| Over the past 60 years, egg yolk has been included as a protective agent in extender used to freeze semen. Recently, there has been increased interest in using extender free of animal products to freeze semen. However, it is difficult to find suitable component in order to replace egg yolk in the extender because the mechanisms involved in sperm protection by egg yolk against damages caused by storage, cooling and freezing remain unclear.;Our studies showed that BSP proteins can bind the low-density lipoprotein fraction (LDF) from egg yolk. This binding of BSP proteins to LDF is rapid, saturable and specific and the binding capacity of LDF is very high. Furthermore, this interaction is stable even after freeze-thawing. Our studies also showed that the interaction between LDF and BSP proteins prevents BSP proteins from binding to sperm and stimulating lipid efflux from the sperm membrane while maintaining sperm motility during sperm storage. These results indicated that egg yolk protects sperm by preventing the stimulation of lipid efflux from the sperm membrane by BSP proteins.;In addition, we verified whether or not deleterious factors (BSP proteins) are present in the SP of other mammalian species. By gelatin- and heparin-affinity chromatography, we isolated four BSP protein homologs from bison SP (BiSV-16kDa, BiSV-17kDa, BiSV-18kDa and BiSV-28kDa) and ram SP (RSP-15kDa, RSP-16kDa, RSP-22kDa and RSP-24kDa). Furthermore, we showed that BiSV and RSP proteins bind to egg yolk LDF. These results confirm that BSP protein homologs are ubiquitous among mammals and suggest that the mechanism of sperm protection by egg yolk is the same in those species.;We believe that the results described in this thesis will aid in the development of novel extenders that do not contain animal products for sperm preservation in liquid or frozen state.;Upon ejaculation, epididymal sperm are mixed with seminal plasma (SP). The BSP (Bovine Seminal Plasma) proteins are the major proteins of bull SP and they stimulate cholesterol and phospholipid efflux from the sperm membrane, which may be detrimental for sperm. We hypothesized that egg yolk protects sperm by binding BSP proteins and preventing them from stimulating lipid efflux from the sperm membrane. |
