Reproduction Abstracts

Introduction: Spermatogenesis is a dynamic process. Its control mechanisms are very complex and many molecules are concerned. Our previous study showed that spermatogonia are abnormally proliferated when p53 (a tumor-suppressor gene) was knocked out in medaka (p53-KO medaka). The p53-KO medaka provides a hint about the regulatory mechanisms of spermatogonial proliferation and differentiation.

Materials and Methods: Since some cytokines have been known as the growth factors of spermatogonia, we analyzed mRNA expression of several cytokines in the medaka testis by quantitative PCR. Leukemia inhibitory factor (LIF)-expressing cells were examined in the testis by in situ hybridization and immunostaining.

To investigate whether LIF promotes spermatogonial proliferation, we produced recombinant LIF protein by baculovirus expression system, and added them directly to a culture system that reproduces all processes (from spermatogonia to spermatozoa) of spermatogenesis in vitro, the system we have previously established.

Results and discussion: We found that the mRNA expression levels of LIF, IL-11b, MIF in p53-KO medaka are higher than these in WT medaka. LIF mRNA and protein expression was detected in Sertoli cells, especially in those surrounding type-A spermatogonia (undifferentiated spermatogonia). The expression was also detected in some of type-A spermatogonia but not in type-B spermatogonia (differentiated spermatogonia).

We also found that spermatogonial proliferation is significantly enhanced in a medium supplemented with the recombinant LIF proteins. These findings reveal that LIF promotes spermatogonial proliferation in the medaka testes, suggesting that LIF functions as a growth factor of spermatogonia in medaka.