2009: Nutriceuticals and other drugs used to enhance fertility in stallions Nutriceuticals and other drugs used to enhance fertility in stallions 1 S. P. Brinsko 2 Department of Large Animal Clinical Sciences, College of Veterinary Medicine, Texas A&M 3 University, College Station, TX, USA 4 Abstract 5 Until recently, there has been little information regarding the ability to improve semen 6 quality or fertility of stallions through alterations in diet. This paper presents evidence from work 7 in humans and other species, including the stallion, which indicates that dietary supplementation 8 with fatty acids, polyamines, vitamins or antioxidants may indeed have the potential of 9 improving semen quality, particularly in those individuals who have marginal semen quality 10 prior to supplementation. 11 Keywords: Stallion, semen quality, dietary supplementation, fatty acids 12 Introduction 13 Over the years, horsemen have been supplementing their animals’ diets with various 14 products in an attempt to enhance performance and overall well being. Most of these products 15 have been geared toward improving stamina, hair coat, joint function and hoof growth. 16 Historically, supplements touted to improve the breeding performance of stallions have not 17 proven to be efficacious. Recently however, supplements have become available that show real 18 promise in this regard. 19 Fatty Acids 20 Most fatty acids are straight-chain compounds, with an even number of carbon atoms. 21 Chain-lengths can range from two to 80 but most commonly range from 12 up to 24. With a 22 chain length from two to six, they are called short-chain, from eight to 10 they are called 23 355 medium-chain and 12 up to 24 they are called long-chain fatty acids. Among straight-chain fatty 24 acids, the simplest are referred to as saturated fatty acids. They have no double bonds and 25 cannot be altered by hydrogenation. When double bonds are present, fatty acids are said to be 26 unsaturated. They are called monounsaturated fatty acids (MUFA) if only one double bond is 27 present and polyunsaturated fatty acids (PUFA) when multiple double bonds are present. 28 The double bonds are counted from the methyl group determining the metabolic family, 29 noted by n-x (with n being the total number of carbon and x the position of the last double bond). 30 For example, linoleic acid is also named 18:2 n-6 in the shorthand nomenclature. Therefore this 31 PUFA has 18 carbon atoms, 2 double bonds and there are 6 carbon atoms from the last double 32 bond to the terminal methyl group. The number following "Omega-" in Omega-3 and Omega-6 33 fatty acids indicates the position of the first double bond, counting from the terminal methyl 34 group on the molecule. Hence, linoleic acid (18:2 n-6) is an Omega-6 fatty acid. Omega-3 fatty 35 acids cannot be converted to Omega-6 fatty acids or vice-versa. 36 Omega-3 fatty acids play an important role as structural membrane lipids in all cells. 37 They are also precursors to reactive substances such as prostaglandins and leukotrienes, and 38 possess anti-inflammatory, antiarrhythmic, antithrombotic and vasodilatory properties. There are 39 also data to indicate that dietary supplementation with Omega-3 fatty acids to horses may modify 40 the response to endotoxin by reducing the synthesis of potentially harmful cellular mediators. A 41 plethora of equine dietary supplements containing Omega-3 fatty acids and their precursors are 42 currently being marketed. 43 Semen from virtually all species examined contains relatively large amounts of lipid. 44 Semen lipids play a major role in motion characteristics, sensitivity to cold shock and fertilizing 45 356 capacity of sperm. Phospholipids are the major lipid components found in semen and they are 46 largely composed of PUFAs.1 While spermatozoa from all mammals contain high 47 concentrations of PUFAs,1,2 the combination and distribution of PUFA in semen varies among 48 species.3 For example, the distribution of long chain PUFAs in stallion spermatozoa is more 49 similar to boars than that of bulls or roosters.3 Major differences in the lipid content of bull 50 spermatozoa compared to those of boars and stallions are the relative amounts of 22:5 and 22:6 51 fatty acids. Spermatozoa of bulls have higher levels of 22:6 fatty acids whereas spermatozoa 52 from stallions and boars have higher levels of 22:5 fatty acids.3 Bulls and roosters produce 53 spermatozoa that are very resistant to cold shock and freeze well, whereas spermatozoa from 54 boars and stallions have very low tolerance to cold shock and in general, freeze poorly. 