الفهرس 
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Abstract
The present study was carried out to investigate: a) Effect of progesterone and/or cyclodextrin on capacitaion and acrosome reaction of bovine spermatozoa during in vitro and b) Effect of capacitated bovine spermatozoa with progesterone and/or cyclodextrin on fertilization rate of bovine oocytes.
In order to achieve that, the following steps were done:
Two types of Nanoparticles were prepared and used in this study, Progesterone Nnaoparticles and Cyclodextrin Nanoparticles
The z-average particle sizes, Zeta potential analysis and Yield of nanoparticles were determined for each formula.
Sperm capacitation was done by thawing of frozen bovin spermatozoa and swim-up technique to separate of motile sperms and then incubated in at 38ºC for 3h in fertilization medium (IVF-TALP) (Control) or Progesterone nanoparticles (PN8) or Cyclodextrin nanoparticles (CDN1, CDN2,CDN3, CDN4) or Progesterone or Cyclodextrin.
Measurement of the hyperactivation motility of spermatozoa, sperm Viability and acrosome reaction were recorded.
Transmission Electron Microscope Analysis of Bovine spermatozoa treated with different Nanoparticles were determined.
Cattle ovaries were collected from slaughterhouse and transported to the laboratory within 2h.
On the laboratory, follicles aspirated from the ovaries and number and state of cumulus-oocyte-complexes (COCs) were recorded.
Oocytes were cultured in each droplet (100 µl) of the maturation medium in a sterile Petri-dish which was covered with mineral oil and incubated for 22-24 h in CO2 incubator (5% CO2) at 39°C and 95% humidity.
The rate of oocyte maturation was evaluated after 22- 24h of incubation by cumulus expansion.
The matured oocytes were washed three times in IVF-TALP and then transferred to droplets of IVF-TALP medium (5 oocytes/100 µl) and incubated in the CO2 incubator for 10 to 24h at 38.5ºC (Control) or inseminated with sperm suspension of best formula of nanopaticles.
The fertilization rate of bovine oocytes was recoreded upon the presence of second polar bodies, 2 pronuclei or cleavage to at least the 2-cell stage.
The obtained data were statistically analyzed using analysis of variance (ANOVA) and least significant difference test (LSD).
The results obtained from this work could be summarized as follows:
Bovine spermatozoa incubated with progesterone nanoparticles at formula of PN8 for 2h revealed increased sperm hyper-motility, viability and acrosome reaction as compared to progesterone or control (spermatozoa without nanoparticles).
Bovine spermatozoa incubated with cyclodextrin nanoparticles at formula of CDN2 for 2h revealed increased sperm hyper-motility and acrosome reaction as compared to others cyclodextrin anoparticles formulae or cyclodextrin or control (spermatozoa without nanoparticles).
Bovine spermatozoa incubated with progesterone and/or cyclodextrin nanoparticles at formula PN8+CDN2 for 2h revealed increased sperm hyper-motility, viability and acrosome reaction as compared to progesterone or cyclodextrin nanoparticles alone or progesterone or cyclodextrin or control (spermatozoa without nanoparticles).
Transmission electron microscopic analysis of spermatozoa incubated with Progesterone and/or cyclodextrin nanoparticles at PN8+CDN2 for 2h revealed presence of nanoparticles both inside the sperm head and tail region.
Bovine spermatozoa treated with cyclodextrin (CDN4) ananoparticles significant deccreased the capacitiaiton and acrosome reaction during the in vitro sperm capacitation at 1, 2 and 3 incubation periods.
Capacitaiton of bovine spermatozoa with progesterone nanoparticles at formula of PN8 produced the highest percentage of fertilized oocytes.
Capacitaiton of bovine spermatozoa with cyclodextrin ananoparticles at formula of CDN2 produced the highest percentage of fertilized oocytes.
Capacitaiton of bovine spermatozoa with progesterone plus cyclodextrin ananoparticles at formula of PN8+CDN2 produced the highest percentage of fertilized oocytes.
The main findings of this thesis are summarised as follows:
We report here that bovine sperm incubated in medium containing both progesterone and/or cyclodextrin nanoparticles have significantly improved hyper-motility and acrosome reaction when compared to sperm incubated without progesterone or cyclodextrin nanparticles or with progesterone or cyclodextrine alone. However, hypermotility and acrosome reaction was significant increased in sperm capacition by addition both progesterone and cyclodextrin when compared to sperm with either component alone. Similarly, Treatment with progesterone plus cyclodextrin nanoparticles has significantly improved the fertilization rate of bovine oocytes.
Collectively, our study has demonstrated that progesterone plus cyclodextrin nanoparticles are the best materials to increase the capacitation of sperms and in vitro fertization in cattle and then cyclodextrin nanoparticles and then progesterone nanoparticles compared to with or without cyclodextrin or progesterone. Also, the study showed that lower concentrations are better than higher concentrations. Therefore, we recommend the addition of PN8 and /or CDN2 to the fertilization medium.
So this study is considered apromise for dissolving many problemes associated with In Vito Fertilization due to bad quality of sperm.
Further studies will be necessary to elucidate the specific biochemical and molecular changes which result from the action of nanoparticles in the capacitation process of bovine sperm.