Reproductive Effects of Feeding Gossypol and Vitamin E to Bulls

Transcription

1 Reproductive Effects of Feeding Gossypol and Vitamin E to Bulls J. Velasquez-Pereira P.J. Chenoweth L.R. McDowell C.A. Risco C.A. Staples D. Prichard F.G. Martin M.C. Calhoun S.N. Williams N.S. Wilkinson 1 Cottonseed meal fed to Holstein bulls at a concentration to provide 6.4 mg of free gossypol per/lb of body weight from 6 to 16 mo of age resulted in increased sperm abnormalities, decreased sperm production, and adversely affected some aspects of sexual behavior, with vitamin E reversing the negative effects of gossypol on reproductive measurements. Summary An experiment was conducted to determine the effects of long-term feeding of cottonseed meal on reproductive traits of Holstein bulls. Twenty-four Holstein bulls approximately 6 mo of age were placed on the following treatments: 1) soybean meal + corn (CON); 2) cottonseed meal + corn (GOS); and 3) cottonseed meal + corn + 4,000 IU vitamin E per bull (G+4E). The GOS and the G+4E diets were formulated to supply 6.4 mg of free gossypol per lb. body weight. These bulls had been in a previous experiment that evaluated the effects of feeding the same type of diets, but from 2 wk to 6 mo of age. Percentage of motility, percentage of normal and live sperm, and daily sperm production were less (P <.05) in the GOS than in the other groups. At 12 and 16 months of age, bulls were given two assessments for sex drive traits. Bulls that received gossypol exhibited less sexual activity (P <.05) at the first test than bulls in other treatments. Vitamin E supplementation in bulls that received gossypol improved the number of mounts in the first test and the time to first service in the second test. There was a trend of gossypol to decrease and vitamin E to improve libido score. The results of GOS first libido test may indicate lack of sexual maturity, which agrees with sperm production data. At the time of first test (12 mo of age), none of GOS, two of CON, and six of G+E4 bulls had reached puberty on the basis of experimental protocol. Long-term feeding of gossypol to Holstein bulls negatively affected some reproductive traits; however, vitamin E supplementation countered these adverse effects and even improved these traits. Introduction Even though whole cottonseed (WCS) and cottonseed meal (CSM) are important sources of protein for ruminants, they contain the toxic polyphenolic pigment gossypol, which adversely affects liver function, erythrocyte oxygen-carrying or -releasing capacity, respiration rate, feed intake, and production and reproductive capacity. Even though mature ruminants seem to have a large capacity to detoxify gossypol, toxicosis resulting from consumption of this compound has been observed. Gossypol intake by mature ruminants may overwhelm ruminal detoxification and become absorbed at potentially toxic concentrations. In ruminants, gossypol has adversely affected sperm production, associated with damage to the spermatogenic epithelium, leading to reduced germinal cell layers. The present study was designed to evaluate the long-term feeding of cottonseed meal and the use of vitamin E to counteract potential gossypol effects on reproductive development of dairy bulls. Procedure This experiment used eight bulls for each of the following dietary treatments: 1) CON, a supplement based on SMB + corn IU of vitamin E/lb of dietary DM; 2) GOS, a supplement based on CSM + corn IU of vitamin E/lb of

