157 Dietary l-arginine supplementation affects boar seminal plasma proteome

Abstract

There are numerous benefit claims for the supplementation of l-arginine in animal production systems. Its positive effect on fertility has been widely characterised in females of various species; however, the effects on the male reproductive system remain debatable with still unfolding molecular mechanisms. Here we investigated the effects of a dietary l-arginine supplementation on boar semen production outcomes, sperm characteristics, and seminal plasma proteome. Nine mature (20 months of age) Nebraska Index Line boars were individually housed and randomly assigned to a soybean meal-based diet containing 0.77% standard l-arginine (control, n = 4) or 1.77% high l-arginine (n = 5). Boar semen was collected weekly during the 6-week dietary arginine trial. Semen production outcomes (volume and total spermatozoa) and sperm motion (total and progressive motility) and morphology characteristics were subsequently analysed using a computer-assisted sperm analyzer (CEROS II). On Week 6, the seminal plasma of each boar was obtained by centrifugation at 4°C for proteome analysis. The clarified seminal plasma was precipitated; the purified proteins were resuspended in an appropriate buffer and loaded (300 μg) onto an immobilized pH gradient strip (isoelectric point 3-10) for the first-dimension electrophoresis. The second-dimension was subsequently performed (4-20% gradient gel), and all gels were stained with Coomassie R250 dye. The PDQuest software (Bio-Rad, Hercules, CA, USA) was used to detect all significantly differentially expressed protein spots (Student’s t-test with P < 0.05). Two-way ANOVA repeated measurements (SPSS Statistics, Chicago, IL, USA) was used to analyse dependent (semen outputs and sperm characteristics) and independent (week) variables. A difference was significant at P < 0.05. During the l-arginine-consumption trial, semen volumes and total spermatozoa were not significantly affected (P > 0.05). Similarly, various sperm motility (total and progressive motility) and velocity characteristics were not affected (P > 0.05). By the end of the feed trial, the average (mean ± standard error of the mean) semen volume (389 ± 40 mL), the total spermatozoa per ejaculate (39 ± 8 × 10⁹), the total (82 ± 2%) and progressive (62 ± 3%) sperm motility, the sperm velocity (e.g. average path: 98 ± 7 μm s⁻¹), and sperm morphology (e.g. distal droplet: 5 ± 1%) parameters of the control group were not significantly different from the group supplemented with high l-arginine: 393 ± 35 mL, 47 ± 4 × 10⁹, 82 ± 2%, 61 ± 3%, 92 ± 8 μm s⁻¹, and 5 ± 0%, respectively (P > 0.05). In contrast, the proteome profiles of seminal plasma harvested on the sixth week of diet revealed various changes, characterised by the significantly increased expression of 8 protein spots in seminal plasma samples derived from arginine-fed boars (P < 0.05). These results indicate that dietary supplementation of l-arginine does not affect the semen outputs, neither the sperm motility characteristics nor their morphology. However, the proteome profile of the seminal plasma was changed by the presence of l-arginine. The findings may have significant implications for boar fertility, and the identification of these proteins is ongoing.

This work was supported by USDA-Agricultural Research Service Biophotonics Initiative #58-6402-3-018.