[Damage of seminiferous tubules in mouse testis caused by active immunization of actin-like protein 7a].

OBJECTIVE

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.

METHODS

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.

RESULTS

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.

CONCLUSIONS

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.