OBJECTIVE: To examine whether human primary spermatocytes will develop into round spermatids after completing meiosis in an in vitro coculture with Vero cells. DESIGN: Prospective, controlled in vitro study. SETTING: A private infertility clinic and a university laboratory. PATIENT(S): Five azoospermic men whose spermatogenesis was proved to be arrested at the level of the primary spermatocyte in open biopsies. INTERVENTION(S): In vitro coculture of isolated primary spermatocytes with Vero cells and chromosomal analysis for assessment of the completion of meiosis. MAIN OUTCOME MEASURE(S): Isolated primary spermatocytes were cocultured with Vero cells under various conditions. The number of chromosomes and chromatids in newly generated cells was determined by Giemsa staining after the cells were injected into mouse oocytes. RESULT(S): The generation rates of round spermatids in six types of in vitro culture with Vero cells were 0%-10% (highest rates of division were in minimum essential medium (MEM) + 50% boar rete testicular fluid or in human synthetic oviduct fluid + 10% human serum). The number of chromosomes and chromatids in the newly developed cells was 23. CONCLUSION(S): A single primary spermatocyte was observed to divide into four cells during in vitro coculture with Vero cells. These newly developed cells were proved to be round spermatids by chromosomal analysis. It was verified that a primary spermatocyte developed into round spermatids after completing two cycles of meiosis through in vitro culture.
OBJECTIVE: To examine whether human primary spermatocytes will develop into round spermatids after completing meiosis in an in vitro coculture with Vero cells. DESIGN: Prospective, controlled in vitro study. SETTING: A private infertility clinic and a university laboratory. PATIENT(S): Five azoospermic men whose spermatogenesis was proved to be arrested at the level of the primary spermatocyte in open biopsies. INTERVENTION(S): In vitro coculture of isolated primary spermatocytes with Vero cells and chromosomal analysis for assessment of the completion of meiosis. MAIN OUTCOME MEASURE(S): Isolated primary spermatocytes were cocultured with Vero cells under various conditions. The number of chromosomes and chromatids in newly generated cells was determined by Giemsa staining after the cells were injected into mouse oocytes. RESULT(S): The generation rates of round spermatids in six types of in vitro culture with Vero cells were 0%-10% (highest rates of division were in minimum essential medium (MEM) + 50% boar rete testicular fluid or in human synthetic oviduct fluid + 10% human serum). The number of chromosomes and chromatids in the newly developed cells was 23. CONCLUSION(S): A single primary spermatocyte was observed to divide into four cells during in vitro coculture with Vero cells. These newly developed cells were proved to be round spermatids by chromosomal analysis. It was verified that a primary spermatocyte developed into round spermatids after completing two cycles of meiosis through in vitro culture.