2 dietary DM; and 3) G+4E, a supplement based on CSM + corn + 4,000 IU of vitamin E per bull per day. These bulls had been in a previous experiment that evaluated the effects of feeding the same type of diets, but from 2 wk to 6 mo of age. Supplements GOS and G+4E were formulated to supply 6.4 mg of free gossypol per lb body weight. The bulls were housed in 12 pens, two bulls per pen and four pens per treatment, from 6 to 16 months of age. The protocol for bull procedures had been approved by the University Animal Use Committee. Testicular Measurements and Semen Evaluation. Beginning at approximately 12 mo of age and continuing at weekly intervals for 9 wk, a breeding soundness evaluation (BSE) program was carried out on each bull. Semen was analyzed for sperm concentration and motility. Age at puberty was determined when bulls provided an ejaculate containing 50 X 106 sperm, with at least 10% showing progressive motility. As soon as sperm production had been established, spermatozoa were assessed for morphological variation. Libido Test. Before the beginning and at the end of the BSE, bulls were evaluated for sex drive. Briefly, two nonpregnant females of size similar to that of the bulls were placed in service crates located approximately 5 yards apart in a pen. Two bulls were admitted to the pen and observed for 10 min. This test was repeated the following day with, in most cases, cohorts and restrained females other than those used the previous day. Results Calculated free gossypol intake per animal in this trial ranged from 2.3 g at the beginning of the experiment to 5.3 g at the end. Testicular Measurements and Semen Evaluation. Average percentages of normal and live sperm were lower (P <.05) in bulls supplemented with GOS than in bulls fed G+4E and CON (Table 1). The reduction of morphologically normal sperm was accompanied by a reduction (P <.1) in the percentage of motile sperm from bulls fed the GOS supplement (Table 1). Feeding vitamin E (G+4E) counteracted the negative effect of CSM on the percentages of normal, live, and motile sperm. The percentage of abnormal sperm midpieces, as evaluated with differential phase microscopy, was greater (P <.05) in GOS fed bulls than in bulls on the other two treatments (Table 1). Electron microscopy from bull sperm in the GOS treatment in this study revealed gaps in the mitochondrial helix of the midpiece. Electron microscopy of the testis revealed that extensive mitochondrial damage had occurred in the testis for bulls supplemented with GOS, which presented as extensive vacuolation and a loss of cristae. Histological examination of the testes revealed that six of eight bulls supplemented with GOS had irregular and degenerative germ cell layers that also showed vacuolation and reduced spermatid numbers. Vitamin E supplemented bulls (G+4E) had less histological damage; only one animal presented vacuolation and reduced spermatid numbers. Daily sperm production per gram (DSPG) of parenchyma (106/g of testicular parenchyma) was reduced (P <.05) in animals supplemented with GOS (Table 2). The same trend was observed for daily sperm production (DSP). Vitamin E supplementation (G + 4E) counteracted (P <.05) the reduction in sperm production caused by gossypol in CSM. The depression of spermatogenesis is a known effect of gossypol on male reproduction. In this study, feeding vitamin E to bulls that received 6.4 mg of free gossypol/lb of body weight increased the percentage of normal sperm, percentage of motility, DSPG, and DSP, and it decreased the percentage of primary and midpiece (DIC) abnormalities. Vitamin E may have reduced the effect of gossypol on these variables by reducing the damage of reactive species of oxygen or by sparing the effect of enzymes involved in the antioxidant system of the testicle. The effect of vitamin E in protecting the sperm membrane against lipid peroxidation may be more important in late spermiogenesis when sperm discards most of their cytoplasm, leading to lowered concentrations of cytoplasmic defensive enzymes. Results of the libido tests are presented in Table 3. Bulls fed GOS exhibited less sexual activity (P <.05) at the first test than bulls in other treatments. Vitamin E supplementation to bulls that were receiving gossypol (G+4E) improved number of mounts (NOM) in the first test and time to first mount (TOS) in the second test. There was a trend for gossypol (GOS) to decrease and for vitamin E (G+4E) to improve libido score (LSC). The lower results from GOS group's first test may indicate a relative lack of sexual maturity in this group; this agrees with sperm production data. At the time of first test (12 mo of age), none of GOS, two of CON, and six of G + 4E bulls had reached puberty on the basis of experimental protocol (ejaculate with sperm and at least 10% motility). The effect of vitamin E supplementation on some measurements of sexual behavior may be related to its antioxidant properties and its accumulation in tissues with high steroidogenic activity and, thus, high reactive oxygen species production, such as in testicular tissue.

