Role of IGFs and insulin in the human testis during postnatal activation: differentiation of steroidogenic cells.

Immunoexpression of IGF-I, IGF-II, type 1 IGF receptor (IGFR), insulin receptor (IR), and GH receptor (GHR) was analyzed in human testis, in three age groups (Gr): Gr1 (neonates), Gr2 (postnatal testicular activation), and Gr3 (early prepuberty). In interstitial cells, low IGF-I and GHR, but moderate IR immunoexpression was observed in all Grs. However, high expression of IGF-II in Gr1, and moderate expression of IGFR in Gr1 and Gr2 were found. In Leydig cell (LC), high expression of IGF-II, moderate expression of IGFR and GHR, and undetectable IGF-I was found. Moreover, IR was highly expressed in Gr2. The effect of IGF-I on cell proliferation (PI) and apoptosis (AI), induction of cytochrome P450 side chain cleavage (cP450scc) immunoexpression, 3beta-hydroxysteroid dehydrogenase mRNA and testosterone (T) secretion was evaluated in human testis cell cultures. IGF-I increased P450scc immunoexpression, 3beta-hydroxysteroid dehydrogenase mRNA, T secretion, and PI, but decreased AI. We propose that IGF-II, mainly through IR, is involved in functional LC differentiation. In some interstitial cells, probably in LC precursors, IGF-II/IR could be involved, among other factors, in the stimulation of PI and/or inhibition of AI, and in LC differentiation.

Expression of aromatase, estrogen receptor alpha and beta, androgen receptor, and cytochrome P-450scc in the human early prepubertal testis.

The expression of aromatase, estrogen receptor alpha (ERalpha) and beta (ERbeta), androgen receptor (AR), and cytochrome P-450 side chain cleavage enzyme (cP450scc) was studied in prepubertal testis. Samples were divided in three age groups (GRs): GR1, newborns (1- to 21-d-old neonates, n = 5); GR2, postnatal activation stage (1- to 7-mo-old infants, n = 6); GR3, childhood (12- to 60-mo-old boys, n = 4). Absent or very poor detection of ERalpha by immunohistochemistry in all cells and by mRNA expression was observed. Leydig cells (LCs) of GR1 and GR2 showed strong immunostaining of aromatase and cP450scc but weak staining of ERbeta and AR. Interstitial cells (ICs) and Sertoli cells (SCs) expressed ERbeta, particularly in GR1 and GR2. Strong expression of AR was found in peritubular cells (PCs). For all markers, expression in GR3 was the weakest. In germ cells (GCs), i.e. gonocytes and spermatogonia, aromatase and ERbeta were immunoexpressed strongly whereas no expression of ERalpha, AR, or cP450scc was detected. It is proposed that in newborn and infantile testis, testosterone acting on PCs might modulate infant LC differentiation, whereas the absence of AR in SCs prevents development of spermatogenesis. The role of estrogen is less clear, but it could modulate the preservation of an adequate pool of precursor LCs and GCs.

Apoptosis and proliferation of human testicular somatic and germ cells during prepuberty: high rate of testicular growth in newborns mediated by decreased apoptosis.

Programmed cell death and proliferation are evolutionary conserved processes that play a major role during normal development and homeostasis. In the testis, during the fetal and newborn periods, they might determine final adult size and fertility potential. In the present study, we have measured the relative number of testicular cells in apoptosis and in active proliferation in the seminiferous cords and in the interstitium, at different age periods of prepubertal testicular development in humans. Testes from 44 prepubertal subjects without endocrine and metabolic abnormalities were collected at necropsy. They were divided in three age groups (Gr): Gr 1, newborn (1- to 21-d-old neonates), n = 18, mean (+/-SD) age 0.3 +/- 0.23 months; Gr 2, post natal activation (1- to 6-month-old infants), n = 13, mean age 3.93 +/- 1.90 months; and Gr 3, early childhood period (1- to <6-yr-old boys), n = 13, mean age 31.5 +/- 18.9 months. Apoptosis was detected in 5- microm tissue sections using a modified terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling assay and cell proliferation was assessed by Ki-67 immunohistochemistry. Evaluation of apoptosis was confirmed by estimation of active caspase-3. Mean (+/-SD) testicular weight was 0.38 +/- 0.20, 0.54 +/- 0.35, and 0.51 +/- 0.11 g in Gr 1, Gr 2, and Gr 3, respectively. In Gr 1, there was a significant positive correlation between age and testis weight (P = 0.02). Mean (+/-SD) germ cell apoptotic index, AI, (% of apoptotic cells out of total cell number) was 15.0 +/- 6.60, 27.0 +/- 8.80 and 33.4 +/- 11.4 in Gr 1, Gr 2, and Gr 3, respectively. In Sertoli cells, it was 6.60 +/- 4.07, 22.0 +/- 14.0 and 27.5 +/- 19.8, respectively. In interstitial cells, it was 10.2 +/- 6.38, 18.0 +/- 6.70 and 25.7 +/- 15.5, respectively. In the three types of cells, AI in Gr 1 was significantly lower than in Gr 2 or Gr 3 (P < 0.05). Mean (+/-SD) germ cell proliferation index, PI, was 18.6 +/- 13.0, 10.0 +/- 6.50 and 10.9 +/- 6.24% in Gr 1, Gr 2, and Gr 3, respectively. In Sertoli cells and in interstitial cells PI was similar in the three age groups. The PI/AI ratio was used to compare relative differences among age groups. The PI/AI ratio of germ cells, Sertoli cells and interstitial cells in Gr 1 was significantly higher than in Gr 2 or Gr 3 (P < 0.05). It is concluded that, in normal subjects, there is a vigorous growth of the testis during the newborn period with subsequent stabilization during the first years of prepuberty. This cell growth seems to be mainly mediated by decreased apoptosis. The factors that modulate apoptosis of testicular cells are not known, but it is remarkable that this change takes place before the testosterone peak of the post natal gonadal activation of the first trimester of life. These changes taking place during the newborn period might be important to define testicular function in adults.