Autocrine/Paracrine Insulin-like Growth Factor Binding Protein-3 Acts asPro-apoptotic Factor for Leydig cells in the Rat Testis

The secretory insulin-like growth factor-binding protein-3 (IGFBP-3) induces apoptosis via both insulin-like growth factor-I (IGF-I)-dependent and -independent mechanisms. Here, we have examined the effects of IGFBP-3 on Leydig cell apoptosis, proliferation and steroidogenesis. Immunohistochemical analysis of testes of rats at different ages revealed that IGFBP-3 is expressed first after 20 days of postnatal life and is present at a high level in the adult testis. In addition, Western blotting showed that the expression of IGFBP-3 in Leydig cells isolated from 60-day-old rats is higher than in 40-day-old animals. The rate of DNA synthesis (as assessed by incorporation of 3H-thymidine in vitro) in Leydig cells from 40-day-old rats is reduced by IGFBP-3, which also blocks the promotion of cell survival by IGF-I. Moreover, IGFBP-3 induces apoptosis in Leydig cells and, at the same time, attenuates the anti-apoptotic action of IGF-I. Furthermore, IGF-I stimulates secretion of IGFBP-3, -4, and -2 by Leydig cells. The pro-inflammatory cytokine tumor necrosis factor- α induces apoptosis in these same cells and increases their secretion of IGFBP-3 and IGFBP-4. These findings provide the first evidence that IGFBP-3 acts as a pro-apoptotic effector of Leydig cells and can also block the positive effect of IGF-I on cell survival. In addition, IGFBPs appear to modulate interactions between IGF-I and pro-inflammatory cytokines in the testis, suggesting possible participation of these proteins in processes such as testicular inflammation and cancer.