Fertility-associated biochemical components in seminal plasma and serum of buffalo (Bubalus bubalis) bulls

The present study looks for components in seminal plasma (SP) and/or serum that are closely related to in vivo fertility of buffalo bulls. Fourteen healthy mature buffalo bulls were classified according to their in vivo fertility into fertile (n = 10) and subfertile (n = 4) groups. Semen and serum samples were collected from all animals for 12 replicates. The collected ejaculates were examined for sperm characteristics before being centrifuged to collect SP for hormonal (FSH, LH, testosterone, and IGF-1), biochemical [total antioxidant capacity (TAC), catalase (CAT), glutathione peroxidase (GPx), nitric oxide (NO), malondialdehyde (MDA), fructose, total protein, albumin, triglycerides, cholesterol, and high-density lipoprotein (HDL)] and proteomic (SDS-PAGE) analyses. Likewise, serum levels of FSH, LH, testosterone, IGF-1, glucose, total protein, albumin, triglycerides, cholesterol, and HDL were determined. All sperm characteristics and the majority of sperm kinematics were (P < 0.01) different between fertile and subfertile groups. Seminal and serum levels of FSH, LH, testosterone, and IGF-1 were higher (P < 0.01) in the fertile group, but only seminal fructose, total protein, albumin, triglycerides, cholesterol, and HDL were higher (P < 0.01) in the fertile group. Moreover, the fertile group had greater TAC, CAT, GPx, and NO, but the subfertile group had greater MDA. Protein bands of 14, 15, 26, 30, and 55 kDa were larger and denser in the SP of the fertile group but were smaller and faint to absent in that of the subfertile group. Also, the protein fractions of detected protein bands demonstrated a substantial influence of fertility on those of 16, 26, 30, and 55 kDa. In conclusion, sperm characteristics and kinematics with serum, and/or seminal hormonal and biochemical components, should be evaluated for reliable prediction of buffalo bull fertility. Furthermore, protein bands of 26, 30, and 55 kDa may represent fertility-associated proteins in buffalo bull SP

Effect of Kisspeptin on the Developmental Competence and Early Transcript Expression in Porcine Oocytes Parthenogenetically Activated with Different Methods

Recent studies showed the modulatory effect of kisspeptin (KP) on calcium waves through the cell membrane and inside the cell. Spermatozoon can induce similar ooplasmic calcium oscillations at fertilization to trigger meiosis II. Here, we evaluated the effect of KP supplementation with 6-dimethylaminopurine (6-DMAP) for 4 h on embryonic development after oocyte activation with single electric pulse, 5 µM ionomycin, or 8% ethanol. Compared to control nonsupplemented groups, KP significantly improved embryo developmental competence electric- and ethanol-activated oocytes in terms of cleavage (75.3% and 58.6% versus 64% and 48%, respectively, p<0.05) and blastocyst development (31.3% and 10% versus 19.3% and 4%, respectively, p<0.05). MOS expression was increased in electrically activated oocytes in presence of KP while it significantly reduced CCNB1 expression. In ionomycin treated group, both MOS and CCNB1 showed significant increase with no difference between KP and control groups. In ethanol-treated group, KP significantly reduced CCNB1 but no effect was observed on MOS expression. The early alterations in MOS and CCNB1 mRNA transcripts caused by KP may explain the significant differences in the developmental competence between the experimental groups. Kisspeptin supplementation may be adopted in protocols for porcine oocyte activation through electric current and ethanol to improve embryonic developmental competence

Desalted and lyophilized seminal plasma increases protein tyrosine-phosphorylation of frozen-thawed bull spermatozoa incubated with a cell-permeable cyclic AMP (cAMP) analog (cBiMPS)

The present study investigates the effect of desalted seminal plasma (SP) added to semen extender on hyperactivated motility and protein tyrosine-phosphorylation (PTP) of bull spermatozoa. The SP was harvested by centrifugation and desalted using Sephadex G-25 columns in order to be added to semen extender at 0 (control), 2.5, 12.5 and 25 mg/ml. Frozen-thawed spermatozoa were incubated with a cellpermeable cyclic AMP (cAMP) analog (cBiMPS) and examined subjectively for hyperactivated motility and for PTP by Western blotting. Although, the added SP sustains sperm motility at all incubation times especially in the presence of cBiMPS but without significant difference from the control samples. Moreover, total sperm motility of 12.5 and 25 mg/ml in the presence of cBiMPS at 60, 120 and 180 min were similar (P ≥ 0.05). Surprisingly, cBiMPS-incubated spermatozoa in the presence of desalted SP were capable of exhibiting hyperactivated motility. Addition of SP increased and prolonged intracellular cAMP-induced PTP and in total 21 phosphorylated proteins with molecular weight ranging from 10 to ＞230 kDa were detected. The most prominent tyrosine-phosphorylated proteins (TPPs) were of 32, 38, 74 and 80 kDa which were more predominant in fertile bulls than subfertile bull. Furthermore, TPPs of 45 and 48 kDa were cBiMPS-dependent in fertile bulls whereas, in subfertile bull the latter was barely detectable and the former was cBiMPS-independent at only 0 min. This increase in PTP not only emphasizing the beneficial roles of desalted SP but excluding any detrimental effect of it on sperm cell functions during storage as well