Postnatal development and testosterone dependence of a rat epididymal protein identified by neonatal tolerization.

A rat epididymal protein of 27 kDa was identified using neonatal tolerization. This study reports the production and characterization of a polyclonal antiserum to this protein. ELISA was used to demonstrate that this antiserum reacts strongly with epididymal sperm proteins, but has little or no reactivity with testicular proteins. Western blot analysis revealed that this polyclonal antiserum recognized a 27 kDa protein extracted from the corpus epididymidis as well as from spermatozoa from the corpus and cauda epididymides, and immunostaining revealed the presence of the protein in the corpus to cauda epididymides. Stronger reactivity was observed in the supranuclear region and stereocilla of principal cells of the corpus epididymidis and in the luminal content of the corpus and cauda epididymides. The testicular section showed no reactivity. Treatment with the antiserum resulted in time- and dose-dependent agglutination of rat spermatozoa. By indirect immunofluorescence, the antiserum localized proteins in the mid-piece region of rat spermatozoa. Studies were carried out to determine the age at which the protein first became apparent during postnatal development. The protein was expressed from day 40 onwards, as demonstrated by western blot analysis. The androgen regulation of this protein was ascertained by castration and supplementation studies. Expression of this protein showed a decline starting at day 14 after castration and by day 21 the protein was absent; however, androgen replacement resulted in the reappearance of the protein. The results of these studies indicate that the protein identified is specific to the epididymis, and is regulated by development and androgens. The importance of epididymis-specific proteins that are regulated by androgens in sperm maturation is discussed, and the need to ascertain the sequence of the protein and clone the cognate gene is indicated.

Immunoregulatory pathways in pregnancy.

Murine pregnancy is characterized by transient thymic atrophy and splenomegally. Several laboratories are investigating the immunoregulatory mechanisms during pregnancy, and the majority of these studies are primarily focused on the immunological changes either in the uterus or the thymus and not much information is available on the immunological changes in the spleen that result in transient splenomegally. An attempt has been made in this review to understand the significance of thymic atrophy, splenomegally and local immune changes in the uterus to understand the overall immunomodulatory mechanisms in pregnant mother. The most significant change which occurs soon after mating is the infiltration of immune cells such as macrophages and gammadelta-T cells into the uterus indicating that the mother's immune system detects the presence of foreign antigens in the reproductive tract. The sensitized cells appear to migrate to the secondary lymphoid organs including the spleen. The microenvironment in the spleen is conducive for the cell-cell contact and generation of immune response. The major changes that occur in the spleen are, the induction of T-cell dependent B-cell response on day-1 post-coitum (P.C.), generation of antibody producing B-cells on day-3 and also proliferation of CD8+ T-cells that peaks on day-3 of pregnancy. The weight of the spleen reaches a peak on day-10 in mice. Thereafter, on day-15 of pregnancy, lymphocyte apoptosis is seen in the spleen indicating the deletion of peripheral sensitized cells. This results in decrease in spleen weight to that of normal non-pregnant mice. The decrease in thymic weight after day-5 pregnancy was associated with the increased apoptosis of cortical thymocytes. This perhaps is due to negative selection of self-reactive thymocytes. Our studies have demonstrated that the pregnancy associated monoclonal antibodies react with antigens of sperm indicating that the mother's immune system recognizes and responds to the constituents of the semen to produce non-precipitating asymmetric auto antibodies (NPAA) or blocking antibodies which have favourable effects on pregnancy. It is postulated that the mother's immune response could be directed to some antigens of sperm along with some conserved antigens such as heat shock proteins (HSP) that are present both in sperm and in the mother. It may be speculated that after the initial priming to some conserved antigens of sperm and due to the presence of similar antigens in the mother, these activated clones are eliminated both in the primary and secondary lymphoid organs to prevent autoimmunity in the mother during pregnancy.