Semen Quality and Sperm Function Loss by Hypercholesterolemic Diet Was Recovered by Addition of Olive Oil to Diet in Rabbit

Fat increment (0.05% cholesterol, chol) in standard diet promoted a significant increase in serum and sperm membrane chol, which ultimately altered membrane-coupled sperm specific functions: osmotic resistance, acrosomal reaction, and sperm capacitation in White New Zealand rabbits. These changes were also associated with a reduction in motility percentage and appearance of abnormal sperm morphology. The present study was carried out to evaluate the effect of dietary olive oil (OO, 7% v/w) administration to several male hypercholesterolemic rabbits (hypercholesterolemic rabbits, HCR) with altered fertility parameters. These HCR males were achieved by feeding normal rabbits with a high-fat diet (0.05% chol). HCR were associated with a modest non-significant increase in body weight (standard diet, 4.08±0.17 Kg, versus high-fat diet, 4.37±0.24 Kg). Hypercholesterolemic rabbits presented a marked decrease in semen volume, sperm cell count, and percentage of sperm motility, associated with a significant increase in sperm cell abnormalities. Moreover, sperm capacitation measured by the characteristic phosphorylated protein pattern in and induced acrosomal reaction were also altered suggesting sperm dysfunction. However, the administration of OO (for 16 weeks) to rabbits that were fed with 50% of the high-fat diet normalized serum chol. Curiously, OO supply succeeded to attenuate the seminal and sperm alterations observed in HCR group. Administration of OO alone did not cause any significant changes in above mentioned parameters. These data suggest that OO administration to HCR male rabbits recovers the loss of semen quality and sperm functionality.Fil: Saez Lancellotti, Tania Emilce Estefania. Universidad del Aconcagua. Facultad de Ciencias Médicas; Argentina; Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Mendoza. Instituto Histología y Embriología de Mendoza; Argentina;Fil: Boarelli, Paola Vanina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Mendoza. Instituto Histología y Embriología de Mendoza; Argentina; Universidad del Aconcagua. Facultad de Ciencias Médicas; Argentina;Fil: Romero, Aida Lorena. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Mendoza. Instituto Histología y Embriología de Mendoza; Argentina;Fil: Funes, Abi Karenina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Mendoza. Instituto Histología y Embriología de Mendoza; Argentina;Fil: Cid Barria, Macarena. Universidad del Aconcagua. Facultad de Ciencias Médicas; Argentina;Fil: Cabrillana, María Eugenia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Mendoza. Instituto Histología y Embriología de Mendoza; Argentina; Universidad del Aconcagua. Facultad de Ciencias Médicas; Argentina;Fil: Monclus, Maria de Los Angeles. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Mendoza. Instituto Histología y Embriología de Mendoza; Argentina; Universidad del Aconcagua. Facultad de Ciencias Médicas; Argentina;Fil: Simón, Layla Yamila. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Mendoza. Instituto Histología y Embriología de Mendoza; Argentina;Fil: Vincenti, Amanda Edith. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Mendoza. Instituto Histología y Embriología de Mendoza; Argentina;Fil: Fornes, Miguel Walter. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Mendoza. Instituto Histología y Embriología de Mendoza; Argentina; Universidad del Aconcagua. Facultad de Ciencias Médicas; Argentina

Experimental groups.

<p>Twenty New Zealand rabbits were initially distributed at random in three groups: NCR, HCR, OO at 5 months of age (corresponding to experimental time (et) = 0 months). After 4 months, NCR continued with normal diet (ND) and HCR was split in two subgroups: HCR and ½ HCR. HCR continued with experimental diet 1 (ED1) and the last group was fed with ED2. Four months later (et = 8 m) ½ HCR was divided again in two (subgroup II): ½ HCR (fed with ED2) and ½ HCR+½ OO (fed with ED3). m = months, n = number of experimental animals. More details in the text.</p

General characteristics of fresh rabbit semen samples (mean ± SEM).

*<p>p<0.05, n = 25.</p

Effect of dietary treatment on sperm membrane cholesterol.

<p>Fluorescence micrographs showing cholesterol content in plasma membrane of ejaculated rabbit spermatozoa detected by filipin probe. Images correspond to filipin-stained sperm cells (×630) from NCR (A), OO (B) HCR (C), ½ HCR (D) and ½ HCR+½ OO (E). Compare the strong signal detected in HCR (C) with the one from the ½ HCR+½ OO (E). The experiment was performed at least three times with 20 sperm from each animal.</p

Impact of diet on sperm functionality: membrane response to hypo-osmotic swelling test.

<p>Bars represent the percentage (mean ± SEM) of spermatozoa swollen from NCR (black bar), OO, HCR, ½ HCR and ½ HCR+½ OO (white bars). The experiment was performed at least three times with each animal. n = 25 samples. *** = significantly different from NCR (p<0.001).</p

Effect of diet on blood cholesterol.

<p>Bars represent the mean ± SEM of plasma cholesterol concentration from NCR, OO, HCR, ½ HCR and ½ HCR+½ OO during the last 3 months of experiment. n = 45 samples. *** indicates significantly different from NCR (p<0.001).</p

Impact of diet on sperm functionality: sperm capacitation and acrosome reaction.

<p>Protein tyrosine phosphorylation (A) and acrosomal exocytosis index (B) of rabbit spermatozoa. A: phospho-Y proteins from control (NCR), HCR and ½ HCR+½ OO showed different patterns ranging from one band (non-capacitated, culture medium without BSA (−), approximately 60 kDa) to many bands (capacitated with BSA (+), from over 20 to 100 kDa). Notice that capacitated sperms from HCR decreased the p-Y protein pattern compared with NCR, and ½ HCR+½ OO shows a p-Y pattern resembling control (NCR) conditions. The experiment was performed at least three times and representative blot is shown. B: Bars represent AR index of spermatozoa from NCR, ½ HCR, ½ HCR+½ OO and OO after 10 µM progesterone incubation. AR index corresponds to normalized data (see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0052386#s2" target="_blank">Materials and Methods</a>). n = 25 samples. *** = significantly different from NCR (p<0.001).</p