that increases in systemic inflammatory cytokines occurduring adulthood.Conclusion: The novel finding that IL-10, an anti-inflammatorycytokine, is higher in the muscle of the aged horse may imply thatsystemic levels of inflammatory cytokines play amore pivotal rolein sarcopenia in the aged than muscular levels; although, furtherstudies are needed to determine this.

References

[1] Degens H. The role of systemic inflammation in age-related muscleweakness and wasting. Scand. J. Med. Sci. Sports 2010;20(1):28-38.

[2] Adams AA, Breathnach CC, Katepalli MP, et al. Advanced age in horsesaffects divisional history of T cells and inflammatory cytokineproduction. Mech. Ageing Dev 2008;129(11):656-64.

[3] Penninx BW, Kritchevsky SB, Newman AB, et al. Inflammatorymarkers and incident mobility limitation in the elderly. J. Am. Geriatr.Soc 2004;52(7):1105-13.

[4] Balage M, Averous J, Remond D, et al. Presence of low-gradeinflammation impaired postprandial stimulation of muscle proteinsynthesis in old rats. J. Nutr. Biochem 2010;21(4):325-31.

[5] Rieu I, Magne H, Savary-Auzeloux I, et al. Reduction of low gradeinflammation restores blunting of postprandial muscle anabolismand limits sarcopenia in old rats. J. Physiol 2009;587(Pt 22):5483-92.

Abstracts / Journal of Equine Veterinary Science 31 (2011) 230-356296

[6] National Research Council. Nutrient Requirements of Horses. 6th reved. Washington D.C.: The National Academies Press; 2007.

[7] Urschel KL, Escobar J, McCutcheon LJ, et al. Feeding a high proteindiet following an 18-hour period of feed withholding increasesmTOR-dependent signaling in skeletal muscle of mature horses. Am.J. Vet. Res; 2010.

[8] Liburt NR, Adams AA, Betancourt A, et al. Exercise-induced increasesin inflammatory cytokines in muscle and blood of horses. Equine VetJ 2010;42(Suppl. 38):280-8.

Horse Intake Estimation of AM versus PM Grazing

S.J. Chavez, P.D. Siciliano, and G.B. HuntingtonNorth Carolina State University, Raleigh, NC, USA

Introduction: Throughout the day, forage composition changesdue to photosynthesis [1]. Soluble carbohydrates increase andcrude protein decreases throughout the day. Cattle and goatsprefer hays cut at sundown rather than sunup. Sheep did notdiffer in intake or digestility of hays cut at different timesthroughout the day; however cutting forage in the afternooncan be beneficial for cattle and goats for increased intake [2].Dry matter intake and dry matter digestibility increasethroughout the day due to decreased neutral detergent fiberand increased nonstructural carbohydrate [3]. The objective ofthis experiment was to estimate equine forage intake duringAM or PM grazing times.Materials and Methods: Protocols were approved by the NorthCarolina State University Animal Care and Use Committee.Six light horse breed geldings (588 � 39 kg) were randomlyassigned to an AM or PM grazing treatment for 14 d and thenswitched treatments for an additional 14 d from mid-April tomid-May 2010. Period 1 was the first 14 d, and period 2 was thefollowing 14 d. The AM treatment grazed from 0700-1300 and thePM group grazed from 1230-2030. Horses were housed in stalls(3.7 m x 12.2 m) when not grazing. Pastures were primarily non-toxic endophyte infected tall fescue (Festuca arundinacea).Pastures were divided into 4 quadrants of approximately 0.4 ha.Grass height measurements were collected and sampled at thebeginning and end of each period. Grass samples were freeze-dried and ground through a 5 mm screen on a Wiley mill, fol-lowed by a 0.5 mm screen on the Cyclotech for alkane analysis.Dotriacontane (C32) was dissolved in warm heptane and sprayedonto oats at approximately 430 mg C32/kg oats. Horses were fed200 g of oats twice daily at 0830 and 2030. Oats were not fed ondays 12-13. Fecal samples (50g) were collected twice daily at 0830and 2030 on days 8-12 of each 14 d period. Fecal samples weremixed for the day and frozen for subsequent analysis. Fecalsamples were then freeze-dried and ground through a 0.5 mmscreen on a Cyclotech. Grass and fecal samples were saponified inethanolic KOH and alkanes were extracted for subsequent GCanalysis [4]. Estimation of intake calculations was completed withC33 and C32 using an equation by Mayes et al [5]. Proc GLMprocedure of SAS was used to analyze the data with P < .05considered significant.Results and Discussion: Estimated DMI of grass was higher(P < .001) for period 2 (6.89 � .20 kg/d) than period 1 (5.31 � .20kg/d). This difference in periods was likely due to the change inforage composition. The fescue had matured and horses atepredominantly seed heads and stems during period 2. Seed headsand stems have lower nutritional value than leaves. PM treatmentintake (6.61 � .23 kg/d) was greater (P < .01) than AM treatment(5.59 � .23 kg/d). Cattle and goats prefer afternoon cut hay thathad greater concentrations of nonstructural carbohydrates thanhay cut in the morning [2]. Horses may have altered their eatingrate to consume more forage in the afternoon rather than spend

