Damage of seminiferous tubules in mouse testis caused by active immunization of actin-like protein 7a / 中国医学科学院学报

<p><b>OBJECTIVE</b>To obtain recombinant sperm-protein actin-like protein 7a (ACTL7a) and detect the damage seminiferous tubules in mouse testis caused by anti-sperm antibodies generated by purified ACTL7a active immunization.</p><p><b>METHODS</b>The recombinant expression plasmid pET30a-ACTL7a was constructed and then transformed into E. coli Rosseta (DE3). The protein expression was induced by isopropyl β-D-1-thiogalactopyranoside (IPTG), and the protein was purified by nickel ions chelating resin. Finally, the protein was separated by sodium dodecyl sulfate poly-acrylamide gel electrophoresis (SDS-PAGE) and harvested by excising the gel containing target. ICR (Institute of Cancer Research) mice were then immunized using purified ACTL7a protein. The antibody titers were determined by ELISA and the development of seminiferous tubules after active immunization was stained by PAS staining.</p><p><b>RESULTS</b>Induced by IPTG, the target protein ACTL7a was expressed in E. coli. After purification, it was used to immunize the ICR mice. As shown by PAS staining, spermatid expulsion, pyknotic cells, absence of germ cells, and germ cells degenerated were seen in the seminiferous tubules in the immunized testes.</p><p><b>CONCLUSIONS</b>The ACTL7a prokaryotic expression vector was successfully constructed. High-purity target protein was obtained after induction and purification. After the active immunization with the target protein, the seminiferous tubules in the mouse testes will be severely damaged.</p>

Cloning and characterization of a novel rat gene RSD-7 differentially expressed in testis / 中国医学科学院学报

<p><b>OBJECTIVE</b>To isolate and identify the differentially expressed genes in spermatogenesis for the understanding molecular mechanism of spermatogenesis.</p><p><b>METHODS</b>Screening of the cDNA library, Northern blot, expression and purification in E. coli with GST expression system, immunocytochemical staining of testis sections were used.</p><p><b>RESULTS</b>(1) A cDNA fragment designated as RSD-7 was isolated from rat testis cDNA library. It was 1,238 bp in length, coding a protein of 232 amino acids with the GenBank accession number AF315467. The encoding protein of RSD-7 cDNA had a Ubiquitin-like domain. (2) Northern blot indicated that RSD-7 was uniquely expressed in rat testis, and in the testis RSD-7 emerged on the 30th postnatal day and expressed until 120th postnatal day. (3) Expression and purification of RSD-7 protein in E. coli with GST expression system and were used to obtain anti-RSD-7 antibody. (4) Immunolocalization of RSD-7 in rat testis revealed that it is expressed only in Sertoli cells.</p><p><b>CONCLUSIONS</b>Transcription pattern of RSD-7 and localization of RSD-7 protein in testis have been made, which established the base for the functional study of RSD-7.</p>