Epsilon Archive for Student Projects

Abstract

The objective of this study was to assess the potential of Hyaluronan Binding Assay (HBA) to predict freezability and fertilization capacity of dromedary camel spermatozoa, and to determine if conventional sperm quality parameters and precursor of A-Kinase Anchoring Protein 4 (proAKAP4) values correlate with HBA results.
Two semen samples were collected from each of six male camels. In experiment 1, the samples were assessed fresh (fresh), pre-freezing (PF) and post-thawing at 0 h (PT0H), and post-thawing at 1.5 h (PT1.5H). HBA slides coated with molecular layer of hyaluronan were used together with a proAKAP4 biomarker-test to validate semen quality of fresh samples and PT0H. Total motility (TM), Progressive motility (PM), motility-related kinematics, morphological abnormalities (MA), vitality (VIT), intact acrosome (IA) and mitochondrial membrane potential (MIT) were evaluated in fresh and PT0H and PT1.5H sperm to find possible correlations with HBA test results. In experiment 2, fertilizing ability of PT0H sperm was evaluated with a heterologous sperm penetration assay (SPA) using zona pellucida free goat oocytes resulting in values for number of oocytes (N), penetration rate (PEN), formation of male pro-nucleus (PN) and number of sperm penetrated per oocyte (SP/OC).
The results showed that dromedary camel spermatozoa bound to hyaluronan; there were no significant differences between males. However, there was no correlation between HBA in fresh sample and PT0H results. Furthermore, the proAKAP4 test results also did not differ between males and did not correlate with HBA results, with the exception of PT0H HBA and proAKAP4 results (r = - 0.62; p-value 0.03). Of the conventional semen analysis parameters (TM, PM, VIT, IA, MIT, Volume, Viscosity, Concentration and MA) none were significantly different between males and only PM correlated with the HBA results in fresh sperm r = 0.65; (p-value 0.02). From LSMean values of computer-assisted semen analysis (CASA) kinematics of fresh spermatozoa, beat cross frequency (p-value 0.01), straightness (p-value 0.01) and curvilinear velocity (p-value 0.05) showed significant variation between males. From the PT0H samples, the lateral head deviation (p-value 0.01), average path velocity (p-value 0.04) and curvilinear velocity (p-value 0.02) varied between males, as did PT1.5H linearity (p-value 0.05). The kinematic straightness correlated with the HBA result for fresh semen (r = 0.69; p-value 0.01). The analysis for freezability was done using HBA results (fresh and PT0H) and post thaw values of different parameters (TM, PM, VIT, IA, MIT, PEN, PN and SP/OC); no significant correlations were found. In the SPA, dromedary camel sperm bound, penetrated, decondensed, and completed pro-nucleus formation in goat oocytes. Significant correlations were identified between fresh sperm HBA result and PEN (p-value 0.03) as well as with PN (p-value 0.03). The proAKAP4 test results from fresh sperm samples and PT0H results did not correlate with penetration assay results. There was an association between proAKAP4 and HBA at PT0H. In conclusion, the results suggested that the HBA score from fresh dromedary camel sperm may predict post-thaw IVF performance, but further investigation is needed.