time in activities other than eating. Consuming high amounts ofnonstructural carbohydrates has been linked with laminitis,thus the increased consumption during PM grazing should beevaluated in more depth [6].This overall low prediction of forage intake was likely due to therestricted time of grazing at 8 h. Horses on average lost 20 kg overthe 28 d experiment. A loss of 20 kg in body weight would be theequivalent of mobilizing 154 Mcal from fat or 5.5 Mcal/d. Theaverage NRC estimate for DE for the horses in the experiment is19.58 Mcal/d [7]. The horses would be receiving approximately12.1 Mcal/d from grazing fescue grass and 1.5 Mcal/d fromsupplemental oats. This loss of body weight would be expectedwhen DE requirements are considered for this group of horses.Conclusion: Horses increased DMI of pasture in the afternoon vs.the morning similar to cattle and goats consuming afternoon cuthays. Horses also increased DMI when consuming seed heads andstems over leaves. Eight hours of grazing either in the morning orevening is not enough time for horses to meet their energyrequirement when grazing tall fescue grass in mid-April to mid-May; however caution should be taken with increased consump-tion of nonstructural carbohydrates in afternoon grazing horses.

References

[1] Fisher DS, Burns JC, Mayland HF. Ruminant selection amongswitchgrass hays cut at either sundown or sunup. Crop Sci 2005;45:1394-402.

[2] Burns JC, Mayland HF, Fisher DS. Dry matter intake and digestion ofalfalfa harvested at sunset and sunrise. J Anim Sci 2005;83:262-70.

[3] Burns JC, Fisher DS, Mayland HF. Diurnal shifts in nutritive value ofalfalfa harvested as hay and evaluated by animal intake and diges-tion. Crop Sci 2007;47:2190-7.

[4] Olivan M, Osoro K. Effect of temperature on alkane extraction fromfaeces and herbage. J Agric Sci 1999;132(3):305-12.

[5] Mayes RW, Lamb CS, Colgrove PM. The use of dosed and herbage n-alkanes as marker for the determination of herbage intake. J Agric Sci1986;107:161-70.

[6] Longland AC, Byrd BM. Pasture nonstructural carbohydrates andequine laminitis. J Nutr 2006;136:2099S-102S.

[7] NRC. Nutrient Requirements of Horses. Sixth ed. Washington, D.C.:National Academy Press; 2007.

Effect of Restricted Grazing on Dry Matter Intake Rate

E.C. Glunk, and P.D. SicilianoNorth Carolina State University, Raleigh, NC, USA

Introduction: Restricting grazing time is a strategy used todecrease caloric intake and related consequences, (e.g., obesity,insulin resistance and laminitis). A general assumption is thatrestriction of grazing time decreases forage intake in directproportionwith time. Preliminary evidence suggests this may notbe true. Dowler et al [1]. reported that the dry matter intake rate(DMIR) in the first 4 h of grazing was approximately double DMIRduring the second consecutive 4 h of grazing. Additionally, thisinitial accelerated DMIR allowed horses to consumeapproximately 55% of their daily maintenance DE in the first 4 hof grazing. It should be noted that these horses did not haveaccess to feed overnight prior to turn out, which likelyinfluenced the results. Nonetheless, an understanding of howDMIR and subsequent dry matter intake (DMI) change inresponse to restricted grazing is necessary to accuratelyregulate pasture intake. Therefore, an experiment was designedto test the hypothesis that pasture DMIR increases withrestriction of time allowed for grazing. The objective of thisstudy was to determine DMIR and DMI in horses having accessto pasture for four different time periods: 3, 6, 9, and 24 h.