Missing Of The H-Y Antigen MRNA In Human Ejaculate Spermatozoa And Possible Function Of Spermatozoal MRNA

The spermatozoon is a highly differentiated terminal cell which transports the paternal genome to the oocyte. It is generally accepted that nuclear gene expression in mature spermatozoa is progressively shutdown during spermiogenesis. While the spermatozoon nucleus seems transcriptionally silent, more recent reports indicate that these cells carry a full complement of mRNAs and that under certain conditions, spermatozoa can translate their mRNAs de novo. This research aimed at whether H-Y antigen mRNA is present in spermatozoal RNA, in order to find out retention mechanism of spermatozoal RNA and its possible function.In our research, human ejaculates were obtained from healthy volunteers. The ejaculate was purified by swim-up technique to get rid of somatic contaminations. Total RNA of purified spermatozoa was extracted by Trizol RNA isolation reagent. In order to determine whether it had effect on H-Y antigen on the membrane of Y-sperm, the IgY polyclonal antibody against H-Y antigen we had obtained was tested by immunohistochemistry and flow cytometry. We also examined whether mRNA from ejaculate spermatozoa contained H-Y antigen mRNA by RT-PCR analysis.By immunohistochemistry, it is clearly shown many spermatozoa were stained by fluorescence-conjugated second antibody. Through flow cytometric analysis, the positive H-Y antigen percentage of human sperm reached at 49.86%. And RT-PCR analysis reported that there was no H-Y antigen-specific band found in the tests. These results approved the presence of H-Y antigen on the membrane of Y-sperm, but mRNA of H-Y antigen is not present in mRNAs persists in the ejaculate spermatozoa. These support that H-Y antigen express on the early stage of spermiogenesis and implicate that spermatozoal RNA is not only residual during spermiogenesis procedure, but also performs an essential function on delivery to the oocyte.