Expression and Effect of Insulin-Like Growth Factor I on Rat Fetal Leydig Cell Function and Differentiation.

Insulin like growth factor I (IGF-I) is believed to be a potent para/autocrine stimulator of Leydig cell function in adult testis. We investigated whether IGF-I is also an intratesticular regulator of fetal Leydig cell function by measuring its production in the fetal testis and its ability to affect testicular steroidogenesis during fetal development. Northern blot analysis revealed one major IGF-I transcript of 7-7.5 kb and two minor transcripts of 3.8 and 1.8 kb in 20.5 day fetal testis. IGF-I was detected by RIA in 16.5 fetal day testes, and the amounts of IGF-I secreted by 16.5 and 20.5 fetal day testes in vitro were much greater than the amounts contained in the testes, indicating active synthesis in culture. The secretion of IGF-I by the fetal testis in vitro was increased with testicular age and time in culture. It was not modified by gonadotropins or (Bu)2cAMP. Testosterone secretion by fetal testes explanted 13.5, 16.5, 18.5, and 20.5 days after conception and cultured in the presence or absence of 100 ng/ml LH for 3 days was not affected by the addition of 50 ng/ml IGF-I to the medium. In contrast, the addition of IGF-I to dispersed fetal testicular cells cultured for 3 days in the presence or absence of LH increased the number of Leydig cells identified by a positive cytochemical reaction for 3beta-hydroxysteroid dehydrogenase (3betaHSD). This was more pronounced with cells from 16.5- day-old fetuses (stage when the fetal Leydig cells are differentiating in vivo) than with 20.5-day-old fetuses cells (stage when the number and the function of fetal Leydig cells are stable or decreasing). It results from both an increased differentiation of mesenchymal cells in fetal Leydig cells and an increase in the mitotic index of the fetal Leydig cells, as inferred from the small increase in the percentage of bromodeoxyuridine/3betaHSD-positive cells. Both LH and IGF-I increased significantly testosterone production by day 16.5 cells. In the presence of LH, a high amount of testosterone was produced per 3betaHSD-positive cell; IGF-I further increased this production. This effect was not observed with day 20.5 cells. The amounts of testosterone produced per 3betaHSD-positive cell cultured in the presence of both LH and IGF-I were more than additive. Like IGF-I, insulin (50 ng/ml) increased testosterone secretion per 3betaHSD-positive cells in cultures of day 16.5 cells, but not in those of day 20.5, cells. Lastly, IGF-I also increased the steroidogenic activity of each Leydig cell in cultures containing (Bu)2cAMP, but its effects were weaker than those observed in the presence of LH. This suggests that IGF-I has sites of action both upstream and downstream cAMP generation. These results suggest that IGF-I acts as paracrine/autocrine factor in the differentiation and activity of fetal Leydig cells.