Transforming growth factor beta3 in the fetal and neonatal rat testis: immunolocalization and effect on fetal Leydig...

The localization of transforming growth factor beta3 (TGFbeta3) in the fetal and neonatal testis (from fetal day 13.5 to postnatal day 6) was investigated by immunohistochemical staining with a specific polyclonal antibody raised against a synthetic peptide corresponding to residues 50-75 of TGFbeta3. This antibody recognized 0.5 ng TGFbeta3 in western blot analysis, but did not detect 25 ng TGFbeta1 or TGFbeta2. The immunolocalization of TGFbeta3 in the fetal and neonatal testis changed throughout development. Immunostaining was present in the gonocytes by fetal day 13.5, persisted until postnatal day 3, and was heterogeneous in spermatogonia on postnatal day 6. The Sertoli cells contained no immunoreactivity at any age. The fetal-type Leydig cells were first immunostained for TGFbeta3 on day 16.5 and staining became very intense from day 18.5 onward. Staining disappeared when the antibody was presaturated with the synthetic peptide, but persisted when the antibody was presaturated with a tenfold excess of the corresponding peptide from TGFbeta2. Furthermore, we researched whether TGFbeta3 could act as a local regulator of fetal Leydig cell function. In a dispersed fetal testicular cell system, TGFbeta3 inhibited the LH-stimulated testosterone production by Leydig cells from 20.5-day-old fetuses. The inhibitory effect of TGFbeta3 was equal to that observed with TGFbeta1 or TGFbeta2. When compared with our previous studies showing the immunolocalization of TGFbeta1 and TGFbeta2, the present study shows that TGFbeta3 may have a specific role in the developing rat testis, but may also overlap the action of TGFbeta1 and TGFbeta2.