55 In particular, docosahexaenoic acid (DHA; 22:6 n-3, an Omega-3 fatty acid) is the major 56 22:6 PUFA in semen and docosapentaenoic acid (DPA; 22:5 n-6, an Omega-6 fatty acid) is the 57 major 22:5 PUFA. The majority of seminal PUFAs reside in spermatozoa. In men with poor 58 sperm motility, the level of DHA in seminal plasma as well as the ratio of Omega-3 to Omega-6 59 fatty acids in their spermatozoa was found to be significantly lower than in men with normal 60 semen quality.4 Studies in the boar and other species have shown that increasing the ratio of 61 DHA to DPA in semen increases fertilizing capacity and semen quality.2,5 Conversely, higher 62 levels of DPA relative to DHA results in reduced fertility.2 The potential role of DHA in 63 spermatozoal motility and membrane stability is supported by the fact that membranes high in 64 DHA are noted for their flexibility, compressibility, elasticity and deformability.6 65 Animals are unable to synthesize PUFAs from saturated or monounsaturated fatty acids. 66 Therefore, they must acquire them from precursor PUFAs in their diet. Transport of PUFAs 67 from the diet to semen has been shown to occur in a number of species including humans,7 fowl,5 68 357 boars,2 and rams.8 Vegetable oils, such as corn and soybean oil, contain high levels of linoleic 69 acid, the parent compound of DPA. Most proprietary equine rations are therefore, very high in 70 linoleic acid, as well as other Omega-6 series fatty acids and their precursors while the 71 precursors for Omega-3 fatty acids, such as DHA, are very low. A diet of this nature would 72 favor the formation of DPA over DHA since conversion of precursors to DPA and DHA uses the 73 same competitive enzymatic pathway. Since high DPA to DHA ratios in semen have been 74 associated with reduced sperm quality and fertility, typical equine diets with an overwhelming 75 availability of n-6 precursors could have a negative impact on quality of stallion semen and its 76 tolerance to cooling and freezing. 77 While simply supplementing the stallion’s diet with precursors to Omega-3 fatty acids 78 such as cod liver oil or flaxseed oil can increase the overall level of Omega-3 fatty acids in 79 semen, this may not result in the desired effects of improved semen quality. For example, 80 supplementing boar diets with cod liver oil did not improve the freezability of semen.9 However, 81 when a supplement containing pre-formed DHA and antioxidants was added to boar rations, 82 significant increases in semen quality and fertility were observed compared to boars fed a control 83 diet.2 Feeding the supplement to boars resulted in a number of benefits including a higher DHA 84 to DPA ratio in semen, as well as an increase in total spermatozoal number, spermatozoal 85 concentration, motility score, percentage of normal spermatozoa, and percentage of viable cells. 86 Cooling and freezing of spermatozoa can induce cellular injury, which is associated with 87 a disruption of membrane lipids, resulting in damage to mitochondria and loss of integrity of 88 both the plasma and acrosomal membranes. These events are accompanied by a loss of motility, 89 viability and fertilizing capacity of sperm, a phenomenon commonly referred to as “cold shock”. 