Ontogenesis of Fas ligand expression in the rat testis

We recently demonstrated that the expression of the Fas/Fas ligand (FasL) system, in the human testis, is developmentally regulated and possibly under gonadotropins control (J. Clin. Endocrinol. Metab. 85: 2692, 2000). In the present study, we investigated the ontogenesis and tissue distribution of FasL expression in rat testis from 18 days post coitum up to 1 year of age by means of semiquantitative RT-PCR, in situ RT-PCR and immunohystochemistry. The in vitro effects of ovine FSH (oFSH) treatment on the expression of FasL in cultured prepubertal Sertoli cells was also evaluated by means of RT-PCR and Western blotting. Semiquantitative RT-PCR experiments were performed on total RNA extracted from rat testis of different age where FasL was coamplified with -actin as internal control. The results of these experiments showed the expression of FasL mRNA at all age analyzed, being 2-3 fold higher at 36 days and 1 year of age. In situ RT-PCR, performed on 4 m section of 4% buffered formalin fixed and paraffin embedded tissue, showed that the expression of FasL mRNA was localized in the Leydig and Sertoli cells from early postnatal days to adult. No expression of FasL mRNA could be detected in germ cells at any stage of development. Immunohystochemistry experiments, performed on 5 m section of Bouin’s fixed and paraffin embedded testis, showed that FasL protein was present in Leydig cells from 2 days of age up to the adult. The expression of FasL protein in Sertoli cells become to be appreciated at 15 days of age to increase at 36 days and 1 year of age. Finally, treatment of prepubertal Sertoli cells with oFSH (200 ng/ml for 18 h) did not affect FasL expression, at both protein and mRNA level. In conclusion, these data appear to confirm a physiological role of the Fas/FasL system in male gonad development and homeostasis.