الفهرس 
IntroductionOnly 14 pages are availabe for public view
 |  | from | 119 |  |  |
| | | from | 119 | | |
Abstract
Our study intended to determine the effect of sperm capacitation on sperm rheotaxis and sperm kinematics within microfluidics in buffalo cryopreserved semen. We used caffeine and heparin as a capacitation media. Semen straws (n = 24) were obtained from the directorate of veterinary medicine in Assiut Governorate from 3 fertile buffalo bulls. Two Frozen semen straws of the same bull were thawed in a water bath at 37 °C for 30 s and pooled in an epindorf tube then, divided into 3 equal parts, each part was diluted with IVF medium in 1:3 ratio, respectively. After mixing the 3 groups: control (IVF medium only), caffeine (IVF medium + 0.125 Mm), and heparin (IVF medium + 20 µl/ml) were incubated in co2 incubator at 38 ℃ and 5% co2 for 45 minutes.
LASER Microchannel was used to record effect of sperm capacitation with caffeine and heparin. The flow of liquid in the microchannel was induced by hydro-static pressure. Level of liquid in inlet of the microchannel was kept higher than that in the output reservoir by a height different Δh. Average liquid velocity in all experiments reported here was kept at 32 that is mimic natural fluid flow velocity inside female genital tract. All groups were examined for sperm rheotaxis percentage and sperm kinematics using computer assisted sperm analysis (CASA). Videos of sperm cells were taken with an Optika XDS-3 inverted microscope with phase contrast (also at 40 × objectives) coupled to a Tucsen ISH1000 camera at 30 frames per second. Conventional CASA systems were found to be not well suited for studying sperm motion inside microchannels. For this reason, an open source CASA plugin for Image-J40 was used with some modifications.
The following parameters were determined: PR%, velocity parameters which include curvilinear velocity (VCL, µm/s), straight-line velocity (VSL, µm/s) and average path velocity (VAP, μm/s). Progression parameters such as linearity (LIN = VSL/VCL, %) and beat cross frequency (BCF, Hz).
The present study revealed that positive rheotaxis percentage PR% was significantly higher in both caffeine and heparin capacitated sperm samples (PR% = 49.4±1 and 48.8±1, respectively) than control (45.1±1). For velocity parameters: the results revealed that caffeine VCL was significantly higher than heparin (0.01).while there was no significant difference in VCL between caffeine and heparin groups and control samples. On the other hand, the VAP of caffeine group was significantly higher (0.05) than both of heparin and control. Also, VSL was significantly higher in caffeine (p<0.001) and heparin (p<0.01) capacitated sperms in comparison with control. For progression parameters There was no significant difference in linearity between groups while BCF were significantly higher in caffeine and heparin groups (p<0.001) than control.
The present study demonstrated that PR% increased by sperm capacitation using caffeine and heparin.