90 Differences in the ability of spermatozoa from various animals to resist cold shock appear to be 91 358 related to their sperm membrane lipid composition.3 The lipid composition of spermatozoal 92 membranes not only influences the response of spermatozoa to cooling and freezing, but also 93 plays a major role in the physiologic changes leading to fertilization 94 Most breed registries have allowed the use of cooled, transported semen, and/or frozen-95 thawed semen for a several years. The use of shipped semen in the cooled-liquid or frozen state 96 offers many advantages to breeders. Unfortunately, there are many stallions that produce semen 97 that is unable to provide acceptable fertility after undergoing the rigors of cooling and storage, 98 and cryopreservation magnifies this reduction in fertility even further. This problem and the 99 promising results observed in pigs prompted equine researchers to investigate whether the 100 addition of a DHA-enriched dietary supplement could result in improvements in equine semen 101 quality. 102 Researchers at Texas A&M used eight stallions in a 2 x 2 crossover study to determine if 103 feeding a supplement rich in DHA would improve semen quality.10 The stallions were randomly 104 assigned to one of two treatment groups (n = 4/group). Within these groups, the stallions were 105 fed either their normal diet (control) or their normal diet top-dressed with 250 grams of a DHA-106 enriched supplement. The feeding trials lasted for 14 weeks after which a 14-week washout 107 period was imposed, during which only the normal diet was fed. Following the wash out period, 108 the treatment groups were reversed for another 14 week feeding trial. Feeding the supplement 109 resulted in a 3-fold increase in semen DHA levels and a doubling of the ratio of DHA to DPA in 110 the stallions’ semen. Spermatozoal motion characteristics in fresh semen were unaffected by 111 feeding the supplement and after 24 h of cooling and semen storage, total and progressive 112 spermatozoal motility also did not differ between treatment groups. However, the spermatozoa 113 from stallions fed the supplement exhibited higher velocity and straighter trajectory than those of 114 359 stallions being fed the control diet. Beneficial effects were more apparent after 48 hours of 115 cooling and storage, where increases in the percentages of spermatozoa exhibiting total motility, 116 progressive motility and rapid motility, were observed in the semen of stallions being fed the 117 supplement. Total sperm numbers and percentage of spermatozoa with normal morphology were 118 unaffected by treatment. However, when stallions were being fed the supplement, the sperm 119 concentration in their ejaculates was almost double that of when they were fed the control diet. 120 Therefore, it is possible that the observed improvements in semen quality after cooling and 121 storage could be attributed, at least in part, to the reduced exposure of sperm to seminal plasma 122 prior to and during processing. 123 In a subset of four stallions, whose progressive spermatozoal motility was <40% after 24 124 hours of cooling and storage when they were fed the control diet, feeding the supplement 125 resulted in improvements in mean progressive spermatozoal motility after both 24 hours and 48 126 hours of cooled storage. Feeding the supplement resulted in similar improvements in motion 127 characteristics being observed in frozen-thawed semen. Total spermatozoal motility, progressive 128 spermatozoal motility, and percentage of spermatozoa exhibiting rapid motility were 129 significantly higher in frozen-thawed semen of stallions being fed the supplement. 130 Despite increasing the level of DHA in semen by feeding the supplement, the level of 131 DPA did not decline. The level of DPA in semen remained higher than that of DHA so that the 132 DHA:DPA ratios were always less than one. Because the stallion’s rations were typical equine 133 formulations containing corn and soybean oils, which are the precursors that favor the pathway 134 to DPA, the authors speculate that more dramatic improvements in semen quality may be 135 observed if modifications in the main fat content of the diet are incorporated with the DHA 136 supplement. 137 360 The authors concluded that supplementing the diet of highly fertile stallions or those that 138 produce sperm that survive cooling does not appear warranted. However, stallions of marginal 139 fertility and those whose spermatozoa have poor tolerance to cooling and freezing would be 140 horses that might benefit most from being fed the supplement. Optimizing levels of DHA and its 141 precursors by altering the diet of marginally fertile stallions, may improve their semen quality 142 sufficiently enough to make them commercially viable for cooling or freezing. 