Abstract: Objective To establish an in vitro model of cultured mouse testis using rotary aerobic culture. Methods Rotary
aerobic incubation with optimized culture conditions was used for in vitro culture of mouse testis, and the morphology of the
cultured testicular tissues was compared with that cultured in Transwell chambers. The changes in the testicular tissue
structure were examined using HE staining, and the cell proliferation was assessed with BrdU staining. Testosterone
concentrations in the culture medium were tested with radioimmunoassay and the expression of the functionally related
proteins in the testis was detected using immunohistochemistry. Results The testicular tissue cultured by optimized rotary
aerobic culture presented with more intact histological structure with the size of the testis ranged from 0.3 to 0.8 mm3. In the
two culture systems, the prolifeation index of the spermatogonia increased and that of Sertoli cells decreased with time, and
such changes in spermatogonia and Sertoli cell proliferation indices became statistically significant at 3 days (P<0.05) and 5
days (P<0.05) of culture, respectively, as compared with those at 1 day. The concentration of testoerone in the culture media
decreased significantly with incubation time (P<0.05). At 3 days of culture, the protein expression of 3β-hydroxysteroid
dehydrogenase, cytochrome P450 17α-hydroxylase and cholesterol side-chain cleavage enzyme was detected in Leydig cell
cytoplasm and vimentin expression in Sertoli cell cytoplasm. Conclusion An in vitro model of cultured mouse testis has been
successfully established using rotary aerobic incubation.