3 Vitamin E's effect on the metabolism of gossypol for growing bulls was investigated. Vitamin E seemed to confer protection against the detrimental effects of gossypol on sperm production and quality. The protection mechanism may be a direct physical interaction of gossypol or its metabolites with vitamin E in lipid membranes. The latter supposition is supported by the fact that no gossypol is present in seminal plasma of G + 4E animals. 1 J. Velasquez-Pereira, Ministry of Agriculture, Managua, Nicaragua; P.J. Chenoweth, Kansas State University; M.C. Calhoun, Texas A&M University; S.N. Williams, Hoffmann-LaRoche L.R. McDowell, C.A. Risco, C.A. Staples, D. Prichard, F.G. Martin, N.S. Wilkinson, Department of Animal Sciences, Gainesville;. Table 1. Effects of cottonseed mean (CSM) and vitamin E on seminal characteristics of dairy bulls a Item CON SE c GOS SE G+4E SE No. of bulls 8 6 d 8 Motility, % 63.7 i j i 6.1 Live, % 42.0 g h g 5.4 Normal, % 68.3 g h g 6.0 Primary abnormalities, % e 24.4 h g h 5.2 Secondary abnormalities, % e Abnormal (DIC), % f 4.1 h g h 1.1 b CON = soybean meal + corn IU of vitamin E/lb of dietary DM; GOS = CSM + corn IU of vitamin E/lb of dietary DM; G+4E = CSM + corn + 4,000 IU of vitamin E/day. c Standard error of the least squares means. d Two of the eight bulls did not produce semen during the experimental period. e Primary = proximal droplet, abnormal acrosome, coiled tail, abnormal head, and abnormal midpiece. Secondary = distal cytoplasmic droplet, kinked tail, and detached head. f Midpiece abnormalities evaluated in isotonic formol saline using differential phase microscopy (Differential Interference Contrast, DIC). g, h Means in a row with different superscripts differ (P <.05). i, j Means in a row with different superscripts differ (P <.1).

4 Table 2. Effects of cottonseed mean (CSM) and vitamin E on sperm production, and testicular micromineral and alpha tocopherol (T) concentrations of dairy bulls a Item CON GOS G+4E SE DSPG 10 6 /g d 14.6 f 10.2 g 17.6 f 1.0 DSP 10 9e 3.2 f 2.2 g 4.1 f.3 Cu, µg/g DM Fe, µg/g DM Zn, µg/g DM T, µg/g wet tissue 4.76 h 6.85 g 16.4 f.59 b CON = soybean meal + corn IU of vitamin E/lb of dietary DM; GOS = CSM + corn IU of vitamin E/lb of dietary DM; G+4E = CSM + corn + 4,000 IU of vitamin E/day. c Standard error of the least squares means. d DSPG = daily sperm production per gram of parenchyma. e DSP = daily sperm production. f,g,h Means in a row with different superscripts differ (P <.05).

5 Table 3. Effects of cottonseed mean (CSM) and vitamin E on sex drive performance of dairy bulls during two libido tests a Performance test c CON SE GOS SE G+4E SE LSC NOM NOS TOM, sec TOS NDX First Second First 5.7 e,f f e 1.2 Second First Second First Second 38.0 e e,f f 5.0 First Second e,f e f 59.8 First 1.2 h g h.8 Second b CON = soybean meal IU of vitamin E/lb of dietary DM; GOS = CSM + corn IU of vitamin E/lb of dietary DM; G+4E = CSM + corn + 4,000 IU of vitamin E/bull/day. c LSC = libido score; NOM = number of mounts; NOS = number of services; TOM = time to first mount; TOS = time to first service; NDX = sexual inaction time (min). Ten minutes allowed per test. d Standard error of the least squares means. e,f Means in a row with different superscripts differ (P <.1). g, h Means in a row with different superscripts differ (P <.05).