143 Similar studies were carried out at the University of Arizona. In those studies, three 144 stallions were fed 550 grams of a DHA-enriched supplement in addition to their normal diet for 145 90 days.11 Dietary supplementation resulted in a 3.5-fold increase in semen DHA 146 concentrations. Unlike the study performed at Texas A&M, motion characteristics of 147 spermatozoa in cooled and frozen-thawed semen did not differ between supplemented and non-148 supplemented stallions. However, in this study dietary n-3 PUFA supplementation did result in a 149 46% increase in daily sperm output and a 15% increase in normal morphology. The most 150 dramatic improvements were observed for one stallion which had the lowest percentages of 151 morphologically normal and progressively motile spermatozoa prior to supplementation. 152 Recently, workers in Uruguay12 performed a 2 x 2 crossover study similar to the one performed 153 at Texas A&M10 with 3 stallions/group and treatment consisting of 30 grams of DHA being fed 154 as a supplement for 80 days. In that study, dietary DHA supplementation improved total sperm 155 numbers, sperm morphology and percentages of live sperm. Progressive motility in semen 156 cooled for 48 hours and frozen-thawed semen improved for some stallions. Consistent with the 157 other studies, the improvements were most noticeable for stallions that initially had poorer semen 158 quality. Improvements in semen quality have also been reported for stallions whose diets were 159 supplemented with rice oil.13 160 361 Polyamines 161 Both spermine and spermidine are polyamines found in all cells and thought to be 162 essential for replication, growth, and differentiation.14 It is believed that spermine and 163 spermidine are produced by the prostate and found in the semen of most mammals. Early 164 investigations documented increased spermatozoal motility in vitro when spermine was added to 165 spermatozoa from mice, rats, guinea pigs and rabbits.12 However, the exact physiologic role of 166 spermine and spermidine in semen remains a matter of debate and their functions may be 167 concentration dependent. Micromolar amounts of spermine in semen appear to enhance the 168 acrosome reaction15 while milimolar amounts appear to inhibit the acrosome reaction.15,16 169 In rams, ejaculates with spermatozoal motility greater than 85% had approximately two-170 fold higher spermine and total sperm polyamine content than ejaculates with lower motility. 171 Compared to spermatozoa from lambs the spermatozoa of mature rams had approximately three-172 fold higher levels of spermidine, spermine, and total polyamines.17 Lower levels of spermidine 173 are found in the seminal plasma of men with idiopathic asthenozoospermia (poor spermatozoal 174 motility) as well as those with asthenozoospermia associated with diabetes compared to 175 normozoospermic men.18 176 To date, no studies have been published which examined the effects of dietary 177 supplementation of polyamines on stallion semen. However, anecdotal information exists from 178 practitioners using an herbal supplement called “SpermAid”, which was originally marketed to 179 increase fertility and libido in stallions, as well as to increase testicular hormone production. The 180 active ingredients in this product are the phytochemicals spermine and spermidine, which are 181 found in radish leaves, radish root, cucumber fruit, and oats. Feeding of the supplement is 182 typically initiated three weeks prior to the breeding season. While significant improvements in 183 362 spermatozoal motility have not been reported with the use of this product, a number of slow 184 breeding stallions have anecdotally shown dramatic improvements in libido. 185 Vitamins and antioxidants 186 Dietary supplementation with antioxidants and vitamins has been shown to have 187 beneficial effect on semen quality. Many of these vitamins exert their beneficial effects through 188 their antioxidant properties. However, as with most supplements examined, conflicting evidence 189 exists regarding their efficacy. The conflicting results can be related to the species of the male 190 subjects in the different studies as well as the doses of individual supplements or combination of 191 supplements investigated. 192 Vitamins C and E are well known for their antioxidant properties and are those that have 193 been the most extensively examined. In rabbits, dietary supplementation with vitamin C, vitamin 194 E or a combination of vitamins C and E increased total spermatozoal output, spermatozoal 195 concentration and spermatozoal motility while decreasing dead and abnormal spermatozoa in the 196 ejaculate.19 Analogous findings have been reported for humans and boars.19,20 In humans, 197 vitamin C was associated with higher spermatozoal numbers and concentrations in ejaculates, 198 whereas vitamin E appeared to exert its effects by improving spermatozoal motility.20 Similarly, 199 there was tendency for semen production to be greater for boars supplemented with water soluble 200 vitamins, with the effect being less in boars supplemented with fat soluble vitamins.21 While the 201 intake of high levels of antioxidant vitamins was associated with better semen quality, moderate 202 intake did not appear to be effective.20 While other investigators were unable to demonstrate any 203 improvements in conventional semen quality parameters from infertile men,22,23 supplementation 204 with Vitamins C and E did result in a significant reduction in DNA fragmentation.23 205 363 Work in the stallion is more limited. In 1978, a German investigator gave 10 stallions of 206 three different breeds an emulsion containing Vitamins A and E for eight weeks.24 207 Improvements in ejaculate volume, spermatozoal concentration, spermatozoal morphology and 208 spermatozoal motility, including post-thaw motility were reported. However, the improvements 209 were not universal and appeared to differ among breeds. More recently, Russian workers 210 formulated a complex feed additive that included vitamins A, D and E.25 They reported 211 improved spermatozoal motility in fresh semen and that spermatozoa remained viable longer 212 after freezing and thawing. 213 Another antioxidant, showing promise for improving semen quality is L-carnitine 214 (levocarnitine). Along with its antioxidant properties, L-carnitine is essential for mitochondrial 215 energy metabolism. Both L-carnitine and L-acetyl-carnintine are found in high concentrations in 216 the epididymis and both forms are accumulated by spermatozoa.26 In men with 217 asthenozoospemia, combined treatment with L-carnitine and L-acetyl-carnitine was effective in 218 increasing spermatozoal motility.24 The most significant improvements were seen in men with 219 the lowest numbers of motile spermatozoa prior to treatment. Feeding L-carnitine to boars 220 resulted in higher semen volumes and spermatozoal concentrations thereby increasing the total 221 number of available spermatozoa in ejaculates for artificial insemination.27 222 223 Conclusions 224 It is clear that dietary alterations can have an effect on semen quality and in some cases, 225 fertility. Controlled studies in stallions are few, but those investigating fatty acids, in particular 226 Omega-3 fatty acids such as DHA, have shown real potential. Based on work in other species, 227 364 further studies involving optimal levels of individual supplements and combinations of 228 supplements which could act synergistically to improve stallion semen quality are needed. 229 References 230 1. Scott JW: Lipid metabolism of spermatozoa. J Reprod Fertil 1973;18 (Suppl):65-76. 231 2. Penny PC, Maldjian A, Noble RC: An enhancement of boar fertility and reproductive 232 performance. Proc14th Intl Cong Anim Reprod 2000; p. 109 (abstr). 233 3. Parks JE, Lynch DV: Lipid composition and thermotrophic phase behavior of boar, bull, 234 stallion, and rooster sperm membranes. Cryobiology 1992;29:255-266. 235 4. Conquer JA, Martin JB, Tummon I, et al: Fatty acid analysis of blood, serum, seminal 236 plasma, and spermatozoa of normozoospermic vs. asthenozoospermic males. Lipids 237 1999;34:793-799. 238 5. Blesbois E, Lessire M, Grasseau I, et al: Effect of dietary fat on the fatty acid composition 239 and fertilizing ability of fowl semen. Biol Reprod 1997;56:1216-1220. 240 6. Salem N, Kim H-Y, Yergey JA: Docosohexaenoic acid: membrane function and metabolism, 241 In: Simopolulos AP, Kifer RR, Martin RE, editors. Health effects of polyunsaturated fatty 242 acids in seafoods. New York: Academic Press; 1986, p 263-321. 243 7. Conquer JA, Martin JB, Tummon I, et al: Effect of DHA supplementation on DHA status and 244 sperm motility in asthenozoospermic males. Lipids 2000;35:149-154. 245 8. Drokin SI, Vaisberg TN, Kopeika EF, et al : Effect of cryopreservation on lipids and some 246 physiological features of spermatozoa from rams pastured in highlands and in valleys. 247 Cytobios 1999;100:27-36. 248 9. Paulenz H, Taugbol O, Kammisrud E, et al: Effect of dietary supplementation with cod liver 249 oil on cold shock and freezability of boar semen. Reprod Domest Anim 1999;34:431-435. 250 365 10. Brinsko SP, Varner DD, Love CC, et al: Effect of feeding a DHA-enriched nutriceutical on 251 the quality of fresh, cooled and frozen stallion semen. Theriogenology 2005;63:1519-1527. 252 11. Harris MA, Baumgard LH, Arns, MJ,et al: Stallion spermatozoa membrane phospholipid 253 dynamics following dietary n-3 supplementation. Anim Reprod Sci 2005;89:234-237 (Abstr). 254 12. Elhordoy DM, Cazales N, Costa G, et al: Effect of dietary supplementation with DHA on the 255 quality of fresh, cooled and frozen stallion semen. Anim Reprod Sci 2008;107:18 (abstr). 256 13. Arlas TR, Pederzolli CD, Terraciano PB, et al: Sperm quality is improved feeding stallions 257 with a rice oil supplement. Anim Reprod Sci 2008;107:5 (abstr). 258 14. Tabor CW, Rosenthal SM: Pharmacology of spermine and spermidine: some effects of 259 animals and bacteria. J Pharmacol Exp Ther 1956;I 16:139-155. 260 15. Rubenstein S, Lax Y Shalev Y, et al: Dual effect of spermine on acrosomal exocytosis in 261 capacitated bovine spermatozoa. Biochimica et Biophysica Acta 1995;1266:196-200. 262 16. Rubenstein S, Breitbart H: Role of spermine in mammalian sperm capacitation and acrosome 263 reaction. Biochem. J 1991;278:25-28. 264 17. Melendrez CS, Ruttle JL, Hallford DM, et al: Polyamines in ejaculated ram spermatozoa and 265 their relationship with sperm motility. J Androl 1992;13:293-296. 266 18. Morales ME: Progressive motility increase caused by L-arginine and polyamines in sperm 267 from patients with idiopathic and diabetic asthenozoospermia. Ginecol Obstet Mex 268 2003;71:297-303. 269 19. Yousef, MI. Effect of ascorbic acid and vitamin E supplementation on semen quality and 270 biochemical parameters of male rabbits. Anim Reprod Sci 2003;76 :99. 271 20. Eskenazi B, Kidd S A, Marks A R, et al: Antioxidant intake is associated with semen quality 272 in healthy men. Human Reprod 2005;20:1006-1012. 273 366 21. Audet I, Laforest J P, Martineau G P, et al: Effect of vitamin supplements on some aspects of 274 performance, vitamin status, and semen quality in boars. J Anim Sci 2004;82:626-633. 275 22. Rolf C, Cooper T G, Yeung C H, et al: Antioxidant treatment of patients with 276 asthenozoospermia or moderate oligoasthenozoospermia with high-dose vitamin C and 277 vitamin E: a randomized, placebo-controlled, double-blind study. Human Reprod 278 1999;14:1028-1033. 279 23. Greco E, Iacobelli M, Rienzi L, et al: Reduction of the incidence of sperm DNA 280 fragmentation by oral antioxidant treatment. J Androl 2005;26:349-353. 281 24. Prinz K: Effect of a vitamin A –E emulsion on stallion semen. Tierarztliche Umschau 282 1978;33:27-30. 283 25. Kalashnikov VV, Ugadchikov ST: The significance of biologically active substances in the 284 feeding of stud stallions. Russian Agricultural Sciences 2001-2002;4:40-43. 285 26. Lenzi A, Sgro P, Salacone P, et al: A placebo-controlled double-blind randomized trial of the 286 use of combined L-carnitine and L-acetyl-carnitine treatment in men with 287 asthenozoospermia. Fertil Steril 2004;81:1578-1584. 288 27. Wahner M,. Geyer M, Hallfarth G, et al: The influence of vitamin emulsion with L-carnitine 289 on the sperm qualities of AI-boars. 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